Patent application title: CHIMERIC RECEPTORS AND METHODS OF USE THEREOF
Inventors:
Jed Wiltzius (Woodland Hills, CA, US)
Ruben Alvarez Rodriguez (Los Angeles, CA, US)
Alice Bakker (Cupertino, CA, US)
Lawren Wu (Foster City, CA, US)
Lawren Wu (Foster City, CA, US)
Tara Arvedson (Moss Beach, CA, US)
IPC8 Class: AC07K1628FI
USPC Class:
1 1
Class name:
Publication date: 2020-04-16
Patent application number: 20200115457
Abstract:
Antigen binding molecules, chimeric receptors, and engineered immune
cells are disclosed in accordance with the invention. The invention
further relates to vectors, compositions, and methods of treatment and/or
detection using the antigen binding molecules and engineered immune
cells.Claims:
1-97. (canceled)
98. A chimeric antigen receptor that binds to C-type lectin-like-1 ("CLL-1"), wherein the chimeric antigen receptor is a polypeptide comprising a single chain Fv (scFv) that binds CLL-1, a transmembrane domain, and an intracellular activating domain, and wherein the scFv comprises a heavy chain variable region (VH) and light chain variable region (VL) comprising: (i) a VH region comprising complementarity determining regions ("CDRs") 1, 2, and 3 with amino acid sequences SEQ ID NO: 95, SEQ ID NO: 52, and SEQ ID NO: 75, respectively; and (ii) a VL region comprising CDRs 1, 2, and 3 with amino acid sequences SEQ ID NO: 78, SEQ ID NO: 57, and SEQ ID NO: 102, respectively.
Description:
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation of U.S. patent application Ser. No. 15/476,699, filed Mar. 31, 2017 and claims the benefit of U.S. Provisional Patent Application 62/317,068, filed Apr. 1, 2016, both of which are hereby incorporated by reference in their entirety.
REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY
[0002] The instant application contains a Sequence Listing which has been submitted electronically in ASCII format and is hereby incorporated by reference in its entirety. Said ASCII copy, created on Nov. 9, 2018, is named K-1029_02US_SL.txt and is 274,746 bytes in size.
BACKGROUND OF THE INVENTION
[0003] C-type lectin-like-1 (CLL-1, also known as CLEC-1, CLEC12A, MICL, Dendritic Cell-Associated Lectin-1 (DCAL-1), and DCAL-2) is a glycoprotein receptor and member of a family of C-type lectin-like receptors involved in the regulation of cell proliferation and immune regulation. CLL-1 is expressed in hematopoietic cells, primarily on innate immune cells including monocytes, granulocytes, dendritic cells, as well as myeloid progenitor cells. Van Rhenen et al., Blood 2007:110(7). CLL-1 has been implicated in the regulation of myeloid cell proliferation and differentiation (Bakker et al., Cancer Res. 64:8443-8450 (2004); Marshall et al., J. Biol. Chem. 279:14792-14802 (2004)), and is present on acute myeloid (myelogenous) leukemia (AML) cells as well as on leukemic stem cells (Zhao et al., Haematologica 2010, 95(1):71-78).
[0004] Accordingly, CLL-1 has been implicated in multiple diseases, including but not limited to, acute myeloid (myelogenous) leukemia (AML), chronic myeloid (myelogenous) leukemia (CML), chronic myelomonocytic leukemia (CMML), juvenile myelomonocytic leukemia, atypical chronic myeloid leukemia, acute promyelocytic leukemia (APL), acute monocytic leukemia, acute monoblastic leukemia, acute erythroid leukemia, acute megakaryoblastic leukemia, myelodysplastic syndrome (MDS), myeloproliferative disorder, myeloid neoplasm, myeloid sarcoma), Blastic Plasmacytoid Dendritic Cell Neoplasm (BPDCN), or combinations thereof.
[0005] CLL-1 may additionally play a role in inflammatory or autoimmune diseases such as rheumatoid arthritis, psoriasis, allergies, asthma, Crohn's disease, IBD, IBS, fibromyalga, mastocytosis, and Celiac disease.
[0006] Human CLL-1 protein comprises a polypeptide of the following amino acid sequence:
TABLE-US-00001 (SEQ ID NO. 140) MSEEVTYADLQFQNSSEMEKIPEIGKFGEKAPPAPSHVWRPAALFLTLLC LLLLIGLGVLASMFHVTLKIEMKKMNKLQNISEELQRNISLQLMSNMNIS NKIRNLSTTLQTIATKLCRELYSKEQEHKCKPCPRRWIWHKDSCYFLSDD VQTWQESKMACAAQNASLLKINNKNALEFIKSQSRSYDYWLGLSPEEDST RGMRVDNIINSSAWVIRNAPDLNNMYCGYINRLYVQYYHCTYKKRMICEK MANPVQLGSTYFREA.
[0007] Additional sequence information is contained in the CLL-1 Uniprot listing at: www.uniprot.org/uniprot/Q5QGZ9, as well as NCBI Reference Sequence NP_612210.4 (www.ncbi.nlm.nih.gov/protein/NP_612210.4).
[0008] When referring to CLL-1, it will be appreciated that reference thereto encompasses fragments thereof, as well as related polypeptides, which include, but are not limited to, allelic variants, splice variants, derivative variants, substitution variants, deletion variants, and/or insertion variants including the addition of an N-terminal methionine, fusion polypeptides, and interspecies homologs. In certain embodiments, a CLL-1 polypeptide includes terminal residues, such as, but not limited to, leader sequence residues, targeting residues, amino terminal methionine residues, lysine residues, tag residues and/or fusion protein residues.
[0009] Certain antibodies to CLL-1 are described in U.S. Pat. No. 8,536,310 and in U.S. Pat. No. 9,163,090.
[0010] Engineered immune cells have been shown to possess desired qualities in therapeutic treatments, particularly in oncology. Two main types of engineered immune cells are those that contain chimeric antigen receptors (termed "CARs" or "CAR-Ts") and T-cell receptors ("TCRs"). These engineered cells are engineered to endow them with antigen specificity while retaining or enhancing their ability to recognize and kill a target cell. Chimeric antigen receptors may comprise, for example, (i) an antigen-specific component ("antigen binding molecule"), (ii) an extracellular domain, (iii) one or more costimulatory domains, and (iv) one or more activating domains. Each domain may be heterogeneous, that is, comprised of sequences derived from (or corresponding to) different protein chains. Chimeric antigen receptor-expressing immune cells (such as T cells) may be used in various therapies, including cancer therapies. It will be appreciated that costimulating domains may be used to enhance the activation of CAR-expressing cells against target antigens, and therefore increase the potency of adoptive immunotherapy.
[0011] Certain CARs to CLL-1 have been described in, e.g., U.S. Patent Application 20160051651 (PCT US2015/041337).
[0012] T cells can be engineered to possess specificity to one or more desired targets. For example, T cells can be transduced with DNA or other genetic material encoding an antigen binding molecule, such as one or more single chain variable fragment ("scFv") of an antibody, in conjunction with one or more signaling molecules, and/or one or more activating domains, such as CD3 zeta.
[0013] In addition to the CAR-T cells' ability to recognize and destroy the targeted cells, successful T cell therapy benefits from the CAR-T cells' ability to persist and maintain the ability to proliferate in response to antigen.
[0014] T cell receptors (TCRs) are molecules found on the surface of T cells that are responsible for recognizing antigen fragments as peptides bound to major histocompatibility complex (MHC) molecules. The TCR is comprised of two different protein chains--in approximately 95% of human TCRs, the TCR consists of an alpha (.alpha.) and beta (.beta.) chain. In approximately 5% of human T cells the TCR consists of gamma and delta (.gamma./.delta.) chains. Each chain is composed of two extracellular domains: a variable (V) region and a constant (C) region, both of the immunoglobulin superfamily. As in other immunoglobulins, the variable domains of the TCR .alpha.-chain and .beta.-chain (or gamma and delta (.gamma./.delta.) chains) each have three hypervariable or complementarity determining regions (CDRs). When the TCR engages with antigenic peptide and MHC (peptide/MHC), the T cell becomes activated, enabling it to attack and destroy the target cell.
[0015] However, current therapies have shown varying levels of effectiveness with undesired side effects. Therefore, a need exists to identify novel and improved therapies for treating CLL-1 related diseases and disorders.
SUMMARY OF THE INVENTION
[0016] The invention relates to engineered immune cells (such as CARs or TCRs), antigen binding molecules (including but not limited to, antibodies, scFvs, heavy and/or light chains, and CDRs of these antigen binding molecules) with specificity to CLL-1.
[0017] The invention further relates to a novel CD28 extracellular (hinge) sequence useful as costimulatory domains in these cells.
[0018] Chimeric antigen receptors of the invention typically comprise: (i) a CLL-1 specific antigen binding molecule, (ii) an extracellular (which may comprise a hinge) domain, (iii) one or more costimulatory domain, and (iv) one or more activating domain. It will be appreciated that each domain may be heterogeneous, thus comprised of sequences derived from (or corresponding to) different protein chains.
[0019] In some embodiments, the invention relates to a chimeric antigen receptor comprising an antigen binding molecule that specifically binds to CLL-1, wherein the antigen binding molecule comprises at least one of: a) a variable heavy chain CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 17, 51, 73, and 95, b) a variable heavy chain CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs 18, 52, 74, and 96, c) a variable heavy chain CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs 19, 53, 75, and 97, d) a variable light chain CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 22, 56, 78, and 100, e) a variable light chain CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs. 23, 57, 79, and 101, and 0 a variable light chain CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs. 24, 58, 80, and 102. The chimeric antigen receptor can further comprise at least one costimulatory domain. The chimeric antigen receptor according to claim 1 further comprising at least one activating domain.
[0020] In certain embodiments, the invention relates to chimeric antigen receptors having at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% identical to the chimeric antigen receptors set forth herein.
[0021] Also encompassed by the invention are chimeric antigen receptors having no more than 8 amino acid substitutions thereto.
[0022] In certain embodiments the costimulatory domain comprises a signaling domain (or other suitable portion) of CD28, OX-40, 4-1BB/CD137, CD2, CD7, CD27, CD30, CD40, Programmed Death-1 (PD-1), inducible T cell costimulator (ICOS), lymphocyte function-associated antigen-1 (LFA-1, CD1-1a/CD18), CD3 gamma, CD3 delta, CD3 epsilon, CD247, CD276 (B7-H3), LIGHT, (TNFSF14), NKG2C, Ig alpha (CD79a), DAP-10, Fc gamma receptor, MHC class 1 molecule, TNF receptor proteins, an Immunoglobulin protein, cytokine receptor, integrins, Signaling Lymphocytic Activation Molecules (SLAM proteins), activating NK cell receptors, BTLA, a Toll ligand receptor, ICAM-1, B7-H3, CDS, ICAM-1, GITR, BAFFR, LIGHT, HVEM (LIGHTR), KIRDS2, SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD19, CD4, CD8alpha, CD8beta, IL-2R beta, IL-2R gamma, IL-7R alpha, ITGA4, VLA1, CD49a, ITGA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD, CD1 1d, ITGAE, CD103, ITGAL, CD1 1a, LFA-1, ITGAM, CD1 1b, ITGAX, CD1 1c, ITGB1, CD29, ITGB2, CD18, LFA-1, ITGB7, NKG2D, TNFR2, TRANCE/RANKL, DNAM1 (CD226), SLAMF4 (CD244, 2B4), CD84, CD96 (Tactile), CEACAM1, CRT AM, Ly9 (CD229), CD160 (BY55), PSGL1, CD100 (SEMA4D), CD69, SLAMF6 (NTB-A, Ly108), SLAM (SLAMF1, CD150, IPO-3), BLAME (SLAMF8), SELPLG (CD162), LTBR, LAT, GADS, SLP-76, PAG/Cbp, CD19a, a ligand that specifically binds with CD83, or any combination thereof.
[0023] In some embodiments, the costimulatory domain can comprise all or a portion of the 4-1BB nucleic acid sequence set forth in SEQ ID NO. 141, and the corresponding amino acid sequence as set forth in SEQ ID NO. 142. In other embodiments, the costimulatory domain can comprise all or a portion of the amino acid sequence of OX40 as set forth in SEQ ID NO. 143. See also Hombach et al., Oncoimmunology. 2012 Jul. 1; 1(4): 458-466. In still other embodiments, the costimulatory domain can comprise all or a portion of the ICOS molecule as described in Guedan et al., Aug. 14, 2014; Blood: 124 (7) and Shen et al., Journal of Hematology & Oncology (2013) 6:33. In still other embodiments, the costimulatory domain can comprise all or a portion of CD27 as described in Song et al., Oncoimmunology. 2012 Jul. 1; 1(4): 547-549.
[0024] Preferred embodiments include incorporation into the CARs of the invention one or more of the following sequences: SEQ ID NO. 2, SEQ ID NO. 4, SEQ ID NO. 6, and SEQ ID NO. 8. Additional preferred embodiments include incorporation into the CARs of the invention the sequence set forth in SEQ ID NO. 14.
[0025] In further embodiments, the activating domain comprises CD3, preferably CD3 zeta, more preferably CD3 zeta having the sequence set forth in SEQ ID NO. 10.
[0026] In other embodiments, the invention relates to a chimeric antigen receptor comprising an antigen binding molecule further comprising SEQ ID NO. 2 and further comprising SEQ ID NO. 10.
[0027] The invention further relates to isolated polynucleotides encoding the chimeric antigen receptors, and vectors comprising the polynucleotides. Any vector known in the art can be suitable for the present invention. In some embodiments, the vector is a viral vector. In some embodiments, the vector is a retroviral vector (such as pMSVG1), a DNA vector, a murine leukemia virus vector, an SFG vector, a plasmid, a RNA vector, an adenoviral vector, a baculoviral vector, an Epstein Barr viral vector, a papovaviral vector, a vaccinia viral vector, a herpes simplex viral vector, an adenovirus associated vector (AAV), a lentiviral vector (such as pGAR), or any combination thereof. The pGAR sequence is as follows:
TABLE-US-00002 (SEQ ID NO: 147) CTGACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGG TTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCT TTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCA AGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGC ACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTGGGCCA TCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTT TAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGG TCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTA AAAAATGAGCTGATTTAACAAAAATTTAACGCGAATTTTAACAAAATATT AACGCTTACAATTTGCCATTCGCCATTCAGGCTGCGCAACTGTTGGGAAG GGCGATCGGTGCGGGCCTCTTCGCTATTACGCCAGCTGGCGAAAGGGGGA TGTGCTGCAAGGCGATTAAGTTGGGTAACGCCAGGGTTTTCCCAGTCACG ACGTTGTAAAACGACGGCCAGTGAATTGTAATACGACTCACTATAGGGCG ACCCGGGGATGGCGCGCCAGTAATCAATTACGGGGTCATTAGTTCATAGC CCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGG CTGACCGCCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTC CCATAGTAACGCCAATAGGGACTTTCCATTGACGTCAATGGGTGGAGTAT TTACGGTAAACTGCCCACTTGGCAGTACATCAAGTGTATCATATGCCAAG TACGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGGCATTATG CCCAGTACATGACCTTATGGGACTTTCCTACTTGGCAGTACATCTACGTA TTAGTCATCGCTATTACCATGCTGATGCGGTTTTGGCAGTACATCAATGG GCGTGGATAGCGGTTTGACTCACGGGGATTTCCAAGTCTCCACCCCATTG ACGTCAATGGGAGTTTGTTTTGGCACCAAAATCAACGGGACTTTCCAAAA TGTCGTAACAACTCCGCCCCATTGACGCAAATGGGCGGTAGGCGTGTACG GTGGGAGGTCTATATAAGCAGAGCTGGTTTAGTGAACCGGGGTCTCTCTG GTTAGACCAGATCTGAGCCTGGGAGCTCTCTGGCTAACTAGGGAACCCAC TGCTTAAGCCTCAATAAAGCTTGCCTTGAGTGCTTCAAGTAGTGTGTGCC CGTCTGTTGTGTGACTCTGGTAACTAGAGATCCCTCAGACCCTTTTAGTC AGTGTGGAAAATCTCTAGCAGTGGCGCCCGAACAGGGACTTGAAAGCGAA AGGGAAACCAGAGGAGCTCTCTCGACGCAGGACTCGGCTTGCTGAAGCGC GCACGGCAAGAGGCGAGGGGCGGCGACTGGTGAGTACGCCAAAAATTTTG ACTAGCGGAGGCTAGAAGGAGAGAGATGGGTGCGAGAGCGTCAGTATTAA GCGGGGGAGAATTAGATCGCGATGGGAAAAAATTCGGTTAAGGCCAGGGG GAAAGAAAAAATATAAATTAAAACATATAGTATGGGCAAGCAGGGAGCTA GAACGATTCGCAGTTAATCCTGGCCTGTTAGAAACATCAGAAGGCTGTAG ACAAATACTGGGACAGCTACAACCATCCCTTCAGACAGGATCAGAAGAAC TTAGATCATTATATAATACAGTAGCAACCCTCTATTGTGTGCATCAAAGG ATAGAGATAAAAGACACCAAGGAAGCTTTAGACAAGATAGAGGAAGAGCA AAACAAAAGTAAGACCACCGCACAGCAAGCCGCCGCTGATCTTCAGACCT GGAGGAGGAGATATGAGGGACAATTGGAGAAGTGAATTATATAAATATAA AGTAGTAAAAATTGAACCATTAGGAGTAGCACCCACCAAGGCAAAGAGAA GAGTGGTGCAGAGAGAAAAAAGAGCAGTGGGAATAGGAGCTTTGTTCCTT GGGTTCTTGGGAGCAGCAGGAAGCACTATGGGCGCAGCGTCAATGACGCT GACGGTACAGGCCAGACAATTATTGTCTGGTATAGTGCAGCAGCAGAACA ATTTGCTGAGGGCTATTGAGGCGCAACAGCATCTGTTGCAACTCACAGTC TGGGGCATCAAGCAGCTCCAGGCAAGAATCCTGGCTGTGGAAAGATACCT AAAGGATCAACAGCTCCTGGGGATTTGGGGTTGCTCTGGAAAACTCATTT GCACCACTGCTGTGCCTTGGAATGCTAGTTGGAGTAATAAATCTCTGGAA CAGATTTGGAATCACACGACCTGGATGGAGTGGGACAGAGAAATTAACAA TTACACAAGCTTAATACACTCCTTAATTGAAGAATCGCAAAACCAGCAAG AAAAGAATGAACAAGAATTATTGGAATTAGATAAATGGGCAAGTTTGTGG AATTGGTTTAACATAACAAATTGGCTGTGGTATATAAAATTATTCATAAT GATAGTAGGAGGCTTGGTAGGTTTAAGAATAGTTTTTGCTGTACTTTCTA TAGTGAATAGAGTTAGGCAGGGATATTCACCATTATCGTTTCAGACCCAC CTCCCAACCCCGAGGGGACCCGACAGGCCCGAAGGAATAGAAGAAGAAGG TGGAGAGAGAGACAGAGACAGATCCATTCGATTAGTGAACGGATCTCGAC GGTATCGGTTAACTTTTAAAAGAAAAGGGGGGATTGGGGGGTACAGTGCA GGGGAAAGAATAGTAGACATAATAGCAACAGACATACAAACTAAAGAATT ACAAAAACAAATTACAAAATTCAAAATTTTATCGCGATCGCGGAATGAAA GACCCCACCTGTAGGTTTGGCAAGCTAGCTTAAGTAACGCCATTTTGCAA GGCATGGAAAATACATAACTGAGAATAGAGAAGTTCAGATCAAGGTTAGG AACAGAGAGACAGCAGAATATGGGCCAAACAGGATATCTGTGGTAAGCAG TTCCTGCCCCGGCTCAGGGCCAAGAACAGATGGTCCCCAGATGCGGTCCC GCCCTCAGCAGTTTCTAGAGAACCATCAGATGTTTCCAGGGTGCCCCAAG GACCTGAAAATGACCCTGTGCCTTATTTGAACTAACCAATCAGTTCGCTT CTCGCTTCTGTTCGCGCGCTTCTGCTCCCCGAGCTCAATAAAAGAGCCCA CAACCCCTCACTCGGCGCGCCAGTCCTTCGAAGTAGATCTTTGTCGATCC TACCATCCACTCGACACACCCGCCAGCGGCCGCTGCCAAGCTTCCGAGCT CTCGAATTAATTCACGGTACCCACCATGGCCTAGGGAGACTAGTCGAATC GATATCAACCTCTGGATTACAAAATTTGTGAAAGATTGACTGGTATTCTT AACTATGTTGCTCCTTTTACGCTATGTGGATACGCTGCTTTAATGCCTTT GTATCATGCTATTGCTTCCCGTATGGCTTTCATTTTCTCCTCCTTGTATA AATCCTGGTTGCTGTCTCTTTATGAGGAGTTGTGGCCCGTTGTCAGGCAA CGTGGCGTGGTGTGCACTGTGTTTGCTGACGCAACCCCCACTGGTTGGGG CATTGCCACCACCTGTCAGCTCCTTTCCGGGACTTTCGCTTTCCCCCTCC CTATTGCCACGGCGGAACTCATCGCCGCCTGCCTTGCCCGCTGCTGGACA GGGGCTCGGCTGTTGGGCACTGACAATTCCGTGGTGTTGTCGGGGAAGCT GACGTCCTTTTCATGGCTGCTCGCCTGTGTTGCCACCTGGATTCTGCGCG GGACGTCCTTCTGCTACGTCCCTTCGGCCCTCAATCCAGCGGACCTTCCT TCCCGCGGCCTGCTGCCGGCTCTGCGGCCTCTTCCGCGTCTTCGCCTTCG CCCTCAGACGAGTCGGATCTCCCTTTGGGCCGCCTCCCCGCCTGGTTAAT TAAAGTACCTTTAAGACCAATGACTTACAAGGCAGCTGTAGATCTTAGCC ACTTTTTAAAAGAAAAGGGGGGACTGGAAGGGCGAATTCACTCCCAACGA AGACAAGATCTGCTTTTTGCTTGTACTGGGTCTCTCTGGTTAGACCAGAT CTGAGCCTGGGAGCTCTCTGGCTAACTAGGGAACCCACTGCTTAAGCCTC AATAAAGCTTGCCTTGAGTGCTTCAAGTAGTGTGTGCCCGTCTGTTGTGT GACTCTGGTAACTAGAGATCCCTCAGACCCTTTTAGTCAGTGTGGAAAAT CTCTAGCAGGCATGCCAGACATGATAAGATACATTGATGAGTTTGGACAA ACCACAACTAGAATGCAGTGAAAAAAATGCTTTATTTGTGAAATTTGTGA TGCTATTGCTTTATTTGTAACCATTATAAGCTGCAATAAACAAGTTAACA ACAACAATTGCATTCATTTTATGTTTCAGGTTCAGGGGGAGGTGTGGGAG GTTTTTTGGCGCGCCATCGTCGAGGTTCCCTTTAGTGAGGGTTAATTGCG AGCTTGGCGTAATCATGGTCATAGCTGTTTCCTGTGTGAAATTGTTATCC GCTCACAATTCCACACAACATACGAGCCGGAAGCATAAAGTGTAAAGCCT GGGGTGCCTAATGAGTGAGCTAACTCACATTAATTGCGTTGCGCTCACTG CCCGCTTTCCAGTCGGGAAACCTGTCGTGCCAGCTGCATTAATGAATCGG CCAACGCGCGGGGAGAGGCGGTTTGCGTATTGGGCGCTCTTCCGCTTCCT CGCTCACTGACTCGCTGCGCTCGGTCGTTCGGCTGCGGCGAGCGGTATCA GCTCACTCAAAGGCGGTAATACGGTTATCCACAGAATCAGGGGATAACGC AGGAAAGAACATGTGAGCAAAAGGCCAGCAAAAGGCCAGGAACCGTAAAA AGGCCGCGTTGCTGGCGTTTTTCCATAGGCTCCGCCCCCCTGACGAGCAT CACAAAAATCGACGCTCAAGTCAGAGGTGGCGAAACCCGACAGGACTATA AAGATACCAGGCGTTTCCCCCTGGAAGCTCCCTCGTGCGCTCTCCTGTTC CGACCCTGCCGCTTACCGGATACCTGTCCGCCTTTCTCCCTTCGGGAAGC GTGGCGCTTTCTCATAGCTCACGCTGTAGGTATCTCAGTTCGGTGTAGGT CGTTCGCTCCAAGCTGGGCTGTGTGCACGAACCCCCCGTTCAGCCCGACC GCTGCGCCTTATCCGGTAACTATCGTCTTGAGTCCAACCCGGTAAGACAC GACTTATCGCCACTGGCAGCAGCCACTGGTAACAGGATTAGCAGAGCGAG GTATGTAGGCGGTGCTACAGAGTTCTTGAAGTGGTGGCCTAACTACGGCT ACACTAGAAGAACAGTATTTGGTATCTGCGCTCTGCTGAAGCCAGTTACC TTCGGAAAAAGAGTTGGTAGCTCTTGATCCGGCAAACAAACCACCGCTGG TAGCGGTGGTTTTTTTGTTTGCAAGCAGCAGATTACGCGCAGAAAAAAAG GATCTCAAGAAGATCCTTTGATCTTTTCTACGGGGTCTGACGCTCAGTGG AACGAAAACTCACGTTAAGGGATTTTGGTCATGAGATTATCAAAAAGGAT CTTCACCTAGATCCTTTTAAATTAAAAATGAAGTTTTAAATCAATCTAAA GTATATATGAGTAAACTTGGTCTGACAGTTACCAATGCTTAATCAGTGAG GCACCTATCTCAGCGATCTGTCTATTTCGTTCATCCATAGTTGCCTGACT CCCCGTCGTGTAGATAACTACGATACGGGAGGGCTTACCATCTGGCCCCA GTGCTGCAATGATACCGCGAGACCCACGCTCACCGGCTCCAGATTTATCA GCAATAAACCAGCCAGCCGGAAGGGCCGAGCGCAGAAGTGGTCCTGCAAC TTTATCCGCCTCCATCCAGTCTATTAATTGTTGCCGGGAAGCTAGAGTAA GTAGTTCGCCAGTTAATAGTTTGCGCAACGTTGTTGCCATTGCTACAGGC ATCGTGGTGTCACGCTCGTCGTTTGGTATGGCTTCATTCAGCTCCGGTTC CCAACGATCAAGGCGAGTTACATGATCCCCCATGTTGTGCAAAAAAGCGG TTAGCTCCTTCGGTCCTCCGATCGTTGTCAGAAGTAAGTTGGCCGCAGTG
TTATCACTCATGGTTATGGCAGCACTGCATAATTCTCTTACTGTCATGCC ATCCGTAAGATGCTTTTCTGTGACTGGTGAGTACTCAACCAAGTCATTCT GAGAATAGTGTATGCGGCGACCGAGTTGCTCTTGCCCGGCGTCAATACGG GATAATACCGCGCCACATAGCAGAACTTTAAAAGTGCTCATCATTGGAAA ACGTTCTTCGGGGCGAAAACTCTCAAGGATCTTACCGCTGTTGAGATCCA GTTCGATGTAACCCACTCGTGCACCCAACTGATCTTCAGCATCTTTTACT TTCACCAGCGTTTCTGGGTGAGCAAAAACAGGAAGGCAAAATGCCGCAAA AAAGGGAATAAGGGCGACACGGAAATGTTGAATACTCATACTCTTCCTTT TTCAATATTATTGAAGCATTTATCAGGGTTATTGTCTCATGAGCGGATAC ATATTTGAATGTATTTAGAAAAATAAACAAATAGGGGTTCCGCGCACATT TCCCCGAAAAGTGCCAC
[0028] The pGAR vector map is set forth in FIG. 11.
[0029] Suitable additional exemplary vectors include e.g., pBABE-puro, pBABE-neo largeTcDNA, pBABE-hygro-hTERT, pMKO.1 GFP, MSCV-IRES-GFP, pMSCV PIG (Puro IRES GFP empty plasmid), pMSCV-loxp-dsRed-loxp-eGFP-Puro-WPRE, MSCV IRES Luciferase, pMIG, MDH1-PGK-GFP 2.0, TtRMPVIR, pMSCV-IRES-mCherry FP, pRetroX GFP T2A Cre, pRXTN, pLncEXP, and pLXIN-Luc.
[0030] Exemplary immune cells include, but are not limited to T cells, tumor infiltrating lymphocytes (TILs), NK cells, TCR-expressing cells, dendritic cells, or NK-T cells. The T cells can be autologous, allogeneic, or heterologous. In other embodiments, the invention relates to pharmaceutical compositions comprising the immune cells of described herein.
[0031] In certain embodiments, the invention relates to antigen binding molecules (and chimeric antigen receptors comprising these molecules) comprising at least one of:
[0032] (a) a VH region comprising the amino acid sequence of SEQ ID NO: 16 and a VL region comprising the amino acid sequence of SEQ ID NO: 21;
[0033] (b) a VH region comprising the amino acid sequence of SEQ ID NO: 50 and a VL region comprising the amino acid sequence of SEQ ID NO: 55;
[0034] (c) a VH region comprising the amino acid sequence of SEQ ID NO: 72 and a VL region comprising the amino acid sequence of SEQ ID NO: 77;
[0035] (d) a VH region comprising the amino acid sequence of SEQ ID NO: 94 and a VL region comprising the amino acid sequence of SEQ ID NO: 99; and wherein the VH and VL region or regions are linked by at least one linker. Also encompassed by the invention are chimeric antigen receptors and/or antigen binding molecules having no more than 8 amino acid substitutions thereto.
[0036] The linker may be, e.g., a poly-Gly linker such as GGGGSGGGGSGGGGS (SEQ ID NO. 130) or GGGGSGGGGSGGGGSGGGGS (SEQ ID NO. 145).
[0037] In other embodiments, the invention relates to antigen binding molecules (and chimeric antigen receptors comprising these molecules) wherein the linker comprises at least one of SEQ ID NO. 130 and SEQ ID NO. 132.
[0038] In certain embodiments, the invention relates to antigen binding molecules and/or chimeric antigen receptors having at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% identical to the antigen binding molecules and/or chimeric antigen receptors set forth herein.
[0039] In other embodiments, the invention relates to isolated polynucleotides comprising at least one of: SEQ ID NO. 27; SEQ ID NO. 31; SEQ ID NO. 35; SEQ ID NO. 39; SEQ ID NO. 43; SEQ ID NO. 47; SEQ ID NO. 61; SEQ ID NO. 65; SEQ ID NO. 69; SEQ ID NO. 83; SEQ ID NO. 87; SEQ ID NO. 91; SEQ ID NO. 105; SEQ ID NO. 109; SEQ ID NO. 113; SEQ ID NO. 117; SEQ ID NO. 121; and SEQ ID NO. 125.
[0040] In certain embodiments, the invention relates to isolated polynucleotides having at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% identical to the polynucleotides set forth herein.
[0041] The invention further relates to vectors comprising these polynucleotides, as well as cells transduced using these vectors.
[0042] In further embodiments, the invention relates to isolated polypeptides comprising the amino acid sequence set forth in at least one of: SEQ ID NO. 28; SEQ ID NO. 32; SEQ ID NO. 36; SEQ ID NO. 40; SEQ ID NO. 44; SEQ ID NO. 48; SEQ ID NO. 62; SEQ ID NO. 66; SEQ ID NO. 70; SEQ ID NO. 84; SEQ ID NO. 88; SEQ ID NO. 92; SEQ ID NO. 106; SEQ ID NO. 110; SEQ ID NO. 114; SEQ ID NO. 118; SEQ ID NO. 122; and SEQ ID NO. 126. In other embodiments, the invention relates to vectors encoding these polypeptides, immune cells comprising these polypeptides. Preferred immune cells include T cells, tumor infiltrating lymphocytes (TILs), NK cells, TCR-expressing cells, dendritic cells, or NK-T cells. The T cells may be autologous, allogeneic, or heterologous. Also encompassed by the invention are chimeric antigen receptors having no more than 8 amino acid substitutions thereto.
[0043] In other embodiments, the invention relates to isolated polynucleotides encoding a chimeric antigen receptor (CAR) or T cell receptor (TCR) comprising an antigen binding molecule that specifically binds to CLL-1, wherein the antigen binding molecule comprises a variable heavy (V.sub.H) chain CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 19, 53, 75, and 97. Also encompassed by the invention are chimeric antigen receptors having no more than 8 amino acid substitutions thereto. The polynucleotides may further comprising an activating domain. In preferred embodiments, the activating domain is CD3, more preferably CD3 zeta, more preferably the amino acid sequence set forth in SEQ ID NO. 9.
[0044] In other embodiments, the invention includes a costimulatory domain comprising the signaling domain (or other suitable portion) of CD28, CD28T, OX40, 4-1BB/CD137, CD2, CD3 (alpha, beta, delta, epsilon, gamma, zeta), CD4, CD5, CD7, CD9, CD16, CD22, CD27, CD30, CD 33, CD37, CD40, CD 45, CD64, CD80, CD86, CD134, CD137, CD154, PD-1, ICOS, lymphocyte function-associated antigen-1 (LFA-1 (CD1 1a/CD18), CD247, CD276 (B7-H3), LIGHT (tumor necrosis factor superfamily member 14; TNFSF14), NKG2C, Ig alpha (CD79a), DAP-10, Fc gamma receptor, MHC class I molecule, TNF, TNFr, integrin, signaling lymphocytic activation molecule, BTLA, Toll ligand receptor, ICAM-1, B7-H3, CDS, ICAM-1, GITR, BAFFR, LIGHT, HVEM (LIGHTR), KIRDS2, SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD19, CD4, CD8alpha, CD8beta, IL-2R beta, IL-2R gamma, IL-7R alpha, ITGA4, VLA1, CD49a, ITGA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD, CD1-1d, ITGAE, CD103, ITGAL, CD1-1a, LFA-1, ITGAM, CD1-1b, ITGAX, CD1-1c, ITGB1, CD29, ITGB2, CD18, LFA-1, ITGB7, NKG2D, TNFR2, TRANCE/RANKL, DNAM1 (CD226), SLAMF4 (CD244, 2B4), CD84, CD96 (Tactile), CEACAM1, CRT AM, Ly9 (CD229), CD160 (BY55), PSGL1, CD100 (SEMA4D), CD69, SLAMF6 (NTB-A, Ly108), SLAM (SLAMF1, CD150, IPO-3), BLAME (SLAMF8), SELPLG (CD162), LTBR, LAT, GADS, SLP-76, PAG/Cbp, CD19a, CD83 ligand, or fragments or combinations thereof. Preferred costimulatory domains are recited hereinbelow.
[0045] In further embodiments, the invention relates to isolated polynucleotides encoding a chimeric antigen receptor (CAR) or T cell receptor (TCR), wherein said CAR or TCR comprises an antigen binding molecule that specifically binds to CLL-1, and wherein the antigen binding molecule comprises a variable light (VL) chain CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 24, 58, 80, and 102. The polynucleotide can further comprise an activating domain. The polynucleotide can further comprise a costimulatory domain.
[0046] In other embodiments, the invention relates to isolated polynucleotides encoding a chimeric antigen receptor (CAR) or T cell receptor (TCR) comprising an antigen binding molecule that specifically binds to CLL-1, wherein the antigen binding molecule heavy chain comprises CDR1 (SEQ ID NO. 17), CDR2 (SEQ ID NO. 18), and CDR3 (SEQ ID NO. 19) and the antigen binding molecule light chain comprises CDR1 (SEQ ID NO. 22), CDR2 (SEQ ID NO. 23), and CDR3 (SEQ ID NO. 24).
[0047] In certain embodiments, the invention relates to isolated polynucleotides having at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% identical to the above sequences.
[0048] In other embodiments, the invention relates to isolated polynucleotides encoding a chimeric antigen receptor (CAR) or T cell receptor (TCR) comprising an antigen binding molecule that specifically binds to CLL-1, wherein the antigen binding molecule heavy chain comprises CDR1 (SEQ ID NO. 51), CDR2 (SEQ ID NO. 52), and CDR3 (SEQ ID NO. 53) and the antigen binding molecule light chain comprises CDR1 (SEQ ID NO. 56), CDR2 (SEQ ID NO. 57), and CDR3 (SEQ ID NO. 58).
[0049] In certain embodiments, the invention relates to isolated polynucleotides having at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% identical to the above sequences.
[0050] In other embodiments, the invention relates to isolated polynucleotides encoding a chimeric antigen receptor (CAR) or T cell receptor (TCR) comprising an antigen binding molecule that specifically binds to CLL-1, wherein the antigen binding molecule heavy chain comprises CDR1 (SEQ ID NO. 73), CDR2 (SEQ ID NO. 74), and CDR3 (SEQ ID NO. 75) and the antigen binding molecule light chain comprises CDR1 (SEQ ID NO. 78), CDR2 (SEQ ID NO. 79), and CDR3 (SEQ ID NO. 80).
[0051] In certain embodiments, the invention relates to isolated polynucleotides having at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% identical to the above sequences.
[0052] In other embodiments, the invention relates to isolated polynucleotides encoding a chimeric antigen receptor (CAR) or T cell receptor (TCR) comprising an antigen binding molecule that specifically binds to CLL-1, wherein the antigen binding molecule heavy chain comprises CDR1 (SEQ ID NO. 95), CDR2 (SEQ ID NO. 96), and CDR3 (SEQ ID NO. 97) and the antigen binding molecule light chain comprises CDR1 (SEQ ID NO. 100), CDR2 (SEQ ID NO. 101), and CDR3 (SEQ ID NO. 102).
[0053] In certain embodiments, the invention relates to isolated polynucleotides having at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% identical to the above sequences.
[0054] In further embodiments, the invention relates to isolated polynucleotides encoding a chimeric antigen receptor (CAR) or T cell receptor (TCR) comprising an antigen binding molecule that specifically binds to CLL-1, and wherein the antigen binding molecule comprises:
[0055] (a) a heavy chain variable region (VH) complementarity determining region (CDR) 1 comprising the amino acid sequence GX.sub.2X.sub.3X.sub.4X.sub.5X.sub.6X.sub.7X.sub.8X.sub.9 (SEQ ID NO: 134), wherein X.sub.2 is G, F, or Y; X.sub.3 is S or T; X.sub.4 is I, F, or L; X.sub.5 is S or T; X.sub.6 is not present or S; X.sub.7 is not present or G; X.sub.8 is not present or E or G; and X.sub.9 is F, L, or Y;
[0056] (b) a heavy chain variable region (VH) complementarity determining region (CDR) 2 comprising the amino acid sequence X.sub.1X.sub.2X.sub.3X.sub.4X.sub.5X.sub.6 (SEQ ID NO: 135), wherein X.sub.1 is D, H, S, or Y; X.sub.2 is H, P, or Y; X.sub.3 is D, E, or S; X.sub.4 is D or G; X.sub.5 is G or S; and X.sub.6 is not present of D or E;
[0057] (c) a heavy chain variable region (VH) complementarity determining region (CDR) 3, comprising the amino acid sequence X.sub.1X.sub.2X.sub.3X.sub.4X.sub.5X.sub.6X.sub.7X.sub.8X.sub.9X.sub.10X.- sub.11X.sub.12DY (SEQ ID NO: 136), wherein X.sub.1 is E or L; X.sub.2 is R, S, or V; X.sub.3 is R or Y; X.sub.4 is C, G, or S; X.sub.5 is not present or G or I; X.sub.6 is not present or G; X.sub.7 is not present or D; X.sub.8 is not present or C; X.sub.9 is not present or W or Y; X.sub.10 is not present or P or S; X.sub.11 is not present or G or Y; and X.sub.12 is F or R;
[0058] (d) a light chain variable region (VL) CDR1 comprising the amino acid sequence X.sub.1ASQX.sub.5X.sub.6X.sub.7X.sub.8X.sub.9LX.sub.11 (SEQ ID NO: 137), wherein X.sub.1 is Q or R; X.sub.5 is D or S; X.sub.6 is I or V; X.sub.7 is N or S; X.sub.8 is N or S; X.sub.9 is F, L, or Y; and X.sub.11 is N or T;
[0059] (e) a light chain variable region (VL) CDR2 comprising the amino acid sequence X.sub.1ASX.sub.4X.sub.5X.sub.6X.sub.7 (SEQ ID NO: 138), wherein X.sub.1 is D or G; X.sub.4 is N, S, or T; X.sub.5 is L or R; X.sub.6 is A, E, or K; and X.sub.7 is S or T; and/or
[0060] (f) a light chain variable region (VL) CDR3 comprising the amino acid sequence QQX.sub.3X.sub.4X.sub.5X.sub.6PX.sub.8T (SEQ ID NO: 139), wherein X.sub.3 is S or Y; X.sub.4 is D, G, or Y; X.sub.5 is N, S, or T; X.sub.6 is L, T, or Y; and X.sub.8 is F or I.
[0061] The invention further relates to antigen binding molecules to CLL-1 comprising at least one variable heavy chain CDR3 or variable light chain CDR3 sequence as set forth herein. The invention further relates to antigen binding molecules to CLL-1 comprising at least one variable heavy chain CDR1, CDR2, and CDR3 sequences as described herein. The invention further relates to antigen binding molecules to CLL-1 comprising at least one variable light chain CDR1, CDR2, and CDR3 sequences as described herein. The invention further relates to antigen binding molecules to CLL-1 comprising both variable heavy chain CDR1, CDR2, CDR3, and variable light chain CDR1, CDR2, and CDR3 sequences as described herein.
[0062] The invention further relates to methods of treating a disease or disorder in a subject in need thereof comprising administering to the subject the antigen binding molecules, the CARs, TCRs, polynucleotides, vectors, cells, or compositions according to the invention. Suitable diseases for treatment include, but are not limited to, acute myeloid leukemia (AML), chronic myelogenous leukemia (CML), chronic myelomonocytic leukemia (CMML), juvenile myelomonocytic leukemia, atypical chronic myeloid leukemia, acute promyelocytic leukemia (APL), acute monoblastic leukemia, acute erythroid leukemia, acute megakaryoblastic leukemia, myelodysplastic syndrome (MDS), myeloproliferative disorder, myeloid neoplasm, myeloid sarcoma), Blastic Plasmacytoid Dendritic Cell Neoplasm (BPDCN), or combinations thereof. Additional diseases include inflammatory and/or autoimmune diseases such as rheumatoid arthritis, psoriasis, allergies, asthma, Crohn's disease, IBD, IBS, fibromyalga, mastocytosis, and Celiac disease.
BRIEF DESCRIPTION OF THE FIGURES
[0063] FIG. 1 shows CLL-1 expression in different cancer cell lines.
[0064] FIG. 2 shows CLL-1 CAR expression determined by protein L 6 hours post mRNA electroporation.
[0065] FIG. 3 shows the results from a cytokine release assay from different CLL-1 CAR-T cell constructs 24 hours after mRNA electroporation.
[0066] FIG. 4 shows cytolytic activity of different CLL-1 CAR-T cell constructs 24 hours after mRNA electroporation.
[0067] FIG. 5 shows cytolytic activity of different CLL-1 CAR-T cell constructs 24 hours after mRNA electroporation.
[0068] FIG. 6 shows CLL-1 CAR expression determined by protein L at day 12 after transduction.
[0069] FIG. 7 shows cytokine release assay from CLL-1 CAR-T cells 16 hours after co-culture with different target cell lines.
[0070] FIG. 8 shows cytolytic activity from CLL-1 CAR-T cells 16 hours and 40 hours after co-culture with different target cell lines.
[0071] FIGS. 9A-9D set forth sequence alignments of the CLL-1 antigen binding molecules of the invention. CDRs are notated in boxes.
[0072] FIG. 10 sets forth bioluminescence results on NSG mice treated with CARs according to the invention.
[0073] FIG. 11 sets forth the pGAR vector map.
DETAILED DESCRIPTION OF THE INVENTION
[0074] It will be appreciated that chimeric antigen receptors (CARs or CAR-Ts) and T cell receptors (TCRs) are genetically engineered receptors. These engineered receptors can be readily inserted into and expressed by immune cells, including T cells in accordance with techniques known in the art. With a CAR, a single receptor can be programmed to both recognize a specific antigen and, when bound to that antigen, activate the immune cell to attack and destroy the cell bearing that antigen. When these antigens exist on tumor cells, an immune cell that expresses the CAR can target and kill the tumor cell.
[0075] CARs can be engineered to bind to an antigen (such as a cell-surface antigen) by incorporating an antigen binding molecule that interacts with that targeted antigen. Preferably, the antigen binding molecule is an antibody fragment thereof, and more preferably one or more single chain antibody fragment ("scFv"). An scFv is a single chain antibody fragment having the variable regions of the heavy and light chains of an antibody linked together. See U.S. Pat. Nos. 7,741,465, and 6,319,494 as well as Eshhar et al., Cancer Immunol Immunotherapy (1997) 45: 131-136. An scFv retains the parent antibody's ability to specifically interact with target antigen. scFvs are preferred for use in chimeric antigen receptors because they can be engineered to be expressed as part of a single chain along with the other CAR components. Id. See also Krause et al., J. Exp. Med., Volume 188, No. 4, 1998 (619-626); Finney et al., Journal of Immunology, 1998, 161: 2791-2797. It will be appreciated that the antigen binding molecule is typically contained within the extracellular portion of the CAR such that it is capable of recognizing and binding to the antigen of interest. Bispecific and multispecific CARs are contemplated within the scope of the invention, with specificity to more than one target of interest.
Costimulatory Domains
[0076] Chimeric antigen receptors may incorporate costimulatory (signaling) domains to increase their potency. See U.S. Pat. Nos. 7,741,465, and 6,319,494, as well as Krause et al. and Finney et al. (supra), Song et al., Blood 119:696-706 (2012); Kalos et al., Sci Transl. Med. 3:95 (2011); Porter et al., N. Engl. J. Med. 365:725-33 (2011), and Gross et al., Annu. Rev. Pharmacol. Toxicol. 56:59-83 (2016). For example, CD28 is a costimulatory protein found naturally on T-cells. A variety of costimulatory molecules are set forth herein, but it will be appreciated that additional costimulatory molecules are also included within the scope of this invention.
[0077] The complete native amino acid sequence of CD28 is described in NCBI Reference Sequence: NP_006130.1. The complete native CD28 nucleic acid sequence is described in NCBI Reference Sequence: NM_006139.1.
[0078] Certain CD28 domains have been used in chimeric antigen receptors. In accordance with the present invention, it has now been found that a novel CD28 extracellular (hinge) construct, termed "CD28T", unexpectedly provides certain benefits when utilized in a CAR construct. This construct demonstrates the ability to retain (and at times exceed) the properties of CD28-containing CARs, despite truncation (removal) of multiple amino acids from the extracellular CD28 sequence. These benefits include equivalent or superior cytokine production, equivalent or superior cytolytic activity, and/or equivalent or superior CAR expression levels.
[0079] The nucleotide sequence of the CD28T molecule, including the extracellular domain, and the CD28 transmembrane and intracellular domains is set forth in SEQ ID NO. 1.
TABLE-US-00003 CTTGATAATGAAAAGTCAAACGGAACAATCATTCACGTGAAGGGCAAGC ACCTCTGTCCGTCACCCTTGTTCCCTGGTCCATCCAAGCCATTCTGGGT GTTGGTCGTAGTGGGTGGAGTCCTCGCTTGTTACTCTCTGCTCGTCACC GTGGCTTTTATAATCTTCTGGGTTAGATCCAAAAGAAGCCGCCTGCTCC ATAGCGATTACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGGAA ACACTACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTATCGGAGC
[0080] The corresponding amino acid sequence is set forth in SEQ ID NO. 2:
[0081] LDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAF IIFWVRSK RSRLLHSDYM NMTPRRPGPT RKHYQPYAPP RDFAAYRS
[0082] The nucleotide sequence of the extracellular portion of CD28T is set forth in SEQ ID NO. 3:
TABLE-US-00004 CTTGATAATGAAAAGTCAAACGGAACAATCATTCACGTGAAGGGCAAGC ACCTCTGTCCGTCACCCTTGTTCCCTGGTCCATCCAAGCCA
[0083] The corresponding amino acid sequence of the CD28T extracellular domain is set forth in SEQ ID NO. 4: LDNEKSNGTI IHVKGKHLCP SPLFPGPSKP
[0084] The nucleotide sequence of the CD28 transmembrane domain is set forth in SEQ ID NO. 5):
TABLE-US-00005 TTCTGGGTGTTGGTCGTAGTGGGTGGAGTCCTCGCTTGTTACTCTCTGCT CGTCACCGTGGCTTTTATAATCTTCTGGGTT
[0085] The amino acid sequence of the CD28 transmembrane domain is set forth in SEQ ID NO. 6: FWVLVVVGGV LACYSLLVTV AFIIFWV
[0086] The nucleotide sequence of the CD28 intracellular signaling domain is set forth in SEQ ID NO. 7:
TABLE-US-00006 AGATCCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATATGACTC CACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTACGCACCACC TAGAGATTTCGCTGCCTATCGGAGC
[0087] The amino acid sequence of the CD28 intracellular signaling domain is set forth in SEQ ID NO. 8:
TABLE-US-00007 RSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS
[0088] Additional CD28 sequences suitable for use in the invention include the CD28 nucleotide sequence set forth in SEQ ID NO. 11:
TABLE-US-00008 ATTGAGGTGATGTATCCACCGCCTTACCTGGATAACGAAAAGAGTAACG GTACCATCATTCACGTGAAAGGTAAACACCTGTGTCCTTCTCCCCTCTTC CCCGGGCCATCAAAGCCC
[0089] The corresponding amino acid sequence is set forth in SEQ ID NO. 12:
TABLE-US-00009 IEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP
[0090] It will be appreciated that the invention relates to antigen binding molecules, CARs, TCRs, and the like comprising at least one isolated nucleic acid sequence of SEQ ID NO. 1 or SEQ ID NO. 3. It will further be appreciated that the invention relates to antigen binding molecules, CARs, TCRs, and the like wherein the extracellular portion consists of at least one isolated nucleic acid sequence of SEQ ID NO. 1 or SEQ ID NO. 3. Additionally, it will be appreciated that the invention relates to antigen binding molecules, CARs, TCRs, and the like wherein the extracellular portion consists essentially of at least one isolated nucleic acid sequence of SEQ ID NO. 1 or SEQ ID NO. 3.
[0091] It will be appreciated that the invention relates to antigen binding molecules, CARs, TCRs, and the like comprising at least one amino acid sequence of SEQ ID NO. 2 or SEQ ID NO. 4. It will further be appreciated that the invention relates to antigen binding molecules, CARs, TCRs, and the like wherein the extracellular portion consists of at least one amino acid sequence of SEQ ID NO. 2 or SEQ ID NO. 4. It will also be appreciated that the invention relates to antigen binding molecules, CARs, TCRs, and the like wherein the extracellular portion consists essentially of at least one amino acid sequence of SEQ ID NO. 2 or SEQ ID NO. 4.
[0092] Another suitable source of extracellular and/or transmembrane domains can be derived from (or correspond to) some or all of CD8. The nucleotide sequence of a suitable CD8 extracellular and transmembrane domain is set forth in SEQ ID NO. 13:
TABLE-US-00010 GCTGCAGCATTGAGCAACTCAATAATGTATTTTAGTCACTTTGTACCAGT GTTCTTGCCGGCTAAGCCTACTACCACACCCGCTCCACGGCCACCTACCC CAGCTCCTACCATCGCTTCACAGCCTCTGTCCCTGCGCCCAGAGGCTTGC CGACCGGCCGCAGGGGGCGCTGTTCATACCAGAGGACTGGATTTCGCCT GCGATATCTATATCTGGGCACCCCTGGCCGGAACCTGCGGCGTACTCCTG CTGTCCCTGGTCATCACGCTCTATTGTAATCACAGGAAC
[0093] The corresponding amino acid sequence is set forth in SEQ ID NO. 14:
TABLE-US-00011 AAALSNSIMYFSHFVPVFLPAKPTTTPAPRPPTPAPTIASQPLSLRPEAC RPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCNHRN
[0094] It will be appreciated that suitable costimulatory domains within the scope of the invention can be derived from (or correspond to) for example, CD28, CD28T, OX40, 4-1BB/CD137, CD2, CD3 (alpha, beta, delta, epsilon, gamma, zeta), CD4, CD5, CD7, CD9, CD16, CD22, CD27, CD30, CD 33, CD37, CD40, CD 45, CD64, CD80, CD86, CD134, CD137, CD154, PD-1, ICOS, lymphocyte function-associated antigen-1 (LFA-1 (CD1 1a/CD18), CD247, CD276 (B7-H3), LIGHT (tumor necrosis factor superfamily member 14; TNFSF14), NKG2C, Ig alpha (CD79a), DAP-10, Fc gamma receptor, MHC class I molecule, TNF, TNFr, integrin, signaling lymphocytic activation molecule, BTLA, Toll ligand receptor, ICAM-1, B7-H3, CDS, ICAM-1, GITR, BAFFR, LIGHT, HVEM (LIGHTR), KIRDS2, SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD19, CD4, CD8alpha, CD8beta, IL-2R beta, IL-2R gamma, IL-7R alpha, ITGA4, VLA1, CD49a, ITGA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD, CD1-1d, ITGAE, CD103, ITGAL, CD1-1a, LFA-1, ITGAM, CD1-1b, ITGAX, CD1-1c, ITGB1, CD29, ITGB2, CD18, LFA-1, ITGB7, NKG2D, TNFR2, TRANCE/RANKL, DNAM1 (CD226), SLAMF4 (CD244, 2B4), CD84, CD96 (Tactile), CEACAM1, CRT AM, Ly9 (CD229), CD160 (BY55), PSGL1, CD100 (SEMA4D), CD69, SLAMF6 (NTB-A, Ly108), SLAM (SLAMF1, CD150, IPO-3), BLAME (SLAMF8), SELPLG (CD162), LTBR, LAT, GADS, SLP-76, PAG/Cbp, CD19a, CD83 ligand, or fragments or combinations thereof. It will be appreciated that additional costimulatory molecules, or fragments thereof, not listed above are within the scope of the invention.
Activating Domains
[0095] CD3 is an element of the T cell receptor on native T cells, and has been shown to be an important intracellular activating element in CARs. In a preferred embodiment, the CD3 is CD3 zeta, the nucleotide sequence of which is set forth in SEQ ID NO. 9:
TABLE-US-00012 AGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATCAGCAGGGCC AGAACCAACTGTATAACGAGCTCAACCTGGGACGCAGGGAAGAGTATG ACGTTTTGGACAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAACC AAGACGAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGA TAAGATGGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGAG AAGGGGAAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACTGCTACG AAGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCACCTAGG
[0096] The corresponding amino acid of intracellular CD3 zeta is set forth in SEQ ID NO. 10:
TABLE-US-00013 RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPR RKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDT YDALHMQALPPR
Domain Orientation Relative to the Cell
[0097] Structurally, it will appreciated that the domains described herein correspond to locations relative to an immune or other cell. These domains thus can be part of the (i) "hinge" or extracellular (EC) domain, (ii) the transmembrane (TM) domain, and/or (iii) the intracellular/cytoplasmic domain (IC). The intracellular component frequently comprises, in part, an activating domain such as a portion of a member of the CD3 family, preferably CD3 zeta. This domain is capable of activating the T cell upon binding of the antigen binding molecule to its target. It will be appreciated that the intracellular domain typically further comprises one or more costimulatory molecules as described herein.
[0098] "Activation" or "Stimulation" as used herein, refers to a primary response induced by binding of an activating molecule with its cognate ligand, wherein the binding mediates a signal transduction event.
[0099] An "activating molecule" or "stimulating molecule" refers to a molecule on a T cell, e.g., the TCR/CD3 complex that specifically binds with a cognate stimulatory ligand present on an antigen present cell. Suitable activating molecules are described herein.
[0100] A "costimulatory molecule" as used herein refers to a molecule that provides a signal which mediates a T cell response, including, but not limited to, proliferation, activation, differentiation, and the like. Costimulatory molecules can provide a signal in addition to the primary signal provided by an activating molecule as described herein.
[0101] Suitable costimulatory molecules include, but are not limited to, all or portions of CD28, CD28T, OX40, 4-1BB/CD137, CD2, CD3 (alpha, beta, delta, epsilon, gamma, zeta), CD4, CD5, CD7, CD9, CD16, CD22, CD27, CD30, CD 33, CD37, CD40, CD 45, CD64, CD80, CD86, CD134, CD137, CD154, PD-1, ICOS, lymphocyte function-associated antigen-1 (LFA-1 (CD1 1a/CD18), CD247, CD276 (B7-H3), LIGHT (tumor necrosis factor superfamily member 14; TNFSF14), NKG2C, Ig alpha (CD79a), DAP-10, Fc gamma receptor, MHC class I molecule, TNF, TNFr, integrin, signaling lymphocytic activation molecule, BTLA, Toll ligand receptor, ICAM-1, B7-H3, CDS, ICAM-1, GITR, BAFFR, LIGHT, HVEM (LIGHTR), KIRDS2, SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD19, CD4, CD8alpha, CD8beta, IL2R beta, IL2R gamma, IL7R alpha, ITGA4, VLA1, CD49a, ITGA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD, CD1-1d, ITGAE, CD103, ITGAL, CD1-1a, LFA-1, ITGAM, CD1-1b, ITGAX, CD1-1c, ITGB1, CD29, ITGB2, CD18, LFA-1, ITGB7, NKG2D, TNFR2, TRANCE/RANKL, DNAM1 (CD226), SLAMF4 (CD244, 2B4), CD84, CD96 (Tactile), CEACAM1, CRT AM, Ly9 (CD229), CD160 (BY55), PSGL1, CD100 (SEMA4D), CD69, SLAMF6 (NTB-A, Ly108), SLAM (SLAMF1, CD150, IPO-3), BLAME (SLAMF8), SELPLG (CD162), LTBR, LAT, GADS, SLP-76, PAG/Cbp, CD19a, CD83 ligand, or fragments or combinations thereof. It will be appreciated that the hinge region may contain some or all of a member of the immunoglobulin family such as IgG1, IgG2, IgG3, IgG4, IgA, IgD, IgE, IgM, or fragment thereof.
[0102] In some embodiments, the extracellular domain is positioned between the antigen binding molecule and the transmembrane domain.
[0103] Exemplary CAR constructs in accordance with the invention are set forth in Table 1.
TABLE-US-00014 TABLE 1 Construct Name scFv Hinge Domain Activating Domain 24C1 CD28T 24C1 CD28T CD3 zeta 24C1 CD28 24C1 CD28 CD3 zeta 24C1 CD8 24C1 CD8 CD3 zeta 24C8 CD28T 24C8 CD28T CD3 zeta 24C8 CD28 24C8 CD28 CD3 zeta 24C8 CD8 24C8 CD8 CD3 zeta 20C5.1 CD28T 20C5.1 CD28T CD3 zeta 20C5.1 CD28 20C5.1 CD28 CD3 zeta 20C5.1 CD8 20C5.1 CD8 CD3 zeta 20C5.2 CD28T 20C5.2 CD28T CD3 zeta 20C5.2 CD28 20C5.2 CD28 CD3 zeta 20C5.2 CD8 20C5.2 CD8 CD3 zeta
[0104] As noted, the engineered T cells of the invention comprise an antigen binding molecule (such as an scFv), an extracellular domain (which may comprise a "hinge" domain), a transmembrane domain, and an intracellular domain. The intracellular domain can comprise at least in part an activating domain, preferably comprised of a CD3 family member such as CD3 zeta, CD3 epsilon, CD3 gamma, or portions thereof.
[0105] It will further be appreciated that the antigen binding molecule (e.g., one or more scFvs) is engineered such that it is located in the extracellular portion of the molecule/construct, such that it is capable of recognizing and binding to its target or targets.
[0106] Extracellular Domain.
[0107] Extracellular domains of particular use in this invention may be derived from (i.e., comprise) all or some of CD28, OX-40, 4-1BB/CD137, CD2, CD7, CD27, CD30, CD40, programmed death-1 (PD-1), inducible T cell costimulator (ICOS), lymphocyte function-associated antigen-1 (LFA-1, CD1-1a/CD18), CD3 gamma, CD3 delta, CD3 epsilon, CD247, CD276 (B7-H3), LIGHT, (TNFSF14), NKG2C, Ig alpha (CD79a), DAP-10, Fc gamma receptor, MHC class 1 molecule, TNF receptor proteins, an Immunoglobulin protein, cytokine receptor, integrins, Signaling Lymphocytic Activation Molecules (SLAM proteins), activating NK cell receptors, BTLA, a Toll ligand receptor, ICAM-1, B7-H3, CDS, ICAM-1, GITR, BAFFR, LIGHT, HVEM (LIGHTR), KIRDS2, SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD19, CD4, CD8alpha, CD8beta, IL-2R beta, IL-2R gamma, IL-7R alpha, ITGA4, VLA1, CD49a, ITGA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD, CD1 1d, ITGAE, CD103, ITGAL, CD1 1a, LFA-1, ITGAM, CD1 1b, ITGAX, CD1 1c, ITGB1, CD29, ITGB2, CD18, LFA-1, ITGB7, NKG2D, TNFR2, TRANCE/RANKL, DNAM1 (CD226), SLAMF4 (CD244, 2B4), CD84, CD96 (Tactile), CEACAM1, CRT AM, Ly9 (CD229), CD160 (BY55), PSGL1, CD100 (SEMA4D), CD69, SLAMF6 (NTB-A, Ly108), SLAM (SLAMF1, CD150, IPO-3), BLAME (SLAMF8), SELPLG (CD162), LTBR, LAT, GADS, SLP-76, PAG/Cbp, CD19a, a ligand that specifically binds with CD83, or any combination thereof. The extracellular domain may be derived either from a natural or from a synthetic source.
[0108] Extracellular domains often comprise the hinge portion, sometimes referred to as the "spacer" region. A variety of hinges can be employed in accordance with the invention, including portions or derivatives of the molecules described herein.
[0109] In certain embodiments, the hinge region comprises an amino acid sequence that is at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% identical to the extracellular domain amino acid sequences set forth herein.
[0110] In certain embodiments, the hinge region comprises an amino acid sequence that is at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% identical to the extracellular nucleotide amino acid sequences set forth herein.
Transmembrane Domain
[0111] The CAR can be designed with a transmembrane domain that is fused to the extracellular domain of the CAR. It can similarly be fused to the intracellular domain of the CAR. In some instances, the transmembrane domain can be selected or modified by amino acid substitution to avoid binding of such domains to the transmembrane domains of the same or different surface membrane proteins to minimize interactions with other members of the receptor complex. The transmembrane domain may be derived either from a natural or from a synthetic source. Where the source is natural, the domain may be derived from any membrane-bound or transmembrane protein. Transmembrane regions of particular use in this invention may be derived from (comprise, or correspond to) CD28, CD28T, OX-40, 4-1BB/CD137, CD2, CD7, CD27, CD30, CD40, programmed death-1 (PD-1), inducible T cell costimulator (ICOS), lymphocyte function-associated antigen-1 (LFA-1, CD1-1a/CD18), CD3 gamma, CD3 delta, CD3 epsilon, CD247, CD276 (B7-H3), LIGHT, (TNFSF14), NKG2C, Ig alpha (CD79a), DAP-10, Fc gamma receptor, MHC class 1 molecule, TNF receptor proteins, an Immunoglobulin protein, cytokine receptor, integrins, Signaling Lymphocytic Activation Molecules (SLAM proteins), activating NK cell receptors, BTLA, a Toll ligand receptor, ICAM-1, B7-H3, CDS, ICAM-1, GITR, BAFFR, LIGHT, HVEM (LIGHTR), KIRDS2, SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD19, CD4, CD8alpha, CD8beta, IL-2R beta, IL-2R gamma, IL-7R alpha, ITGA4, VLA1, CD49a, ITGA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD, CD1 1d, ITGAE, CD103, ITGAL, CD1 1a, LFA-1, ITGAM, CD1 1b, ITGAX, CD1 1c, ITGB1, CD29, ITGB2, CD18, LFA-1, ITGB7, NKG2D, TNFR2, TRANCE/RANKL, DNAM1 (CD226), SLAMF4 (CD244, 2B4), CD84, CD96 (Tactile), CEACAM1, CRT AM, Ly9 (CD229), CD160 (BY55), PSGL1, CD100 (SEMA4D), CD69, SLAMF6 (NTB-A, Ly108), SLAM (SLAMF1, CD150, IPO-3), BLAME (SLAMF8), SELPLG (CD162), LTBR, LAT, GADS, SLP-76, PAG/Cbp, CD19a, a ligand that specifically binds with CD83, or any combination thereof.
[0112] Optionally, short linkers may form linkages between any or some of the extracellular, transmembrane, and intracellular domains of the CAR.
[0113] In other embodiments, the transmembrane domain in the CAR of the invention is a CD8 transmembrane domain. In one embodiment, the CD8 transmembrane domain comprises the transmembrane portion of the nucleic acid sequence of SEQ ID NO: 13. In another embodiment, the CD8 transmembrane domain comprises the nucleic acid sequence that encodes the transmembrane amino acid sequence contained within SEQ ID NO: 14.
[0114] In certain embodiments, the transmembrane domain in the CAR of the invention is the CD28 transmembrane domain. In one embodiment, the CD28 transmembrane domain comprises the nucleic acid sequence of SEQ ID NO: 5. In one embodiment, the CD28 transmembrane domain comprises the nucleic acid sequence that encodes the amino acid sequence of SEQ ID NO: 6. In another embodiment, the CD28 transmembrane domain comprises the amino acid sequence of SEQ ID NO: 6.
[0115] Intracellular (Cytoplasmic) Domain.
[0116] The intracellular (cytoplasmic) domain of the engineered T cells of the invention can provide activation of at least one of the normal effector functions of the immune cell. Effector function of a T cell, for example, may refer to cytolytic activity or helper activity, including the secretion of cytokines.
[0117] It will be appreciated that suitable intracellular molecules include (i.e., comprise), but are not limited to signaling domains derived from (or corresponding to) CD28, to CD28T, OX-40, 4-1BB/CD137, CD2, CD7, CD27, CD30, CD40, programmed death-1 (PD-1), inducible T cell costimulator (ICOS), lymphocyte function-associated antigen-1 (LFA-1, CD1-1a/CD18), CD3 gamma, CD3 delta, CD3 epsilon, CD247, CD276 (B7-H3), LIGHT, (TNFSF14), NKG2C, Ig alpha (CD79a), DAP-10, Fc gamma receptor, MHC class 1 molecule, TNF receptor proteins, an Immunoglobulin protein, cytokine receptor, integrins, Signaling Lymphocytic Activation Molecules (SLAM proteins), activating NK cell receptors, BTLA, a Toll ligand receptor, ICAM-1, B7-H3, CDS, ICAM-1, GITR, BAFFR, LIGHT, HVEM (LIGHTR), KIRDS2, SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD19, CD4, CD8alpha, CD8beta, IL-2R beta, IL-2R gamma, IL-7R alpha, ITGA4, VLA1, CD49a, ITGA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD, CD1 1d, ITGAE, CD103, ITGAL, CD1 1a, LFA-1, ITGAM, CD1 1b, ITGAX, CD1 1c, ITGB1, CD29, ITGB2, CD18, LFA-1, ITGB7, NKG2D, TNFR2, TRANCE/RANKL, DNAM1 (CD226), SLAMF4 (CD244, 2B4), CD84, CD96 (Tactile), CEACAM1, CRT AM, Ly9 (CD229), CD160 (BY55), PSGL1, CD100 (SEMA4D), CD69, SLAMF6 (NTB-A, Ly108), SLAM (SLAMF1, CD150, IPO-3), BLAME (SLAMF8), SELPLG (CD162), LTBR, LAT, GADS, SLP-76, PAG/Cbp, CD19a, a ligand that specifically binds with CD83, or any combination thereof.
[0118] In a preferred embodiment, the intracellular/cytoplasmic domain of the CAR can be designed to comprise the CD3 zeta domain by itself or combined with any other desired intracellular domain(s) useful in the context of the CAR of the invention. For example, the intracellular domain of the CAR can comprise a CD3 zeta chain portion and a portion of a costimulatory signaling molecule. The intracellular signaling sequences within the intracellular signaling portion of the CAR of the invention may be linked to each other in a random or specified order.
[0119] In another preferred embodiment, the intracellular domain is designed to comprise the activating domain of CD3 zeta and a signaling domain of CD28. In another embodiment, the intracellular domain is designed to comprise the activating domain of CD3 zeta and a signaling domain of 4-1BB. In another embodiment, the intracellular domain in the CAR is designed to comprise a portion of CD28 and CD3 zeta, wherein the intracellular CD28 comprises the nucleic acid sequence set forth in SEQ ID NO: 7 and the amino acid sequence set forth in SEQ ID NO. 8. The CD3 zeta nucleic acid sequence is set forth in SEQ ID NO: 9, and the amino acid sequence is set forth in SEQ ID NO. 8.
[0120] It will be appreciated that one preferred orientation of the CARs in accordance with the invention comprises an antigen binding molecule (such as scFv) in tandem with an extracellular and/or hinge domain, a costimulatory domain, and an activating domain. It will be further appreciated that multiple domains can be utilized in tandem.
[0121] In some embodiments, isolated nucleic acids are provided comprising a promoter operably linked to a first polynucleotide encoding an antigen binding molecule, at least one costimulatory molecule, and an activating domain. In some embodiments, the nucleic acid construct is contained within a viral vector. In some embodiments, the viral vector is selected from the group consisting of retroviral vectors, murine leukemia virus vectors, SFG vectors, adenoviral vectors, lentiviral vectors, adeno-associated virus (AAV) vectors, Herpes virus vectors, and vaccinia virus vectors. In some embodiments, the nucleic acid is contained within a plasmid.
[0122] In some embodiments, the engineered immune cell is a T cell, tumor infiltrating lymphocyte (TIL), NK cell, TCR-expressing cell, dendritic cell, or NK-T cell. In some embodiments, the cell is obtained or prepared from peripheral blood. In some embodiments, the cell is obtained or prepared from peripheral blood mononuclear cells (PBMCs). In some embodiments, the cell is obtained or prepared from bone marrow. In some embodiments, the cell is obtained or prepared from umbilical cord blood. In some embodiments, the cell is a human cell. In some embodiments, the cell is transfected or transduced by the nucleic acid vector using a method selected from the group consisting of electroporation, sonoporation, biolistics (e.g., Gene Gun), lipid transfection, polymer transfection, nanoparticles, or polyplexes.
[0123] In some embodiments, chimeric antigen receptors are expressed in the engineered immune cells that comprise the nucleic acids of the present application. These chimeric antigen receptors of the present application may comprise, in some embodiments, (i) an antigen binding molecule (such as an scFv), (ii) a transmembrane region, and (iii) a T cell activation molecule or region.
[0124] It is to be further understood that wherever aspects are described herein with the language "comprising," otherwise analogous aspects described in terms of "consisting of" and/or "consisting essentially of" are also provided.
[0125] Additionally, the terms "about" or "comprising essentially of" refer to a value or composition that is within an acceptable error range for the particular value or composition as determined by one of ordinary skill in the art, which will depend in part on how the value or composition is measured or determined, i.e., the limitations of the measurement system. For example, "about" or "comprising essentially of" can mean within 1 or more than 1 standard deviation per the practice in the art. Alternatively, "about" or "comprising essentially of" can mean a range of up to 10% (i.e., .+-.10%). For example, about 3 mg can include any number between 2.7 mg and 3.3 mg (for 10%). Furthermore, particularly with respect to biological systems or processes, the terms can mean up to an order of magnitude or up to 5-fold of a value. When particular values or compositions are provided in the application and claims, unless otherwise stated, the meaning of "about" or "comprising essentially of" should be assumed to be within an acceptable error range for that particular value or composition.
Antigen Binding Molecules
[0126] Antigen binding molecules are within the scope of the invention. An "antigen binding molecule" as used herein means any protein that binds a specified target antigen. In the instant application, the specified target antigen is the CLL-1 protein or fragment thereof. Antigen binding molecules include, but are not limited to antibodies and binding parts thereof, such as immunologically functional fragments. Peptibodies (i.e., Fc fusion molecules comprising peptide binding domains) are another example of suitable antigen binding molecules.
[0127] In certain embodiments, the invention is directed to an antigen binding molecule comprising:
[0128] (a) a heavy chain variable region (VH) complementarity determining region (CDR) 1 comprising the amino acid sequence GX.sub.2X.sub.3X.sub.4X.sub.5X.sub.6X.sub.7X.sub.8X.sub.9 (SEQ ID NO: 134), wherein X.sub.2 is G, F, or Y; X.sub.3 is S or T; X.sub.4 is I, F, or L; X.sub.5 is S or T; X.sub.6 is not present or S; X.sub.7 is not present or G; X.sub.8 is not present or E or G; and X.sub.9 is F, L, or Y;
[0129] (b) a heavy chain variable region (VH) complementarity determining region (CDR) 2 comprising the amino acid sequence X.sub.1X.sub.2X.sub.3X.sub.4X.sub.5X.sub.6 (SEQ ID NO: 135), wherein X.sub.1 is D, H, S, or Y; X.sub.2 is H, P, or Y; X.sub.3 is D, E, or S; X.sub.4 is D or G; X.sub.5 is G or S; and X.sub.6 is not present of D or E;
[0130] (c) a heavy chain variable region (VH) complementarity determining region (CDR) 3, comprising the amino acid sequence X.sub.1X.sub.2X.sub.3X.sub.4X.sub.5X.sub.6X.sub.7X.sub.8X.sub.9X.sub.10X.- sub.11X.sub.12DY (SEQ ID NO: 136), wherein X.sub.1 is E or L; X.sub.2 is R, S, or V; X.sub.3 is R or Y; X.sub.4 is C, G, or S; X.sub.5 is not present or G or I; X.sub.6 is not present or G; X.sub.7 is not present or D; X.sub.8 is not present or C; X.sub.9 is not present or W or Y; X.sub.10 is not present or P or S; X.sub.11 is not present or G or Y; and X.sub.12 is F or R;
[0131] (d) a light chain variable region (VL) CDR1 comprising the amino acid sequence X.sub.1ASQX.sub.5X.sub.6X.sub.7X.sub.8X.sub.9LX.sub.11 (SEQ ID NO: 137), wherein X.sub.1 is Q or R; X.sub.5 is D or S; X.sub.6 is I or V; X.sub.7 is N or S; X.sub.8 is N or S; X.sub.9 is F, L, or Y; and X.sub.11 is N or T;
[0132] (e) a light chain variable region (VL) CDR2 comprising the amino acid sequence X.sub.1ASX.sub.4X.sub.5X.sub.6X.sub.7 (SEQ ID NO: 138), wherein X.sub.1 is D or G; X.sub.4 is N, S, or T; X.sub.5 is L or R; X.sub.6 is A, E, or K; and X.sub.7 is S or T; and/or
[0133] (f) a light chain variable region (VL) CDR3 comprising the amino acid sequence QQX.sub.3X.sub.4X.sub.5X.sub.6PX.sub.8T (SEQ ID NO: 139), wherein X.sub.3 is S or Y; X.sub.4 is D, G, or Y; X.sub.5 is N, S, or T; X.sub.6 is L, T, or Y; and X.sub.8 is F or I.
[0134] In some embodiments, the invention relates to antigen binding molecules comprising at least one of: (a) a variable heavy chain CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 17, 51, 73, 95, 5, and 97; (b) a variable heavy chain CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs 18, 52, 74, 96; (c) a variable heavy chain CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs SEQ ID NO: 19, 53, 75, and 97; (d) a variable light chain CDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 22, 56, 78, and 100; (e) a variable light chain CDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs. 23, 57, 79, and 101; (f) a variable light chain CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs. 24, 58, 80, and 102.
[0135] In other embodiments, the invention relates to antigen binding molecules (and chimeric antigen receptors comprising these molecules) comprising at least one of: (a) a VH region comprising the amino acid sequence of SEQ ID NO: 16 and a VL region comprising the amino acid sequence of SEQ ID NO: 21; (b) a VH region comprising the amino acid sequence of SEQ ID NO: 50 and a VL region comprising the amino acid sequence of SEQ ID NO: 55; (c) a VH region comprising the amino acid sequence of SEQ ID NO: 72 and a VL region comprising the amino acid sequence of SEQ ID NO: 77; (d) a VH region comprising the amino acid sequence of SEQ ID NO: 94 and a VL region comprising the amino acid sequence of SEQ ID NO: 99; and wherein the VH and VL region or regions are linked by at least one linker. In other embodiments, the invention relates to antigen binding molecules (and chimeric antigen receptors comprising these molecules) wherein the linker comprises at least one of SEQ ID NO. 130 and SEQ ID NO. 132.
[0136] In further embodiments, the invention relates to antigen binding molecules comprising a variable light (VL) chain CDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 24, 58, 80, and 102.
[0137] In other embodiments, the invention relates to isolated polynucleotides encoding an antigen binding molecule that specifically binds to CLL-1, wherein the antigen binding molecule heavy chain comprises CDR1 (SEQ ID NO. 17), CDR2 (SEQ ID NO. 18), and CDR3 (SEQ ID NO. 19) and the antigen binding molecule light chain comprises CDR1 (SEQ ID NO. 22), CDR2 (SEQ ID NO. 23), and CDR3 (SEQ ID NO. 24).
[0138] In other embodiments, the invention relates to an antigen binding molecule that specifically binds to CLL-1, wherein the antigen binding molecule heavy chain comprises CDR1 (SEQ ID NO. 51), CDR2 (SEQ ID NO. 52), and CDR3 (SEQ ID NO. 53) and the antigen binding molecule light chain comprises CDR1 (SEQ ID NO. 56), CDR2 (SEQ ID NO. 57), and CDR3 (SEQ ID NO. 58).
[0139] In other embodiments, the invention relates an antigen binding molecule that specifically binds to CLL-1, wherein the antigen binding molecule heavy chain comprises CDR1 (SEQ ID NO. 73), CDR2 (SEQ ID NO. 74), and CDR3 (SEQ ID NO. 75) and the antigen binding molecule light chain comprises CDR1 (SEQ ID NO. 78), CDR2 (SEQ ID NO. 79), and CDR3 (SEQ ID NO. 80).
[0140] In other embodiments, the invention relates to isolated polynucleotides encoding an antigen binding molecule that specifically binds to CLL-1, wherein the antigen binding molecule heavy chain comprises CDR1 (SEQ ID NO. 95), CDR2 (SEQ ID NO. 96), and CDR3 (SEQ ID NO. 97) and the antigen binding molecule light chain comprises CDR1 (SEQ ID NO. 100), CDR2 (SEQ ID NO. 101), and CDR3 (SEQ ID NO. 102).
[0141] In certain embodiments, the present invention is directed to an isolated polynucleotide encoding an anti-CLL-1 antigen binding molecule which cross competes with one or more antibodies described herein or an antigen binding molecule thereof encoded by the polynucleotide. In one embodiment, the invention is directed to isolated polynucleotides encoding an anti-CLL-1 antigen binding molecule thereof which binds to the same epitope as one or more of the antigen binding molecules described herein.
[0142] In some embodiments, the antigen binding molecule binds to an antigen on a tumor cell. In some embodiments, the antigen binding molecule binds to an antigen on a cell involved in a hyperproliferative disease or to a viral or bacterial antigen. In further embodiments, the antigen binding molecule is an antibody of fragment thereof, including one or more of the complementarity determining regions (CDRs) thereof. In further embodiments, the antigen binding molecule is a single chain variable fragment (scFv).
[0143] The term "immunologically functional fragment" (or "fragment") of an antigen binding molecule is a species of antigen binding molecule comprising a portion (regardless of how that portion is obtained or synthesized) of an antibody that lacks at least some of the amino acids present in a full-length chain but which is still capable of specifically binding to an antigen. Such fragments are biologically active in that they bind to the target antigen and can compete with other antigen binding molecules, including intact antibodies, for binding to a given epitope. In some embodiments, the fragments are neutralizing fragments. In some embodiments, the fragments can block or reduce the activity of CLL-1. In one aspect, such a fragment will retain at least one CDR present in the full-length light or heavy chain, and in some embodiments will comprise a single heavy chain and/or light chain or portion thereof. These fragments can be produced by recombinant DNA techniques, or can be produced by enzymatic or chemical cleavage of antigen binding molecules, including intact antibodies.
[0144] Immunologically functional immunoglobulin fragments include, but are not limited to, scFv fragments, Fab fragments (Fab', F(ab').sub.2, and the like), one or more CDR, a diabody (heavy chain variable domain on the same polypeptide as a light chain variable domain, connected via a short peptide linker that is too short to permit pairing between the two domains on the same chain), domain antibodies, and single-chain antibodies. These fragments can be derived from any mammalian source, including but not limited to human, mouse, rat, camelid or rabbit. As will be appreciated by one of skill in the art, an antigen binding molecule can include non-protein components.
[0145] Variants of the antigen binding molecules are also within the scope of the invention, e.g., variable light and/or variable heavy chains that each have at least 70-80%, 80-85%, 85-90%, 90-95%, 95-97%, 97-99%, or above 99% identity to the amino acid sequences of the sequences described herein. In some instances, such molecules include at least one heavy chain and one light chain, whereas in other instances the variant forms contain two identical light chains and two identical heavy chains (or subparts thereof). A skilled artisan will be able to determine suitable variants of the antigen binding molecules as set forth herein using well-known techniques. In certain embodiments, one skilled in the art can identify suitable areas of the molecule that may be changed without destroying activity by targeting regions not believed to be important for activity.
[0146] In certain embodiments, the polypeptide structure of the antigen binding molecules is based on antibodies, including, but not limited to, monoclonal antibodies, bispecific antibodies, minibodies, domain antibodies, synthetic antibodies (sometimes referred to herein as "antibody mimetics"), chimeric antibodies, humanized antibodies, human antibodies, antibody fusions (sometimes referred to herein as "antibody conjugates"), and fragments thereof, respectively. In some embodiments, the antigen binding molecule comprises or consists of avimers.
[0147] In some embodiments, an antigen binding molecule to CLL-1 is administered as part of a CAR, TCR, or other immune cell. In such immune cells, the antigen binding molecule to CLL-1 can be under the control of the same promoter region, or a separate promoter. In certain embodiments, the genes encoding protein agents and/or an antigen binding molecule to CLL-1 can be in separate vectors.
[0148] The invention further provides for pharmaceutical compositions comprising an antigen binding molecule to CLL-1 together with a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative and/or adjuvant. In certain embodiments, pharmaceutical compositions will include more than one different antigen binding molecule to CLL-1. In certain embodiments, pharmaceutical compositions will include more than one antigen binding molecule to CLL-1 wherein the antigen binding molecules to CLL-1 bind more than one epitope. In some embodiments, the various antigen binding molecules will not compete with one another for binding to CLL-1.
[0149] In other embodiments, the pharmaceutical composition can be selected for parenteral delivery, for inhalation, or for delivery through the digestive tract, such as orally. The preparation of such pharmaceutically acceptable compositions is within the ability of one skilled in the art. In certain embodiments, buffers are used to maintain the composition at physiological pH or at a slightly lower pH, typically within a pH range of from about 5 to about 8. In certain embodiments, when parenteral administration is contemplated, a therapeutic composition can be in the form of a pyrogen-free, parenterally acceptable aqueous solution comprising a desired antigen binding molecule to CLL-1, with or without additional therapeutic agents, in a pharmaceutically acceptable vehicle. In certain embodiments, a vehicle for parenteral injection is sterile distilled water in which an antigen binding molecule to CLL-1, with or without at least one additional therapeutic agent, is formulated as a sterile, isotonic solution, properly preserved. In certain embodiments, the preparation can involve the formulation of the desired molecule with polymeric compounds (such as polylactic acid or polyglycolic acid), beads or liposomes that can provide for the controlled or sustained release of the product which can then be delivered via a depot injection. In certain embodiments, implantable drug delivery devices can be used to introduce the desired molecule.
[0150] In some embodiments, the antigen binding molecule is used as a diagnostic or validation tool. The antigen binding molecule can be used to assay the amount of CLL-1 present in a sample and/or subject. In some embodiments, the diagnostic antigen binding molecule is not neutralizing. In some embodiments, the antigen binding molecules disclosed herein are used or provided in an assay kit and/or method for the detection of CLL-1 in mammalian tissues or cells in order to screen/diagnose for a disease or disorder associated with changes in levels of CLL-1. The kit can comprise an antigen binding molecule that binds CLL-1, along with means for indicating the binding of the antigen binding molecule with CLL-1, if present, and optionally CLL-1 protein levels.
[0151] The antigen binding molecules will be further understood in view of the definitions and descriptions below.
[0152] An "Fc" region comprises two heavy chain fragments comprising the CH1 and CH2 domains of an antibody. The two heavy chain fragments are held together by two or more disulfide bonds and by hydrophobic interactions of the CH3 domains.
[0153] A "Fab fragment" comprises one light chain and the CH1 and variable regions of one heavy chain. The heavy chain of a Fab molecule cannot form a disulfide bond with another heavy chain molecule. A "Fab' fragment" comprises one light chain and a portion of one heavy chain that contains the VH domain and the CH1 domain and also the region between the CH1 and CH2 domains, such that an interchain disulfide bond can be formed between the two heavy chains of two Fab' fragments to form an F(ab')2 molecule. An "F(ab')2 fragment" contains two light chains and two heavy chains containing a portion of the constant region between the CH1 and CH2 domains, such that an interchain disulfide bond is formed between the two heavy chains. An F(ab')2 fragment thus is composed of two Fab' fragments that are held together by a disulfide bond between the two heavy chains.
[0154] The "Fv region" comprises the variable regions from both the heavy and light chains, but lacks the constant regions.
[0155] "Single chain variable fragment" ("scFv", also termed "single-chain antibody") refers to Fv molecules in which the heavy and light chain variable regions have been connected by a flexible linker to form a single polypeptide chain, which forms an antigen binding region. See PCT application WO88/01649 and U.S. Pat. Nos. 4,946,778 and 5,260,203, the disclosures of which are incorporated by reference in their entirety.
[0156] A "bivalent antigen binding molecule" comprises two antigen binding sites. In some instances, the two binding sites have the same antigen specificities. Bivalent antigen binding molecules can be bispecific. A "multispecific antigen binding molecule" is one that targets more than one antigen or epitope. A "bispecific," "dual-specific" or "bifunctional" antigen binding molecule is a hybrid antigen binding molecule or antibody, respectively, having two different antigen binding sites. The two binding sites of a bispecific antigen binding molecule will bind to two different epitopes, which can reside on the same or different protein targets.
[0157] An antigen binding molecule is said to "specifically bind" its target antigen when the dissociation constant (K.sub.d) is .about.1.times.10.sup.-7 M. The antigen binding molecule specifically binds antigen with "high affinity" when the K.sub.d is 1-5.times.10.sup.-9 M, and with "very high affinity" when the K.sub.d is 1-5.times.10.sup.-10 M. In one embodiment, the antigen binding molecule has a K.sub.d of 10.sup.-9 M. In one embodiment, the off-rate is <1.times.10.sup.-5. In other embodiments, the antigen binding molecules will bind to human CLL-1 with a K.sub.d of between about 10.sup.-7M and 10.sup.-13 M, and in yet another embodiment the antigen binding molecules will bind with a K.sub.d 1.0-5.times.10.sup.-10.
[0158] In some embodiments, the antibody or antigen binding molecules of the present invention specifically bind CLL-1 (e.g., hCLL-1). In certain embodiments, an anti-CLL-1 antibody or antigen binding molecule of the present invention binds human CLL-1 with a K.sub.D of less than 1.times.10.sup.-6 M, less than 1.times.10.sup.-7M, less than 1.times.10.sup.-8 M, or less than 1.times.10.sup.-9M. In one particular embodiment, the anti-CLL-1 antibody or antigen binding molecules binds human CLL-1 with a K.sub.D of less than 1.times.10.sup.-7M. In another embodiment, the anti-CLL-1 antibody or antigen binding molecules binds human CLL-1 with a K.sub.D of less than 1.times.10.sup.-8 M. In some embodiments, the anti-CLL-1 antibody or antigen binding molecules binds human CLL-1 with a K.sub.D of about 1.times.10.sup.-7M, about 2.times.10.sup.-7M, about 3.times.10.sup.-7M, about 4.times.10.sup.-7M, about 5.times.10.sup.-7M, about 6.times.10.sup.-7M, about 7.times.10.sup.-7M, about 8.times.10.sup.-7M, about 9.times.10.sup.-7 M, about 1.times.10.sup.-8 M, about 2.times.10.sup.-8 M, about 3.times.10.sup.-8 M, about 4.times.10.sup.-8 M, about 5.times.10.sup.-8 M, about 6.times.10.sup.-8 M, about 7.times.10.sup.-8 M, about 8.times.10.sup.-8 M, about 9.times.10.sup.-8 M, about 1.times.10.sup.-8 M, about 2.times.10.sup.-9M, about 3.times.10.sup.-9M, about 4.times.10.sup.-9M, about 5.times.10.sup.-9M, about 6.times.10.sup.-9M, about 7.times.10.sup.-9M, about 8.times.10.sup.-9 M, about 9.times.10.sup.-9 M, about 1.times.10.sup.-10 M, or about 5.times.10.sup.-10 M. In certain embodiments, the K.sub.D is calculated as the quotient of k.sub.off/k.sub.on, and the k.sub.on and k.sub.off are determined using a monovalent antibody, such as a Fab fragment, as measured by, e.g., BIAcore.RTM. surface plasmon resonance technology. In other embodiments, the K.sub.D is calculated as the quotient of k.sub.off/k.sub.on, and the k.sub.on and k.sub.off are determined using a bivalent antibody, such as a Fab fragment, as measured by, e.g., BIAcore.RTM. surface plasmon resonance technology.
[0159] In other embodiments, the anti-CLL-1 antibody or antigen binding molecule binds human CLL-1-Fc with a K.sub.D of less than 1.times.10.sup.-9M, less than 3.times.10.sup.-9M, less than 5.times.10.sup.-9 M, less than 1.times.10.sup.-10 M, less than 3.times.10.sup.-10 M, or less than 5.times.10.sup.-10 M. In other embodiments, the anti-CLL-1 antibody or antigen binding molecules binds cyno CLL-1-Fc with a K.sub.D of less than 1.times.10.sup.-5M, less than 1.times.10.sup.-6M, less than 1.times.10.sup.-7 M, less than 1.times.10.sup.-8 M, less than 1.times.10.sup.-9M, or less than 1.times.10.sup.-10 M.
[0160] In some embodiments, the anti-CLL-1 antibody or antigen binding molecule binds human CLL-1 with an association rate (k.sub.on) of less than 1.times.10.sup.-4 M.sup.-1 s.sup.-1, less than 2.times.10.sup.-4M.sup.-1 s.sup.-1, less than 3.times.10.sup.-4 M.sup.-1 s.sup.-1 less than 4.times.10.sup.-4 M.sup.-1 s.sup.-1 less than 5.times.10.sup.-4M.sup.-1 s.sup.-1, less than 6.times.10.sup.-4 M.sup.-1 s.sup.-1, less than 7.times.10.sup.-4 M.sup.-1 s.sup.-1, less than 8.times.10.sup.-4M.sup.-1 s.sup.-1, less than 9.times.10.sup.-4M.sup.-1 s.sup.-1, less than 1.times.10.sup.-5M.sup.-1 s.sup.-1, less than 2.times.10.sup.-5 M.sup.-1 s.sup.-1, less than 3.times.10.sup.-5 M.sup.-1 s.sup.-1, less than 4.times.10.sup.-5 M.sup.-1 s.sup.-1, less than 5.times.10.sup.-5 M.sup.-1 s.sup.-1, less than 6.times.10.sup.-5 M.sup.-1 s.sup.-1, less than 7.times.10.sup.-5 M.sup.-1 s.sup.-1, less than 8.times.10.sup.-5 M.sup.-1 s.sup.-1, less than 9.times.10.sup.-5 M.sup.-1 s.sup.-1, less than 1.times.10.sup.-6 M.sup.-1 s.sup.-1, less than 2.times.10.sup.-6 M.sup.-1s.sup.-1, less than 3.times.10.sup.-6 M.sup.-1 s.sup.-1, less than 4.times.10.sup.-6 M.sup.-1 s.sup.-1, less than 5.times.10.sup.-6 M.sup.-1 s.sup.-1, less than 6.times.10.sup.-6 M.sup.-1 s.sup.-1, less than 7.times.10.sup.-6 M.sup.-1 s.sup.-1, less than 8.times.10.sup.-6 M.sup.-1 s.sup.-1, less than 9.times.10.sup.-6 M.sup.-1 s.sup.-1, or less than 1.times.10.sup.-7M.sup.-1 s.sup.-1. In certain embodiments, the k.sub.on is determined using a monovalent antibody, such as a Fab fragment, as measured by, e.g., BIAcore.RTM. surface plasmon resonance technology. In other embodiments, the k.sub.on is determined using a bivalent antibody as measured by, e.g., BIAcore.RTM. surface plasmon resonance technology.
[0161] In some embodiments, the anti-CLL-1 antibody or antigen binding molecule binds human CLL-1 with an dissociation rate (k.sub.off) of less than 1.times.10's.sup.-1, less than 2.times.10.sup.-2 s.sup.-1, less than 3.times.10.sup.-2 s.sup.-1, less than 4.times.10.sup.-2 s.sup.-1, less than 5.times.10.sup.-2 s.sup.-1, less than 6.times.10.sup.-2 s.sup.-1, less than 7.times.10.sup.-2 s.sup.-1, less than 8.times.10.sup.-2 s.sup.-1, less than 9.times.10.sup.-2 s.sup.-1, less than 1.times.10.sup.-3 s.sup.-1, less than 2.times.10.sup.-3 s.sup.-1, less than 3.times.10.sup.-3 s.sup.-1, less than 4.times.10.sup.-3 s.sup.-1, less than 5.times.10.sup.-3 s.sup.-1, less than 6.times.10.sup.-3 s.sup.-1, less than 7.times.10.sup.-3 s.sup.-1, less than 8.times.10.sup.-3 s.sup.-1, less than 9.times.10.sup.-3 s.sup.-1, less than 1.times.10.sup.-4 s.sup.-1, less than 2.times.10.sup.-4 s.sup.-1, less than 3.times.10.sup.-4 s.sup.-1, less than 4.times.10.sup.-4 s.sup.-1, less than 5.times.10.sup.-4 s.sup.-1, less than 6.times.10.sup.-4 s.sup.-1, less than 7.times.10.sup.-4 s.sup.-1, less than 8.times.10.sup.-4 s.sup.-1, less than 9.times.10.sup.-4 s.sup.-1, less than 1.times.10.sup.-4 s.sup.-1, or less than 5.times.10.sup.-4 s.sup.-1. In certain embodiments, the k.sub.off is determined using a monovalent antibody, such as a Fab fragment, as measured by, e.g., BIAcore.RTM. surface plasmon resonance technology. In other embodiments, the k.sub.off is determined using a bivalent antibody as measured by, e.g., BIAcore.RTM. surface plasmon resonance technology.
[0162] An antigen binding molecule is said to be "selective" when it binds to one target more tightly than it binds to a second target.
[0163] The term "antibody" refers to an intact immunoglobulin of any isotype, or a fragment thereof that can compete with the intact antibody for specific binding to the target antigen, and includes, for instance, chimeric, humanized, fully human, and bispecific antibodies. An "antibody" is a species of an antigen binding molecule as defined herein. An intact antibody will generally comprise at least two full-length heavy chains and two full-length light chains, but in some instances can include fewer chains such as antibodies naturally occurring in camelids which can comprise only heavy chains. Antibodies can be derived solely from a single source, or can be chimeric, that is, different portions of the antibody can be derived from two different antibodies as described further below. The antigen binding molecules, antibodies, or binding fragments can be produced in hybridomas, by recombinant DNA techniques, or by enzymatic or chemical cleavage of intact antibodies. Unless otherwise indicated, the term "antibody" includes, in addition to antibodies comprising two full-length heavy chains and two full-length light chains, derivatives, variants, fragments, and muteins thereof, examples of which are described below. Furthermore, unless explicitly excluded, antibodies include monoclonal antibodies, bispecific antibodies, minibodies, domain antibodies, synthetic antibodies (sometimes referred to herein as "antibody mimetics"), chimeric antibodies, humanized antibodies, human antibodies, antibody fusions (sometimes referred to herein as "antibody conjugates") and fragments thereof, respectively.
[0164] The variable regions typically exhibit the same general structure of relatively conserved framework regions (FR) joined by the 3 hypervariable regions (i.e., "CDRs"). The CDRs from the two chains of each pair typically are aligned by the framework regions, which can enable binding to a specific epitope. From N-terminal to C-terminal, both light and heavy chain variable regions typically comprise the domains FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4. By convention, CDR regions in the heavy chain are typically referred to as HC CDR1, CDR2, and CDR3. The CDR regions in the light chain are typically referred to as LC CDR1, CDR2, and CDR3. The assignment of amino acids to each domain is typically in accordance with the definitions of Kabat, Chothia, or the AbM definition.
[0165] The term "Kabat numbering" and like terms are recognized in the art and refer to a system of numbering amino acid residues in the heavy and light chain variable regions of an antibody, or an antigen-binding portion thereof. In certain aspects, the CDRs of an antibody can be determined according to the Kabat numbering system (see, e.g., Kabat E A & Wu T T (1971) Ann NY Acad Sci 190: 382-391 and Kabat E A et al., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No. 91-3242). Using the Kabat numbering system, CDRs within an antibody heavy chain molecule are typically present at amino acid positions 31 to 35, which optionally can include one or two additional amino acids, following 35 (referred to in the Kabat numbering scheme as 35A and 35B) (CDR1), amino acid positions 50 to 65 (CDR2), and amino acid positions 95 to 102 (CDR3). Using the Kabat numbering system, CDRs within an antibody light chain molecule are typically present at amino acid positions 24 to 34 (CDR1), amino acid positions 50 to 56 (CDR2), and amino acid positions 89 to 97 (CDR3). In a specific embodiment, the CDRs of the antibodies described herein have been determined according to the Kabat numbering scheme.
[0166] In certain aspects, the CDRs of an antibody can be determined according to the Chothia numbering scheme, which refers to the location of immunoglobulin structural loops (see, e.g., Chothia C & Lesk A M, (1987), J Mol Biol 196: 901-917; Al-Lazikani B et al., (1997) J Mol Biol 273: 927-948; Chothia C et al., (1992) J Mol Biol 227: 799-817; Tramontano A et al., (1990) J Mol Biol 215(1): 175-82; and U.S. Pat. No. 7,709,226). Typically, when using the Kabat numbering convention, the Chothia CDR-H1 loop is present at heavy chain amino acids 26 to 32, 33, or 34, the Chothia CDR-H2 loop is present at heavy to chain amino acids 52 to 56, and the Chothia CDR-H3 loop is present at heavy chain amino acids 95 to 102, while the Chothia CDR-L1 loop is present at light chain amino acids 24 to 34, the Chothia CDR-L2 loop is present at light chain amino acids 50 to 56, and the Chothia CDR-L3 loop is present at light chain amino acids 89 to 97. The end of the Chothia CDR-HI loop when numbered using the Kabat numbering convention varies between H32 and H34 depending on the length of the loop (this is because the Kabat numbering scheme places the insertions at H35A and H35B; if neither 35A nor 35B is present, the loop ends at 32; if only 35A is present, the loop ends at 33; if both 35A and 35B are present, the loop ends at 34).
[0167] In a specific embodiment, the CDRs of the antibodies described herein have been determined according to the Chothia numbering scheme.
[0168] A number of definitions of the CDRs are commonly in use: Kabat numbering, Chothia numbering, AbM numbering, or contact numbering. The AbM definition is a compromise between the two used by Oxford Molecular's AbM antibody modelling software. The contact definition is based on an analysis of the available complex crystal structures.
TABLE-US-00015 TABLE 2 CDR Numbering Loop Kabat AbM Chothia Contact L1 L24--L34 L24--L34 L24--L34 L30--L36 L2 L50--L56 L50--L56 L50--L56 L46--L55 L3 L89--L97 L89--L97 L89--L97 L89--L96 H1 H31--H35B H26--H35B H26--H32 . . . 34 H30--H35B (Kabat Numbering) H1 H31--H35 H26--H35 H26--H32 H30--H35 (Chothia Numbering) H2 H50--H65 H50--H58 H52--H56 H47--H58 H3 H95--H102 H95--H102 H95--H102 H93--H101
[0169] As used herein, the term "heavy chain" when used in reference to an antibody can refer to any distinct type, e.g., alpha (.alpha.), delta (.delta.), epsilon (.epsilon.), gamma (.gamma.) and mu (.mu.), based on the amino acid sequence of the constant domain, which give rise to IgA, IgD, IgE, IgG and IgM classes of antibodies, respectively, including subclasses of IgG, e.g., IgG.sub.1, IgG.sub.2, IgG.sub.3 and IgG.sub.4.
[0170] As used herein, the term "light chain" when used in reference to an antibody can refer to any distinct type, e.g., kappa (.kappa.) or lambda (.lamda.) based on the amino acid sequence of the constant domains. Light chain amino acid sequences are well known in the art. In specific embodiments, the light chain is a human light chain.
[0171] The term "variable region" or "variable domain" refers to a portion of the light and/or heavy chains of an antibody, typically including approximately the amino-terminal 120 to 130 amino acids in the heavy chain and about 100 to 110 amino terminal amino acids in the light chain. The variable region of an antibody typically determines specificity of a particular antibody for its target.
[0172] Variability is not evenly distributed throughout the variable domains of antibodies or antigen binding molecules; it is concentrated in sub-domains of each of the heavy and light chain variable regions. These subdomains are called "hypervariable regions" or "complementarity determining regions" (CDRs) as further described herein. The more conserved (i.e., non-hypervariable) portions of the variable domains are called the "framework" regions (FRM or FR) and provide a scaffold for the six CDRs in three dimensional space to form an antigen-binding surface. The variable domains of naturally occurring heavy and light chains each comprise four FRM regions (FR1, FR2, FR3, and FR4), largely adopting a .beta.-sheet configuration, connected by three hypervariable regions, which form loops connecting, and in some cases forming part of, the .beta.-sheet structure. The hypervariable regions in each chain are held together in close proximity by the FRM and, with the hypervariable regions from the other chain, contribute to the formation of the antigen-binding site (see Kabat et al., described further herein.
[0173] Typically, CDRs form a loop structure that can be classified as a canonical structure. The term "canonical structure" refers to the main chain conformation that is adopted by the antigen binding (CDR) loops. From comparative structural studies, it has been found that five of the six antigen binding loops have only a limited repertoire of available conformations. Each canonical structure can be characterized by the torsion angles of the polypeptide backbone. Correspondent loops between antibodies may, therefore, have very similar three dimensional structures, despite high amino acid sequence variability in most parts of the loops (Chothia and Lesk, J. Mol. Biol., 1987, 196: 901; Chothia et al., Nature, 1989, 342: 877; Martin and Thornton, J. Mol. Biol, 1996, 263: 800). Furthermore, there is a relationship between the adopted loop structure and the amino acid sequences surrounding it. The conformation of a particular canonical class is determined by the length of the loop and the amino acid residues residing at key positions within the loop, as well as within the conserved framework (i.e., outside of the loop). Assignment to a particular canonical class can therefore be made based on the presence of these key amino acid residues.
[0174] The term "canonical structure" may also include considerations as to the linear sequence of the antibody, for example, as catalogued by Kabat (Kabat et al., herein). The Kabat numbering scheme (system) is a widely adopted standard for numbering the amino acid residues of an antibody variable domain in a consistent manner and is the preferred scheme applied in the present invention as also mentioned elsewhere herein. Additional structural considerations can also be used to determine the canonical structure of an antibody. For example, those differences not fully reflected by Kabat numbering can be described by the numbering system of Chothia et al. and/or revealed by other techniques, for example, crystallography and two- or three-dimensional computational modeling. Accordingly, a given antibody sequence may be placed into a canonical class which allows for, among other things, identifying appropriate chassis sequences (e.g., based on a desire to include a variety of canonical structures in a library). Kabat numbering of antibody amino acid sequences and structural considerations as described by Chothia et al. (herein) and their implications for construing canonical aspects of antibody structure, are described in the literature. The subunit structures and three-dimensional configurations of different classes of immunoglobulins are well known in the art. For a review of the antibody structure, see Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory, eds. Harlow et al., 1988.
[0175] The CDR3 of the light chain and, particularly, the CDR3 of the heavy chain may constitute the most important determinants in antigen binding within the light and heavy chain variable regions. In some antibody constructs, the heavy chain CDR3 appears to constitute the major area of contact between the antigen and the antibody. In vitro selection schemes in which CDR3 alone is varied can be used to vary the binding properties of an antibody or determine which residues contribute to the binding of an antigen. Hence, CDR3 is typically the greatest source of molecular diversity within the antibody-binding site. H3, for example, can be as short as two amino acid residues or greater than 26 amino acids.
[0176] As used herein, the terms "constant region" and "constant domain" are interchangeable and have a meaning common in the art. The constant region is an antibody portion, e.g., a carboxyl terminal portion of a light and/or heavy chain which is not directly involved in binding of an antibody to antigen but which can exhibit various effector functions, such as interaction with the Fc receptor. The constant region of an immunoglobulin molecule generally has a more conserved amino acid sequence relative to an immunoglobulin variable domain.
[0177] "Binding affinity" generally refers to the strength of the sum total of noncovalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). Unless indicated otherwise, as used herein, "binding affinity" refers to intrinsic binding affinity which reflects a 1:1 interaction between members of a binding pair (e.g., antibody and antigen). The affinity of a molecule X for its partner Y can generally be represented by the dissociation constant (K.sub.D). Affinity can be measured and/or expressed in a number of ways known in the art, including, but not limited to, equilibrium dissociation constant (K.sub.D), and equilibrium association constant (K.sub.A). The K.sub.D is calculated from the quotient of k.sub.off/k.sub.on, whereas K.sub.A is calculated from the quotient of k.sub.on/k.sub.off. k.sub.on refers to the association rate constant of, e.g., an antibody to an antigen, and k.sub.off refers to the dissociation of, e.g., an antibody to an antigen. The k.sub.on and k.sub.off can be determined by techniques known to one of ordinary skill in the art, such as BIAcore.RTM. or KinExA.
[0178] The term "neutralizing" refers to an antigen binding molecule, scFv, or antibody, respectively, that binds to a ligand and prevents or reduces the biological effect of that ligand. This can be done, for example, by directly blocking a binding site on the ligand or by binding to the ligand and altering the ligand's ability to bind through indirect means (such as structural or energetic alterations in the ligand). In some embodiments, the term can also denote an antigen binding molecule that prevents the protein to which it is bound from performing a biological function.
[0179] The term "target" or "antigen" refers to a molecule or a portion of a molecule capable of being bound by an antigen binding molecule. In certain embodiments, a target can have one or more epitopes.
[0180] The term "compete" when used in the context of antigen binding molecules that compete for the same epitope means competition between antigen binding molecules as determined by an assay in which the antigen binding molecule (e.g., antibody or immunologically functional fragment thereof) being tested prevents or inhibits (e.g., reduces) specific binding of a reference antigen binding molecule to an antigen. Numerous types of competitive binding assays can be used to determine if one antigen binding molecule competes with another, for example: solid phase direct or indirect radioimmunoassay (RIA), solid phase direct or indirect enzyme immunoassay (EIA), sandwich competition assay (Stahli et al., 1983, Methods in Enzymology 9:242-253); solid phase direct biotin-avidin EIA (Kirkland et al., 1986, J. Immunol. 137:3614-3619), solid phase direct labeled assay, solid phase direct labeled sandwich assay (Harlow and Lane, 1988, Antibodies, A Laboratory Manual, Cold Spring Harbor Press); solid phase direct label RIA using 1-125 label (Morel et al., 1988, Molec. Immunol. 25:7-15); solid phase direct biotin-avidin EIA (Cheung, et al., 1990, Virology 176:546-552); and direct labeled RIA (Moldenhauer et al., 1990, Scand. J. Immunol. 32:77-82).
[0181] As used herein, the term "epitope" refers to a localized region of an antigen to which an antibody can specifically bind. An epitope can be, for example, contiguous amino acids of a polypeptide (linear or contiguous epitope) or an epitope can, for example, come together from two or more non-contiguous regions of a polypeptide or polypeptides (conformational, non-linear, discontinuous, or non-contiguous epitope). In certain embodiments, the epitope to which an antibody binds can be determined by, e.g., NMR spectroscopy, X-ray diffraction crystallography studies, ELISA assays, hydrogen/deuterium exchange coupled with mass spectrometry (e.g., liquid chromatography electrospray mass spectrometry), array-based oligo-peptide scanning assays, and/or mutagenesis mapping (e.g., site-directed mutagenesis mapping). For X-ray crystallography, crystallization may be accomplished using any of the known methods in the art (e.g., Giege R. et al., (1994) Acta Crystallogr D Biol Crystallogr 50(Pt 4): 339-350; McPherson A (1990) Eur J Biochem 189: 1-23; Chayen N E (1997) Structure 5: 1269-1274; McPherson A (1976) J Biol Chem 251: 6300-6303). Antibody:antigen crystals may be studied using well known X-ray diffraction techniques and may be refined using computer software such as X-PLOR (Yale University, 1992, distributed by Molecular Simulations, Inc.; see e.g. Meth Enzymol (1985) volumes 114 & 115, eds Wyckoff H W et al.; U.S. 2004/0014194), and BUSTER (Bricogne G (1993) Acta Crystallogr D Biol Crystallogr 49(Pt 1): 37-60; Bricogne G (1997) Meth Enzymol 276A: 361-423, ed Carter C W; Roversi P et al., (2000) Acta Crystallogr D Biol Crystallogr 56(Pt 10): 1316-1323). Mutagenesis mapping studies may be accomplished using any method known to one of skill in the art. See, e.g., Champe M et al., (1995) J Biol Chem 270: 1388-1394 and Cunningham B C & Wells J A (1989) Science 244: 1081-1085 for a description of mutagenesis techniques, including alanine scanning mutagenesis techniques.
[0182] As used herein, the terms "label" or "labeled" refers to incorporation of a detectable marker, e.g., by incorporation of a radiolabeled amino acid or attachment to a polypeptide of biotin moieties that can be detected by marked avidin (e.g., streptavidin containing a fluorescent marker or enzymatic activity that can be detected by optical or colorimetric methods). In certain embodiments, the label or marker can also be therapeutic. Various methods of labeling polypeptides and glycoproteins are known in the art and can be used.
Methods of Treatment
[0183] Using adoptive immunotherapy, native T cells can be (i) removed from a patient, (ii) genetically engineered to express a chimeric antigen receptor (CAR) that binds to at least one tumor antigen (iii) expanded ex vivo into a larger population of engineered T cells, and (iv) reintroduced into the patient. See e.g., U.S. Pat. Nos. 7,741,465, and 6,319,494, Eshhar et al. (Cancer Immunol, supra); Krause et al. (supra); Finney et al. (supra). After the engineered T cells are reintroduced into the patient, they mediate an immune response against cells expressing the tumor antigen. See e.g., Krause et al., J. Exp. Med., Volume 188, No. 4, 1998 (619-626). This immune response includes secretion of IL-2 and other cytokines by T cells, the clonal expansion of T cells recognizing the tumor antigen, and T cell-mediated specific killing of target-positive cells. See Hombach et al., Journal of Immun. 167: 6123-6131 (2001).
[0184] The term "lymphocyte" as used herein includes natural killer (NK) cells, T cells, or B cells. NK cells are a type of cytotoxic (cell toxic) lymphocyte that represent a major component of the inherent immune system. NK cells reject tumors and cells infected by viruses. It works through the process of apoptosis or programmed cell death. They were termed "natural killers" because they do not require activation in order to kill cells. T-cells play a major role in cell-mediated-immunity (no antibody involvement). Its T-cell receptors (TCR) differentiate themselves from other lymphocyte types. The thymus, a specialized organ of the immune system, is primarily responsible for the T cell's maturation. There are six types of T-cells, namely: Helper T-cells (e.g., CD4+ cells), Cytotoxic T-cells (also known as TC, cytotoxic T lymphocyte, CTL, T-killer cell, cytolytic T cell, CD8+ T-cells or killer T cell), Memory T-cells ((i) stem memory T.sub.SCM cells, like naive cells, are CD45RO-, CCR7+, CD45RA+, CD62L+ (L-selectin), CD27+, CD28+ and IL-7R.alpha.+, but they also express large amounts of CD95, IL-2R.beta., CXCR3, and LFA-1, and show numerous functional attributes distinctive of memory cells); (ii) central memory T.sub.CM cells express L-selectin and the CCR7, they secrete IL-2, but not IFN.gamma. or IL-4, and (iii) effector memory TEM cells, however, do not express L-selectin or CCR7 but produce effector cytokines like IFN.gamma. and IL-4), Regulatory T-cells (Tregs, suppressor T cells, or CD4+CD25+ regulatory T cells), Natural Killer T-cells (NKT) and Gamma Delta T-cells. B-cells, on the other hand, play a principal role in humoral immunity (with antibody involvement). It makes antibodies and antigens and performs the role of antigen-presenting cells (APCs) and turns into memory B-cells after activation by antigen interaction. In mammals, immature B-cells are formed in the bone marrow, where its name is derived from.
[0185] The term "genetically engineered" or "engineered" refers to a method of modifying the genome of a cell, including, but not limited to, deleting a coding or non-coding region or a portion thereof or inserting a coding region or a portion thereof. In some embodiments, the cell that is modified is a lymphocyte, e.g., a T cell, which can either be obtained from a patient or a donor. The cell can be modified to express an exogenous construct, such as, e.g., a chimeric antigen receptor (CAR) or a T cell receptor (TCR), which is incorporated into the cell's genome.
[0186] An "immune response" refers to the action of a cell of the immune system (for example, T lymphocytes, B lymphocytes, natural killer (NK) cells, macrophages, eosinophils, mast cells, dendritic cells and neutrophils) and soluble macromolecules produced by any of these cells or the liver (including Abs, cytokines, and complement) that results in selective targeting, binding to, damage to, destruction of, and/or elimination from a vertebrate's body of invading pathogens, cells or tissues infected with pathogens, cancerous or other abnormal cells, or, in cases of autoimmunity or pathological inflammation, normal human cells or tissues.
[0187] The term "immunotherapy" refers to the treatment of a subject afflicted with, or at risk of contracting or suffering a recurrence of, a disease by a method comprising inducing, enhancing, suppressing or otherwise modifying an immune response. Examples of immunotherapy include, but are not limited to, T cell therapies. T cell therapy can include adoptive T cell therapy, tumor-infiltrating lymphocyte (TIL) immunotherapy, autologous cell therapy, engineered autologous cell therapy (eACT), and allogeneic T cell transplantation. However, one of skill in the art would recognize that the conditioning methods disclosed herein would enhance the effectiveness of any transplanted T cell therapy. Examples of T cell therapies are described in U.S. Patent Publication Nos. 2014/0154228 and 2002/0006409, U.S. Pat. No. 5,728,388, and International Publication No. WO 2008/081035.
[0188] The T cells of the immunotherapy can come from any source known in the art. For example, T cells can be differentiated in vitro from a hematopoietic stem cell population, or T cells can be obtained from a subject. T cells can be obtained from, e.g., peripheral blood mononuclear cells (PBMCs), bone marrow, lymph node tissue, cord blood, thymus tissue, tissue from a site of infection, ascites, pleural effusion, spleen tissue, and tumors. In addition, the T cells can be derived from one or more T cell lines available in the art. T cells can also be obtained from a unit of blood collected from a subject using any number of techniques known to the skilled artisan, such as FICOLL.TM. separation and/or apheresis. Additional methods of isolating T cells for a T cell therapy are disclosed in U.S. Patent Publication No. 2013/0287748, which is herein incorporated by references in its entirety.
[0189] The term "engineered Autologous Cell Therapy," which can be abbreviated as "eACT.TM.," also known as adoptive cell transfer, is a process by which a patient's own T cells are collected and subsequently genetically altered to recognize and target one or more antigens expressed on the cell surface of one or more specific tumor cells or malignancies. T cells can be engineered to express, for example, chimeric antigen receptors (CAR) or T cell receptor (TCR). CAR positive (+) T cells are engineered to express an extracellular single chain variable fragment (scFv) with specificity for a particular tumor antigen linked to an intracellular signaling part comprising at least one costimulatory domain and at least one activating domain. The costimulatory domain can be derived from (or correspond to), e.g., CD28, and the activating domain can be derived from (or correspond to) e.g., CD3-zeta. In certain embodiments, the CAR is designed to have two, three, four, or more costimulatory domains.
[0190] The term "autologous" refers to any material derived from the same individual to which it is later to be re-introduced. For example, the engineered autologous cell therapy (eACT.TM.) method described herein involves collection of lymphocytes from a patient, which are then engineered to express, e.g., a CAR construct, and then administered back to the same patient.
[0191] The term "allogeneic" refers to any material derived from one individual which is then introduced to another individual of the same species, e.g., allogeneic T cell transplantation.
[0192] In some aspects, the invention therefore comprises a method for treating or preventing a condition associated with undesired and/or elevated CLL-1 levels in a patient, comprising administering to a patient in need thereof an effective amount of at least one isolated antigen binding molecule, CAR, or TCR disclosed herein.
[0193] Methods are provided for treating diseases or disorders, including cancer. In some embodiments, the invention relates to creating a T cell-mediated immune response in a subject, comprising administering an effective amount of the engineered immune cells of the present application to the subject. In some embodiments, the T cell-mediated immune response is directed against a target cell or cells. In some embodiments, the engineered immune cell comprises a chimeric antigen receptor (CAR), or a T cell receptor (TCR). In some embodiments, the target cell is a tumor cell. In some aspects, the invention comprises a method for treating or preventing a malignancy, said method comprising administering to a subject in need thereof an effective amount of at least one isolated antigen binding molecule described herein. In some aspects, the invention comprises a method for treating or preventing a malignancy, said method comprising administering to a subject in need thereof an effective amount of at least one immune cell, wherein the immune cell comprises at least one chimeric antigen receptor, T cell receptor, and/or isolated antigen binding molecule as described herein.
[0194] In some aspects, the invention comprises a pharmaceutical composition comprising at least one antigen binding molecule as described herein and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition further comprises an additional active agent.
[0195] The antigen binding molecules, CARs, TCRs, immune cells, and the like of the invention can be used to treat myeloid diseases including but not limited to acute myeloid leukemia (AML), chronic myelogenous leukemia (CML), chronic myelomonocytic leukemia (CMML), juvenile myelomonocytic leukemia, atypical chronic myeloid leukemia, acute promyelocytic leukemia (APL), acute monoblastic leukemia, acute erythroid leukemia, acute megakaryoblastic leukemia, myelodysplastic syndrome (MDS), myeloproliferative disorder, myeloid neoplasm, myeloid sarcoma), Blastic Plasmacytoid Dendritic Cell Neoplasm (BPDCN), or combinations thereof. Additional diseases include inflammatory and/or autoimmune diseases such as rheumatoid arthritis, psoriasis, allergies, asthma, Crohn's disease, IBD, IBS, fibromyalga, mastocytosis, and Celiac disease.
[0196] It will be appreciated that target doses for CAR.sup.+/CAR-T.sup.+/TCR.sup.+ cells can range from 1.times.10.sup.6-2.times.10.sup.10 cells/kg, preferably 2.times.10.sup.6 cells/kg, more preferably. It will be appreciated that doses above and below this range may be appropriate for certain subjects, and appropriate dose levels can be determined by the healthcare provider as needed. Additionally, multiple doses of cells can be provided in accordance with the invention.
[0197] Also provided are methods for reducing the size of a tumor in a subject, comprising administering to the subject an engineered cell of the present invention to the subject, wherein the cell comprises a chimeric antigen receptor, a T cell receptor, or a T cell receptor based chimeric antigen receptor comprising an antigen binding molecule binds to an antigen on the tumor. In some embodiments, the subject has a solid tumor, or a blood malignancy such as lymphoma or leukemia. In some embodiments, the engineered cell is delivered to a tumor bed. In some embodiments, the cancer is present in the bone marrow of the subject. In some embodiments, the engineered cells are autologous T cells. In some embodiments, the engineered cells are allogeneic T cells. In some embodiments, the engineered cells are heterologous T cells. In some embodiments, the engineered cells of the present application are transfected or transduced in vivo. In other embodiments, the engineered cells are transfected or transduced ex vivo. As used herein, the term "in vitro cell" refers to any cell which is cultured ex vivo. In particular, an in vitro cell can include a T cell.
[0198] The methods can further comprise administering one or more chemotherapeutic agent. In certain embodiments, the chemotherapeutic agent is a lymphodepleting (preconditioning) chemotherapeutic. Beneficial preconditioning treatment regimens, along with correlative beneficial biomarkers are described in U.S. Provisional Patent Applications 62/262,143 and 62/167,750 which are hereby incorporated by reference in their entirety herein. These describe, e.g., methods of conditioning a patient in need of a T cell therapy comprising administering to the patient specified beneficial doses of cyclophosphamide (between 200 mg/m.sup.2/day and 2000 mg/m.sup.2/day) and specified doses of fludarabine (between 20 mg/m.sup.2/day and 900 mg/m.sup.2/day). A preferred dose regimen involves treating a patient comprising administering daily to the patient about 500 mg/m.sup.2/day of cyclophosphamide and about 60 mg/m.sup.2/day of fludarabine for three days prior to administration of a therapeutically effective amount of engineered T cells to the patient.
[0199] In other embodiments, the antigen binding molecule, transduced (or otherwise engineered) cells (such as CARs or TCRs), and the chemotherapeutic agent are administered each in an amount effective to treat the disease or condition in the subject.
[0200] In certain embodiments, compositions comprising CAR-expressing immune effector cells disclosed herein may be administered in conjunction with any number of chemotherapeutic agents. Examples of chemotherapeutic agents include alkylating agents such as thiotepa and cyclophosphamide (CYTOXAN.TM.); alkyl sulfonates such as busulfan, improsulfan and piposulfan; aziridines such as benzodopa, carboquone, meturedopa, and uredopa; ethylenimines and methylamelamines including altretamine, triethylenemelamine, trietylenephosphoramide, triethylenethiophosphaoramide and trimethylolomelamine resume; nitrogen mustards such as chlorambucil, chlornaphazine, cholophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembichin, phenesterine, prednimustine, trofosfamide, uracil mustard; nitrosureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, ranimustine; antibiotics such as aclacinomysins, actinomycin, authramycin, azaserine, bleomycins, cactinomycin, calicheamicin, carabicin, carminomycin, carzinophilin, chromomycins, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, epirubicin, esorubicin, idarubicin, marcellomycin, mitomycins, mycophenolic acid, nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin, quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; anti-metabolites such as methotrexate and 5-fluorouracil (5-FU); folic acid analogues such as denopterin, methotrexate, pteropterin, trimetrexate; purine analogs such as fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogs such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine, 5-FU; androgens such as calusterone, dromostanolone propionate, epitiostanol, mepitiostane, testolactone; anti-adrenals such as aminoglutethimide, mitotane, trilostane; folic acid replenisher such as frolinic acid; aceglatone; aldophosphamide glycoside; aminolevulinic acid; amsacrine; bestrabucil; bisantrene; edatraxate; defofamine; demecolcine; diaziquone; elformithine; elliptinium acetate; etoglucid; gallium nitrate; hydroxyurea; lentinan; lonidamine; mitoguazone; mitoxantrone; mopidamol; nitracrine; pentostatin; phenamet; pirarubicin; podophyllinic acid; 2-ethylhydrazide; procarbazine; PSK.RTM.; razoxane; sizofiran; spirogermanium; tenuazonic acid; triaziquone; 2, 2',2''-trichlorotriethylamine; urethan; vindesine; dacarbazine; mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine; arabinoside ("Ara-C"); cyclophosphamide; thiotepa; taxoids, e.g. paclitaxel (TAXOL, Bristol-Myers Squibb) and doxetaxel (TAXOTERE.RTM., Rhone-Poulenc Rorer); chlorambucil; gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; platinum analogs such as cisplatin and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitomycin C; mitoxantrone; vincristine; vinorelbine; navelbine; novantrone; teniposide; daunomycin; aminopterin; xeloda; ibandronate; CPT-11; topoisomerase inhibitor RFS2000; difluoromethylomithine (DMFO); retinoic acid derivatives such as Targretin (bexarotene), Panretin, (alitretinoin); ONTAK.TM. (denileukin diftitox); esperamicins; capecitabine; and pharmaceutically acceptable salts, acids or derivatives of any of the above. Also included in this definition are anti-hormonal agents that act to regulate or inhibit hormone action on tumors such as anti-estrogens including for example tamoxifen, raloxifene, aromatase inhibiting 4(5)-imidazoles, 4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone, and toremifene (Fareston); and anti-androgens such as flutamide, nilutamide, bicalutamide, leuprolide, and goserelin; and pharmaceutically acceptable salts, acids or derivatives of any of the above. Combinations of chemotherapeutic agents are also administered where appropriate, including, but not limited to CHOP, i.e., Cyclophosphamide (Cytoxan.RTM.), Doxorubicin (hydroxydoxorubicin), Vincristine (Oncovin.RTM.), and Prednisone.
[0201] In some embodiments, the chemotherapeutic agent is administered at the same time or within one week after the administration of the engineered cell, polypeptide, or nucleic acid. In other embodiments, the chemotherapeutic agent is administered from 1 to 4 weeks or from 1 week to 1 month, 1 week to 2 months, 1 week to 3 months, 1 week to 6 months, 1 week to 9 months, or 1 week to 12 months after the administration of the engineered cell, polypeptide, or nucleic acid. In other embodiments, the chemotherapeutic agent is administered at least 1 month before administering the cell, polypeptide, or nucleic acid. In some embodiments, the methods further comprise administering two or more chemotherapeutic agents.
[0202] A variety of additional therapeutic agents may be used in conjunction with the compositions described herein. For example, potentially useful additional therapeutic agents include PD-1 inhibitors such as nivolumab (Opdivo.RTM.), pembrolizumab (Keytruda.RTM.), pembrolizumab, pidilizumab, and atezolizumab.
[0203] Additional therapeutic agents suitable for use in combination with the invention include, but are not limited to, ibrutinib (Imbruvica.RTM.), ofatumumab (Arzerra.RTM.), rituximab (Rituxan.RTM.), bevacizumab (Avastin.RTM.), trastuzumab (Herceptin.RTM.), trastuzumab emtansine (KADCYLA.RTM.), imatinib (Gleevec.RTM.), cetuximab (Erbitux.RTM.), panitumumab (Vectibix.RTM.), catumaxomab, ibritumomab, ofatumumab, tositumomab, brentuximab, alemtuzumab, gemtuzumab, erlotinib, gefitinib, vandetanib, afatinib, lapatinib, neratinib, axitinib, masitinib, pazopanib, sunitinib, sorafenib, toceranib, lestaurtinib, axitinib, cediranib, lenvatinib, nintedanib, pazopanib, regorafenib, semaxanib, sorafenib, sunitinib, tivozanib, toceranib, vandetanib, entrectinib, cabozantinib, imatinib, dasatinib, nilotinib, ponatinib, radotinib, bosutinib, lestaurtinib, ruxolitinib, pacritinib, cobimetinib, selumetinib, trametinib, binimetinib, alectinib, ceritinib, crizotinib, aflibercept,adipotide, denileukin diftitox, mTOR inhibitors such as Everolimus and Temsirolimus, hedgehog inhibitors such as sonidegib and vismodegib, CDK inhibitors such as CDK inhibitor (palbociclib).
[0204] In additional embodiments, the composition comprising CAR-containing immune can be administered with an anti-inflammatory agent. Anti-inflammatory agents or drugs include, but are not limited to, steroids and glucocorticoids (including betamethasone, budesonide, dexamethasone, hydrocortisone acetate, hydrocortisone, hydrocortisone, methylprednisolone, prednisolone, prednisone, triamcinolone), nonsteroidal anti-inflammatory drugs (NSAIDS) including aspirin, ibuprofen, naproxen, methotrexate, sulfasalazine, leflunomide, anti-TNF medications, cyclophosphamide and mycophenolate. Exemplary NSAIDs include ibuprofen, naproxen, naproxen sodium, Cox-2 inhibitors, and sialylates. Exemplary analgesics include acetaminophen, oxycodone, tramadol of proporxyphene hydrochloride. Exemplary glucocorticoids include cortisone, dexamethasone, hydrocortisone, methylprednisolone, prednisolone, or prednisone. Exemplary biological response modifiers include molecules directed against cell surface markers (e.g., CD4, CD5, etc.), cytokine inhibitors, such as the TNF antagonists, (e.g., etanercept (ENBREL.RTM.), adalimumab (HUMIRA.RTM.) and infliximab (REMICADE.RTM.), chemokine inhibitors and adhesion molecule inhibitors. The biological response modifiers include monoclonal antibodies as well as recombinant forms of molecules. Exemplary DMARDs include azathioprine, cyclophosphamide, cyclosporine, methotrexate, penicillamine, leflunomide, sulfasalazine, hydroxychloroquine, Gold (oral (auranofin) and intramuscular) and minocycline.
[0205] In certain embodiments, the compositions described herein are administered in conjunction with a cytokine. "Cytokine" as used herein is meant to refer to proteins released by one cell population that act on another cell as intercellular mediators. Examples of cytokines are lymphokines, monokines, and traditional polypeptide hormones. Included among the cytokines are growth hormones such as human growth hormone, N-methionyl human growth hormone, and bovine growth hormone; parathyroid hormone; thyroxine; insulin; proinsulin; relaxin; prorelaxin; glycoprotein hormones such as follicle stimulating hormone (FSH), thyroid stimulating hormone (TSH), and luteinizing hormone (LH); hepatic growth factor (HGF); fibroblast growth factor (FGF); prolactin; placental lactogen; mullerian-inhibiting substance; mouse gonadotropin-associated peptide; inhibin; activin; vascular endothelial growth factor; integrin; thrombopoietin (TPO); nerve growth factors (NGFs) such as NGF-beta; platelet-growth factor; transforming growth factors (TGFs) such as TGF-alpha and TGF-beta; insulin-like growth factor-I and -II; erythropoietin (EPO); osteoinductive factors; interferons such as interferon-alpha, beta, and -gamma; colony stimulating factors (CSFs) such as macrophage-CSF (M-CSF); granulocyte-macrophage-CSF (GM-CSF); and granulocyte-CSF (G-CSF); interleukins (ILs) such as IL-1, IL-1 alpha, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12; IL-15, a tumor necrosis factor such as TNF-alpha or TNF-beta; and other polypeptide factors including LIF and kit ligand (KL). As used herein, the term cytokine includes proteins from natural sources or from recombinant cell culture, and biologically active equivalents of the native sequence cytokines.
[0206] In some aspects, the invention comprises an antigen binding molecule that binds to CLL-1 with a K.sub.d that is smaller than 100 pM. In some embodiments, the antigen binding molecule binds with a K.sub.d that is smaller than 10 pM. In other embodiments, the antigen binding molecule binds with a K.sub.d that is less than 5 pM.
Methods of Making
[0207] A variety of known techniques can be utilized in making the polynucleotides, polypeptides, vectors, antigen binding molecules, immune cells, compositions, and the like according to the invention.
[0208] Prior to the in vitro manipulation or genetic modification of the immune cells described herein, the cells may be obtained from a subject. In some embodiments, the immune cells comprise T cells. T cells can be obtained from a number of sources, including peripheral blood mononuclear cells (PBMCs), bone marrow, lymph nodes tissue, cord blood, thymus tissue, tissue from a site of infection, ascites, pleural effusion, spleen tissue, and tumors. In certain embodiments, T cells can be obtained from a unit of blood collected from the subject using any number of techniques known to the skilled person, such as FICOLL.TM. separation. Cells may preferably be obtained from the circulating blood of an individual by apheresis. The apheresis product typically contains lymphocytes, including T cells, monocytes, granulocytes, B cells, other nucleated white blood cells, red blood cells, and platelets. In certain embodiments, the cells collected by apheresis may be washed to remove the plasma fraction, and placed in an appropriate buffer or media for subsequent processing. The cells may be washed with PBS. As will be appreciated, a washing step may be used, such as by using a semiautomated flowthrough centrifuge--for example, the Cobe.TM. 2991 cell processor, the Baxter CytoMate.TM., or the like. After washing, the cells may be resuspended in a variety of biocompatible buffers, or other saline solution with or without buffer. In certain embodiments, the undesired components of the apheresis sample may be removed.
[0209] In certain embodiments, T cells are isolated from PBMCs by lysing the red blood cells and depleting the monocytes, for example, using centrifugation through a PERCOLL.TM. gradient. A specific subpopulation of T cells, such as CD28.sup.+, CD4.sup.+, CD8.sup.+, CD45RA.sup.+, and CD45RO.sup.+ T cells can be further isolated by positive or negative selection techniques known in the art. For example, enrichment of a T cell population by negative selection can be accomplished with a combination of antibodies directed to surface markers unique to the negatively selected cells. One method for use herein is cell sorting and/or selection via negative magnetic immunoadherence or flow cytometry that uses a cocktail of monoclonal antibodies directed to cell surface markers present on the cells negatively selected. For example, to enrich for CD4.sup.+ cells by negative selection, a monoclonal antibody cocktail typically includes antibodies to CD14, CD20, CD1 1b, CD16, HLA-DR, and CD8. Flow cytometry and cell sorting may also be used to isolate cell populations of interest for use in the present invention.
[0210] PBMCs may be used directly for genetic modification with the immune cells (such as CARs or TCRs) using methods as described herein. In certain embodiments, after isolating the PBMCs, T lymphocytes can be further isolated and both cytotoxic and helper T lymphocytes can be sorted into naive, memory, and effector T cell subpopulations either before or after genetic modification and/or expansion.
[0211] In some embodiments, CD8.sup.+ cells are further sorted into naive, central memory, and effector cells by identifying cell surface antigens that are associated with each of these types of CD8.sup.+ cells. In some embodiments, the expression of phenotypic markers of central memory T cells include CD45RO, CD62L, CCR7, CD28, CD3, and CD127 and are negative for granzyme B. In some embodiments, central memory T cells are CD45RO.sup.+, CD62L.sup.+, CD8.sup.+ T cells. In some embodiments, effector T cells are negative for CD62L, CCR7, CD28, and CD127, and positive for granzyme B and perforin. In certain embodiments, CD4.sup.+ T cells are further sorted into subpopulations. For example, CD4.sup.+ T helper cells can be sorted into naive, central memory, and effector cells by identifying cell populations that have cell surface antigens.
[0212] The immune cells, such as T cells, can be genetically modified following isolation using known methods, or the immune cells can be activated and expanded (or differentiated in the case of progenitors) in vitro prior to being genetically modified. In another embodiment, the immune cells, such as T cells, are genetically modified with the chimeric antigen receptors described herein (e.g., transduced with a viral vector comprising one or more nucleotide sequences encoding a CAR) and then are activated and/or expanded in vitro. Methods for activating and expanding T cells are known in the art and are described, for example, in U.S. Pat. Nos. 6,905,874; 6,867,041; 6,797,514; and PCT WO2012/079000, the contents of which are hereby incorporated by reference in their entirety. Generally, such methods include contacting PBMC or isolated T cells with a stimulatory molecule and a costimulatory molecule, such as anti-CD3 and anti-CD28 antibodies, generally attached to a bead or other surface, in a culture medium with appropriate cytokines, such as IL-2. Anti-CD3 and anti-CD28 antibodies attached to the same bead serve as a "surrogate" antigen presenting cell (APC). One example is The Dynabeads.RTM. system, a CD3/CD28 activator/stimulator system for physiological activation of human T cells. In other embodiments, the T cells may be activated and stimulated to proliferate with feeder cells and appropriate antibodies and cytokines using methods such as those described in U.S. Pat. Nos. 6,040,177; 5,827,642; and WO2012129514, the contents of which are hereby incorporated by reference in their entirety.
[0213] Certain methods for making the constructs and engineered immune cells of the invention are described in PCT application PCT/US15/14520, the contents of which are hereby incorporated by reference in their entirety. Additional methods of making the constructs and cells can be found in U.S. provisional patent application No. 62/244,036 the contents of which are hereby incorporated by reference in their entirety.
[0214] It will be appreciated that PBMCs can further include other cytotoxic lymphocytes such as NK cells or NKT cells. An expression vector carrying the coding sequence of a chimeric receptor as disclosed herein can be introduced into a population of human donor T cells, NK cells or NKT cells. Successfully transduced T cells that carry the expression vector can be sorted using flow cytometry to isolate CD3 positive T cells and then further propagated to increase the number of these CAR expressing T cells in addition to cell activation using anti-CD3 antibodies and IL-2 or other methods known in the art as described elsewhere herein. Standard procedures are used for cryopreservation of T cells expressing the CAR for storage and/or preparation for use in a human subject. In one embodiment, the in vitro transduction, culture and/or expansion of T cells are performed in the absence of non-human animal derived products such as fetal calf serum and fetal bovine serum.
[0215] For cloning of polynucleotides, the vector may be introduced into a host cell (an isolated host cell) to allow replication of the vector itself and thereby amplify the copies of the polynucleotide contained therein. The cloning vectors may contain sequence components generally include, without limitation, an origin of replication, promoter sequences, transcription initiation sequences, enhancer sequences, and selectable markers. These elements may be selected as appropriate by a person of ordinary skill in the art. For example, the origin of replication may be selected to promote autonomous replication of the vector in the host cell.
[0216] In certain embodiments, the present disclosure provides isolated host cells containing the vector provided herein. The host cells containing the vector may be useful in expression or cloning of the polynucleotide contained in the vector. Suitable host cells can include, without limitation, prokaryotic cells, fungal cells, yeast cells, or higher eukaryotic cells such as mammalian cells. Suitable prokaryotic cells for this purpose include, without limitation, eubacteria, such as Gram-negative or Gram-positive organisms, for example, Enterobactehaceae such as Escherichia, e.g., E. coli, Enterobacter, Erwinia, Klebsiella, Proteus, Salmonella, e.g., Salmonella typhimurium, Serratia, e.g., Serratia marcescans, and Shigella, as well as Bacilli such as B. subtilis and B. licheniformis, Pseudomonas such as P. aeruginosa, and Streptomyces.
[0217] The vector can be introduced to the host cell using any suitable methods known in the art, including, without limitation, DEAE-dextran mediated delivery, calcium phosphate precipitate method, cationic lipids mediated delivery, liposome mediated transfection, electroporation, microprojectile bombardment, receptor-mediated gene delivery, delivery mediated by polylysine, histone, chitosan, and peptides. Standard methods for transfection and transformation of cells for expression of a vector of interest are well known in the art. In a further embodiment, a mixture of different expression vectors can be used in genetically modifying a donor population of immune effector cells wherein each vector encodes a different CAR as disclosed herein. The resulting transduced immune effector cells form a mixed population of engineered cells, with a proportion of the engineered cells expressing more than one different CARs.
[0218] In one embodiment, the invention provides a method of storing genetically engineered cells expressing CARs or TCRs which target a CLL-1 protein. This involves cryopreserving the immune cells such that the cells remain viable upon thawing. A fraction of the immune cells expressing the CARs can be cryopreserved by methods known in the art to provide a permanent source of such cells for the future treatment of patients afflicted with a malignancy. When needed, the cryopreserved transformed immune cells can be thawed, grown and expanded for more such cells.
[0219] As used herein, "cryopreserve" refers to the preservation of cells by cooling to sub-zero temperatures, such as (typically) 77 Kelvin or -196.degree. C. (the boiling point of liquid nitrogen). Cryoprotective agents are often used at sub-zero temperatures to prevent the cells being preserved from damage due to freezing at low temperatures or warming to room temperature. Cryopreservative agents and optimal cooling rates can protect against cell injury. Cryoprotective agents which can be used in accordance with the invention include but are not limited to: dimethyl sulfoxide (DMSO) (Lovelock & Bishop, Nature (1959); 183: 1394-1395; Ashwood-Smith, Nature (1961); 190: 1204-1205), glycerol, polyvinylpyrrolidine (Rinfret, Ann. N.Y. Acad. Sci. (1960); 85: 576), and polyethylene glycol (Sloviter & Ravdin, Nature (1962); 196: 48). The preferred cooling rate is 1.degree.-3.degree. C./minute.
[0220] The term, "substantially pure," is used to indicate that a given component is present at a high level. The component is desirably the predominant component present in a composition. Preferably it is present at a level of more than 30%, of more than 50%, of more than 75%, of more than 90%, or even of more than 95%, said level being determined on a dry weight/dry weight basis with respect to the total composition under consideration. At very high levels (e.g. at levels of more than 90%, of more than 95% or of more than 99%) the component can be regarded as being in "pure form." Biologically active substances of the present invention (CARs, TCRs, isolated polypeptides, isolated nucleic acid molecules, antigen binding molecules, moieties) can be provided in a form that is substantially free of one or more contaminants with which the substance might otherwise be associated. When a composition is substantially free of a given contaminant, the contaminant will be at a low level (e.g., at a level of less than 10%, less than 5%, or less than 1% on the dry weight/dry weight basis set out above).
[0221] In some embodiments, the cells are formulated by first harvesting them from their culture medium, and then washing and concentrating the cells in a medium and container system suitable for administration (a "pharmaceutically acceptable" carrier) in a treatment-effective amount. Suitable infusion media can be any isotonic medium formulation, typically normal saline, Normosol.TM. R (Abbott) or Plasma-Lyte.TM. A (Baxter), but also 5% dextrose in water or Ringer's lactate can be utilized. The infusion medium can be supplemented with human serum albumin.
[0222] Desired treatment amounts of cells in the composition is generally at least 2 cells (for example, at least 1 CD8.sup.+ central memory T cell and at least 1 CD4.sup.+ helper T cell subset) or is more typically greater than 10.sup.2 cells, and up to 10.sup.6, up to and including 10.sup.8 or 10.sup.9 cells and can be more than 10.sup.10 cells. The number of cells will depend upon the desired use for which the composition is intended, and the type of cells included therein. The density of the desired cells is typically greater than 10.sup.6 cells/ml and generally is greater than 10.sup.7 cells/ml, generally 10.sup.8 cells/ml or greater. The clinically relevant number of immune cells can be apportioned into multiple infusions that cumulatively equal or exceed 10.sup.5, 10.sup.6, 10.sup.7, 10.sup.8, 10.sup.9, 10.sup.10, 10.sup.11, or 10.sup.12 cells. In some aspects of the present invention, particularly since all the infused cells will be redirected to a particular target antigen (CLL-1), lower numbers of cells, in the range of 10.sup.6/kilogram (10.sup.6-10.sup.11 per patient) may be administered. CAR treatments may be administered multiple times at dosages within these ranges. The cells may be autologous, allogeneic, or heterologous to the patient undergoing therapy.
[0223] The CAR expressing cell populations of the present invention may be administered either alone, or as a pharmaceutical composition in combination with diluents and/or with other components such as IL-2 or other cytokines or cell populations. Pharmaceutical compositions of the present invention may comprise a CAR or TCR expressing cell population, such as T cells, as described herein, in combination with one or more pharmaceutically or physiologically acceptable carriers, diluents or excipients. Such compositions may comprise buffers such as neutral buffered saline, phosphate buffered saline and the like; carbohydrates such as glucose, mannose, sucrose or dextrans, mannitol; proteins; polypeptides or amino acids such as glycine; antioxidants; chelating agents such as EDTA or glutathione; adjuvants (e.g., aluminum hydroxide); and preservatives. Compositions of the present invention are preferably formulated for intravenous administration.
[0224] The pharmaceutical compositions (solutions, suspensions or the like), may include one or more of the following: sterile diluents such as water for injection, saline solution, preferably physiological saline, Ringer's solution, isotonic sodium chloride, fixed oils such as synthetic mono- or diglycerides which may serve as the solvent or suspending medium, polyethylene glycols, glycerin, propylene glycol or other solvents; antibacterial agents such as benzyl alcohol or methyl paraben; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose. The parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic. An injectable pharmaceutical composition is preferably sterile.
[0225] It will be appreciated that adverse events may be minimized by transducing the immune cells (containing one or more CARs or TCRs) with a suicide gene. It may also be desired to incorporate an inducible "on" or "accelerator" switch into the immune cells. Suitable techniques include use of inducible caspase-9 (U.S. Appl. 2011/0286980) or a thymidine kinase, before, after or at the same time, as the cells are transduced with the CAR construct of the present invention. Additional methods for introducing suicide genes and/or "on" switches include TALENS, zinc fingers, RNAi, siRNA, shRNA, antisense technology, and other techniques known in the art.
[0226] In accordance with the invention, additional on-off or other types of control switch techniques may be incorporated herein. These techniques may employ the use of dimerization domains and optional activators of such domain dimerization. These techniques include, e.g., those described by Wu et al., Science 2014 350 (6258) utilizing FKBP/Rapalog dimerization systems in certain cells, the contents of which are incorporated by reference herein in their entirety. Additional dimerization technology is described in, e.g., Fegan et al. Chem. Rev. 2010, 110, 3315-3336 as well as U.S. Pat. Nos. 5,830,462; 5,834,266; 5,869,337; and 6,165,787, the contents of which are also incorporated by reference herein in their entirety. Additional dimerization pairs may include cyclosporine-A/cyclophilin, receptor, estrogen/estrogen receptor (optionally using tamoxifen), glucocorticoids/glucocorticoid receptor, tetracycline/tetracycline receptor, vitamin D/vitamin D receptor. Further examples of dimerization technology can be found in e.g., WO 2014/127261, WO 2015/090229, US 2014/0286987, US2015/0266973, US2016/0046700, U.S. Pat. No. 8,486,693, US 2014/0171649, and US 2012/0130076, the contents of which are further incorporated by reference herein in their entirety.
[0227] It will be understood that descriptions herein are exemplary and explanatory only and are not restrictive of the invention as claimed. In this application, the use of the singular includes the plural unless specifically stated otherwise.
[0228] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described. All documents, or portions of documents, cited in this application, including but not limited to patents, patent applications, articles, books, and treatises, are hereby expressly incorporated by reference in their entirety for any purpose. As utilized in accordance with the present disclosure, the following terms, unless otherwise indicated, shall be understood to have the following meanings:
[0229] In this application, the use of "or" means "and/or" unless stated otherwise. Furthermore, the use of the term "including", as well as other forms, such as "includes" and "included", is not limiting. Also, terms such as "element" or "component" encompass both elements and components comprising one unit and elements and components that comprise more than one subunit unless specifically stated otherwise.
[0230] The term "CLL-1 activity" includes any biological effect of CLL-1. In certain embodiments, CLL-1 activity includes the ability of CLL-1 to interact or bind to a substrate or receptor.
[0231] The term "polynucleotide," "nucleotide," or "nucleic acid" includes both single-stranded and double-stranded nucleotide polymers. This preferably includes isolated polynucleotides, nucleotides or nucleic acids as defined herein. The nucleotides comprising the polynucleotide can be ribonucleotides or deoxyribonucleotides or a modified form of either type of nucleotide. Said modifications include base modifications such as bromouridine and inosine derivatives, ribose modifications such as 2',3'-dideoxyribose, and internucleotide linkage modifications such as phosphorothioate, phosphorodithioate, phosphoroselenoate, phosphoro-diselenoate, phosphoro-anilothioate, phoshoraniladate and phosphoroamidate.
[0232] The term "oligonucleotide" refers to a polynucleotide comprising 200 or fewer nucleotides. Oligonucleotides can be single stranded or double stranded, e.g., for use in the construction of a mutant gene. Oligonucleotides can be sense or antisense oligonucleotides. An oligonucleotide can include a label, including a radiolabel, a fluorescent label, a hapten or an antigenic label, for detection assays. Oligonucleotides can be used, for example, as PCR primers, cloning primers or hybridization probes.
[0233] The term "control sequence" refers to a polynucleotide sequence that can affect the expression and processing of coding sequences to which it is ligated. The nature of such control sequences can depend upon the host organism. In particular embodiments, control sequences for prokaryotes can include a promoter, a ribosomal binding site, and a transcription termination sequence. For example, control sequences for eukaryotes can include promoters comprising one or a plurality of recognition sites for transcription factors, transcription enhancer sequences, and transcription termination sequence. "Control sequences" can include leader sequences (signal peptides) and/or fusion partner sequences.
[0234] In some embodiments, the polynucleotide of the present invention encodes a CAR or a TCR can further comprises a leader sequence or peptide (also referred to herein as a "signal peptide"). In certain embodiments, the leader peptide comprises an amino acid sequence that is at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% identical to the amino acid sequence MALPVTALLLPLALLLHAARP (SEQ ID NO: 144). In some embodiments, the leader peptide comprises the amino acid sequence of SEQ ID NO: 144.
[0235] As used herein, "operably linked" means that the components to which the term is applied are in a relationship that allows them to carry out their inherent functions under suitable conditions.
[0236] The term "vector" means any molecule or entity (e.g., nucleic acid, plasmid, bacteriophage or virus) used to transfer protein coding information into a host cell. The term "expression vector" or "expression construct" refers to a vector that is suitable for transformation of a host cell and contains nucleic acid sequences that direct and/or control (in conjunction with the host cell) expression of one or more heterologous coding regions operatively linked thereto. An expression construct can include, but is not limited to, sequences that affect or control transcription, translation, and, if introns are present, affect RNA splicing of a coding region operably linked thereto.
[0237] The term "host cell" refers to a cell that has been transformed, or is capable of being transformed, with a nucleic acid sequence and thereby expresses a gene of interest. The term includes the progeny of the parent cell, whether or not the progeny is identical in morphology or in genetic make-up to the original parent cell, so long as the gene of interest is present.
[0238] The term "transformation" refers to a change in a cell's genetic characteristics, and a cell has been transformed when it has been modified to contain new DNA or RNA. For example, a cell is transformed where it is genetically modified from its native state by introducing new genetic material via transfection, transduction, or other techniques. Following transfection or transduction, the transforming DNA can recombine with that of the cell by physically integrating into a chromosome of the cell, or can be maintained transiently as an episomal element without being replicated, or can replicate independently as a plasmid. A cell is considered to have been "stably transformed" when the transforming DNA is replicated with the division of the cell.
[0239] The term "transfection" refers to the uptake of foreign or exogenous DNA by a cell. A number of transfection techniques are well known in the art and are disclosed herein. See, e.g., Graham et al., 1973, Virology 52:456; Sambrook et al., 2001, Molecular Cloning: A Laboratory Manual, supra; Davis et al., 1986, Basic Methods in Molecular Biology, Elsevier; Chu et al., 1981, Gene 13:197.
[0240] The term "transduction" refers to the process whereby foreign DNA is introduced into a cell via viral vector. See Jones et al., (1998). Genetics: principles and analysis. Boston: Jones & Bartlett Publ.
[0241] The terms "polypeptide" or "protein" refer to a macromolecule having the amino acid sequence of a protein, including deletions from, additions to, and/or substitutions of one or more amino acids of the native sequence, and preferably no more than 8 amino acid substitutions therein. Preferably, the polypeptides or proteins are isolated as defined herein. The terms "polypeptide" and "protein" specifically encompass CLL-1 antigen binding molecules, antibodies, or sequences that have deletions from, additions to, and/or substitutions of one or more amino acid of antigen-binding protein, and preferably no more than 8 amino acid substitutions therein. The term "polypeptide fragment" refers to an isolated polypeptide that has an amino-terminal deletion, a carboxyl-terminal deletion, and/or an internal deletion as compared with the full-length native protein. Such fragments can also contain modified amino acids as compared with the native protein. Useful polypeptide fragments include immunologically functional fragments of antigen binding molecules. Useful fragments include but are not limited to one or more CDR regions, variable domains of a heavy and/or light chain, a portion of other portions of an antibody chain, and the like.
[0242] The term "isolated" means (i) free of at least some other proteins with which it would normally be found, (ii) is essentially free of other proteins from the same source, e.g., from the same species, (iii) separated from at least about 50 percent of polynucleotides, lipids, carbohydrates, or other materials with which it is associated in nature, (iv) operably associated (by covalent or noncovalent interaction) with a polypeptide with which it is not associated in nature, or (v) does not occur in nature.
[0243] A "variant" of a polypeptide (e.g., an antigen binding molecule, or an antibody) comprises an amino acid sequence wherein one or more amino acid residues are inserted into, deleted from and/or substituted into the amino acid sequence relative to another polypeptide sequence. Variants include fusion proteins.
[0244] The term "identity" refers to a relationship between the sequences of two or more polypeptide molecules or two or more nucleic acid molecules, as determined by aligning and comparing the sequences. "Percent identity" means the percent of identical residues between the amino acids or nucleotides in the compared molecules and is calculated based on the size of the smallest of the molecules being compared. For these calculations, gaps in alignments (if any) are preferably addressed by a particular mathematical model or computer program (i.e., an "algorithm").
[0245] To calculate percent identity, the sequences being compared are typically aligned in a way that gives the largest match between the sequences. One example of a computer program that can be used to determine percent identity is the GCG program package, which includes GAP (Devereux et al., 1984, Nucl. Acid Res. 12:387; Genetics Computer Group, University of Wisconsin, Madison, Wis.). The computer algorithm GAP is used to align the two polypeptides or polynucleotides for which the percent sequence identity is to be determined. The sequences are aligned for optimal matching of their respective amino acid or nucleotide (the "matched span", as determined by the algorithm). In certain embodiments, a standard comparison matrix (see, Dayhoff et al., 1978, Atlas of Protein Sequence and Structure 5:345-352 for the PAM 250 comparison matrix; Henikoff et al., 1992, Proc. Natl. Acad. Sci. U.S.A. 89:10915-10919 for the BLOSUM 62 comparison matrix) is also used by the algorithm.
[0246] As used herein, the twenty conventional (e.g., naturally occurring) amino acids and their abbreviations follow conventional usage. See Immunology--A Synthesis (2nd Edition, Golub and Gren, Eds., Sinauer Assoc., Sunderland, Mass. (1991)), which is incorporated herein by reference for any purpose. Stereoisomers (e.g., D-amino acids) of the twenty conventional amino acids, unnatural amino acids such as alpha-, alpha-disubstituted amino acids, N-alkyl amino acids, lactic acid, and other unconventional amino acids can also be suitable components for polypeptides of the present invention. Examples of unconventional amino acids include: 4-hydroxyproline, .gamma.-carboxyglutamate, epsilon-N,N,N-trimethyllysine, e-N-acetyllysine, O-phosphoserine, N-acetylserine, N-formylmethionine, 3-methylhistidine, 5-hydroxylysine, .sigma.-N-methylarginine, and other similar amino acids and imino acids (e.g., 4-hydroxyproline). In the polypeptide notation used herein, the left-hand direction is the amino terminal direction and the right-hand direction is the carboxy-terminal direction, in accordance with standard usage and convention.
[0247] Conservative amino acid substitutions can encompass non-naturally occurring amino acid residues, which are typically incorporated by chemical peptide synthesis rather than by synthesis in biological systems. These include peptidomimetics and other reversed or inverted forms of amino acid moieties. Naturally occurring residues can be divided into classes based on common side chain properties:
[0248] a) hydrophobic: norleucine, Met, Ala, Val, Leu, Ile;
[0249] b) neutral hydrophilic: Cys, Ser, Thr, Asn, Gln;
[0250] c) acidic: Asp, Glu;
[0251] d) basic: His, Lys, Arg;
[0252] e) residues that influence chain orientation: Gly, Pro; and
[0253] f) aromatic: Trp, Tyr, Phe.
[0254] For example, non-conservative substitutions can involve the exchange of a member of one of these classes for a member from another class. Such substituted residues can be introduced, for example, into regions of a human antibody that are homologous with non-human antibodies, or into the non-homologous regions of the molecule. Exemplary amino acid substitutions are set forth in Table 3.
TABLE-US-00016 TABLE 3 Original Residues Exemplary Substitutions Preferred Substitutions Ala Val, Leu, Ile Val Arg Lys, Gln, Asn Lys Asn Gln Gln Asp Glu Glu Cys Ser, Ala Ser Gln Asn Asn Glu Asp Asp Gly Pro, Ala Ala His Asn, Gln, Lys, Arg Arg Ile Leu, Val, Met, Ala, Phe, Leu Norleucine Leu Norleucine, Ile, Val, Met, Ile Ala, Phe Lys Arg, 1,4 Diamino-butyric Arg Acid, Gln, Asn Met Leu, Phe, Ile Leu Phe Leu, Val, Ile, Ala, Leu Tyr Pro Ala Gly Ser Thr, Ala, Cys Thr Thr Ser Ser Trp Tyr, Phe Tyr Tyr Trp, Phe, Thr, Ser Phe Val Ile, Met, Leu, Phe, Leu Ala, Norleucine
[0255] The term "derivative" refers to a molecule that includes a chemical modification other than an insertion, deletion, or substitution of amino acids (or nucleic acids). In certain embodiments, derivatives comprise covalent modifications, including, but not limited to, chemical bonding with polymers, lipids, or other organic or inorganic moieties. In certain embodiments, a chemically modified antigen binding molecule can have a greater circulating half-life than an antigen binding molecule that is not chemically modified. In some embodiments, a derivative antigen binding molecule is covalently modified to include one or more water soluble polymer attachments, including, but not limited to, polyethylene glycol, polyoxyethylene glycol, or polypropylene glycol.
[0256] Peptide analogs are commonly used in the pharmaceutical industry as non-peptide drugs with properties analogous to those of the template peptide. These types of non-peptide compound are termed "peptide mimetics" or "peptidomimetics." Fauchere, J., Adv. Drug Res., 15:29 (1986); Veber & Freidinger, TINS, p. 392 (1985); and Evans et al., J. Med. Chem., 30:1229 (1987), which are incorporated herein by reference for any purpose.
[0257] A "therapeutically effective amount," "effective dose," "effective amount," or "therapeutically effective dosage" of a therapeutic agent, e.g., engineered CART cells, is any amount that, when used alone or in combination with another therapeutic agent, protects a subject against the onset of a disease or promotes disease regression evidenced by a decrease in severity of disease symptoms, an increase in frequency and duration of disease symptom-free periods, or a prevention of impairment or disability due to the disease affliction. The ability of a therapeutic agent to promote disease regression can be evaluated using a variety of methods known to the skilled practitioner, such as in human subjects during clinical trials, in animal model systems predictive of efficacy in humans, or by assaying the activity of the agent in in vitro assays.
[0258] The terms "patient" and "subject" are used interchangeably and include human and non-human animal subjects as well as those with formally diagnosed disorders, those without formally recognized disorders, those receiving medical attention, those at risk of developing the disorders, etc.
[0259] The term "treat" and "treatment" includes therapeutic treatments, prophylactic treatments, and applications in which one reduces the risk that a subject will develop a disorder or other risk factor. Treatment does not require the complete curing of a disorder and encompasses embodiments in which one reduces symptoms or underlying risk factors. The term "prevent" does not require the 100% elimination of the possibility of an event. Rather, it denotes that the likelihood of the occurrence of the event has been reduced in the presence of the compound or method.
[0260] Standard techniques can be used for recombinant DNA, oligonucleotide synthesis, and tissue culture and transformation (e.g., electroporation, lipofection). Enzymatic reactions and purification techniques can be performed according to manufacturer's specifications or as commonly accomplished in the art or as described herein. The foregoing techniques and procedures can be generally performed according to conventional methods well known in the art and as described in various general and more specific references that are cited and discussed throughout the present specification. See, e.g., Sambrook et al., Molecular Cloning: A Laboratory Manual (2d ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1989)), which is incorporated herein by reference for any purpose.
INCORPORATION BY REFERENCE
[0261] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference. However, the citation of a reference herein should not be construed as an acknowledgement that such reference is prior art to the present invention. To the extent that any of the definitions or terms provided in the references incorporated by reference differ from the terms and discussion provided herein, the present terms and definitions control.
[0262] The foregoing written specification is considered to be sufficient to enable one skilled in the art to practice the invention. The foregoing description and examples detail certain preferred embodiments of the invention and describe the best mode contemplated by the inventors. It will be appreciated, however, that no matter how detailed the foregoing may appear in text, the invention may be practiced in many ways and the invention should be construed in accordance with the appended claims and any equivalents thereof.
[0263] The following examples, including the experiments conducted and results achieved, are provided for illustrative purposes only and are not to be construed as limiting the present invention.
Example 1
[0264] Determining CLL-1 CAR activity by mRNA electroporation in human PBMCs. Plasmids encoding a T7 promoter, CAR construct and a beta globin stabilizing sequence were linearize by overnight digestion of Mug of DNA with EcoRI and BamHI (NEB). DNA was then digested for 2 hours at 50.degree. C. with proteinase K (Thermo Fisher.TM., 600 U/ml) purified with phenol/chloroform and precipitated by adding sodium acetate and two volumes of ethanol. Pellets were then dried, resuspended in RNAse/DNAse free water and quantified. 1 .mu.g of the linear DNA was then use to generate the in vitro transcription using the mMES SAGE mMACHINE T7 Ultra (Thermo Fisher.TM.) following the manufacturer instructions. RNA was further purified using the MEGAClear Kit (Thermo Fisher.TM.) following the manufacturer instructions, and quantified using NanoDrop.TM.. mRNA integrity was assessed by running an agarose gel.
[0265] Different cancer cell lines were evaluated for CLL-1 expression. Namalwa (ATCC), U937 (ATCC), HL-60 (ATCC), EoL-1 (Sigma), KG1a (ATCC) and MV4;11 (ATCC) cells were stained with anti-CLL-1 antibody conjugated to PE (BD Pharmingen.TM.) in stain buffer (BD Pharmingen.TM.) for 30 minutes at 4.degree. C. Cells were then washed and resuspended in stain buffer with propidium iodide (BD Pharmingen.TM.) prior to data acquisition. Samples were then acquired by flow cytometry and data analyzed and plotted in histograms using FlowJo.TM.. Results for the CLL-1 expression can be seen in FIG. 1.
[0266] PBMCs were isolated from healthy donor leukopaks (Hemacare.TM.) using ficoll-paque density centrifugation per manufacturer's instructions. PBMCs were stimulated using OKT3 (50 ng/ml, Miltenyi Biotec.TM.) in R10 media+IL-2 (300 IU/ml, Proleukin.RTM., Prometheus.RTM. Therapeutics and Diagnostics). Seven days after stimulation, T cells were washed twice in Opti-MEM.TM. (Thermo Fisher Scientific.TM.) and resuspended at a final concentration of 2.5.times.10.sup.7 cells/ml in Opti-MEM. 10 .mu.g of mRNA was used per electroporation. Electroporation of cells was performed using a Gemini X2 system (Harvard Apparatus BTX.TM.) set to deliver a single 400V pulse for 0.5 ms in 2 mm cuvettes (Harvard Apparatus BTX.TM.). Cells were immediately transferred to R10+IL-2 media and allowed to recover. Cells were maintained at 0.5-2.0.times.10.sup.6 cells/ml prior to use in activity assays.
[0267] Six hours after mRNA electroporation, T cells were stained with biotinylated Protein L (Thermo Scientific.TM.) in stain buffer (BD Pharmingen.TM.) for 30 minutes at 4.degree. C. Cells were then washed and stained with PE Streptavidin (BD Pharmingen.TM.) in stain buffer for 30 minutes at 4.degree. C. Cells were then washed and resuspended in stain buffer with propidium iodide (BD Pharmingen.TM.) prior to data acquisition. Results for CAR detection are shown in FIG. 2.
[0268] Effector cells were cultured with target cells at a 1:1 E:T ratio in R10 media 6 hours after mRNA electroporation. Cell lines tested included Namalwa, U937, HL-60, EoL-1, KG1 a and MV4;11. Sixteen hours post-coculture, supernatants were analyzed by Luminex (EMD Millipore) following manufacturer instructions and target cell viability was assessed by flow cytometric analysis of propidium iodide (PI) uptake. Results corresponding to the cytokine release assay can be found in FIG. 3. Results of the cytolytic activity assay can be found in FIG. 4 and FIG. 5.
Example 2
[0269] Determining CLL-1 CAR activity by lentiviral transduction of human PBMCs. A third generation lentiviral transfer vector containing the different CLL-1 CAR construct was used along with the ViraPower.TM. Lentiviral Packaging Mix (Life Technologies.TM.) to generate the lentiviral supernatants. Briefly, a transfection mix was generated by mixing 15 ug of DNA and 22.5 ul of polyethileneimine (Polysciences.TM., 1 mg/ml) in 600 ul of OptiMEM.TM. media. The mix was incubated for 5 minutes at room temperature. Simultaneously, 293T cells (ATCC) were trypsinized, counted and a total of 10.times.10.sup.6 total cells were plated in a T75 flask along the transfection mix. Three days after the transfection, supernatants were collected and filtered through a 0.45 um filter and stored at -80 C until used.
[0270] PBMCs were isolated from healthy donor leukopaks (Hemacare.TM.) using ficoll-paque density centrifugation per manufacturer's instructions. PBMCs were stimulated using OKT3 (50 ng/ml, Miltenyi Biotec.TM.) in R10 media+IL-2 (300 IU/ml, Proleukin.RTM., Prometheus.RTM. Therapeutics and Diagnostics). Forty eight hours post-stimulation, cells were transduced using lentivirus at a MOI=10. Cells were maintained at 0.5-2.0.times.10.sup.6 cells/ml prior to use in activity assays.
[0271] At day 12 post stimulation, T cells were stained with biotinylated Protein L (Thermo Scientific.TM.) in stain buffer (BD Pharmingen.TM.) for 30 minutes at 4.degree. C. Cells were then washed and stained with PE Streptavidin (BD Pharmingen.TM.) in stain buffer for 30 minutes at 4.degree. C. Cells were then washed and resuspended in stain buffer with propidium iodide (BD Pharmingen.TM.) prior to data acquisition. Results for CAR detection are shown in FIG. 6.
[0272] Effector cells were cultured with target cells at a 1:1 E:T ratio in R10 media 12 days after T cell stimulation. Cell lines tested included Namalwa, U937, HL-60, EoL-1, KG1a and MV4;11. 16 hours post-coculture, supernatants were analyzed by Luminex (EMD Millipore.TM.) following manufacturer instructions and target cell viability was assessed by flow cytometric analysis of propidium iodide (PI) uptake. Results corresponding to the cytokine release assay can be found in FIG. 7. Results of the cytolytic activity assay can be found in FIG. 8.
Example 3
[0273] Female Jackson NSG mice (NOD.Cg-Prkdc.sup.scidIl2rg.sup.tm1Wjl/SzJ) aged 5-6 weeks old were used in this study. The mice were fed an irradiated Harlan 2918.15 Rodent Diet and water ad libitum. The mice were housed in Innovive.TM. disposable ventilated caging with corn cob bedding inside Biobubble.RTM. Clean Rooms that provide H.E.P.A filtered air into the bubble environment at 100 complete air changes per hour. All treatments, body weight determinations, and tumor measurements were carried out in the bubble environment. The environment was controlled to a temperature range of 70.degree..+-.2.degree. F. and a humidity range of 30-70%. All procedures were conducted in compliance with all the laws, regulations and guidelines of the National Institutes of Health (NIH) and with the approval of Molecular Imaging, Inc.'s Animal Care and Use Committee.
Tumor Cell Preparation
[0274] U937-luc cells were obtained in Lifor.RTM. Preservation Solution. The cells were centrifuged at 200 rcf for 8 minutes at 4.degree. C., the supernatant was aspirated, and the pellet was re-suspended in cold Dulbecco's Phosphate Buffered Saline (DPBS) by pipetting. An aliquot of the homogeneous cell suspension was diluted in a trypan blue solution and counted using a Luna.TM. automated cell counter. The cell suspension was centrifuged at 200 rcf for 8 minutes at 4.degree. C. The supernatant was aspirated and the cell pellet was re-suspended in cold serum-free medium to generate the final concentrations of trypan-excluding cells/ml. The cell suspension was maintained on wet ice during implantation. Test animals were implanted with 1.00E+06 cells intravenously via the lateral tail vein on Day 0 in 0.2 ml using a 27-gauge needle and syringe.
CAR T-Cell Preparation
[0275] T cells according to the invention were obtained, frozen on dry ice, and stored in liquid nitrogen. On the day of treatment, the provided cryovials were removed from cryostorage and thawed in a 37.degree. C. water bath. For each group, the provided T cells were combined into a single 50 ml conical tube with warm RPMI 1640 supplemented with 10% FBS. The cryovial tubes were rinsed with warm RPMI 1640 with 10% FBS to minimize loss of cells to reach a total volume of 50 ml in each conical tube. Each 50 ml conical tube was centrifuged at 200 rcf for 8 minutes at 4.degree. C. The supernatants were aspirated, and the cell pellets re-suspended in 10 ml of room temperature DPBS. An aliquot of the homogeneous cell suspension was diluted in a trypan blue solution and manually counted using a hemacytometer. The cell suspensions were again centrifuged at 200 rcf for 8 minutes at 4.degree. C. The supernatants were aspirated and the cell pellets were re-suspended in room temperature DPBS to generate the required final concentrations. The cell suspensions were maintained on wet ice during treatment administration.
Bioluminescence Imaging
[0276] In vivo bioluminescence imaging (BLI) was performed using an IVIS Spectrum (Perkin Elmer, Hopkinton, Mass.). Animals were imaged up to 5 at a time under .about.1-2% isoflurane gas anesthesia. Each mouse was injected IP with 150 mg/kg (15 mg/ml) D-luciferin and imaged in the prone, then supine positions, 10 minutes following injection. Large to small binning of the CCD chip was used, and exposure time adjusted (2 seconds to 2 minutes) to obtain at least several hundred counts per image, and further to avoid saturation of the CCD chip. BLI images were collected on Days 3, 11, 18, and 25. Images were analyzed using the Living Image version 4.5 (Perkin Elmer, Hopkinton, Mass.) software. Whole body fixed-volume ROIs were placed on prone and supine images for each individual animal, and labeled based on animal identification. Total radiance expressed in photon/sec (p/s) was calculated and exported for all ROIs to facilitate analyses between groups. The prone and supine ROIs were summed together to estimate whole body tumor burden.
Treatment
[0277] All mice were sorted into study groups based on BLI-derived estimation of whole body tumor burden. The mice were distributed to ensure that the mean tumor burden for all groups was within 10% of the overall mean tumor burden for the study population. Treatment with CAR T cells began on Day 3. All mice were dosed with a fixed volume of 0.2 mL. The results are set forth in FIG. 10.
Assessment of Side Effects
[0278] All animals were observed for clinical signs at least once daily. Animals were weighed on each day of treatment. Individual body weights were recorded 3 times weekly.
[0279] The following sequences will further exemplify the invention.
TABLE-US-00017 CD28T DNA Extracellular, transmembrane, intracellular CTTGATAATGAAAAGTCAAACGGAACAATCATTCACGTGAAGGG CAAGCACCTCTGTCCGTCACCCTTGTTCCCTGGTCCATCCAAGCCA TTCTGGGTGTTGGTCGTAGTGGGTGGAGTCCTCGCTTGTTACTCTC TGCTCGTCACCGTGGCTTTTATAATCTTCTGGGTTAGATCCAAAAG AAGCCGCCTGCTCCATAGCGATTACATGAATATGACTCCACGCCG CCCTGGCCCCACAAGGAAACACTACCAGCCTTACGCACCACCTAG AGATTTCGCTGCCTATCGGAGC (SEQ ID NO. 1) CD28T Extracellular, transmembrane, intracellular AA LDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLL VTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFA AYRS (SEQ ID NO. 2) CD28T DNA-Extracellular CTTGATAATGAAAAGTCAAACGGAACAATCATTCACGTGAAGGG CAAGCACCTCTGTCCGTCACCCTTGTTCCCTGGTCCATCCAAGCCA (SEQ ID NO. 3) CD28T AA-Extracellular LDNEKSNGTI IHVKGKHLCP SPLFPGPSKP (SEQ ID NO. 4) CD28 DNA Transmembrane Domain TTCTGGGTGTTGGTCGTAGTGGGTGGAGTCCTCGCTTGTTACTCTC TGCTCGTCACCGTGGCTTTTATAATCTTCTGGGTT (SEQ ID NO. 5) CD28 AA Transmembrane Domain FWVLVVVGGV LACYSLLVTV AFIIFWV (SEQ ID NO. 6) CD28 DNA Intracellular Domain AGATCCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATATG ACTCCACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTAC GCACCACCTAGAGATTTCGCTGCCTATCGGAGC (SEQ ID NO. 7) CD28 AA Intracellular Domain RSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS (SEQ ID NO. 8) CD3 zeta DNA AGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATCAGCAG GGCCAGAACCAACTGTATAACGAGCTCAACCTGGGACGCAGGGA AGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACCCTGAGA TGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGTCTCTAT AATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCTGAAAT AGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACGACGGT TTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATGACGCT CTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 9) CD3 zeta AA RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEM GGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLY QGLSTATKDTYDALHMQALPPR (SEQ ID NO. 10) CD3 zeta variant AA RVKFSRSADAPAYKQGQNQLYNELNLGRREEYDVLDKRRGRDPEM GGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLY QGLSTATKDTYDALHMQALPPR (SEQ ID NO. 146) CD28 DNA ATTGAGGTGATGTATCCACCGCCTTACCTGGATAACGAAAAGAGT AACGGTACCATCATTCACGTGAAAGGTAAACACCTGTGTCCTTCT CCCCTCTTCCCCGGGCCATCAAAGCCC (SEQ ID NO. 11) CD28 AA IEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO. 12) CD8 DNA extracellular & transmembrane domain GCTGCAGCATTGAGCAACTCAATAATGTATTTTAGTCACTTTGTAC CAGTGTTCTTGCCGGCTAAGCCTACTACCACACCCGCTCCACGGC CACCTACCCCAGCTCCTACCATCGCTTCACAGCCTCTGTCCCTGCG CCCAGAGGCTTGCCGACCGGCCGCAGGGGGCGCTGTTCATACCAG AGGACTGGATTTCGCCTGCGATATCTATATCTGGGCACCCCTGGC CGGAACCTGCGGCGTACTCCTGCTGTCCCTGGTCATCACGCTCTAT TGTAATCACAGGAAC (SEQ ID NO. 13) CD8 AA extracellular & transmembrane Domain AAALSNSIMYFSHFVPVFLPAKPTTTPAPRPPTPAPTIASQPLSLRPEA CRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLYCNHRN (SEQ ID NO. 14) Clone 24C1 HC DNA CAGGTGCAGCTGCAGGAATCCGGACCGGGGCTGGTGAAGCCCAG CGAGACTCTGAGTCTCACGTGTACAGTTTCTGGAGGTAGCATTAG CTCCTACTATTGGTCATGGATAAGGCAGCCCCCCGGGAAGGGATT GGAATGGATCGGCTATATTTACTACAGTGGGAGCACCAATTACAA CCCCTCACTGAAGTCTAGAGTTACAATCAGCGTTGACACCTCAAA GAATCAGTTCAGTTTGAAATTGTCTAGCGTCACAGCAGCTGATAC AGCCGTCTATTATTGTGTTTCTCTGGTCTATTGCGGTGGGGATTGT TACAGTGGCTTTGACTATTGGGGGCAGGGTACTCTGGTTACAGTT TCTTCC (SEQ ID NO. 15) Clone 24C1 HC AA (CDRs Underlined) QVQLQESGPGLVKPSETLSLTCTVSGGSISSYYWSWIRQPPGKGLEWI GY IYYSGSTNYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCVSL VYCGGDCYSGFDYWGQGTLVTVSS (SEQ ID NO. 16) Clone 24C1 HC AA CDR1: GGSISSY (SEQ ID NO. 17) Clone 24C1 HC AA CDR2: YYSGS (SEQ ID NO. 18) Clone 24C1 HC AA CDR3: LVYCGGDCYS GFDY (SEQ ID NO. 19) Clone 24C1 LC DNA GACATCCAGTTGACACAGAGCCCGAGTTCCTTGTCCGCCTCCGTC GGGGATAGAGTGTCATTTACCTGTCAGGCCTCTCAGGATATTAAT AACTTTCTGAATTGGTATCAGCAAAAGCCCGGAAAGGCACCCAAG CTGTTGATTTACGACGCCAGTAACCTGGAGACAGGCGTGCCCTCC CGGTTTAGTGGTAGCGGAAGCGGTACGGATTTTACCTTTACTATC AGCTCTCTCCAACCCGAAGACATTGCAACCTACTATTGTCAACAA TATGGAAACCTGCCTTTTACATTTGGCGGCGGCACCAAGGTGGAG ATTAAGCGG (SEQ ID NO. 20) Clone 24C1 LC AA (CDRs Underlined) DIQLTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLI YDASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFT FGGGTKVEIKR (SEQ ID NO. 21) Clone 24C1 LC CDR1 AA: QASQDINNFLN (SEQ ID NO. 22) Clone 24C1 LC CDR2 AA: DASNLET (SEQ ID NO. 23) Clone 24C1 LC CDR3 AA: QQYGNLPFT (SEQ ID NO. 24) Clone 24C1 CD28T CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTCCAACTGCAAGAAAGCGGACCC GGACTGGTGAAGCCTTCTGAGACACTTAGTCTGACGTGCACGGTC AGTGGCGGCTCCATCTCCTCCTATTATTGGTCATGGATACGACAA CCCCCAGGTAAGGGCCTGGAATGGATTGGCTATATCTACTATTCA GGAAGCACGAACTACAATCCCAGCCTGAAGTCCCGAGTGACAATT TCAGTAGATACCAGTAAAAACCAGTTCAGTCTTAAACTGTCAAGC GTGACAGCTGCCGACACCGCTGTGTATTACTGCGTCTCACTGGTG TATTGTGGAGGGGATTGTTATAGCGGGTTCGATTATTGGGGACAG GGAACCCTGGTGACTGTATCTTCCGGCGGCGGCGGCTCAGGGGGT GGCGGTAGTGGCGGTGGGGGTTCCGATATTCAACTGACACAATCC CCCAGCTCACTCAGCGCCAGCGTGGGGGACAGGGTTAGCTTTACC TGTCAAGCCTCTCAGGATATAAATAACTTTCTGAACTGGTATCAA CAGAAGCCTGGGAAGGCGCCCAAACTCCTGATCTATGATGCGTCC AACCTGGAAACTGGCGTGCCTTCACGCTTTAGCGGCTCTGGCAGT GGTACAGACTTCACTTTTACCATCTCTTCACTTCAGCCGGAGGACA TCGCCACATATTACTGTCAACAGTACGGAAACTTGCCCTTTACTTT TGGAGGCGGCACCAAAGTTGAAATCAAAAGGGCCGCTGCCCTGG ATAACGAAAAGAGCAATGGGACTATAATACATGTTAAAGGAAAA CACCTGTGTCCATCTCCCCTGTTCCCTGGACCGTCAAAGCCATTTT GGGTGCTCGTGGTTGTCGGTGGCGTTCTCGCCTGTTATAGCTTGCT GGTGACAGTAGCCTTCATTATCTTTTGGGTGAGATCCAAAAGAAG CCGCCTGCTCCATAGCGATTACATGAATATGACTCCACGCCGCCC TGGCCCCACAAGGAAACACTACCAGCCTTACGCACCACCTAGAGA TTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCTGCAGA TGCACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAACGAGCT CAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACAAGCGCA GAGGACGGGACCCTGAGATGGGTGGCAAACCAAGACGAAAAAAC CCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAAGATGGCT GAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGAGAAGGGG AAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACTGCTACGA AGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCACCTAGGT AA (SEQ ID NO. 25) Clone 24C1 CD28T CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in bold) MALPVTALLLPLALLLHAARPQVQLQESGPGLVKPSETLSLTCTVS GGSISSYYWSWIRQPPGKGLEWIGYIYYSGSTNYNPSLKSRVTISVDT SKNQFSLKLSSVTAADTAVYYCVSLVYCGGDCYSGFDYWGQGTLV TVSSGGGGSGGGGSGGGGSDIQLTQSPSSLSASVGDRVSFTCQASQDI NNFLNWYQQKPGKAPKLLIYDASNLETGVPSRFSGSGSGTDFTFTISS LQPEDIATYYCQQYGNLPFTFGGGTKVEIKRAAALDNEKSNGTIIHV KGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSK RSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSA DAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKN PQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATK DTYDALHMQALPPR (SEQ ID NO. 26) Clone 24C1 CD28T CD3 zeta CAR DNA Heavy & Light Chains CAGGTCCAACTGCAAGAAAGCGGACCCGGACTGGTGAAGCCTTCT GAGACACTTAGTCTGACGTGCACGGTCAGTGGCGGCTCCATCTCC TCCTATTATTGGTCATGGATACGACAACCCCCAGGTAAGGGCCTG GAATGGATTGGCTATATCTACTATTCAGGAAGCACGAACTACAAT CCCAGCCTGAAGTCCCGAGTGACAATTTCAGTAGATACCAGTAAA AACCAGTTCAGTCTTAAACTGTCAAGCGTGACAGCTGCCGACACC GCTGTGTATTACTGCGTCTCACTGGTGTATTGTGGAGGGGATTGTT ATAGCGGGTTCGATTATTGGGGACAGGGAACCCTGGTGACTGTAT CTTCCGGCGGCGGCGGCTCAGGGGGTGGCGGTAGTGGCGGTGGG GGTTCCGATATTCAACTGACACAATCCCCCAGCTCACTCAGCGCC AGCGTGGGGGACAGGGTTAGCTTTACCTGTCAAGCCTCTCAGGAT ATAAATAACTTTCTGAACTGGTATCAACAGAAGCCTGGGAAGGCG CCCAAACTCCTGATCTATGATGCGTCCAACCTGGAAACTGGCGTG CCTTCACGCTTTAGCGGCTCTGGCAGTGGTACAGACTTCACTTTTA CCATCTCTTCACTTCAGCCGGAGGACATCGCCACATATTACTGTCA ACAGTACGGAAACTTGCCCTTTACTTTTGGAGGCGGCACCAAAGT TGAAATCAAAAGGGCCGCTGCCCTGGATAACGAAAAGAGCAATG GGACTATAATACATGTTAAAGGAAAACACCTGTGTCCATCTCCCC TGTTCCCTGGACCGTCAAAGCCATTTTGGGTGCTCGTGGTTGTCGG TGGCGTTCTCGCCTGTTATAGCTTGCTGGTGACAGTAGCCTTCATT ATCTTTTGGGTGAGATCCAAAAGAAGCCGCCTGCTCCATAGCGAT TACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGGAAACAC TACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTATCGGAGC AGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATCAGCAG GGCCAGAACCAACTGTATAACGAGCTCAACCTGGGACGCAGGGA AGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACCCTGAGA TGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGTCTCTAT AATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCTGAAAT AGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACGACGGT TTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATGACGCT CTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 27) Clone 24C1 CD28T CD3 zeta CAR AA Heavy & Light Chains QVQLQESGPGLVKPSETLSLTCTVSGGSISSYYWSWIRQPPGKGLEWI GYIYYSGSTNYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYC VSLVYCGGDCYSGFDYWGQGTLVTVSSGGGGSGGGGSGGGGSDIQ LTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLIYD ASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFTFG GGTKVEIKRAAALDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLV VVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRK HYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYNELNLGRRE EYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIG MKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 28) Clone 24C1 CD28 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTGCAGCTGCAGGAATCCGGACCG GGGCTGGTGAAGCCCAGCGAGACTCTGAGTCTCACGTGTACAGTT TCTGGAGGTAGCATTAGCTCCTACTATTGGTCATGGATAAGGCAG CCCCCCGGGAAGGGATTGGAATGGATCGGCTATATTTACTACAGT GGGAGCACCAATTACAACCCCTCACTGAAGTCTAGAGTTACAATC AGCGTTGACACCTCAAAGAATCAGTTCAGTTTGAAATTGTCTAGC GTCACAGCAGCTGATACAGCCGTCTATTATTGTGTTTCTCTGGTCT ATTGCGGTGGGGATTGTTACAGTGGCTTTGACTATTGGGGGCAGG GTACTCTGGTTACAGTTTCTTCCGGGGGGGGAGGCTCTGGGGGCG GAGGCTCAGGTGGTGGAGGCAGCGACATCCAGTTGACACAGAGC CCGAGTTCCTTGTCCGCCTCCGTCGGGGATAGAGTGTCATTTACCT GTCAGGCCTCTCAGGATATTAATAACTTTCTGAATTGGTATCAGC AAAAGCCCGGAAAGGCACCCAAGCTGTTGATTTACGACGCCAGT AACCTGGAGACAGGCGTGCCCTCCCGGTTTAGTGGTAGCGGAAGC GGTACGGATTTTACCTTTACTATCAGCTCTCTCCAACCCGAAGACA TTGCAACCTACTATTGTCAACAATATGGAAACCTGCCTTTTACATT TGGCGGCGGCACCAAGGTGGAGATTAAGCGGGCGGCAGCTATTG AGGTGATGTATCCACCGCCTTACCTGGATAACGAAAAGAGTAACG GTACCATCATTCACGTGAAAGGTAAACACCTGTGTCCTTCTCCCCT CTTCCCCGGGCCATCAAAGCCCTTCTGGGTTCTTGTGGTCGTGGGA GGCGTGCTTGCTTGTTATTCTCTGCTCGTTACCGTGGCGTTTATCA TTTTTTGGGTTAGATCCAAAAGAAGCCGCCTGCTCCATAGCGATT ACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGGAAACACT ACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTATCGGAGCA GGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATCAGCAGG GCCAGAACCAACTGTATAACGAGCTCAACCTGGGACGCAGGGAA GAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACCCTGAGAT
GGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGTCTCTATA ATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCTGAAATA GGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACGACGGTT TGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATGACGCTC TCCACATGCAAGCCCTGCCACCTAGGTAA (SEQ ID NO. 29) Clone 24C1 CD28 CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPQVQLQESGPGLVKPSETLSLTCTVS GGSISSYYWSWIRQPPGKGLEWIGYIYYSGSTNYNPSLKSRVTISVDT SKNQFSLKLSSVTAADTAVYYCVSLVYCGGDCYSGFDYWGQGTLV TVSSGGGGSGGGGSGGGGSDIQLTQSPSSLSASVGDRVSFTCQASQDI NNFLNWYQQKPGKAPKLLIYDASNLETGVPSRFSGSGSGTDFTFTISS LQPEDIATYYCQQYGNLPFTFGGGTKVEIKRAAAIEVMYPPPYLDNE KSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVA FIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEM GGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLY QGLSTATKDTYDALHMQALPPR (SEQ ID NO. 30) Clone 24C1 CD28 CD3 zeta CAR DNA Heavy & Light Chains CAGGTGCAGCTGCAGGAATCCGGACCGGGGCTGGTGAAGCCCAG CGAGACTCTGAGTCTCACGTGTACAGTTTCTGGAGGTAGCATTAG CTCCTACTATTGGTCATGGATAAGGCAGCCCCCCGGGAAGGGATT GGAATGGATCGGCTATATTTACTACAGTGGGAGCACCAATTACAA CCCCTCACTGAAGTCTAGAGTTACAATCAGCGTTGACACCTCAAA GAATCAGTTCAGTTTGAAATTGTCTAGCGTCACAGCAGCTGATAC AGCCGTCTATTATTGTGTTTCTCTGGTCTATTGCGGTGGGGATTGT TACAGTGGCTTTGACTATTGGGGGCAGGGTACTCTGGTTACAGTT TCTTCCGGGGGGGGAGGCTCTGGGGGCGGAGGCTCAGGTGGTGG AGGCAGCGACATCCAGTTGACACAGAGCCCGAGTTCCTTGTCCGC CTCCGTCGGGGATAGAGTGTCATTTACCTGTCAGGCCTCTCAGGA TATTAATAACTTTCTGAATTGGTATCAGCAAAAGCCCGGAAAGGC ACCCAAGCTGTTGATTTACGACGCCAGTAACCTGGAGACAGGCGT GCCCTCCCGGTTTAGTGGTAGCGGAAGCGGTACGGATTTTACCTT TACTATCAGCTCTCTCCAACCCGAAGACATTGCAACCTACTATTGT CAACAATATGGAAACCTGCCTTTTACATTTGGCGGCGGCACCAAG GTGGAGATTAAGCGGGCGGCAGCTATTGAGGTGATGTATCCACCG CCTTACCTGGATAACGAAAAGAGTAACGGTACCATCATTCACGTG AAAGGTAAACACCTGTGTCCTTCTCCCCTCTTCCCCGGGCCATCAA AGCCCTTCTGGGTTCTTGTGGTCGTGGGAGGCGTGCTTGCTTGTTA TTCTCTGCTCGTTACCGTGGCGTTTATCATTTTTTGGGTTAGATCC AAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATATGACTCCA CGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTACGCACCA CCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGA TCTGCAGATGCACCAGCGTATCAGCAGGGCCAGAACCAACTGTAT AACGAGCTCAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGA CAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAACCAAGAC GAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGAT AAGATGGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCG GAGAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGACTCAGCA CTGCTACGAAGGATACTTATGACGCTCTCCACATGCAAGCCCTGC CACCTAGG (SEQ ID NO. 31) Clone 24C1 CD28 CD3 zeta CAR AA Heavy & Light Chains QVQLQESGPGLVKPSETLSLTCTVSGGSISSYYWSWIRQPPGKGLEWI GYIYYSGSTNYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYC VSLVYCGGDCYSGFDYWGQGTLVTVSSGGGGSGGGGSGGGGSDIQ LTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLIYD ASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFTFG GGTKVEIKRAAAIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGP SKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMT PRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLY NELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDK MAEAYSEIGMKGERRRGKGHDGLYQGLST ATKDTYDALHMQALPPR (SEQ ID NO. 32) Clone 24C1 CD8 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTGCAATTGCAAGAGTCCGGCCCCG GACTCGTTAAACCCAGTGAGACGCTTAGCCTGACCTGTACCGTCT CAGGGGGCAGCATCTCCTCTTATTACTGGAGCTGGATCAGGCAGC CTCCAGGAAAAGGCCTTGAATGGATTGGGTACATCTACTACTCTG GCTCAACAAATTATAATCCATCCCTGAAGTCCCGCGTGACTATCT CTGTGGACACCAGCAAGAATCAGTTTTCACTGAAGTTGTCTAGTG TTACCGCGGCCGACACCGCCGTATACTACTGTGTGTCTCTTGTGTA CTGTGGCGGCGACTGCTATTCCGGGTTCGACTACTGGGGCCAAGG GACTCTGGTAACCGTGTCCTCAGGCGGCGGCGGGTCAGGAGGAG GCGGCAGTGGAGGTGGCGGCTCCGACATCCAGCTGACACAATCA CCATCTTCCCTTTCAGCTTCAGTCGGGGACAGAGTGTCCTTCACAT GCCAGGCCAGCCAGGATATCAATAACTTCCTGAACTGGTACCAAC AGAAACCCGGAAAGGCTCCAAAGCTCCTGATCTATGATGCTTCCA ACCTGGAGACCGGCGTGCCCTCCAGGTTCAGTGGTTCAGGATCAG GCACTGACTTTACGTTCACCATATCCAGTCTTCAGCCCGAAGACA TTGCAACCTATTACTGCCAACAATACGGGAACCTTCCCTTTACATT CGGAGGCGGCACCAAGGTGGAAATCAAAAGGGCTGCAGCATTGA GCAACTCAATAATGTATTTTAGTCACTTTGTACCAGTGTTCTTGCC GGCTAAGCCTACTACCACACCCGCTCCACGGCCACCTACCCCAGC TCCTACCATCGCTTCACAGCCTCTGTCCCTGCGCCCAGAGGCTTGC CGACCGGCCGCAGGGGGCGCTGTTCATACCAGAGGACTGGATTTC GCCTGCGATATCTATATCTGGGCACCCCTGGCCGGAACCTGCGGC GTACTCCTGCTGTCCCTGGTCATCACGCTCTATTGTAATCACAGGA ACAGATCCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATA TGACTCCACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTT ACGCACCACCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGT TTTCCAGATCTGCAGATGCACCAGCGTATCAGCAGGGCCAGAACC AACTGTATAACGAGCTCAACCTGGGACGCAGGGAAGAGTATGAC GTTTTGGACAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAA ACCAAGACGAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGC AGAAGGATAAGATGGCTGAAGCCTATTCTGAAATAGGCATGAAA GGAGAGCGGAGAAGGGGAAAAGGGCACGACGGTTTGTACCAGGG ACTCAGCACTGCTACGAAGGATACTTATGACGCTCTCCACATGCA AGCCCTGCCACCTAGGTAA (SEQ ID NO. 33) Clone 24C1 CD8 CD3 zeta CAR AA Heavy & Light Chains (Signal peptide in bold) MALPVTALLLPLALLLHAARPQVQLQESGPGLVKPSETLSLTCTVS GGSISSYYWSWIRQPPGKGLEWIGYIYYSGSTNYNPSLKSRVTISVDT SKNQFSLKLSSVTAADTAVYYCVSLVYCGGDCYSGFDYWGQGTLV TVSSGGGGSGGGGSGGGGSDIQLTQSPSSLSASVGDRVSFTCQASQDI NNFLNWYQQKPGKAPKLLIYDASNLETGVPSRFSGSGSGTDFTFTISS LQPEDIATYYCQQYGNLPFTFGGGTKVEIKRAAALSNSIMYFSHFVPV FLPAKPTTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFA CDIYIWAPLAGTCGVLLLSLVITLYCNHRNRSKRSRLLHSDYMNMTP RRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKM AEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 34) Clone 24C1 CD8 CD3 zeta CAR DNA Heavy & Light Chains CAGGTGCAATTGCAAGAGTCCGGCCCCGGACTCGTTAAACCCAGT GAGACGCTTAGCCTGACCTGTACCGTCTCAGGGGGCAGCATCTCC TCTTATTACTGGAGCTGGATCAGGCAGCCTCCAGGAAAAGGCCTT GAATGGATTGGGTACATCTACTACTCTGGCTCAACAAATTATAAT CCATCCCTGAAGTCCCGCGTGACTATCTCTGTGGACACCAGCAAG AATCAGTTTTCACTGAAGTTGTCTAGTGTTACCGCGGCCGACACC GCCGTATACTACTGTGTGTCTCTTGTGTACTGTGGCGGCGACTGCT ATTCCGGGTTCGACTACTGGGGCCAAGGGACTCTGGTAACCGTGT CCTCAGGCGGCGGCGGGTCAGGAGGAGGCGGCAGTGGAGGTGGC GGCTCCGACATCCAGCTGACACAATCACCATCTTCCCTTTCAGCTT CAGTCGGGGACAGAGTGTCCTTCACATGCCAGGCCAGCCAGGATA TCAATAACTTCCTGAACTGGTACCAACAGAAACCCGGAAAGGCTC CAAAGCTCCTGATCTATGATGCTTCCAACCTGGAGACCGGCGTGC CCTCCAGGTTCAGTGGTTCAGGATCAGGCACTGACTTTACGTTCA CCATATCCAGTCTTCAGCCCGAAGACATTGCAACCTATTACTGCC AACAATACGGGAACCTTCCCTTTACATTCGGAGGCGGCACCAAGG TGGAAATCAAAAGGGCTGCAGCATTGAGCAACTCAATAATGTATT TTAGTCACTTTGTACCAGTGTTCTTGCCGGCTAAGCCTACTACCAC ACCCGCTCCACGGCCACCTACCCCAGCTCCTACCATCGCTTCACA GCCTCTGTCCCTGCGCCCAGAGGCTTGCCGACCGGCCGCAGGGGG CGCTGTTCATACCAGAGGACTGGATTTCGCCTGCGATATCTATATC TGGGCACCCCTGGCCGGAACCTGCGGCGTACTCCTGCTGTCCCTG GTCATCACGCTCTATTGTAATCACAGGAACAGATCCAAAAGAAGC CGCCTGCTCCATAGCGATTACATGAATATGACTCCACGCCGCCCT GGCCCCACAAGGAAACACTACCAGCCTTACGCACCACCTAGAGAT TTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCTGCAGAT GCACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAACGAGCT CAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACAAGCGCA GAGGACGGGACCCTGAGATGGGTGGCAAACCAAGACGAAAAAAC CCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAAGATGGCT GAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGAGAAGGGG AAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACTGCTACGA AGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 35) Clone 24C1 CD8 CD3 zeta CAR AA Heavy & Light Chains QVQLQESGPGLVKPSETLSLTCTVSGGSISSYYWSWIRQPPGKGLEWI GYIYYSGSTNYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYC VSLVYCGGDCYSGFDYWGQGTLVTVSSGGGGSGGGGSGGGGSDIQ LTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLIYD ASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFTFG GGTKVEIKRAAALSNSIMYFSHFVPVFLPAKPTTTPAPRPPTPAPTIAS QPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVI TLYCNHRNRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAA YRSRVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDP EMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDG LYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 36) Clone 24C1 CD28T CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGGATATCCAGCTCACGCAATCCCCCTCAA GCTTGAGTGCCTCCGTGGGCGACCGGGTGTCCTTCACATGTCAGG CAAGCCAAGACATAAATAATTTCCTGAATTGGTACCAACAAAAAC CCGGCAAGGCTCCCAAACTCCTGATTTATGATGCCTCCAATCTGG AGACCGGGGTCCCTTCTAGATTCAGCGGAAGTGGCAGCGGCACA GACTTTACATTTACTATCTCTTCTCTGCAACCAGAGGACATCGCCA CATACTATTGCCAGCAATACGGCAATCTGCCCTTCACCTTCGGAG GCGGAACCAAGGTAGAAATTAAAAGGGGCGGTGGAGGCTCCGGA GGGGGGGGCTCTGGCGGAGGGGGCTCCCAAGTACAATTGCAGGA GTCAGGGCCTGGACTCGTGAAGCCTTCAGAAACTTTGTCACTGAC ATGTACAGTGTCCGGCGGAAGCATTTCCAGTTACTATTGGTCCTG GATTAGACAGCCACCCGGCAAAGGACTGGAATGGATTGGATATA TCTACTACTCTGGATCTACAAACTATAATCCCAGCCTCAAATCCA GGGTCACTATTAGTGTGGATACATCAAAGAATCAGTTCTCCTTGA AGCTGAGCTCAGTCACTGCTGCCGACACCGCAGTGTACTATTGTG TGAGCCTGGTCTACTGCGGCGGAGATTGCTACAGCGGTTTCGATT ACTGGGGCCAGGGCACCCTGGTTACCGTTAGTTCCGCGGCTGCTC TTGATAACGAGAAGTCCAACGGTACGATTATCCACGTTAAGGGTA AGCACCTTTGCCCTAGCCCGCTGTTCCCAGGCCCCAGTAAGCCCTT TTGGGTCCTCGTTGTGGTAGGTGGGGTACTCGCCTGCTACTCCCTG CTCGTCACTGTCGCATTCATCATCTTCTGGGTCAGATCCAAAAGA AGCCGCCTGCTCCATAGCGATTACATGAATATGACTCCACGCCGC CCTGGCCCCACAAGGAAACACTACCAGCCTTACGCACCACCTAGA GATTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCTGCA GATGCACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAACGA GCTCAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACAAGC GCAGAGGACGGGACCCTGAGATGGGTGGCAAACCAAGACGAAAA AACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAAGAT GGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGAGAA GGGGAAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACTGCT ACGAAGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCACCT AGGTAA (SEQ ID NO. 37) Clone 24C1 CD28T CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPDIQLTQSPSSLSASVGDRVSFTCQAS QDINNFLNWYQQKPGKAPKLLIYDASNLETGVPSRFSGSGSGTDFTF TISSLQPEDIATYYCQQYGNLPFTFGGGTKVEIKRGGGGSGGGGSGG GGSQVQLQESGPGLVKPSETLSLTCTVSGGSISSYYWSWIRQPPGKGL EWIGYIYYSGSTNYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVY YCVSLVYCGGDCYSGFDYWGQGTLVTVSSAAALDNEKSNGTIIHVK GKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRS RLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADA PAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDT YDALHMQALPPR (SEQ ID NO. 38) Clone 24C1 CD28T CD3 zeta CAR DNA Heavy & Light Chains GATATCCAGCTCACGCAATCCCCCTCAAGCTTGAGTGCCTCCGTG GGCGACCGGGTGTCCTTCACATGTCAGGCAAGCCAAGACATAAAT AATTTCCTGAATTGGTACCAACAAAAACCCGGCAAGGCTCCCAAA CTCCTGATTTATGATGCCTCCAATCTGGAGACCGGGGTCCCTTCTA GATTCAGCGGAAGTGGCAGCGGCACAGACTTTACATTTACTATCT CTTCTCTGCAACCAGAGGACATCGCCACATACTATTGCCAGCAAT ACGGCAATCTGCCCTTCACCTTCGGAGGCGGAACCAAGGTAGAAA TTAAAAGGGGCGGTGGAGGCTCCGGAGGGGGGGGCTCTGGCGGA GGGGGCTCCCAAGTACAATTGCAGGAGTCAGGGCCTGGACTCGTG AAGCCTTCAGAAACTTTGTCACTGACATGTACAGTGTCCGGCGGA AGCATTTCCAGTTACTATTGGTCCTGGATTAGACAGCCACCCGGC AAAGGACTGGAATGGATTGGATATATCTACTACTCTGGATCTACA AACTATAATCCCAGCCTCAAATCCAGGGTCACTATTAGTGTGGAT ACATCAAAGAATCAGTTCTCCTTGAAGCTGAGCTCAGTCACTGCT GCCGACACCGCAGTGTACTATTGTGTGAGCCTGGTCTACTGCGGC GGAGATTGCTACAGCGGTTTCGATTACTGGGGCCAGGGCACCCTG GTTACCGTTAGTTCCGCGGCTGCTCTTGATAACGAGAAGTCCAAC GGTACGATTATCCACGTTAAGGGTAAGCACCTTTGCCCTAGCCCG CTGTTCCCAGGCCCCAGTAAGCCCTTTTGGGTCCTCGTTGTGGTAG GTGGGGTACTCGCCTGCTACTCCCTGCTCGTCACTGTCGCATTCAT CATCTTCTGGGTCAGATCCAAAAGAAGCCGCCTGCTCCATAGCGA TTACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGGAAACA CTACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTATCGGAG CAGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATCAGCA GGGCCAGAACCAACTGTATAACGAGCTCAACCTGGGACGCAGGG AAGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACCCTGAG ATGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGTCTCTA TAATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCTGAAAT AGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACGACGGT TTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATGACGCT CTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 39) Clone 24C1 CD28T CD3 zeta CAR AA Heavy & Light Chains
DIQLTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLI YDASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFT FGGGTKVEIKRGGGGSGGGGSGGGGSQVQLQESGPGLVKPSETLSLT CTVSGGSISSYYWSWIRQPPGKGLEWIGYIYYSGSTNYNPSLKSRVTI SVDTSKNQFSLKLSSVTAADTAVYYCVSLVYCGGDCYSGFDYWGQ GTLVTVSSAAALDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVV VGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKH YQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYNELNLGRREE YDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGM KGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 40) Clone 24C1 CD28 CD3 zeta CAR DNA AA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGGATATCCAGCTGACCCAGTCTCCATCCT CTTTGAGTGCCTCCGTGGGTGACCGCGTCTCTTTCACTTGCCAAGC CAGCCAAGACATCAACAACTTTCTGAATTGGTACCAGCAGAAACC AGGCAAAGCACCAAAGCTCCTCATCTACGACGCCTCCAACCTGGA AACCGGGGTGCCCAGCAGGTTTAGCGGGAGCGGTTCTGGCACGG ATTTTACGTTCACCATCTCCTCTCTGCAGCCCGAGGATATAGCTAC TTATTACTGTCAGCAGTACGGGAATCTGCCATTTACTTTTGGGGGT GGAACTAAGGTGGAAATCAAAAGGGGCGGCGGGGGAAGCGGGG GCGGGGGCTCAGGTGGCGGAGGGAGCCAGGTGCAACTCCAGGAA AGTGGCCCAGGATTGGTGAAGCCCAGCGAGACCCTTTCCCTTACT TGTACTGTTAGCGGAGGCAGCATAAGCAGCTACTATTGGTCCTGG ATCAGACAGCCACCAGGGAAAGGGCTTGAATGGATTGGCTACATT TACTATTCCGGGTCCACCAACTACAACCCATCCCTCAAGTCCCGC GTGACAATTTCCGTCGACACAAGCAAGAACCAGTTCTCCCTGAAA CTTAGTAGCGTCACTGCTGCAGATACAGCAGTGTACTATTGTGTC AGCCTTGTCTACTGTGGCGGCGACTGCTACAGTGGCTTTGATTACT GGGGACAGGGCACGCTCGTGACAGTGTCCAGCGCTGCGGCTATCG AGGTAATGTATCCGCCACCGTATCTGGACAACGAGAAGTCTAATG GGACAATCATTCACGTGAAGGGGAAGCACCTGTGTCCATCCCCCC TGTTTCCGGGTCCCAGTAAACCCTTCTGGGTGCTTGTTGTCGTTGG CGGGGTGCTGGCCTGCTATTCCCTGCTGGTGACCGTCGCGTTTATT ATTTTCTGGGTTAGATCCAAAAGAAGCCGCCTGCTCCATAGCGAT TACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGGAAACAC TACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTATCGGAGC AGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATCAGCAG GGCCAGAACCAACTGTATAACGAGCTCAACCTGGGACGCAGGGA AGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACCCTGAGA TGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGTCTCTAT AATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCTGAAAT AGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACGACGGT TTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATGACGCT CTCCACATGCAAGCCCTGCCACCTAGGTAA (SEQ ID NO. 41) Clone 24C1 CD28 CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPDIQLTQSPSSLSASVGDRVSFTCQAS QDINNFLNWYQQKPGKAPKLLIYDASNLETGVPSRFSGSGSGTDFTF TISSLQPEDIATYYCQQYGNLPFTFGGGTKVEIKRGGGGSGGGGSGG GGSQVQLQESGPGLVKPSETLSLTCTVSGGSISSYYWSWIRQPPGKGL EWIGYIYYSGSTNYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVY YCVSLVYCGGDCYSGFDYWGQGTLVTVSSAAAIEVMYPPPYLDNEK SNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFI IFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSR VKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMG GKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQ GLSTATKDTYDALHMQALPPR (SEQ ID NO. 42) Clone 24C1 CD28 CD3 zeta CAR DNA Heavy & Light Chains GATATCCAGCTGACCCAGTCTCCATCCTCTTTGAGTGCCTCCGTGG GTGACCGCGTCTCTTTCACTTGCCAAGCCAGCCAAGACATCAACA ACTTTCTGAATTGGTACCAGCAGAAACCAGGCAAAGCACCAAAG CTCCTCATCTACGACGCCTCCAACCTGGAAACCGGGGTGCCCAGC AGGTTTAGCGGGAGCGGTTCTGGCACGGATTTTACGTTCACCATC TCCTCTCTGCAGCCCGAGGATATAGCTACTTATTACTGTCAGCAGT ACGGGAATCTGCCATTTACTTTTGGGGGTGGAACTAAGGTGGAAA TCAAAAGGGGCGGCGGGGGAAGCGGGGGCGGGGGCTCAGGTGGC GGAGGGAGCCAGGTGCAACTCCAGGAAAGTGGCCCAGGATTGGT GAAGCCCAGCGAGACCCTTTCCCTTACTTGTACTGTTAGCGGAGG CAGCATAAGCAGCTACTATTGGTCCTGGATCAGACAGCCACCAGG GAAAGGGCTTGAATGGATTGGCTACATTTACTATTCCGGGTCCAC CAACTACAACCCATCCCTCAAGTCCCGCGTGACAATTTCCGTCGA CACAAGCAAGAACCAGTTCTCCCTGAAACTTAGTAGCGTCACTGC TGCAGATACAGCAGTGTACTATTGTGTCAGCCTTGTCTACTGTGGC GGCGACTGCTACAGTGGCTTTGATTACTGGGGACAGGGCACGCTC GTGACAGTGTCCAGCGCTGCGGCTATCGAGGTAATGTATCCGCCA CCGTATCTGGACAACGAGAAGTCTAATGGGACAATCATTCACGTG AAGGGGAAGCACCTGTGTCCATCCCCCCTGTTTCCGGGTCCCAGT AAACCCTTCTGGGTGCTTGTTGTCGTTGGCGGGGTGCTGGCCTGCT ATTCCCTGCTGGTGACCGTCGCGTTTATTATTTTCTGGGTTAGATC CAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATATGACTCC ACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTACGCACC ACCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAG ATCTGCAGATGCACCAGCGTATCAGCAGGGCCAGAACCAACTGTA TAACGAGCTCAACCTGGGACGCAGGGAAGAGTATGACGTTTTGG ACAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAACCAAGA CGAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGA TAAGATGGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAGAGC GGAGAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGACTCAGC ACTGCTACGAAGGATACTTATGACGCTCTCCACATGCAAGCCCTG CCACCTAGG (SEQ ID NO. 43) Clone 24C1 CD28 CD3 zeta CAR AA Heavy & Light Chains DIQLTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLI YDASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFT FGGGTKVEIKRGGGGSGGGGSGGGGSQVQLQESGPGLVKPSETLSLT CTVSGGSISSYYWSWIRQPPGKGLEWIGYIYYSGSTNYNPSLKSRVTI SVDTSKNQFSLKLSSVTAADTAVYYCVSLVYCGGDCYSGFDYWGQ GTLVTVSSAAAIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPS KPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTP RRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKM AEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 44) Clone 24C1 CD8 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGGACATTCAATTGACCCAGTCCCCTAGCA GTCTCTCAGCAAGTGTGGGAGATAGGGTGTCATTCACCTGTCAGG CTTCACAGGACATCAACAACTTCCTCAATTGGTATCAGCAGAAGC CAGGGAAGGCACCAAAGCTGCTCATATATGACGCTTCAAACCTTG AAACCGGAGTACCTAGCCGCTTCAGCGGAAGCGGATCAGGGACT GACTTCACTTTTACCATCTCTTCACTGCAGCCCGAAGACATCGCCA CATACTACTGCCAGCAGTACGGAAACTTGCCTTTTACATTTGGGG GCGGCACCAAAGTGGAGATTAAGCGAGGGGGAGGCGGCTCAGGA GGCGGTGGCTCCGGAGGCGGGGGTTCCCAGGTCCAGCTCCAGGA ATCCGGCCCAGGTCTGGTTAAGCCCAGTGAAACTTTGTCCCTCAC GTGTACTGTGAGCGGTGGTTCAATCTCCTCATACTATTGGTCTTGG ATACGGCAACCTCCTGGAAAGGGCCTCGAGTGGATCGGCTATATC TACTATAGTGGCTCCACTAATTACAACCCTTCCCTCAAGTCCAGA GTCACCATTTCCGTGGACACATCTAAGAACCAGTTCAGTCTGAAG TTGTCCAGCGTTACAGCCGCAGACACAGCCGTTTATTACTGTGTGT CTCTTGTTTACTGCGGGGGAGACTGTTATAGCGGCTTCGATTACTG GGGCCAGGGCACCTTGGTCACAGTCTCTTCCGCGGCCGCCCTCTC TAACAGTATTATGTACTTTTCTCATTTTGTACCCGTGTTCCTTCCCG CTAAGCCAACTACTACCCCGGCCCCACGGCCGCCTACCCCTGCAC CCACAATAGCCAGTCAGCCTTTGAGCCTGAGACCTGAGGCTTGTC GGCCGGCTGCTGGGGGTGCAGTGCACACACGAGGTCTTGATTTTG CTTGCGACATATACATCTGGGCCCCTCTGGCCGGGACCTGTGGGG TGCTGCTTCTGAGCTTGGTCATCACGCTCTATTGCAACCATCGCAA CAGATCCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATAT GACTCCACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTA CGCACCACCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTT TTCCAGATCTGCAGATGCACCAGCGTATCAGCAGGGCCAGAACCA ACTGTATAACGAGCTCAACCTGGGACGCAGGGAAGAGTATGACG TTTTGGACAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAA CCAAGACGAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCA GAAGGATAAGATGGCTGAAGCCTATTCTGAAATAGGCATGAAAG GAGAGCGGAGAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGA CTCAGCACTGCTACGAAGGATACTTATGACGCTCTCCACATGCAA GCCCTGCCACCTAGGTAA (SEQ ID NO. 45) Clone 24C1 CD8 CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPDIQLTQSPSSLSASVGDRVSFTCQAS QDINNFLNWYQQKPGKAPKLLIYDASNLETGVPSRFSGSGSGTDFTF TISSLQPEDIATYYCQQYGNLPFTFGGGTKVEIKRGGGGSGGGGSGG GGSQVQLQESGPGLVKPSETLSLTCTVSGGSISSYYWSWIRQPPGKGL EWIGYIYYSGSTNYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVY YCVSLVYCGGDCYSGFDYWGQGTLVTVSSAAALSNSIMYFSHFVPV FLPAKPTTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFA CDIYIWAPLAGTCGVLLLSLVITLYCNHRNRSKRSRLLHSDYMNMTP RRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKM AEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 46) Clone 24C1 CD8 CD3 zeta CAR DNA Heavy & Light Chains GACATTCAATTGACCCAGTCCCCTAGCAGTCTCTCAGCAAGTGTG GGAGATAGGGTGTCATTCACCTGTCAGGCTTCACAGGACATCAAC AACTTCCTCAATTGGTATCAGCAGAAGCCAGGGAAGGCACCAAA GCTGCTCATATATGACGCTTCAAACCTTGAAACCGGAGTACCTAG CCGCTTCAGCGGAAGCGGATCAGGGACTGACTTCACTTTTACCAT CTCTTCACTGCAGCCCGAAGACATCGCCACATACTACTGCCAGCA GTACGGAAACTTGCCTTTTACATTTGGGGGCGGCACCAAAGTGGA GATTAAGCGAGGGGGAGGCGGCTCAGGAGGCGGTGGCTCCGGAG GCGGGGGTTCCCAGGTCCAGCTCCAGGAATCCGGCCCAGGTCTGG TTAAGCCCAGTGAAACTTTGTCCCTCACGTGTACTGTGAGCGGTG GTTCAATCTCCTCATACTATTGGTCTTGGATACGGCAACCTCCTGG AAAGGGCCTCGAGTGGATCGGCTATATCTACTATAGTGGCTCCAC TAATTACAACCCTTCCCTCAAGTCCAGAGTCACCATTTCCGTGGAC ACATCTAAGAACCAGTTCAGTCTGAAGTTGTCCAGCGTTACAGCC GCAGACACAGCCGTTTATTACTGTGTGTCTCTTGTTTACTGCGGGG GAGACTGTTATAGCGGCTTCGATTACTGGGGCCAGGGCACCTTGG TCACAGTCTCTTCCGCGGCCGCCCTCTCTAACAGTATTATGTACTT TTCTCATTTTGTACCCGTGTTCCTTCCCGCTAAGCCAACTACTACC CCGGCCCCACGGCCGCCTACCCCTGCACCCACAATAGCCAGTCAG CCTTTGAGCCTGAGACCTGAGGCTTGTCGGCCGGCTGCTGGGGGT GCAGTGCACACACGAGGTCTTGATTTTGCTTGCGACATATACATC TGGGCCCCTCTGGCCGGGACCTGTGGGGTGCTGCTTCTGAGCTTG GTCATCACGCTCTATTGCAACCATCGCAACAGATCCAAAAGAAGC CGCCTGCTCCATAGCGATTACATGAATATGACTCCACGCCGCCCT GGCCCCACAAGGAAACACTACCAGCCTTACGCACCACCTAGAGAT TTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCTGCAGAT GCACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAACGAGCT CAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACAAGCGCA GAGGACGGGACCCTGAGATGGGTGGCAAACCAAGACGAAAAAAC CCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAAGATGGCT GAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGAGAAGGGG AAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACTGCTACGA AGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 47) Clone 24C1 CD8 CD3 zeta CAR AA Heavy & Light Chains DIQLTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLI YDASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFT FGGGTKVEIKRGGGGSGGGGSGGGGSQVQLQESGPGLVKPSETLSLT CTVSGGSISSYYWSWIRQPPGKGLEWIGYIYYSGSTNYNPSLKSRVTI SVDTSKNQFSLKLSSVTADTAVYYCVSLVYCGGDCYSGFDYWGQGT LVTVSSAAALSNSIMYFSHFVPVFLPAKPTTTPAPRPPTPAPTIASQPLS LRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITLY CNHRNRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS RVKFSRSADAPAYQQGQNQLYNELNLGREEYDVLDKRRGRDPEMG GKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQ GLSTATKDTYDALHMQALPPR (SEQ ID NO. 48) Clone 24C8 Heavy Chain (HC) DNA CAGGTACAGCTGCAGGAATCTGGGCCCGGACTTGTCAAGCCAAGT CAGACACTTTCTCTTACATGTACCGTGAGCGGCGGAAGTATAAGC AGTGGAGGCTTTTACTGGTCTTGGATACGGCAGCACCCAGGCAAA GGCTTGGAGTGGATTGGATACATTCATCATTCAGGATCTACACAC TATAATCCATCCCTTAAGTCCCGGGTCACCATTAGCATTGATACGT CTAAGAATCTGTTCAGTCTCAGGCTGTCCTCCGTCACTGCTGCCGA CACAGCCGTGTACTACTGCGCCTCCTTGGTTTACTGCGGAGGCGA CTGTTATAGCGGCTTTGATTATTGGGGGCAGGGGACCCTCGTAAC CGTGAGCTCT (SEQ ID NO. 48) Clone 24C8 AA HC (CDRs in Underline) QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGFYWSWIRQHPGKGL EWIGYIHHSGSTHYNPSLKSRVTISIDTSKNLFSLRLSSVTAADTAVYY CASLVYCGGDCYSGFDYWGQGTLVTVSS (SEQ ID NO. 50) Clone 24C8 HC CDR1 AA: GGSISSGGF (SEQ ID NO. 51) Clone 24C8 HC CDR2 AA: HHSGS (SEQ ID NO. 52) Clone 24C8 HC CDR3 AA: LVYCGGDCYS GFDY (SEQ ID NO. 53) Clone 24C8 Light Chain (LC) DNA GATATCCAGCTCACTCAAAGCCCCTCTAGTCTCTCTGCCTCAGTGG GGGATCGGGTCAGTTTTACTTGTCAAGCTTCACAGGATATCAACA ACTTCCTTAATTGGTATCAGCAGAAGCCAGGAAAAGCACCCAAGC TGCTCATCTATGATGCCTCAAATTTGGAGACGGGTGTTCCCAGTC GATTCTCTGGGTCAGGGTCCGGGACCGACTTTACGTTTACGATCTC CTCTCTGCAGCCCGAAGACATCGCCACATACTATTGTCAACAGTA CGGCAACTTGCCTTTCACATTTGGGGGCGGGACTAAGGTTGAAAT CAAGAGG (SEQ ID NO. 54) Clone 24C8 LC AA (CDRs in Underline) DIQLTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLI YDASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFT FGGGTKVEIKR (SEQ ID NO. 55) Clone 24C8 LC CDR1 AA: QASQDINNFLN (SEQ ID NO. 56) Clone 24C8 LC CDR2 AA: DASNLET (SEQ ID NO. 57) Clone 24C8 LC CDR3 AA: QQYGNLPFT (SEQ ID NO. 58)
Clone 24C8 CD28T CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTACAGCTGCAGGAATCTGGGCCCG GACTTGTCAAGCCAAGTCAGACACTTTCTCTTACATGTACCGTGA GCGGCGGAAGTATAAGCAGTGGAGGCTTTTACTGGTCTTGGATAC GGCAGCACCCAGGCAAAGGCTTGGAGTGGATTGGATACATTCATC ATTCAGGATCTACACACTATAATCCATCCCTTAAGTCCCGGGTCA CCATTAGCATTGATACGTCTAAGAATCTGTTCAGTCTCAGGCTGTC CTCCGTCACTGCTGCCGACACAGCCGTGTACTACTGCGCCTCCTTG GTTTACTGCGGAGGCGACTGTTATAGCGGCTTTGATTATTGGGGG CAGGGGACCCTCGTAACCGTGAGCTCTGGAGGGGGTGGGAGCGG GGGAGGAGGTTCAGGGGGGGGCGGCTCCGATATCCAGCTCACTC AAAGCCCCTCTAGTCTCTCTGCCTCAGTGGGGGATCGGGTCAGTT TTACTTGTCAAGCTTCACAGGATATCAACAACTTCCTTAATTGGTA TCAGCAGAAGCCAGGAAAAGCACCCAAGCTGCTCATCTATGATGC CTCAAATTTGGAGACGGGTGTTCCCAGTCGATTCTCTGGGTCAGG GTCCGGGACCGACTTTACGTTTACGATCTCCTCTCTGCAGCCCGAA GACATCGCCACATACTATTGTCAACAGTACGGCAACTTGCCTTTC ACATTTGGGGGCGGGACTAAGGTTGAAATCAAGAGGGCCGCTGC ACTGGACAATGAGAAGTCCAACGGCACCATCATCCACGTGAAGG GCAAGCACCTGTGCCCTAGTCCTCTGTTCCCAGGCCCATCCAAAC CTTTTTGGGTTCTTGTTGTGGTCGGGGGGGTGCTGGCCTGCTATTC TCTGCTGGTCACGGTGGCCTTCATAATTTTCTGGGTTAGATCCAAA AGAAGCCGCCTGCTCCATAGCGATTACATGAATATGACTCCACGC CGCCCTGGCCCCACAAGGAAACACTACCAGCCTTACGCACCACCT AGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCT GCAGATGCACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAA CGAGCTCAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACA AGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAACCAAGACGA AAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAA GATGGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGA GAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACT GCTACGAAGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCA CCTAGGTAA (SEQ ID NO. 59) Clone 24C8 CD28T CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPQVQLQESGPGLVKPSQTLSLTCTVS GGSISSGGFYWSWIRQHPGKGLEWIGYIHHSGSTHYNPSLKSRVTISI DTSKNLFSLRLSSVTAADTAVYYCASLVYCGGDCYSGFDYWGQGTL VTVSSGGGGSGGGGSGGGGSDIQLTQSPSSLSASVGDRVSFTCQASQ DINNFLNWYQQKPGKAPKLLIYDASNLETGVPSRFSGSGSGTDFTFTI SSLQPEDIATYYCQQYGNLPFTFGGGTKVEIKRAAALDNEKSNGTIIH VKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRS KRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRS ADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRK NPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTAT KDTYDALHMQALPPR (SEQ ID NO. 60) Clone 24C8 CD28T CD3 zeta CAR DNA Heavy & Light Chains CAGGTACAGCTGCAGGAATCTGGGCCCGGACTTGTCAAGCCAAGT CAGACACTTTCTCTTACATGTACCGTGAGCGGCGGAAGTATAAGC AGTGGAGGCTTTTACTGGTCTTGGATACGGCAGCACCCAGGCAAA GGCTTGGAGTGGATTGGATACATTCATCATTCAGGATCTACACAC TATAATCCATCCCTTAAGTCCCGGGTCACCATTAGCATTGATACGT CTAAGAATCTGTTCAGTCTCAGGCTGTCCTCCGTCACTGCTGCCGA CACAGCCGTGTACTACTGCGCCTCCTTGGTTTACTGCGGAGGCGA CTGTTATAGCGGCTTTGATTATTGGGGGCAGGGGACCCTCGTAAC CGTGAGCTCTGGAGGGGGTGGGAGCGGGGGAGGAGGTTCAGGGG GGGGCGGCTCCGATATCCAGCTCACTCAAAGCCCCTCTAGTCTCT CTGCCTCAGTGGGGGATCGGGTCAGTTTTACTTGTCAAGCTTCAC AGGATATCAACAACTTCCTTAATTGGTATCAGCAGAAGCCAGGAA AAGCACCCAAGCTGCTCATCTATGATGCCTCAAATTTGGAGACGG GTGTTCCCAGTCGATTCTCTGGGTCAGGGTCCGGGACCGACTTTA CGTTTACGATCTCCTCTCTGCAGCCCGAAGACATCGCCACATACT ATTGTCAACAGTACGGCAACTTGCCTTTCACATTTGGGGGCGGGA CTAAGGTTGAAATCAAGAGGGCCGCTGCACTGGACAATGAGAAG TCCAACGGCACCATCATCCACGTGAAGGGCAAGCACCTGTGCCCT AGTCCTCTGTTCCCAGGCCCATCCAAACCTTTTTGGGTTCTTGTTG TGGTCGGGGGGGTGCTGGCCTGCTATTCTCTGCTGGTCACGGTGG CCTTCATAATTTTCTGGGTTAGATCCAAAAGAAGCCGCCTGCTCC ATAGCGATTACATGAATATGACTCCACGCCGCCCTGGCCCCACAA GGAAACACTACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCT ATCGGAGCAGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGT ATCAGCAGGGCCAGAACCAACTGTATAACGAGCTCAACCTGGGA CGCAGGGAAGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGA CCCTGAGATGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGG GTCTCTATAATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATT CTGAAATAGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCA CGACGGTTTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTA TGACGCTCTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 61) Clone 24C8 CD28T CD3 zeta CAR AA Heavy & Light Chains QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGFYWSWIRQHPGKGL EWIGYIHHSGSTHYNPSLKSRVTISIDTSKNLFSLRLSSVTAADTAVYY CASLVYCGGDCYSGFDYWGQGTLVTVSSGGGGSGGGGSGGGGSDI QLTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLIY DASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFTF GGGTKVEIKRAAALDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVL VVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTR KHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYNELNLGRR EEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEI GMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 62) Clone 24C8 CD28 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTGCAGCTGCAGGAAAGCGGTCCG GGACTTGTCAAGCCGTCCCAAACGCTGAGTCTGACGTGTACTGTC TCTGGTGGCTCTATTTCTTCCGGGGGCTTTTATTGGTCTTGGATCA GACAACACCCTGGCAAAGGGCTGGAGTGGATAGGGTATATTCAC CACTCTGGGTCCACTCACTACAACCCATCATTGAAATCCAGAGTG ACTATCTCAATCGACACATCCAAGAACCTTTTCAGCCTGAGGTTG TCATCAGTTACCGCCGCTGACACCGCGGTGTATTATTGCGCCTCTC TCGTGTACTGCGGTGGCGATTGTTATAGTGGCTTTGACTACTGGG GGCAGGGGACATTGGTTACCGTTTCAAGTGGAGGCGGTGGGTCTG GCGGGGGCGGTAGCGGAGGTGGGGGGAGCGACATACAGCTTACG CAGAGCCCCTCCAGCCTTTCAGCCTCCGTGGGGGATAGGGTGTCC TTTACCTGCCAGGCTTCCCAGGACATAAACAACTTCCTCAATTGGT ATCAGCAAAAGCCCGGGAAAGCACCAAAGCTGCTCATCTACGAT GCCAGCAACCTGGAAACCGGAGTGCCGTCTCGCTTCTCTGGAAGT GGCAGTGGGACCGATTTCACTTTTACAATCTCAAGTTTGCAGCCA GAAGACATTGCAACATACTACTGTCAACAGTACGGCAATCTCCCC TTTACATTTGGGGGGGGAACTAAAGTGGAGATTAAGCGCGCTGCA GCCATTGAAGTTATGTATCCGCCCCCGTATCTGGATAACGAGAAA TCTAATGGTACCATAATACATGTGAAGGGGAAGCACCTCTGTCCA TCACCGCTGTTCCCCGGCCCTTCAAAACCTTTCTGGGTACTCGTTG TCGTGGGTGGAGTTCTGGCCTGCTATAGTCTGCTGGTGACCGTGG CGTTTATCATCTTCTGGGTAAGATCCAAAAGAAGCCGCCTGCTCC ATAGCGATTACATGAATATGACTCCACGCCGCCCTGGCCCCACAA GGAAACACTACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCT ATCGGAGCAGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGT ATCAGCAGGGCCAGAACCAACTGTATAACGAGCTCAACCTGGGA CGCAGGGAAGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGA CCCTGAGATGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGG GTCTCTATAATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATT CTGAAATAGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCA CGACGGTTTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTA TGACGCTCTCCACATGCAAGCCCTGCCACCTAGGTAA (SEQ ID NO. 63) Clone 24C8 CD28 CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPQVQLQESGPGLVKPSQTLSLTCTVS GGSISSGGFYWSWIRQHPGKGLEWIGYIHHSGSTHYNPSLKSRVTISI DTSKNLFSLRLSSVTAADTAVYYCASLVYCGGDCYSGFDYWGQGTL VTVSSGGGGSGGGGSGGGGSDIQLTQSPSSLSASVGDRVSFTCQASQ DINNFLNWYQQKPGKAPKLLIYDASNLETGVPSRFSGSGSGTDFTFTI SSLQPEDIATYYCQQYGNLPFTFGGGTKVEIKRAAAIEVMYPPPYLD NEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVT VAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAY RSRVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPE MGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGL YQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 64) Clone 24C8 CD28 CD3 zeta CAR DNA Heavy & Light Chains CAGGTGCAGCTGCAGGAAAGCGGTCCGGGACTTGTCAAGCCGTCC CAAACGCTGAGTCTGACGTGTACTGTCTCTGGTGGCTCTATTTCTT CCGGGGGCTTTTATTGGTCTTGGATCAGACAACACCCTGGCAAAG GGCTGGAGTGGATAGGGTATATTCACCACTCTGGGTCCACTCACT ACAACCCATCATTGAAATCCAGAGTGACTATCTCAATCGACACAT CCAAGAACCTTTTCAGCCTGAGGTTGTCATCAGTTACCGCCGCTG ACACCGCGGTGTATTATTGCGCCTCTCTCGTGTACTGCGGTGGCG ATTGTTATAGTGGCTTTGACTACTGGGGGCAGGGGACATTGGTTA CCGTTTCAAGTGGAGGCGGTGGGTCTGGCGGGGGCGGTAGCGGA GGTGGGGGGAGCGACATACAGCTTACGCAGAGCCCCTCCAGCCTT TCAGCCTCCGTGGGGGATAGGGTGTCCTTTACCTGCCAGGCTTCC CAGGACATAAACAACTTCCTCAATTGGTATCAGCAAAAGCCCGGG AAAGCACCAAAGCTGCTCATCTACGATGCCAGCAACCTGGAAACC GGAGTGCCGTCTCGCTTCTCTGGAAGTGGCAGTGGGACCGATTTC ACTTTTACAATCTCAAGTTTGCAGCCAGAAGACATTGCAACATAC TACTGTCAACAGTACGGCAATCTCCCCTTTACATTTGGGGGGGGA ACTAAAGTGGAGATTAAGCGCGCTGCAGCCATTGAAGTTATGTAT CCGCCCCCGTATCTGGATAACGAGAAATCTAATGGTACCATAATA CATGTGAAGGGGAAGCACCTCTGTCCATCACCGCTGTTCCCCGGC CCTTCAAAACCTTTCTGGGTACTCGTTGTCGTGGGTGGAGTTCTGG CCTGCTATAGTCTGCTGGTGACCGTGGCGTTTATCATCTTCTGGGT AAGATCCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATAT GACTCCACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTA CGCACCACCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTT TTCCAGATCTGCAGATGCACCAGCGTATCAGCAGGGCCAGAACCA ACTGTATAACGAGCTCAACCTGGGACGCAGGGAAGAGTATGACG TTTTGGACAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAA CCAAGACGAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCA GAAGGATAAGATGGCTGAAGCCTATTCTGAAATAGGCATGAAAG GAGAGCGGAGAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGA CTCAGCACTGCTACGAAGGATACTTATGACGCTCTCCACATGCAA GCCCTGCCACCTAGG (SEQ ID NO. 65) Clone 24C8 CD28 CD3 zeta CAR AA Heavy & Light Chains QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGFYWSWIRQHPGKGL EWIGYIHHSGSTHYNPSLKSRVTISIDTSKNLFSLRLSSVTAADTAVYY CASLVYCGGDCYSGFDYWGQGTLVTVSSGGGGSGGGGSGGGGSDI QLTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLIY DASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFTF GGGTKVEIKRAAAIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPG PSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNM TPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQL YNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKD KMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALP PR(SEQ ID NO. 66) Clone 24C8 CD8 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTGCAGTTGCAGGAAAGCGGGCCT GGCCTTGTGAAACCAAGCCAGACACTGAGCCTGACATGCACTGTG TCCGGCGGGTCCATATCTTCCGGGGGTTTTTATTGGTCCTGGATAC GCCAGCATCCCGGGAAAGGACTTGAATGGATTGGATATATCCACC ATTCCGGAAGCACCCACTACAATCCAAGCCTTAAATCCCGGGTGA CAATCTCCATCGACACCTCAAAGAATCTTTTTTCCCTGCGGTTGTC TTCAGTAACTGCCGCCGATACCGCTGTGTACTACTGTGCCAGCCTC GTCTATTGCGGCGGAGATTGTTATTCTGGGTTCGATTATTGGGGTC AAGGCACACTGGTAACTGTCAGCAGCGGAGGCGGCGGTTCCGGG GGCGGGGGCAGTGGAGGGGGCGGATCTGACATTCAGCTTACGCA GTCCCCATCTTCACTTAGCGCCAGCGTTGGCGATCGGGTCAGCTTC ACGTGTCAAGCAAGTCAGGATATCAACAACTTTCTTAACTGGTAC CAGCAGAAGCCAGGCAAGGCACCCAAGTTGCTGATTTACGATGCT TCTAACCTCGAGACGGGAGTGCCTAGCCGCTTCTCCGGGAGCGGC AGCGGCACAGACTTTACCTTTACGATTTCCAGTCTGCAGCCAGAG GATATAGCAACTTATTACTGTCAGCAGTATGGCAACCTCCCTTTTA CCTTCGGTGGTGGCACAAAGGTCGAGATTAAAAGAGCCGCAGCG TTGTCCAACTCCATAATGTATTTTTCTCATTTTGTGCCCGTCTTTCT GCCTGCCAAACCTACCACCACCCCCGCCCCACGACCACCTACTCC AGCCCCCACCATCGCCTCCCAGCCCCTCAGCCTGAGGCCAGAGGC TTGTCGCCCTGCTGCGGGGGGCGCTGTCCATACCAGAGGACTCGA CTTCGCCTGCGATATTTATATATGGGCCCCCCTCGCCGGCACCTGC GGAGTCTTGCTCCTGAGCCTTGTGATCACGCTTTATTGTAACCATC GGAATAGATCCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGA ATATGACTCCACGCCGCCCTGGCCCCACAAGGAAACACTACCAGC CTTACGCACCACCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGA AGTTTTCCAGATCTGCAGATGCACCAGCGTATCAGCAGGGCCAGA ACCAACTGTATAACGAGCTCAACCTGGGACGCAGGGAAGAGTAT GACGTTTTGGACAAGCGCAGAGGACGGGACCCTGAGATGGGTGG CAAACCAAGACGAAAAAACCCCCAGGAGGGTCTCTATAATGAGC TGCAGAAGGATAAGATGGCTGAAGCCTATTCTGAAATAGGCATG AAAGGAGAGCGGAGAAGGGGAAAAGGGCACGACGGTTTGTACCA GGGACTCAGCACTGCTACGAAGGATACTTATGACGCTCTCCACAT GCAAGCCCTGCCACCTAGGTAA (SEQ ID NO. 67) Clone 24C8 CD8 CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPQVQLQESGPGLVKPSQTLSLTCTVS GGSISSGGFYWSWIRQHPGKGLEWIGYIHHSGSTHYNPSLKSRVTISI DTSKNLFSLRLSSVTAADTAVYYCASLVYCGGDCYSGFDYWGQGTL VTVSSGGGGSGGGGSGGGGSDIQLTQSPSSLSASVGDRVSFTCQASQ DINNFLNWYQQKPGKAPKLLIYDASNLETGVPRFSGSGSGTDFTFTIS SLQPEDIATYYCQYGNLPFTFGGGTKVEIKRAAALSNSIMYFSHFVPV FLPAKPTTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFA CDIYIWAPLAGTCGVLLLSLVITLYCNHRNRSKRSRLLHSDYMNMTP RRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKM AEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 68) Clone 24C8 CD8 CD3 zeta CAR DNA Heavy & Light Chains CAGGTGCAGTTGCAGGAAAGCGGGCCTGGCCTTGTGAAACCAAG CCAGACACTGAGCCTGACATGCACTGTGTCCGGCGGGTCCATATC TTCCGGGGGTTTTTATTGGTCCTGGATACGCCAGCATCCCGGGAA AGGACTTGAATGGATTGGATATATCCACCATTCCGGAAGCACCCA CTACAATCCAAGCCTTAAATCCCGGGTGACAATCTCCATCGACAC
CTCAAAGAATCTTTTTTCCCTGCGGTTGTCTTCAGTAACTGCCGCC GATACCGCTGTGTACTACTGTGCCAGCCTCGTCTATTGCGGCGGA GATTGTTATTCTGGGTTCGATTATTGGGGTCAAGGCACACTGGTA ACTGTCAGCAGCGGAGGCGGCGGTTCCGGGGGCGGGGGCAGTGG AGGGGGCGGATCTGACATTCAGCTTACGCAGTCCCCATCTTCACT TAGCGCCAGCGTTGGCGATCGGGTCAGCTTCACGTGTCAAGCAAG TCAGGATATCAACAACTTTCTTAACTGGTACCAGCAGAAGCCAGG CAAGGCACCCAAGTTGCTGATTTACGATGCTTCTAACCTCGAGAC GGGAGTGCCTAGCCGCTTCTCCGGGAGCGGCAGCGGCACAGACTT TACCTTTACGATTTCCAGTCTGCAGCCAGAGGATATAGCAACTTA TTACTGTCAGCAGTATGGCAACCTCCCTTTTACCTTCGGTGGTGGC ACAAAGGTCGAGATTAAAAGAGCCGCAGCGTTGTCCAACTCCATA ATGTATTTTTCTCATTTTGTGCCCGTCTTTCTGCCTGCCAAACCTAC CACCACCCCCGCCCCACGACCACCTACTCCAGCCCCCACCATCGC CTCCCAGCCCCTCAGCCTGAGGCCAGAGGCTTGTCGCCCTGCTGC GGGGGGCGCTGTCCATACCAGAGGACTCGACTTCGCCTGCGATAT TTATATATGGGCCCCCCTCGCCGGCACCTGCGGAGTCTTGCTCCTG AGCCTTGTGATCACGCTTTATTGTAACCATCGGAATAGATCCAAA AGAAGCCGCCTGCTCCATAGCGATTACATGAATATGACTCCACGC CGCCCTGGCCCCACAAGGAAACACTACCAGCCTTACGCACCACCT AGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCT GCAGATGCACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAA CGAGCTCAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACA AGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAACCAAGACGA AAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAA GATGGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGA GAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACT GCTACGAAGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCA CCTAGG (SEQ ID NO. 69) Clone 24C8 CD8 CD3 zeta CAR AA Heavy & Light Chains QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGFYWSWIRQHPGKGL EWIGYIHHSGSTHYNPSLKSRVTISIDTSKNLFSLRLSSVTAADTAVYY CASLVYCGGDCYSGFDYWGQGTLVTVSSGGGGSGGGGSGGGGSDI QLTQSPSSLSASVGDRVSFTCQASQDINNFLNWYQQKPGKAPKLLIY DASNLETGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQYGNLPFTF GGGTKVEIKRAAALSNSIMYFSHFVPVFLPAKPTTTPAPRPPTPAPTIA SQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSL VITLYCNHRNRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDF AAYRSRVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRG RDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKG HDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 70) Clone 2005.1 HC DNA CAGGTCCAACTGGTGCAGTCCGGAGCCGAAGTCAAGAAACCAGG TGCCTCCGTTAAAGTGAGTTGCAAAGTCTCTGGATACACTCTGAC CGAGCTCTCTATGCACTGGGTCCGGCAGGCCCCCGGCAAGGGATT GGAATGGATGGGCGGGTTCGATCCTGAGGACGGAGAGACTATCT ACGCTCAAAAATTCCAGGGACGAGTGACTGTGACCGAAGACACT AGTACCGACACTGCCTACATGGAACTTTCCTCTCTGCGATCAGAA GATACCGCAGTGTACTACTGTGCTACTGAATCTAGGGGCATTGGA TGGCCCTACTTCGATTACTGGGGTCAGGGAACTCTGGTGACTGTC TCCAGC (SEQ ID NO. 71) Clone 205.1 AA HC (CDRs in Underline) QVQLVQSGAEVKKPGASVKVSCKVSGYTLTELSMHWVRQAPGKGL EWMGGFDPEDGETIYAQKFQGRVTVTEDTSTDTAYMELSSLRSEDT AVYYCATESRGIGWPYFDYWGQGTLVTVSS (SEQ ID NO. 72) Clone 205.1 HC AA CDR1: GYTLTEL (SEQ ID NO. 73) Clone 205.1 HC AA CDR2: DPEDGE (SEQ ID NO. 74) Clone 205.1 HC AA CDR3: ESRGIGWPYFDY (SEQ ID NO. 75) Clone 2005.1 LC DNA GATATTCAGATGACTCAATCTCCTTCTTCTCTGTCCGCTTCCGTGG GCGATAGAGTGACCATTACTTGTAGGGCGTCCCAGTCAATCTCCA GTTATTTGAATTGGTATCAGCAGAAGCCCGGGAAAGCACCTAAGC TGTTGATCAGCGGGGCTTCTAGCCTGAAGAGTGGGGTACCTTCAC GGTTCAGCGGAAGCGGAAGCGGAACCGATTTCACCCTGACTATCA GCAGCCTGCCACCTGAGGACTTTGCAACTTACTACTGCCAACAGT CATACAGCACTCCGATCACTTTCGGCCAGGGCACCCGGCTCGAAA TCAAGCGC (SEQ ID NO. 76) Clone 2005.1 AA LC (CDRs in Underline) DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLI SGASSLKSGVPSRFSGSGSGTDFTLTISSLPPEDFATYYCQQSYSTPITF GQGTRLEIKR (SEQ ID NO. 77) Clone 2005.1 AA LC CDR1: RASQSISSYLN (SEQ ID NO. 78) Clone 205.1 AA LC CDR2: GASSLKS (SEQ ID NO. 79) Clone 205.1 AA LC CDR3: QQSYSTPIT (SEQ ID NO. 80) Clone 2005.1 CD28T CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTCCAACTGGTGCAGTCCGGAGCCG AAGTCAAGAAACCAGGTGCCTCCGTTAAAGTGAGTTGCAAAGTCT CTGGATACACTCTGACCGAGCTCTCTATGCACTGGGTCCGGCAGG CCCCCGGCAAGGGATTGGAATGGATGGGCGGGTTCGATCCTGAG GACGGAGAGACTATCTACGCTCAAAAATTCCAGGGACGAGTGAC TGTGACCGAAGACACTAGTACCGACACTGCCTACATGGAACTTTC CTCTCTGCGATCAGAAGATACCGCAGTGTACTACTGTGCTACTGA ATCTAGGGGCATTGGATGGCCCTACTTCGATTACTGGGGTCAGGG AACTCTGGTGACTGTCTCCAGCGGTGGAGGTGGCAGCGGTGGTGG CGGAAGCGGGGGGGGCGGCTCTGATATTCAGATGACTCAATCTCC TTCTTCTCTGTCCGCTTCCGTGGGCGATAGAGTGACCATTACTTGT AGGGCGTCCCAGTCAATCTCCAGTTATTTGAATTGGTATCAGCAG AAGCCCGGGAAAGCACCTAAGCTGTTGATCAGCGGGGCTTCTAGC CTGAAGAGTGGGGTACCTTCACGGTTCAGCGGAAGCGGAAGCGG AACCGATTTCACCCTGACTATCAGCAGCCTGCCACCTGAGGACTT TGCAACTTACTACTGCCAACAGTCATACAGCACTCCGATCACTTTC GGCCAGGGCACCCGGCTCGAAATCAAGCGCGCTGCTGCTTTGGAC AATGAGAAGTCAAACGGCACCATCATACATGTTAAAGGTAAACA TCTGTGTCCCTCCCCGCTGTTCCCCGGCCCTTCCAAACCGTTCTGG GTTCTGGTGGTGGTCGGAGGCGTACTCGCTTGCTATAGTCTGCTG GTAACTGTCGCCTTCATCATCTTTTGGGTGAGATCCAAAAGAAGC CGCCTGCTCCATAGCGATTACATGAATATGACTCCACGCCGCCCT GGCCCCACAAGGAAACACTACCAGCCTTACGCACCACCTAGAGAT TTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCTGCAGAT GCACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAACGAGCT CAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACAAGCGCA GAGGACGGGACCCTGAGATGGGTGGCAAACCAAGACGAAAAAAC CCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAAGATGGCT GAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGAGAAGGGG AAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACTGCTACGA AGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCACCTAGGT AA (SEQ ID NO. 81) Clone 2005.1 CD28T CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPQVQLVQSGAEVKKPGASVKVSCKV SGYTLTELSMHWVRQAPGKGLEWMGGFDPEDGETIYAQKFQGRVT VTEDTSTDTAYMELSSLRSEDTAVYYCATESRGIGWPYFDYWGQGT LVTVSSGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRAS QSISSYLNWYQQKPGKAPKLLISGASSLKSGVPSRFSGSGSGTDFTLTI SSLPPEDFATYYCQQSYSTPITFGQGTRLEIKRAAALDNEKSNGTIIHV KGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSK RSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSA DAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKN PQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATK DTYDALHMQALPPR (SEQ ID NO. 82) Clone 2005.1 CD28T CD3 zeta CAR DNA Heavy & Light Chains CAGGTCCAACTGGTGCAGTCCGGAGCCGAAGTCAAGAAACCAGG TGCCTCCGTTAAAGTGAGTTGCAAAGTCTCTGGATACACTCTGAC CGAGCTCTCTATGCACTGGGTCCGGCAGGCCCCCGGCAAGGGATT GGAATGGATGGGCGGGTTCGATCCTGAGGACGGAGAGACTATCT ACGCTCAAAAATTCCAGGGACGAGTGACTGTGACCGAAGACACT AGTACCGACACTGCCTACATGGAACTTTCCTCTCTGCGATCAGAA GATACCGCAGTGTACTACTGTGCTACTGAATCTAGGGGCATTGGA TGGCCCTACTTCGATTACTGGGGTCAGGGAACTCTGGTGACTGTC TCCAGCGGTGGAGGTGGCAGCGGTGGTGGCGGAAGCGGGGGGGG CGGCTCTGATATTCAGATGACTCAATCTCCTTCTTCTCTGTCCGCT TCCGTGGGCGATAGAGTGACCATTACTTGTAGGGCGTCCCAGTCA ATCTCCAGTTATTTGAATTGGTATCAGCAGAAGCCCGGGAAAGCA CCTAAGCTGTTGATCAGCGGGGCTTCTAGCCTGAAGAGTGGGGTA CCTTCACGGTTCAGCGGAAGCGGAAGCGGAACCGATTTCACCCTG ACTATCAGCAGCCTGCCACCTGAGGACTTTGCAACTTACTACTGC CAACAGTCATACAGCACTCCGATCACTTTCGGCCAGGGCACCCGG CTCGAAATCAAGCGCGCTGCTGCTTTGGACAATGAGAAGTCAAAC GGCACCATCATACATGTTAAAGGTAAACATCTGTGTCCCTCCCCG CTGTTCCCCGGCCCTTCCAAACCGTTCTGGGTTCTGGTGGTGGTCG GAGGCGTACTCGCTTGCTATAGTCTGCTGGTAACTGTCGCCTTCAT CATCTTTTGGGTGAGATCCAAAAGAAGCCGCCTGCTCCATAGCGA TTACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGGAAACA CTACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTATCGGAG CAGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATCAGCA GGGCCAGAACCAACTGTATAACGAGCTCAACCTGGGACGCAGGG AAGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACCCTGAG ATGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGTCTCTA TAATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCTGAAAT AGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACGACGGT TTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATGACGCT CTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 83) Clone 2005.1 CD28T CD3 zeta CAR AA Heavy & Light Chains QVQLVQSGAEVKKPGASVKVSCKVSGYTLTELSMHWVRQAPGKGL EWMGGFDPEDGETIYAQKFQGRVTVTEDTSTDTAYMELSSLRSEDT AVYYCATESRGIGWPYFDYWGQGTLVTVSSGGGGSGGGGSGGGGS DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLI SGASSLKSGVPSRFSGSGSGTDFTLTISSLPPEDFATYYCQQSYSTPITF GQGTRLEIKRAAALDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLV VVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRK HYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYNELNLGRRE EYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIG MKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 84) Clone 2005.1 CD28 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTGCAGCTTGTGCAGAGCGGGGCC GAGGTGAAGAAGCCCGGGGCCAGCGTCAAAGTGTCCTGTAAGGT CAGCGGTTACACCCTCACCGAGCTGAGCATGCACTGGGTACGGCA GGCTCCCGGCAAAGGTCTTGAGTGGATGGGTGGATTTGATCCAGA AGATGGAGAGACTATCTACGCCCAGAAGTTCCAGGGCCGGGTCA CCGTAACAGAAGACACCTCAACTGACACCGCTTACATGGAGCTGA GTTCACTGCGGTCCGAGGACACGGCCGTGTATTATTGTGCCACCG AGAGCCGCGGAATCGGATGGCCTTACTTCGACTACTGGGGACAGG GTACACTTGTTACAGTATCATCCGGGGGTGGCGGCTCTGGTGGGG GCGGCTCCGGAGGGGGTGGATCAGATATCCAAATGACTCAAAGT CCAAGTTCCCTGTCTGCCTCAGTCGGAGATAGAGTCACCATAACC TGCAGGGCAAGTCAGTCCATCTCCTCCTATCTGAACTGGTACCAA CAGAAACCTGGAAAGGCGCCTAAGCTCCTGATCTCCGGAGCCTCA TCTTTGAAATCCGGTGTCCCATCTCGCTTCAGTGGCTCTGGAAGCG GTACAGATTTTACTTTGACCATTAGCAGCCTCCCACCGGAAGACT TTGCTACATATTACTGCCAGCAGTCTTACTCAACCCCAATCACCTT CGGGCAAGGCACCAGACTCGAAATAAAAAGAGCAGCTGCTATCG AGGTTATGTACCCACCGCCGTACTTGGATAACGAAAAAAGCAATG GGACCATCATTCATGTGAAGGGTAAGCACCTTTGCCCTAGCCCAC TGTTTCCTGGCCCGAGTAAACCCTTTTGGGTACTTGTGGTCGTCGG CGGCGTGCTGGCCTGCTACTCACTCCTGGTTACCGTCGCATTCATC ATCTTTTGGGTGAGATCCAAAAGAAGCCGCCTGCTCCATAGCGAT TACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGGAAACAC TACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTATCGGAGC AGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATCAGCAG GGCCAGAACCAACTGTATAACGAGCTCAACCTGGGACGCAGGGA AGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACCCTGAGA TGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGTCTCTAT AATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCTGAAAT AGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACGACGGT TTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATGACGCT CTCCACATGCAAGCCCTGCCACCTAGGTAA (SEQ ID NO. 85) Clone 2005.1 CD28 CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPQVQLVQSGAEVKKPGASVKVSCKV SGYTLTELSMHWVRQAPGKGLEWMGGFDPEDGETIYAQKFQGRVT VTEDTSTDTAYMELSSLRSEDTAVYYCATESRGIGWPYFDYWGQGT LVTVSSGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRAS QSISSYLNWYQQKPGKAPKLLISGASSLKSGVPSRFSGSGSGTDFTLTI SSLPPEDFATYYCQQSYSTPITFGQGTRLEIKRAAAIEVMYPPPYLDNE KSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVA FIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS RVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEM GGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLY QGLSTATKDTYDALHMQALPPR (SEQ ID NO. 86) Clone 2005.1 CD28 CD3 zeta CAR DNA Heavy & Light Chains CAGGTGCAGCTTGTGCAGAGCGGGGCCGAGGTGAAGAAGCCCGG GGCCAGCGTCAAAGTGTCCTGTAAGGTCAGCGGTTACACCCTCAC CGAGCTGAGCATGCACTGGGTACGGCAGGCTCCCGGCAAAGGTCT TGAGTGGATGGGTGGATTTGATCCAGAAGATGGAGAGACTATCTA CGCCCAGAAGTTCCAGGGCCGGGTCACCGTAACAGAAGACACCT CAACTGACACCGCTTACATGGAGCTGAGTTCACTGCGGTCCGAGG ACACGGCCGTGTATTATTGTGCCACCGAGAGCCGCGGAATCGGAT GGCCTTACTTCGACTACTGGGGACAGGGTACACTTGTTACAGTAT CATCCGGGGGTGGCGGCTCTGGTGGGGGCGGCTCCGGAGGGGGT GGATCAGATATCCAAATGACTCAAAGTCCAAGTTCCCTGTCTGCC TCAGTCGGAGATAGAGTCACCATAACCTGCAGGGCAAGTCAGTCC ATCTCCTCCTATCTGAACTGGTACCAACAGAAACCTGGAAAGGCG CCTAAGCTCCTGATCTCCGGAGCCTCATCTTTGAAATCCGGTGTCC CATCTCGCTTCAGTGGCTCTGGAAGCGGTACAGATTTTACTTTGAC CATTAGCAGCCTCCCACCGGAAGACTTTGCTACATATTACTGCCA GCAGTCTTACTCAACCCCAATCACCTTCGGGCAAGGCACCAGACT CGAAATAAAAAGAGCAGCTGCTATCGAGGTTATGTACCCACCGCC GTACTTGGATAACGAAAAAAGCAATGGGACCATCATTCATGTGAA GGGTAAGCACCTTTGCCCTAGCCCACTGTTTCCTGGCCCGAGTAA ACCCTTTTGGGTACTTGTGGTCGTCGGCGGCGTGCTGGCCTGCTAC
TCACTCCTGGTTACCGTCGCATTCATCATCTTTTGGGTGAGATCCA AAAGAAGCCGCCTGCTCCATAGCGATTACATGAATATGACTCCAC GCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTACGCACCAC CTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGAT CTGCAGATGCACCAGCGTATCAGCAGGGCCAGAACCAACTGTATA ACGAGCTCAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGAC AAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAACCAAGACG AAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATA AGATGGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGG AGAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGACTCAGCAC TGCTACGAAGGATACTTATGACGCTCTCCACATGCAAGCCCTGCC ACCTAGG (SEQ ID NO. 87) Clone 2005.1 CD28 CD3 zeta CAR AA Heavy & Light Chains QVQLVQSGAEVKKPGASVKVSCKVSGYTLTELSMHWVRQAPGKGL EWMGGFDPEDGETIYAQKFQGRVTVTEDTSTDTAYMELSSLRSEDT AVYYCATESRGIGWPYFDYWGQGTLVTVSSGGGGSGGGGSGGGGS DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLI SGASSLKSGVPSRFSGSGSGTDFTLTISSLPPEDFATYYCQQSYSTPITF GQGTRLEIKRAAAIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPG PSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNM TPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQL YNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKD KMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHM QALPPR (SEQ ID NO. 88) Clone 205.1 CD8 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTGCAGTTGGTGCAAAGCGGCGCA GAAGTTAAGAAACCTGGGGCGTCAGTTAAGGTGTCTTGCAAAGTA TCTGGCTATACCCTCACTGAGCTGTCCATGCATTGGGTAAGGCAG GCTCCTGGAAAGGGGCTCGAATGGATGGGAGGATTTGACCCTGA AGACGGAGAGACCATCTACGCCCAGAAATTCCAGGGTAGAGTAA CAGTGACTGAGGACACTAGCACTGACACAGCGTACATGGAGCTG AGTTCTCTGAGAAGTGAGGACACAGCCGTTTACTACTGCGCTACC GAGTCCAGAGGTATTGGCTGGCCATACTTCGACTATTGGGGTCAG GGCACCCTGGTTACAGTGAGTTCAGGAGGCGGGGGCTCTGGGGG GGGCGGTTCCGGAGGGGGGGGCTCAGATATACAGATGACGCAGA GTCCATCAAGTCTCTCAGCCAGCGTGGGAGATCGCGTGACTATTA CTTGCCGCGCCAGCCAGAGTATTAGCTCCTATCTGAATTGGTACC AGCAAAAGCCCGGGAAGGCCCCTAAGCTTCTGATTTCTGGCGCCT CCTCTTTGAAGTCAGGTGTGCCAAGCAGATTTAGCGGGTCTGGAA GTGGCACTGACTTTACACTTACTATCTCCAGCCTGCCCCCAGAGG ATTTTGCCACATATTACTGTCAGCAAAGCTACTCTACTCCAATCAC TTTCGGCCAGGGCACAAGATTGGAGATTAAGAGGGCTGCCGCACT TTCAAATTCCATCATGTATTTCAGCCATTTTGTGCCTGTTTTTCTTC CGGCCAAACCTACAACCACTCCCGCCCCACGCCCACCTACTCCCG CCCCTACCATTGCCTCCCAGCCTCTGTCTCTTAGACCTGAGGCTTG TAGACCTGCTGCCGGCGGAGCCGTGCACACTCGCGGTCTGGACTT CGCCTGCGACATCTATATCTGGGCCCCTCTGGCCGGCACCTGCGG CGTTCTCCTTCTCTCACTCGTAATCACACTCTATTGCAATCACAGG AACAGATCCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAAT ATGACTCCACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCT TACGCACCACCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAG TTTTCCAGATCTGCAGATGCACCAGCGTATCAGCAGGGCCAGAAC CAACTGTATAACGAGCTCAACCTGGGACGCAGGGAAGAGTATGA CGTTTTGGACAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCA AACCAAGACGAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTG CAGAAGGATAAGATGGCTGAAGCCTATTCTGAAATAGGCATGAA AGGAGAGCGGAGAAGGGGAAAAGGGCACGACGGTTTGTACCAGG GACTCAGCACTGCTACGAAGGATACTTATGACGCTCTCCACATGC AAGCCCTGCCACCTAGGTAA (SEQ ID NO. 89) Clone 205.1 CD8 CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPQVQLVQSGAEVKKPGASVKVSCKV SGYTLTELSMHWVRQAPGKGLEWMGGFDPEDGETIYAQKFQGRVT VTEDTSTDTAYMELSSLRSEDTAVYYCATESRGIGWPYFDYWGQGT LVTVSSGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRAS QSISSYLNWYQQKPGKAPKLLISGASSLKSGVPSRFSGSGSGTDFTLTI SSLPPEDFATYYCQQSYSTPITFGQGTRLEIKRAAALSNSIMYFSHFVP VFLPAKPTTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDF ACDIYIWAPLAGTCGVLLLSLVITLYCNHRNRSKRSRLLHSDYMNMT PRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLY NELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDK MAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPP R (SEQ ID NO. 90) Clone 2005.1 CD8 CD3 zeta CAR DNA Heavy & Light Chains CAGGTGCAGTTGGTGCAAAGCGGCGCAGAAGTTAAGAAACCTGG GGCGTCAGTTAAGGTGTCTTGCAAAGTATCTGGCTATACCCTCAC TGAGCTGTCCATGCATTGGGTAAGGCAGGCTCCTGGAAAGGGGCT CGAATGGATGGGAGGATTTGACCCTGAAGACGGAGAGACCATCT ACGCCCAGAAATTCCAGGGTAGAGTAACAGTGACTGAGGACACT AGCACTGACACAGCGTACATGGAGCTGAGTTCTCTGAGAAGTGAG GACACAGCCGTTTACTACTGCGCTACCGAGTCCAGAGGTATTGGC TGGCCATACTTCGACTATTGGGGTCAGGGCACCCTGGTTACAGTG AGTTCAGGAGGCGGGGGCTCTGGGGGGGGCGGTTCCGGAGGGGG GGGCTCAGATATACAGATGACGCAGAGTCCATCAAGTCTCTCAGC CAGCGTGGGAGATCGCGTGACTATTACTTGCCGCGCCAGCCAGAG TATTAGCTCCTATCTGAATTGGTACCAGCAAAAGCCCGGGAAGGC CCCTAAGCTTCTGATTTCTGGCGCCTCCTCTTTGAAGTCAGGTGTG CCAAGCAGATTTAGCGGGTCTGGAAGTGGCACTGACTTTACACTT ACTATCTCCAGCCTGCCCCCAGAGGATTTTGCCACATATTACTGTC AGCAAAGCTACTCTACTCCAATCACTTTCGGCCAGGGCACAAGAT TGGAGATTAAGAGGGCTGCCGCACTTTCAAATTCCATCATGTATT TCAGCCATTTTGTGCCTGTTTTTCTTCCGGCCAAACCTACAACCAC TCCCGCCCCACGCCCACCTACTCCCGCCCCTACCATTGCCTCCCAG CCTCTGTCTCTTAGACCTGAGGCTTGTAGACCTGCTGCCGGCGGA GCCGTGCACACTCGCGGTCTGGACTTCGCCTGCGACATCTATATCT GGGCCCCTCTGGCCGGCACCTGCGGCGTTCTCCTTCTCTCACTCGT AATCACACTCTATTGCAATCACAGGAACAGATCCAAAAGAAGCC GCCTGCTCCATAGCGATTACATGAATATGACTCCACGCCGCCCTG GCCCCACAAGGAAACACTACCAGCCTTACGCACCACCTAGAGATT TCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCTGCAGATG CACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAACGAGCTC AACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACAAGCGCAG AGGACGGGACCCTGAGATGGGTGGCAAACCAAGACGAAAAAACC CCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAAGATGGCT GAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGAGAAGGGG AAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACTGCTACGA AGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 91) Clone 205.1 CD8 CD3 zeta CAR AA Heavy & Light Chains QVQLVQSGAEVKKPGASVKVSCKVSGYTLTELSMHWVRQAPGKGL EWMGGFDPEDGETIYAQKFQGRVTVTEDTSTDTAYMELSSLRSEDT AVYYCATESRGIGWPYFDYWGQGTLVTVSSGGGGSGGGGSGGGGS DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLI SGASSLKSGVPSRFSGSGSGTDFTLTISSLPPEDFATYYCQQSYSTPITF GQGTRLEIKRAAALSNSIMYFSHFVPVFLPAKPTTTPAPRPPTPAPTIA SQPLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSL VITLYCNHRNRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDF AAYRSRVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRG RDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKG HDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 92) Clone 2005.2 HC DNA CAGGTCCAGTTGGTCGAAAGTGGCGGTGGTGTAGTGCAGCCGGGC CGCAGTTTGAGGCTTTCCTGTGCGGCTTCAGGCTTTACTTTTTCCA GCTATGGAATGCACTGGGTGCGGCAGGCCCCCGGCAAAGGACTT GAGTGGGTGGCCGTCATTTCTTATGACGGATCAGATAAGTACTAC GTGGACAGCGTCAAGGGCAGATTCACCATCTCTAGGGACAACAGT AAAAATAGACTCTACCTCCAGATGAATAGCCTCAGAGCTGAAGAC ACGGCCGTCTACTATTGTGCTCGGGAGCGGTATAGTGGCAGAGAC TACTGGGGGCAGGGCACACTCGTTACAGTGAGTAGC (SEQ ID NO. 93) Clone 2005.2 AA HC (CDRs in Underline) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLE WVAVISYDGSDKYYVDSVKGRFTISRDNSKNRLYLQMNSLRAEDTA VYYCARERYSGRDYWGQGTLVTVSS (SEQ ID NO. 94) Clone 2005.2 HC AA CDR1: GFTFSSY (SEQ ID NO. 95) Clone 2005.2 HC AA CDR2: SYDGSD (SEQ ID NO. 96) Clone 2005.2 HC AA CDR3: ERYSGRDY (SEQ ID NO. 97) Clone 2005.2 LC DNA GAGATTGTTATGACCCAGAGTCCTGCGACCCTCTCAGTCAGCCCC GGGGAGCGCGCAACTTTGTCTTGCAGAGCTAGTCAGTCCGTGTCC TCTCTTCTGACATGGTACCAGCAAAAGCCCGGGCAGGCTCCGCGC CTTTTGATCTTTGGGGCTTCAACAAGAGCCACTGGGATTCCCGCA CGATTCTCTGGCTCCGGGAGCGGTACTGGTTTCACCCTGACGATT AGCAGTCTCCAGAGCGAGGACTTCGCCGTATACTACTGCCAGCAG TACGATACGTGGCCATTCACTTTTGGACCAGGGACTAAAGTGGAT TTTAAGCGC (SEQ ID NO. 98) Clone 2005.2 AA LC (CDRs in Underline) EIVMTQSPATLSVSPGERATLSCRASQSVSSLLTWYQQKPGQAPRLLI FGASTRATGIPARFSGSGSGTGFTLTISSLQSEDFAVYYCQQYDTWPF TFGPGTKVDFKR (SEQ ID NO. 99) Clone 2005.2 AA LC CDR1: RASQSVSSLLT (SEQ ID NO. 100) Clone 2005.2 AA LC CDR2: GASTRAT (SEQ ID NO. 101) Clone 2005.2 AA LC CDR3: QQYDTWPFT (SEQ ID NO. 102) Clone 2005.2 CD28T CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTCCAGTTGGTCGAAAGTGGCGGTG GTGTAGTGCAGCCGGGCCGCAGTTTGAGGCTTTCCTGTGCGGCTT CAGGCTTTACTTTTTCCAGCTATGGAATGCACTGGGTGCGGCAGG CCCCCGGCAAAGGACTTGAGTGGGTGGCCGTCATTTCTTATGACG GATCAGATAAGTACTACGTGGACAGCGTCAAGGGCAGATTCACC ATCTCTAGGGACAACAGTAAAAATAGACTCTACCTCCAGATGAAT AGCCTCAGAGCTGAAGACACGGCCGTCTACTATTGTGCTCGGGAG CGGTATAGTGGCAGAGACTACTGGGGGCAGGGCACACTCGTTAC AGTGAGTAGCGGCGGAGGAGGGAGTGGGGGCGGTGGCTCCGGTG GAGGAGGTTCTGAGATTGTTATGACCCAGAGTCCTGCGACCCTCT CAGTCAGCCCCGGGGAGCGCGCAACTTTGTCTTGCAGAGCTAGTC AGTCCGTGTCCTCTCTTCTGACATGGTACCAGCAAAAGCCCGGGC AGGCTCCGCGCCTTTTGATCTTTGGGGCTTCAACAAGAGCCACTG GGATTCCCGCACGATTCTCTGGCTCCGGGAGCGGTACTGGTTTCA CCCTGACGATTAGCAGTCTCCAGAGCGAGGACTTCGCCGTATACT ACTGCCAGCAGTACGATACGTGGCCATTCACTTTTGGACCAGGGA CTAAAGTGGATTTTAAGCGCGCCGCCGCTCTCGATAACGAAAAGT CAAATGGCACCATAATCCACGTCAAAGGCAAGCACCTGTGCCCTT CCCCGCTCTTCCCCGGACCCAGTAAACCATTTTGGGTGCTGGTTGT TGTGGGGGGCGTGCTGGCCTGCTATAGCCTTTTGGTCACTGTAGC CTTCATTATTTTTTGGGTCAGATCCAAAAGAAGCCGCCTGCTCCAT AGCGATTACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGG AAACACTACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTAT CGGAGCAGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTAT CAGCAGGGCCAGAACCAACTGTATAACGAGCTCAACCTGGGACG CAGGGAAGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACC CTGAGATGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGT CTCTATAATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCT GAAATAGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACG ACGGTTTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATG ACGCTCTCCACATGCAAGCCCTGCCACCTAGGTAA (SEQ ID NO. 103) Clone 2005.2 CD28T CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPQVQLVESGGGVVQPGRSLRLSCAA SGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTI SRDNSKNRLYLQMNSLRAEDTAVYYCARERYSGRDYWGQGTLVTV SSGGGGSGGGGSGGGGSEIVMTQSPATLSVSPGERATLSCRASQSVSS LLTWYQQKPGQAPRLLIFGASTRATGIPARFSGSGSGTGFTLTISSLQS EDFAVYYCQQYDTWPFTFGPGTKVDFKRAAALDNEKSNGTIIHVKG KHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSR LLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAP AYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQE GLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATK DTYDALHMQALPPR (SEQ ID NO. 104) Clone 2005.2 CD28T CD3 zeta CAR DNA Heavy & Light Chains CAGGTCCAGTTGGTCGAAAGTGGCGGTGGTGTAGTGCAGCCGGGC CGCAGTTTGAGGCTTTCCTGTGCGGCTTCAGGCTTTACTTTTTCCA GCTATGGAATGCACTGGGTGCGGCAGGCCCCCGGCAAAGGACTT GAGTGGGTGGCCGTCATTTCTTATGACGGATCAGATAAGTACTAC GTGGACAGCGTCAAGGGCAGATTCACCATCTCTAGGGACAACAGT AAAAATAGACTCTACCTCCAGATGAATAGCCTCAGAGCTGAAGAC ACGGCCGTCTACTATTGTGCTCGGGAGCGGTATAGTGGCAGAGAC TACTGGGGGCAGGGCACACTCGTTACAGTGAGTAGCGGCGGAGG AGGGAGTGGGGGCGGTGGCTCCGGTGGAGGAGGTTCTGAGATTG TTATGACCCAGAGTCCTGCGACCCTCTCAGTCAGCCCCGGGGAGC GCGCAACTTTGTCTTGCAGAGCTAGTCAGTCCGTGTCCTCTCTTCT GACATGGTACCAGCAAAAGCCCGGGCAGGCTCCGCGCCTTTTGAT CTTTGGGGCTTCAACAAGAGCCACTGGGATTCCCGCACGATTCTC TGGCTCCGGGAGCGGTACTGGTTTCACCCTGACGATTAGCAGTCT CCAGAGCGAGGACTTCGCCGTATACTACTGCCAGCAGTACGATAC GTGGCCATTCACTTTTGGACCAGGGACTAAAGTGGATTTTAAGCG CGCCGCCGCTCTCGATAACGAAAAGTCAAATGGCACCATAATCCA CGTCAAAGGCAAGCACCTGTGCCCTTCCCCGCTCTTCCCCGGACC CAGTAAACCATTTTGGGTGCTGGTTGTTGTGGGGGGCGTGCTGGC CTGCTATAGCCTTTTGGTCACTGTAGCCTTCATTATTTTTTGGGTC AGATCCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATATG ACTCCACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTAC GCACCACCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTTT TCCAGATCTGCAGATGCACCAGCGTATCAGCAGGGCCAGAACCA ACTGTATAACGAGCTCAACCTGGGACGCAGGGAAGAGTATGACG TTTTGGACAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAA CCAAGACGAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCA GAAGGATAAGATGGCTGAAGCCTATTCTGAAATAGGCATGAAAG GAGAGCGGAGAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGA CTCAGCACTGCTACGAAGGATACTTATGACGCTCTCCACATGCAA GCCCTGCCACCTAGG (SEQ ID NO. 105)
Clone 2005.2 CD28T CD3 zeta CAR AA Heavy & Light Chains QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLE WVAVISYDGSDKYYVDSVKGRFTISRDNSKNRLYLQMNSLRAEDTA VYYCARERYSGRDYWGQGTLVTVSSGGGGSGGGGSGGGGSEIVMT QSPATLSVSPGERATLSCRASQSVSSLLTWYQQKPGQAPRLLIFGAST RATGIPARFSGSGSGTGFTLTISSLQSEDFAVYYCQQYDTWPFTFGPG TKVDFKRAAALDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVV GGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHY QPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYNELNLGRREEY DVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMK GERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 106) Clone 2005.2 CD28 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTGCAGCTCGTGGAGTCTGGCGGCG GCGTGGTCCAGCCCGGCCGGTCCCTGCGCCTGTCCTGCGCCGCCA GCGGGTTTACTTTTTCCTCCTACGGCATGCACTGGGTGCGCCAGGC TCCCGGCAAGGGCCTCGAGTGGGTCGCCGTGATCTCATACGATGG GTCAGACAAATACTATGTCGATTCTGTTAAAGGGCGGTTTACCAT TTCAAGAGATAACTCTAAGAATAGGCTGTATTTGCAGATGAACAG CCTGAGGGCTGAAGATACCGCAGTGTACTATTGCGCTAGGGAGCG GTATAGTGGCCGCGATTACTGGGGACAGGGTACACTGGTGACCGT GAGCTCTGGGGGTGGCGGAAGCGGGGGTGGCGGAAGCGGCGGAG GGGGTAGTGAAATTGTGATGACCCAGTCTCCGGCTACACTTTCAG TCTCCCCTGGGGAGAGAGCTACACTGTCATGCAGAGCGTCCCAGT CCGTCTCTTCTCTCCTTACCTGGTATCAGCAGAAGCCCGGCCAGGC TCCTCGACTGCTGATCTTCGGTGCCTCCACAAGGGCGACCGGGAT TCCAGCCCGCTTCTCAGGTTCTGGGAGCGGAACTGGTTTCACTTTG ACAATCAGTTCACTGCAGTCAGAGGATTTCGCCGTGTACTACTGC CAGCAATACGACACATGGCCATTCACTTTCGGACCCGGTACCAAA GTCGATTTCAAGAGAGCCGCGGCCATCGAGGTTATGTACCCACCA CCATATCTGGACAATGAAAAAAGCAATGGAACCATTATCCATGTG AAGGGTAAACACCTCTGCCCTAGCCCACTTTTCCCTGGCCCATCA AAGCCCTTCTGGGTCTTGGTGGTCGTGGGGGGTGTGCTGGCCTGT TACAGCCTTCTGGTGACGGTTGCTTTCATTATCTTCTGGGTTAGAT CCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATATGACTC CACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTACGCAC CACCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCA GATCTGCAGATGCACCAGCGTATCAGCAGGGCCAGAACCAACTGT ATAACGAGCTCAACCTGGGACGCAGGGAAGAGTATGACGTTTTG GACAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAACCAAG ACGAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGG ATAAGATGGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAGAG CGGAGAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGACTCAG CACTGCTACGAAGGATACTTATGACGCTCTCCACATGCAAGCCCT GCCACCTAGGTAA (SEQ ID NO. 107) Clone 2005.2 CD28 CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPQVQLVESGGGVVQPGRSLRLSCAA SGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTI SRDNSKNRLYLQMNSLRAEDTAVYYCARERYSGRDYWGQGTLVTV SSGGGGSGGGGSGGGGSEIVMTQSPATLSVSPGERATLSCRASQSVSS LLTWYQQKPGQAPRLLIFGASTRATGIPARFSGSGSGTGFTLTISSLQS EDFAVYYCQQYDTWPFTFGPGTKVDFKRAAAIEVMYPPPYLDNEKS NGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFII FWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRV KFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGG KPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQG LSTATKDTYDALHMQALPPR (SEQ ID NO. 108) Clone 2005.2 CD28 CD3 zeta CAR DNA Heavy & Light Chains CAGGTGCAGCTCGTGGAGTCTGGCGGCGGCGTGGTCCAGCCCGGC CGGTCCCTGCGCCTGTCCTGCGCCGCCAGCGGGTTTACTTTTTCCT CCTACGGCATGCACTGGGTGCGCCAGGCTCCCGGCAAGGGCCTCG AGTGGGTCGCCGTGATCTCATACGATGGGTCAGACAAATACTATG TCGATTCTGTTAAAGGGCGGTTTACCATTTCAAGAGATAACTCTA AGAATAGGCTGTATTTGCAGATGAACAGCCTGAGGGCTGAAGAT ACCGCAGTGTACTATTGCGCTAGGGAGCGGTATAGTGGCCGCGAT TACTGGGGACAGGGTACACTGGTGACCGTGAGCTCTGGGGGTGGC GGAAGCGGGGGTGGCGGAAGCGGCGGAGGGGGTAGTGAAATTGT GATGACCCAGTCTCCGGCTACACTTTCAGTCTCCCCTGGGGAGAG AGCTACACTGTCATGCAGAGCGTCCCAGTCCGTCTCTTCTCTCCTT ACCTGGTATCAGCAGAAGCCCGGCCAGGCTCCTCGACTGCTGATC TTCGGTGCCTCCACAAGGGCGACCGGGATTCCAGCCCGCTTCTCA GGTTCTGGGAGCGGAACTGGTTTCACTTTGACAATCAGTTCACTG CAGTCAGAGGATTTCGCCGTGTACTACTGCCAGCAATACGACACA TGGCCATTCACTTTCGGACCCGGTACCAAAGTCGATTTCAAGAGA GCCGCGGCCATCGAGGTTATGTACCCACCACCATATCTGGACAAT GAAAAAAGCAATGGAACCATTATCCATGTGAAGGGTAAACACCT CTGCCCTAGCCCACTTTTCCCTGGCCCATCAAAGCCCTTCTGGGTC TTGGTGGTCGTGGGGGGTGTGCTGGCCTGTTACAGCCTTCTGGTG ACGGTTGCTTTCATTATCTTCTGGGTTAGATCCAAAAGAAGCCGC CTGCTCCATAGCGATTACATGAATATGACTCCACGCCGCCCTGGC CCCACAAGGAAACACTACCAGCCTTACGCACCACCTAGAGATTTC GCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCTGCAGATGCA CCAGCGTATCAGCAGGGCCAGAACCAACTGTATAACGAGCTCAA CCTGGGACGCAGGGAAGAGTATGACGTTTTGGACAAGCGCAGAG GACGGGACCCTGAGATGGGTGGCAAACCAAGACGAAAAAACCCC CAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAAGATGGCTGA AGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGAGAAGGGGAA AAGGGCACGACGGTTTGTACCAGGGACTCAGCACTGCTACGAAG GATACTTATGACGCTCTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 109) Clone 2005.2 CD28 CD3 zeta CAR AA Heavy & Light Chains QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLE WVAVISYDGSDKYYVDSVKGRFTISRDNSKNRLYLQMNSLRAEDTA VYYCARERYSGRDYWGQGTLVTVSSGGGGSGGGGSGGGGSEIVMT QSPATLSVSPGERATLSCRASQSVSSLLTWYQQKPGQAPRLLIFGAST RATGIPARFSGSGSGTGFTLTISSLQSEDFAVYYCQQYDTWPFTFGPG TKVDFKRAAAIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSK PFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPR RPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYNE LNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMA EAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 110) Clone 2005.2 CD8 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGCAGGTGCAGTTGGTTGAATCAGGAGGG GGTGTGGTGCAACCCGGTCGGTCACTGCGCCTCAGTTGTGCTGCT TCCGGGTTTACTTTCAGCTCATATGGGATGCACTGGGTACGGCAG GCTCCAGGTAAAGGCTTGGAATGGGTGGCGGTGATCAGCTATGAC GGCTCTGACAAATATTATGTGGACTCCGTGAAAGGCAGATTCACC ATCAGTCGAGACAACTCAAAGAATAGACTCTACTTGCAGATGAAT AGCCTCCGGGCCGAAGATACTGCAGTCTATTATTGCGCCCGGGAG CGCTACAGTGGAAGAGACTATTGGGGGCAAGGAACTCTTGTCACA GTCTCATCTGGCGGCGGCGGCAGCGGTGGGGGCGGATCTGGCGG GGGCGGCAGCGAAATCGTTATGACTCAGAGTCCTGCCACACTGAG CGTTAGCCCTGGTGAGAGAGCAACACTTAGCTGCAGAGCTAGTCA GAGTGTTTCCAGTCTTTTGACATGGTACCAACAGAAGCCCGGTCA AGCTCCACGACTGCTCATCTTCGGTGCATCCACCCGCGCAACCGG GATACCCGCCCGGTTTTCCGGTTCTGGAAGTGGCACAGGATTCAC GCTCACCATTTCTTCTCTGCAGTCTGAAGACTTTGCCGTGTATTAC TGCCAGCAGTACGATACCTGGCCCTTTACCTTTGGCCCAGGTACT AAAGTGGATTTTAAACGAGCTGCTGCACTTTCCAATAGTATTATG TACTTTTCACATTTTGTGCCCGTGTTCCTGCCTGCGAAGCCTACGA CAACCCCAGCCCCTAGGCCGCCCACACCGGCCCCAACTATTGCCT CCCAGCCATTGTCTCTGAGACCCGAAGCTTGCAGACCTGCTGCTG GAGGCGCCGTTCACACCCGAGGATTGGATTTCGCATGTGACATTT ACATCTGGGCCCCTTTGGCCGGAACCTGCGGTGTGCTGCTGCTGT CACTCGTGATTACACTTTACTGCAACCACCGAAACAGATCCAAAA GAAGCCGCCTGCTCCATAGCGATTACATGAATATGACTCCACGCC GCCCTGGCCCCACAAGGAAACACTACCAGCCTTACGCACCACCTA GAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCTG CAGATGCACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAAC GAGCTCAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACAA GCGCAGAGGACGGGACCCTGAGATGGGTGGCAAACCAAGACGAA AAAACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAAG ATGGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGAG AAGGGGAAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACTG CTACGAAGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCAC CTAGGTAA (SEQ ID NO. 111) Clone 2005.2 CD8 CD3 zeta CAR AA Heavy & Light Chains (Signal peptide in Bold) MALPVTALLLPLALLLHAARPQVQLVESGGGVVQPGRSLRLSCAA SGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTI SRDNSKNRLYLQMNSLRAEDTAVYYCARERYSGRDYWGGTLVTVS SGGGGSGGGGSGGGGSEIVMTQSPATLSVSPGERATLSCRASQSVSS LLTWYQQKPGQAPRLLIFGASTRATGIPARFSGSGSGTGFTLTISSLQS EDFAVYYCQQYDTWPFTFGPGTKVDFKRAAALSNSIMYFSHFVPVFL PAKPTTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFAC DIYIWAPLAGTCGVLLLSLVITLYCNHRNRSKRSRLLHSDYMNMTPR RPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYNE LNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMA EAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 112) Clone 2005.2 CD8 CD3 zeta CAR DNA Heavy & Light Chains CAGGTGCAGTTGGTTGAATCAGGAGGGGGTGTGGTGCAACCCGGT CGGTCACTGCGCCTCAGTTGTGCTGCTTCCGGGTTTACTTTCAGCT CATATGGGATGCACTGGGTACGGCAGGCTCCAGGTAAAGGCTTGG AATGGGTGGCGGTGATCAGCTATGACGGCTCTGACAAATATTATG TGGACTCCGTGAAAGGCAGATTCACCATCAGTCGAGACAACTCAA AGAATAGACTCTACTTGCAGATGAATAGCCTCCGGGCCGAAGATA CTGCAGTCTATTATTGCGCCCGGGAGCGCTACAGTGGAAGAGACT ATTGGGGGCAAGGAACTCTTGTCACAGTCTCATCTGGCGGCGGCG GCAGCGGTGGGGGCGGATCTGGCGGGGGCGGCAGCGAAATCGTT ATGACTCAGAGTCCTGCCACACTGAGCGTTAGCCCTGGTGAGAGA GCAACACTTAGCTGCAGAGCTAGTCAGAGTGTTTCCAGTCTTTTG ACATGGTACCAACAGAAGCCCGGTCAAGCTCCACGACTGCTCATC TTCGGTGCATCCACCCGCGCAACCGGGATACCCGCCCGGTTTTCC GGTTCTGGAAGTGGCACAGGATTCACGCTCACCATTTCTTCTCTGC AGTCTGAAGACTTTGCCGTGTATTACTGCCAGCAGTACGATACCT GGCCCTTTACCTTTGGCCCAGGTACTAAAGTGGATTTTAAACGAG CTGCTGCACTTTCCAATAGTATTATGTACTTTTCACATTTTGTGCC CGTGTTCCTGCCTGCGAAGCCTACGACAACCCCAGCCCCTAGGCC GCCCACACCGGCCCCAACTATTGCCTCCCAGCCATTGTCTCTGAG ACCCGAAGCTTGCAGACCTGCTGCTGGAGGCGCCGTTCACACCCG AGGATTGGATTTCGCATGTGACATTTACATCTGGGCCCCTTTGGCC GGAACCTGCGGTGTGCTGCTGCTGTCACTCGTGATTACACTTTACT GCAACCACCGAAACAGATCCAAAAGAAGCCGCCTGCTCCATAGC GATTACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGGAAA CACTACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTATCGG AGCAGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATCAG CAGGGCCAGAACCAACTGTATAACGAGCTCAACCTGGGACGCAG GGAAGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACCCTG AGATGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGTCTC TATAATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCTGAA ATAGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACGACG GTTTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATGACG CTCTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 113) Clone 2005.2 CD8 CD3 zeta CAR AA Heavy & Light Chains QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLE WVAVISYDGSDKYYVDSVKGRFTISRDNSKNRLYLQMNSLRAEDTA VYYCARERYSGRDYWGQGTLVTVSSGGGGSGGGGSGGGGSEIVMT QSPATLSVSPGERATLSCRASQSVSSLLTWYQQKPGQAPRLLIFGAST RATGIPARFSGSGSGTGFTLTISSLQSEDFAVYYCQQYDTWPFTFGPG TKVDFKRAAALSNSIMYFSHFVPVFLPAKPTTTPAPRPPTPAPTIASQP LSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVITL YCNHRNRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYR SRVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPE MGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGL YQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 114) Clone 2005.2 CD28T CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGGAGATTGTGATGACCCAGTCCCCTGCTA CCCTGTCCGTCAGTCCGGGCGAGAGAGCCACCTTGTCATGCCGGG CCAGCCAGTCCGTCAGCAGTCTCCTGACTTGGTATCAGCAAAAAC CAGGGCAGGCACCGCGGCTTTTGATTTTTGGTGCAAGCACACGCG CCACTGGCATTCCAGCTAGGTTTTCTGGAAGTGGATCTGGGACAG GCTTCACTCTGACAATCAGTAGCCTGCAGAGTGAGGACTTTGCTG TTTACTACTGTCAACAGTACGACACCTGGCCATTCACATTCGGGC CCGGCACCAAGGTCGACTTCAAGAGGGGCGGTGGAGGTTCAGGT GGTGGCGGGTCAGGCGGCGGTGGGTCTCAGGTTCAACTGGTGGA ATCAGGTGGCGGCGTTGTCCAACCGGGGCGATCACTTCGACTTTC CTGTGCTGCCTCAGGCTTTACTTTTTCATCCTATGGGATGCACTGG GTTCGGCAGGCTCCCGGAAAAGGACTCGAGTGGGTTGCAGTGATC TCTTACGATGGCTCAGACAAGTATTATGTGGACTCAGTCAAGGGG AGATTCACAATAAGCCGAGACAACTCCAAAAACCGGCTTTATCTC CAGATGAACAGCCTTAGAGCGGAAGATACCGCGGTATACTACTGT GCCCGCGAGAGGTATTCCGGCAGAGACTACTGGGGACAGGGCAC ACTGGTCACCGTGAGTTCTGCCGCAGCGCTCGATAACGAAAAGAG CAACGGAACCATTATCCACGTTAAGGGCAAGCACCTGTGCCCCAG TCCCCTCTTCCCAGGACCATCTAAACCCTTCTGGGTTCTGGTAGTA GTTGGAGGGGTCCTTGCATGTTACTCCCTTTTGGTCACCGTCGCCT TCATTATTTTCTGGGTGAGATCCAAAAGAAGCCGCCTGCTCCATA GCGATTACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGGA AACACTACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTATC GGAGCAGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATC AGCAGGGCCAGAACCAACTGTATAACGAGCTCAACCTGGGACGC AGGGAAGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACCC TGAGATGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGTC TCTATAATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCTG AAATAGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACGA CGGTTTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATGA CGCTCTCCACATGCAAGCCCTGCCACCTAGGTAA (SEQ ID NO. 115) Clone 2005.2 CD28T CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPEIVMTQSPATLSVSPGERATLSCRA SQSVSSLLTWYQQKPGQAPRLLIFGASTRATGIPARFSGSGSGTGFTL TISSLQSEDFAVYYCQQYDTWPFTFGPGTKVDFKRGGGGSGGGGSG GGGSQVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPG KGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNRLYLQMNSLRA
EDTAVYYCARERYSGRDYWGQGTLVTVSSAAALDNEKSNGTIIHVK GKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRS RLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADA PAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQ EGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQGLSTATKDT YDALHMQALPPR (SEQ ID NO. 116) Clone 2005.2 CD28T CD3 zeta CAR DNA Heavy & Light Chains GAGATTGTGATGACCCAGTCCCCTGCTACCCTGTCCGTCAGTCCG GGCGAGAGAGCCACCTTGTCATGCCGGGCCAGCCAGTCCGTCAGC AGTCTCCTGACTTGGTATCAGCAAAAACCAGGGCAGGCACCGCGG CTTTTGATTTTTGGTGCAAGCACACGCGCCACTGGCATTCCAGCTA GGTTTTCTGGAAGTGGATCTGGGACAGGCTTCACTCTGACAATCA GTAGCCTGCAGAGTGAGGACTTTGCTGTTTACTACTGTCAACAGT ACGACACCTGGCCATTCACATTCGGGCCCGGCACCAAGGTCGACT TCAAGAGGGGCGGTGGAGGTTCAGGTGGTGGCGGGTCAGGCGGC GGTGGGTCTCAGGTTCAACTGGTGGAATCAGGTGGCGGCGTTGTC CAACCGGGGCGATCACTTCGACTTTCCTGTGCTGCCTCAGGCTTTA CTTTTTCATCCTATGGGATGCACTGGGTTCGGCAGGCTCCCGGAA AAGGACTCGAGTGGGTTGCAGTGATCTCTTACGATGGCTCAGACA AGTATTATGTGGACTCAGTCAAGGGGAGATTCACAATAAGCCGAG ACAACTCCAAAAACCGGCTTTATCTCCAGATGAACAGCCTTAGAG CGGAAGATACCGCGGTATACTACTGTGCCCGCGAGAGGTATTCCG GCAGAGACTACTGGGGACAGGGCACACTGGTCACCGTGAGTTCTG CCGCAGCGCTCGATAACGAAAAGAGCAACGGAACCATTATCCAC GTTAAGGGCAAGCACCTGTGCCCCAGTCCCCTCTTCCCAGGACCA TCTAAACCCTTCTGGGTTCTGGTAGTAGTTGGAGGGGTCCTTGCAT GTTACTCCCTTTTGGTCACCGTCGCCTTCATTATTTTCTGGGTGAG ATCCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATATGAC TCCACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTACGC ACCACCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTC CAGATCTGCAGATGCACCAGCGTATCAGCAGGGCCAGAACCAAC TGTATAACGAGCTCAACCTGGGACGCAGGGAAGAGTATGACGTTT TGGACAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAACCA AGACGAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAA GGATAAGATGGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAG AGCGGAGAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGACTC AGCACTGCTACGAAGGATACTTATGACGCTCTCCACATGCAAGCC CTGCCACCTAGG (SEQ ID NO. 117) Clone 2005.2 CD28T CD3 zeta CAR AA Heavy & Light Chains EIVMTQSPATLSVSPGERATLSCRASQSVSSLLTWYQQKPGQAPRLLI FGASTRATGIPARFSGSGSGTGFTLTISSLQSEDFAVYYCQQYDTWPF TFGPGTKVDFKRGGGGSGGGGSGGGGSQVQLVESGGGVVQPGRSLR LSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSV KGRFTISRDNSKNRLYLQMNSLRAEDTAVYYCARERYSGRDYWGQ GTLVTVSSAAALDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVV VGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKH YQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYNELNLGRREE YDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKMAEAYSEIGM KGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 118) Clone 2005.2 CD28 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGGAGATCGTCATGACACAGAGTCCAGCTA CCCTGAGCGTGTCCCCTGGAGAGAGAGCCACCCTGTCCTGTAGGG CTAGTCAGAGTGTGTCCAGCCTCCTCACCTGGTATCAACAGAAGC CTGGTCAAGCTCCCCGGCTGCTTATCTTCGGGGCCAGCACGCGAG CCACAGGCATCCCGGCCAGATTCTCTGGCTCTGGCAGTGGCACCG GGTTCACTCTCACGATCTCATCCCTGCAGTCAGAGGATTTCGCTGT GTATTACTGTCAGCAGTACGATACATGGCCCTTCACCTTCGGCCC GGGCACAAAAGTAGATTTCAAGCGCGGCGGCGGGGGTAGTGGGG GCGGGGGATCAGGAGGAGGGGGCTCCCAAGTACAGCTGGTTGAG AGCGGCGGCGGGGTGGTTCAGCCCGGGCGCAGCCTCAGGCTGAG TTGCGCAGCATCAGGATTCACATTCAGTTCTTATGGAATGCATTG GGTCAGACAGGCTCCCGGGAAGGGCCTTGAATGGGTGGCAGTCA TTAGCTACGACGGAAGCGATAAGTACTATGTGGACTCAGTTAAAG GGAGATTTACTATCAGCCGCGACAATTCCAAAAACAGATTGTATT TGCAGATGAACTCCCTCAGGGCGGAGGACACTGCTGTATATTACT GCGCACGAGAGAGATACTCCGGCCGAGACTATTGGGGCCAAGGA ACATTGGTAACTGTGAGCTCCGCCGCAGCTATTGAGGTCATGTAC CCCCCACCTTATCTCGATAATGAGAAGAGTAATGGGACTATAATT CACGTAAAGGGCAAACACCTGTGCCCTTCCCCGCTGTTTCCAGGT CCAAGTAAGCCGTTCTGGGTCCTGGTTGTGGTGGGAGGGGTGCTG GCCTGCTATTCTCTGTTGGTTACCGTGGCCTTTATCATTTTCTGGGT GAGATCCAAAAGAAGCCGCCTGCTCCATAGCGATTACATGAATAT GACTCCACGCCGCCCTGGCCCCACAAGGAAACACTACCAGCCTTA CGCACCACCTAGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTT TTCCAGATCTGCAGATGCACCAGCGTATCAGCAGGGCCAGAACCA ACTGTATAACGAGCTCAACCTGGGACGCAGGGAAGAGTATGACG TTTTGGACAAGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAA CCAAGACGAAAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCA GAAGGATAAGATGGCTGAAGCCTATTCTGAAATAGGCATGAAAG GAGAGCGGAGAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGA CTCAGCACTGCTACGAAGGATACTTATGACGCTCTCCACATGCAA GCCCTGCCACCTAGGTAA (SEQ ID NO. 119) Clone 2005.2 CD28 CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPEIVMTQSPATLSVSPGERATLSCRA SQSVSSLLTWYQQKPGQAPRLLIFGASTRATGIPARFSGSGSGTGFTL TISSLQSEDFAVYYCQQYDTWPFTFGPGTKVDFKRGGGGSGGGGSG GGGSQVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPG KGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNRLYLQMNSLRA EDTAVYYCARERYSGRDYWGQGTLVTVSSAAAIEVMYPPPYLDNEK SNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFI IFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRSR VKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPEMG GKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGLYQ GLSTATKDTYDALHMQALPPR (SEQ ID NO. 120) Clone 2005.2 CD28 CD3 zeta CAR DNA Heavy & Light Chains GAGATCGTCATGACACAGAGTCCAGCTACCCTGAGCGTGTCCCCT GGAGAGAGAGCCACCCTGTCCTGTAGGGCTAGTCAGAGTGTGTCC AGCCTCCTCACCTGGTATCAACAGAAGCCTGGTCAAGCTCCCCGG CTGCTTATCTTCGGGGCCAGCACGCGAGCCACAGGCATCCCGGCC AGATTCTCTGGCTCTGGCAGTGGCACCGGGTTCACTCTCACGATCT CATCCCTGCAGTCAGAGGATTTCGCTGTGTATTACTGTCAGCAGT ACGATACATGGCCCTTCACCTTCGGCCCGGGCACAAAAGTAGATT TCAAGCGCGGCGGCGGGGGTAGTGGGGGCGGGGGATCAGGAGGA GGGGGCTCCCAAGTACAGCTGGTTGAGAGCGGCGGCGGGGTGGT TCAGCCCGGGCGCAGCCTCAGGCTGAGTTGCGCAGCATCAGGATT CACATTCAGTTCTTATGGAATGCATTGGGTCAGACAGGCTCCCGG GAAGGGCCTTGAATGGGTGGCAGTCATTAGCTACGACGGAAGCG ATAAGTACTATGTGGACTCAGTTAAAGGGAGATTTACTATCAGCC GCGACAATTCCAAAAACAGATTGTATTTGCAGATGAACTCCCTCA GGGCGGAGGACACTGCTGTATATTACTGCGCACGAGAGAGATACT CCGGCCGAGACTATTGGGGCCAAGGAACATTGGTAACTGTGAGCT CCGCCGCAGCTATTGAGGTCATGTACCCCCCACCTTATCTCGATA ATGAGAAGAGTAATGGGACTATAATTCACGTAAAGGGCAAACAC CTGTGCCCTTCCCCGCTGTTTCCAGGTCCAAGTAAGCCGTTCTGGG TCCTGGTTGTGGTGGGAGGGGTGCTGGCCTGCTATTCTCTGTTGGT TACCGTGGCCTTTATCATTTTCTGGGTGAGATCCAAAAGAAGCCG CCTGCTCCATAGCGATTACATGAATATGACTCCACGCCGCCCTGG CCCCACAAGGAAACACTACCAGCCTTACGCACCACCTAGAGATTT CGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCTGCAGATGC ACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAACGAGCTCA ACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACAAGCGCAGA GGACGGGACCCTGAGATGGGTGGCAAACCAAGACGAAAAAACCC CCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAAGATGGCTG AAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGAGAAGGGGA AAAGGGCACGACGGTTTGTACCAGGGACTCAGCACTGCTACGAA GGATACTTATGACGCTCTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 121) Clone 2005.2 CD28 CD3 zeta CAR AA Heavy & Light Chains EIVMTQSPATLSVSPGERATLSCRASQSVSSLLTWYQQKPGQAPRLLI FGASTRATGIPARFSGSGSGTGFTLTISSLQSEDFAVYYCQQYDTWPF TFGPGTKVDFKRGGGGSGGGGSGGGGSQVQLVESGGGVVQPGRSLR LSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSV KGRFTISRDNSKNRLYLQMNSLRAEDTAVYYCARERYSGRDYWGQ GTLVTVSSAAAIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPS KPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTP RRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKM AEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 122) Clone 2005.2 CD8 CD3 zeta CAR DNA Heavy & Light Chains ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC TGCACGCCGCACGCCCGGAAATAGTGATGACTCAGTCCCCGGCCA CCCTCAGCGTGTCCCCCGGGGAGCGAGCGACCCTGTCATGCAGGG CTTCCCAGAGTGTCAGCTCCCTGCTCACTTGGTATCAGCAAAAGC CGGGGCAGGCTCCCCGCCTCCTCATCTTCGGGGCATCAACTAGGG CCACCGGCATTCCTGCAAGATTTTCCGGGTCTGGCAGCGGCACCG GCTTCACCCTTACCATTAGCTCTCTGCAGTCTGAGGACTTCGCCGT TTACTATTGTCAGCAGTATGATACTTGGCCCTTTACCTTCGGTCCC GGAACTAAGGTGGACTTCAAGCGCGGGGGGGGTGGATCTGGAGG TGGTGGCTCCGGGGGCGGTGGAAGCCAGGTCCAGTTGGTTGAGA GCGGCGGCGGAGTGGTGCAGCCCGGGAGGTCCTTGCGGCTGAGC TGTGCAGCCTCCGGTTTTACTTTTTCTAGCTATGGAATGCATTGGG TAAGACAGGCTCCCGGAAAAGGCCTCGAGTGGGTGGCGGTCATT AGCTATGATGGATCTGATAAATACTATGTGGACTCAGTTAAGGGG CGCTTCACAATCTCAAGAGACAATAGCAAAAATAGACTGTACCTG CAGATGAATAGTCTGCGCGCCGAGGACACTGCCGTGTACTACTGC GCCCGCGAGAGATACAGCGGACGGGATTACTGGGGCCAGGGTAC CCTCGTAACGGTGTCCTCCGCTGCCGCCCTTAGCAACAGCATTAT GTACTTTTCTCATTTCGTGCCAGTCTTTCTCCCAGCAAAGCCCACC ACTACCCCGGCCCCCAGGCCGCCTACTCCTGCCCCCACTATCGCG TCTCAGCCTCTCTCCTTGCGGCCCGAGGCCTGCCGGCCAGCCGCA GGGGGCGCCGTACATACTCGGGGTTTGGATTTCGCTTGCGACATA TATATTTGGGCCCCCCTCGCCGGCACATGTGGAGTGCTGCTCCTG AGTCTCGTTATAACCCTCTATTGCAACCATAGAAACAGATCCAAA AGAAGCCGCCTGCTCCATAGCGATTACATGAATATGACTCCACGC CGCCCTGGCCCCACAAGGAAACACTACCAGCCTTACGCACCACCT AGAGATTTCGCTGCCTATCGGAGCAGGGTGAAGTTTTCCAGATCT GCAGATGCACCAGCGTATCAGCAGGGCCAGAACCAACTGTATAA CGAGCTCAACCTGGGACGCAGGGAAGAGTATGACGTTTTGGACA AGCGCAGAGGACGGGACCCTGAGATGGGTGGCAAACCAAGACGA AAAAACCCCCAGGAGGGTCTCTATAATGAGCTGCAGAAGGATAA GATGGCTGAAGCCTATTCTGAAATAGGCATGAAAGGAGAGCGGA GAAGGGGAAAAGGGCACGACGGTTTGTACCAGGGACTCAGCACT GCTACGAAGGATACTTATGACGCTCTCCACATGCAAGCCCTGCCA CCTAGGTAA (SEQ ID NO. 123) Clone 2005.2 CD8 CD3 zeta CAR AA Heavy & Light Chains (Signal Peptide in Bold) MALPVTALLLPLALLLHAARPEIVMTQSPATLSVSPGERATLSCRA SQSVSSLLTWYQQKPGQAPRLLIFGASTRATGIPARFSGSGSGTGFTL TISSLQSEDFAVYYCQQYDTWPFTFGPGTKVDFKRGGGGSGGGGSG GGGSQVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPG KGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNRLYLQMNSLRA EDTAVYYCARERYSGRDYWGQGTLVTVSSAAALSNSIMYFSHFVPV FLPAKPTTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFA CDIYIWAPLAGTCGVLLLSLVITLYCNHRNRSKRSRLLHSDYMNMTP RRPGPTRKHYQPYAPPRDFAAYRSRVKFSRSADAPAYQQGQNQLYN ELNLGRREEYDVLDKRRGRDPEMGGKPRRKNPQEGLYNELQKDKM AEAYSEIGMKGERRRGKGHDGLYQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 124) Clone 2005.2 CD8 CD3 zeta CAR DNA Heavy & Light Chains GAAATAGTGATGACTCAGTCCCCGGCCACCCTCAGCGTGTCCCCC GGGGAGCGAGCGACCCTGTCATGCAGGGCTTCCCAGAGTGTCAGC TCCCTGCTCACTTGGTATCAGCAAAAGCCGGGGCAGGCTCCCCGC CTCCTCATCTTCGGGGCATCAACTAGGGCCACCGGCATTCCTGCA AGATTTTCCGGGTCTGGCAGCGGCACCGGCTTCACCCTTACCATT AGCTCTCTGCAGTCTGAGGACTTCGCCGTTTACTATTGTCAGCAGT ATGATACTTGGCCCTTTACCTTCGGTCCCGGAACTAAGGTGGACTT CAAGCGCGGGGGGGGTGGATCTGGAGGTGGTGGCTCCGGGGGCG GTGGAAGCCAGGTCCAGTTGGTTGAGAGCGGCGGCGGAGTGGTG CAGCCCGGGAGGTCCTTGCGGCTGAGCTGTGCAGCCTCCGGTTTT ACTTTTTCTAGCTATGGAATGCATTGGGTAAGACAGGCTCCCGGA AAAGGCCTCGAGTGGGTGGCGGTCATTAGCTATGATGGATCTGAT AAATACTATGTGGACTCAGTTAAGGGGCGCTTCACAATCTCAAGA GACAATAGCAAAAATAGACTGTACCTGCAGATGAATAGTCTGCGC GCCGAGGACACTGCCGTGTACTACTGCGCCCGCGAGAGATACAGC GGACGGGATTACTGGGGCCAGGGTACCCTCGTAACGGTGTCCTCC GCTGCCGCCCTTAGCAACAGCATTATGTACTTTTCTCATTTCGTGC CAGTCTTTCTCCCAGCAAAGCCCACCACTACCCCGGCCCCCAGGC CGCCTACTCCTGCCCCCACTATCGCGTCTCAGCCTCTCTCCTTGCG GCCCGAGGCCTGCCGGCCAGCCGCAGGGGGCGCCGTACATACTC GGGGTTTGGATTTCGCTTGCGACATATATATTTGGGCCCCCCTCGC CGGCACATGTGGAGTGCTGCTCCTGAGTCTCGTTATAACCCTCTAT TGCAACCATAGAAACAGATCCAAAAGAAGCCGCCTGCTCCATAG CGATTACATGAATATGACTCCACGCCGCCCTGGCCCCACAAGGAA ACACTACCAGCCTTACGCACCACCTAGAGATTTCGCTGCCTATCG GAGCAGGGTGAAGTTTTCCAGATCTGCAGATGCACCAGCGTATCA GCAGGGCCAGAACCAACTGTATAACGAGCTCAACCTGGGACGCA GGGAAGAGTATGACGTTTTGGACAAGCGCAGAGGACGGGACCCT GAGATGGGTGGCAAACCAAGACGAAAAAACCCCCAGGAGGGTCT CTATAATGAGCTGCAGAAGGATAAGATGGCTGAAGCCTATTCTGA AATAGGCATGAAAGGAGAGCGGAGAAGGGGAAAAGGGCACGAC GGTTTGTACCAGGGACTCAGCACTGCTACGAAGGATACTTATGAC GCTCTCCACATGCAAGCCCTGCCACCTAGG (SEQ ID NO. 125) Clone 2005.2 CD8 CD3 zeta CAR AA Heavy & Light Chains EIVMTQSPATLSVSPGERATLSCRASQSVSSLLTWYQQKPGQAPRLLI FGASTRATGIPARFSGSGSGTGFTLTISSLQSEDFAVYYCQQYDTWPF TFGPGTKVDFKRGGGGSGGGGSGGGGSQVQLVESGGGVVQPGRSLR LSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSV KGRFTISRDNSKNRLYLQMNSLRAEDTAVYYCARERYSGRDYWGQ GTLVTVSSAAALSNSIMYFSHFVPVFLPAKPTTTPAPRPPTPAPTIASQ PLSLRPEACRPAAGGAVHTRGLDFACDIYIWAPLAGTCGVLLLSLVIT LYCNHRNRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAY RSRVKFSRSADAPAYQQGQNQLYNELNLGRREEYDVLDKRRGRDPE MGGKPRRKNPQEGLYNELQKDKMAEAYSEIGMKGERRRGKGHDGL YQGLSTATKDTYDALHMQALPPR (SEQ ID NO. 126) CAR Signal Peptide DNA ATGGCACTCCCCGTAACTGCTCTGCTGCTGCCGTTGGCATTGCTCC
TGCACGCCGCACGCCCG (SEQ ID NO. 127) CAR Signal Peptide: MALPVTALLLPLALLLHAARP (SEQ ID NO. 128) scFv GLIS linker DNA GGCGGTGGAGGCTCCGGAGGGGGGGGCTCTGGCGGAGGGGGCTC C (SEQ ID NO. 129) scFv G4s linker: GGGGSGGGGSGGGGS (SEQ ID NO. 130) Additional G45 linker: GGGGSGGGGSGGGGSGGGGS (SEQ ID NO. 145) scFv Whitlow linker DNA GGGTCTACATCCGGCTCCGGGAAGCCCGGAAGTGGCGAAGGTAG TACAAAGGGG (SEQ ID NO. 131) scFv Whitlow linker: GSTSGSGKPGSGEGSTKG (SEQ ID NO. 132) CD28 AA Extracellular Domain MLRLLLALNLFPSIQVTGNKILVKQSPMLVAYDNAVNLSCKYSYNLF SREFRASLHKGLDSAVEVCVVYGNYSQQLQVYSKTGFNCDGKLGNE SVTFYLQNLYVNQTDIYFCKIEVMYPPPYLDNEKSNGTIIHVKGKHLC PSPLFPGPSKP (SEQ ID NO. 133) GX.sub.2X.sub.3X.sub.4X.sub.5X.sub.6X.sub.7X.sub.8X.sub.9 (SEQ ID NO: 134) X.sub.1X.sub.2X.sub.3X.sub.4X.sub.5X.sub.6 (SEQ ID NO: 135) X.sub.1X.sub.2X.sub.3X.sub.4X.sub.5X.sub.6X.sub.7X.sub.8X.sub.9X.sub.10X.s- ub.11X.sub.12DY (SEQ ID NO: 136) X.sub.1ASQX.sub.5X.sub.6X.sub.7X.sub.8X.sub.9LX.sub.11 (SEQ ID NO: 137) X.sub.1ASX.sub.4X.sub.5X.sub.6X.sub.7 (SEQ ID NO: 138) QQX.sub.3X.sub.4X.sub.5X.sub.6PX.sub.8T (SEQ ID NO: 139) CLL-1 AA (a.k.a., CLEC12A) MSEEVTYADLQFQNSSEMEKIPEIGKFGEKAPPAPSHVWRPAALFLT LLCLLLLIGLGVLASMFHVTLKIEMKKMNKLQNISEELQRNISLQLMS NMNISNKIRNLSTTLQTIATKLCRELYSKEQEHKCKPCPRRWIWHKD SCYFLSDDVQTWQESKMACAAQNASLLKINNKNALEFIKSQSRSYD YWLGLSPEEDSTRGMRVDNIINSSAWVIRNAPDLNNMYCGYINRLY VQYYHCTYKKRMICEKMANPVQLGSTYFREA (SEQ ID NO. 140) 4-1BB Nucleic Acid Sequence (intracellular domain) AAGCGCGGCAGGAAGAAGCTCCTCTACATTTTTAAGCAGCCTTTT ATGAGGCCCGTACAGACAACACAGGAGGAAGATGGCTGTAGCTG CAGATTTCCCGAGGAGGAGGAAGGTGGGTGCGAGCTG (SEQ ID NO. 141) 4-1BB AA (intracellular domain) KRGRKKLLYIFKQPFMRPVQTTQEEDGCSCRFPEEEEGGCEL (SEQ ID NO. 142) OX40 AA RRDQRLPPDAHKPPGGGSFRTPIQEEQADAHSTLAKI (SEQ ID NO. 143) Leader Sequence AA MALPVTALLLPLALLLHAARP (SEQ ID NO: 144)
Sequence CWU
1
1
1471294DNAHomo sapiens 1cttgataatg aaaagtcaaa cggaacaatc attcacgtga
agggcaagca cctctgtccg 60tcacccttgt tccctggtcc atccaagcca ttctgggtgt
tggtcgtagt gggtggagtc 120ctcgcttgtt actctctgct cgtcaccgtg gcttttataa
tcttctgggt tagatccaaa 180agaagccgcc tgctccatag cgattacatg aatatgactc
cacgccgccc tggccccaca 240aggaaacact accagcctta cgcaccacct agagatttcg
ctgcctatcg gagc 294298PRTHomo sapiens 2Leu Asp Asn Glu Lys Ser
Asn Gly Thr Ile Ile His Val Lys Gly Lys1 5
10 15His Leu Cys Pro Ser Pro Leu Phe Pro Gly Pro Ser
Lys Pro Phe Trp 20 25 30Val
Leu Val Val Val Gly Gly Val Leu Ala Cys Tyr Ser Leu Leu Val 35
40 45Thr Val Ala Phe Ile Ile Phe Trp Val
Arg Ser Lys Arg Ser Arg Leu 50 55
60Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro Gly Pro Thr65
70 75 80Arg Lys His Tyr Gln
Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala Tyr 85
90 95Arg Ser390DNAHomo sapiens 3cttgataatg
aaaagtcaaa cggaacaatc attcacgtga agggcaagca cctctgtccg 60tcacccttgt
tccctggtcc atccaagcca 90430PRTHomo
sapiens 4Leu Asp Asn Glu Lys Ser Asn Gly Thr Ile Ile His Val Lys Gly Lys1
5 10 15His Leu Cys Pro
Ser Pro Leu Phe Pro Gly Pro Ser Lys Pro 20 25
30581DNAHomo sapiens 5ttctgggtgt tggtcgtagt gggtggagtc
ctcgcttgtt actctctgct cgtcaccgtg 60gcttttataa tcttctgggt t
81627PRTHomo sapiens 6Phe Trp Val Leu
Val Val Val Gly Gly Val Leu Ala Cys Tyr Ser Leu1 5
10 15Leu Val Thr Val Ala Phe Ile Ile Phe Trp
Val 20 257123DNAHomo sapiens 7agatccaaaa
gaagccgcct gctccatagc gattacatga atatgactcc acgccgccct 60ggccccacaa
ggaaacacta ccagccttac gcaccaccta gagatttcgc tgcctatcgg 120agc
123841PRTHomo
sapiens 8Arg Ser Lys Arg Ser Arg Leu Leu His Ser Asp Tyr Met Asn Met Thr1
5 10 15Pro Arg Arg Pro
Gly Pro Thr Arg Lys His Tyr Gln Pro Tyr Ala Pro 20
25 30Pro Arg Asp Phe Ala Ala Tyr Arg Ser 35
409336DNAHomo sapiens 9agggtgaagt tttccagatc tgcagatgca
ccagcgtatc agcagggcca gaaccaactg 60tataacgagc tcaacctggg acgcagggaa
gagtatgacg ttttggacaa gcgcagagga 120cgggaccctg agatgggtgg caaaccaaga
cgaaaaaacc cccaggaggg tctctataat 180gagctgcaga aggataagat ggctgaagcc
tattctgaaa taggcatgaa aggagagcgg 240agaaggggaa aagggcacga cggtttgtac
cagggactca gcactgctac gaaggatact 300tatgacgctc tccacatgca agccctgcca
cctagg 33610112PRTHomo sapiens 10Arg Val Lys
Phe Ser Arg Ser Ala Asp Ala Pro Ala Tyr Gln Gln Gly1 5
10 15Gln Asn Gln Leu Tyr Asn Glu Leu Asn
Leu Gly Arg Arg Glu Glu Tyr 20 25
30Asp Val Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu Met Gly Gly Lys
35 40 45Pro Arg Arg Lys Asn Pro Gln
Glu Gly Leu Tyr Asn Glu Leu Gln Lys 50 55
60Asp Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg65
70 75 80Arg Arg Gly Lys
Gly His Asp Gly Leu Tyr Gln Gly Leu Ser Thr Ala 85
90 95Thr Lys Asp Thr Tyr Asp Ala Leu His Met
Gln Ala Leu Pro Pro Arg 100 105
11011117DNAHomo sapiens 11attgaggtga tgtatccacc gccttacctg gataacgaaa
agagtaacgg taccatcatt 60cacgtgaaag gtaaacacct gtgtccttct cccctcttcc
ccgggccatc aaagccc 1171239PRTHomo sapiens 12Ile Glu Val Met Tyr Pro
Pro Pro Tyr Leu Asp Asn Glu Lys Ser Asn1 5
10 15Gly Thr Ile Ile His Val Lys Gly Lys His Leu Cys
Pro Ser Pro Leu 20 25 30Phe
Pro Gly Pro Ser Lys Pro 3513288DNAHomo sapiens 13gctgcagcat
tgagcaactc aataatgtat tttagtcact ttgtaccagt gttcttgccg 60gctaagccta
ctaccacacc cgctccacgg ccacctaccc cagctcctac catcgcttca 120cagcctctgt
ccctgcgccc agaggcttgc cgaccggccg cagggggcgc tgttcatacc 180agaggactgg
atttcgcctg cgatatctat atctgggcac ccctggccgg aacctgcggc 240gtactcctgc
tgtccctggt catcacgctc tattgtaatc acaggaac 2881496PRTHomo
sapiens 14Ala Ala Ala Leu Ser Asn Ser Ile Met Tyr Phe Ser His Phe Val
Pro1 5 10 15Val Phe Leu
Pro Ala Lys Pro Thr Thr Thr Pro Ala Pro Arg Pro Pro 20
25 30Thr Pro Ala Pro Thr Ile Ala Ser Gln Pro
Leu Ser Leu Arg Pro Glu 35 40
45Ala Cys Arg Pro Ala Ala Gly Gly Ala Val His Thr Arg Gly Leu Asp 50
55 60Phe Ala Cys Asp Ile Tyr Ile Trp Ala
Pro Leu Ala Gly Thr Cys Gly65 70 75
80Val Leu Leu Leu Ser Leu Val Ile Thr Leu Tyr Cys Asn His
Arg Asn 85 90
9515366DNAArtificial SequenceDescription of Artificial Sequence Synthetic
polynucleotide 15caggtgcagc tgcaggaatc cggaccgggg ctggtgaagc
ccagcgagac tctgagtctc 60acgtgtacag tttctggagg tagcattagc tcctactatt
ggtcatggat aaggcagccc 120cccgggaagg gattggaatg gatcggctat atttactaca
gtgggagcac caattacaac 180ccctcactga agtctagagt tacaatcagc gttgacacct
caaagaatca gttcagtttg 240aaattgtcta gcgtcacagc agctgataca gccgtctatt
attgtgtttc tctggtctat 300tgcggtgggg attgttacag tggctttgac tattgggggc
agggtactct ggttacagtt 360tcttcc
36616122PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 16Gln Val Gln Leu Gln Glu
Ser Gly Pro Gly Leu Val Lys Pro Ser Glu1 5
10 15Thr Leu Ser Leu Thr Cys Thr Val Ser Gly Gly Ser
Ile Ser Ser Tyr 20 25 30Tyr
Trp Ser Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile 35
40 45Gly Tyr Ile Tyr Tyr Ser Gly Ser Thr
Asn Tyr Asn Pro Ser Leu Lys 50 55
60Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu65
70 75 80Lys Leu Ser Ser Val
Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Val 85
90 95Ser Leu Val Tyr Cys Gly Gly Asp Cys Tyr Ser
Gly Phe Asp Tyr Trp 100 105
110Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115
120177PRTArtificial SequenceDescription of Artificial Sequence Synthetic
peptide 17Gly Gly Ser Ile Ser Ser Tyr1
5185PRTArtificial SequenceDescription of Artificial Sequence Synthetic
peptide 18Tyr Tyr Ser Gly Ser1 51914PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 19Leu
Val Tyr Cys Gly Gly Asp Cys Tyr Ser Gly Phe Asp Tyr1 5
1020324DNAArtificial SequenceDescription of Artificial
Sequence Synthetic polynucleotide 20gacatccagt tgacacagag cccgagttcc
ttgtccgcct ccgtcgggga tagagtgtca 60tttacctgtc aggcctctca ggatattaat
aactttctga attggtatca gcaaaagccc 120ggaaaggcac ccaagctgtt gatttacgac
gccagtaacc tggagacagg cgtgccctcc 180cggtttagtg gtagcggaag cggtacggat
tttaccttta ctatcagctc tctccaaccc 240gaagacattg caacctacta ttgtcaacaa
tatggaaacc tgccttttac atttggcggc 300ggcaccaagg tggagattaa gcgg
32421108PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
21Asp Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1
5 10 15Asp Arg Val Ser Phe Thr
Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe 20 25
30Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Leu Leu Ile 35 40 45Tyr Asp Ala
Ser Asn Leu Glu Thr Gly Val Pro Ser Arg Phe Ser Gly 50
55 60Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser
Ser Leu Gln Pro65 70 75
80Glu Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Gly Asn Leu Pro Phe
85 90 95Thr Phe Gly Gly Gly Thr
Lys Val Glu Ile Lys Arg 100
1052211PRTArtificial SequenceDescription of Artificial Sequence Synthetic
peptide 22Gln Ala Ser Gln Asp Ile Asn Asn Phe Leu Asn1
5 10237PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 23Asp Ala Ser Asn Leu Glu Thr1
5249PRTArtificial SequenceDescription of Artificial Sequence Synthetic
peptide 24Gln Gln Tyr Gly Asn Leu Pro Phe Thr1
5251440DNAArtificial SequenceDescription of Artificial Sequence Synthetic
polynucleotide 25atggcactcc ccgtaactgc tctgctgctg ccgttggcat
tgctcctgca cgccgcacgc 60ccgcaggtcc aactgcaaga aagcggaccc ggactggtga
agccttctga gacacttagt 120ctgacgtgca cggtcagtgg cggctccatc tcctcctatt
attggtcatg gatacgacaa 180cccccaggta agggcctgga atggattggc tatatctact
attcaggaag cacgaactac 240aatcccagcc tgaagtcccg agtgacaatt tcagtagata
ccagtaaaaa ccagttcagt 300cttaaactgt caagcgtgac agctgccgac accgctgtgt
attactgcgt ctcactggtg 360tattgtggag gggattgtta tagcgggttc gattattggg
gacagggaac cctggtgact 420gtatcttccg gcggcggcgg ctcagggggt ggcggtagtg
gcggtggggg ttccgatatt 480caactgacac aatcccccag ctcactcagc gccagcgtgg
gggacagggt tagctttacc 540tgtcaagcct ctcaggatat aaataacttt ctgaactggt
atcaacagaa gcctgggaag 600gcgcccaaac tcctgatcta tgatgcgtcc aacctggaaa
ctggcgtgcc ttcacgcttt 660agcggctctg gcagtggtac agacttcact tttaccatct
cttcacttca gccggaggac 720atcgccacat attactgtca acagtacgga aacttgccct
ttacttttgg aggcggcacc 780aaagttgaaa tcaaaagggc cgctgccctg gataacgaaa
agagcaatgg gactataata 840catgttaaag gaaaacacct gtgtccatct cccctgttcc
ctggaccgtc aaagccattt 900tgggtgctcg tggttgtcgg tggcgttctc gcctgttata
gcttgctggt gacagtagcc 960ttcattatct tttgggtgag atccaaaaga agccgcctgc
tccatagcga ttacatgaat 1020atgactccac gccgccctgg ccccacaagg aaacactacc
agccttacgc accacctaga 1080gatttcgctg cctatcggag cagggtgaag ttttccagat
ctgcagatgc accagcgtat 1140cagcagggcc agaaccaact gtataacgag ctcaacctgg
gacgcaggga agagtatgac 1200gttttggaca agcgcagagg acgggaccct gagatgggtg
gcaaaccaag acgaaaaaac 1260ccccaggagg gtctctataa tgagctgcag aaggataaga
tggctgaagc ctattctgaa 1320ataggcatga aaggagagcg gagaagggga aaagggcacg
acggtttgta ccagggactc 1380agcactgcta cgaaggatac ttatgacgct ctccacatgc
aagccctgcc acctaggtaa 144026479PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 26Met Ala Leu Pro Val Thr
Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1 5
10 15His Ala Ala Arg Pro Gln Val Gln Leu Gln Glu Ser
Gly Pro Gly Leu 20 25 30Val
Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Thr Val Ser Gly Gly 35
40 45Ser Ile Ser Ser Tyr Tyr Trp Ser Trp
Ile Arg Gln Pro Pro Gly Lys 50 55
60Gly Leu Glu Trp Ile Gly Tyr Ile Tyr Tyr Ser Gly Ser Thr Asn Tyr65
70 75 80Asn Pro Ser Leu Lys
Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys 85
90 95Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr
Ala Ala Asp Thr Ala 100 105
110Val Tyr Tyr Cys Val Ser Leu Val Tyr Cys Gly Gly Asp Cys Tyr Ser
115 120 125Gly Phe Asp Tyr Trp Gly Gln
Gly Thr Leu Val Thr Val Ser Ser Gly 130 135
140Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Asp
Ile145 150 155 160Gln Leu
Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg
165 170 175Val Ser Phe Thr Cys Gln Ala
Ser Gln Asp Ile Asn Asn Phe Leu Asn 180 185
190Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile
Tyr Asp 195 200 205Ala Ser Asn Leu
Glu Thr Gly Val Pro Ser Arg Phe Ser Gly Ser Gly 210
215 220Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser Ser Leu
Gln Pro Glu Asp225 230 235
240Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Gly Asn Leu Pro Phe Thr Phe
245 250 255Gly Gly Gly Thr Lys
Val Glu Ile Lys Arg Ala Ala Ala Leu Asp Asn 260
265 270Glu Lys Ser Asn Gly Thr Ile Ile His Val Lys Gly
Lys His Leu Cys 275 280 285Pro Ser
Pro Leu Phe Pro Gly Pro Ser Lys Pro Phe Trp Val Leu Val 290
295 300Val Val Gly Gly Val Leu Ala Cys Tyr Ser Leu
Leu Val Thr Val Ala305 310 315
320Phe Ile Ile Phe Trp Val Arg Ser Lys Arg Ser Arg Leu Leu His Ser
325 330 335Asp Tyr Met Asn
Met Thr Pro Arg Arg Pro Gly Pro Thr Arg Lys His 340
345 350Tyr Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala
Ala Tyr Arg Ser Arg 355 360 365Val
Lys Phe Ser Arg Ser Ala Asp Ala Pro Ala Tyr Gln Gln Gly Gln 370
375 380Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly
Arg Arg Glu Glu Tyr Asp385 390 395
400Val Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu Met Gly Gly Lys
Pro 405 410 415Arg Arg Lys
Asn Pro Gln Glu Gly Leu Tyr Asn Glu Leu Gln Lys Asp 420
425 430Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly
Met Lys Gly Glu Arg Arg 435 440
445Arg Gly Lys Gly His Asp Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr 450
455 460Lys Asp Thr Tyr Asp Ala Leu His
Met Gln Ala Leu Pro Pro Arg465 470
475271374DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 27caggtccaac tgcaagaaag cggacccgga
ctggtgaagc cttctgagac acttagtctg 60acgtgcacgg tcagtggcgg ctccatctcc
tcctattatt ggtcatggat acgacaaccc 120ccaggtaagg gcctggaatg gattggctat
atctactatt caggaagcac gaactacaat 180cccagcctga agtcccgagt gacaatttca
gtagatacca gtaaaaacca gttcagtctt 240aaactgtcaa gcgtgacagc tgccgacacc
gctgtgtatt actgcgtctc actggtgtat 300tgtggagggg attgttatag cgggttcgat
tattggggac agggaaccct ggtgactgta 360tcttccggcg gcggcggctc agggggtggc
ggtagtggcg gtgggggttc cgatattcaa 420ctgacacaat cccccagctc actcagcgcc
agcgtggggg acagggttag ctttacctgt 480caagcctctc aggatataaa taactttctg
aactggtatc aacagaagcc tgggaaggcg 540cccaaactcc tgatctatga tgcgtccaac
ctggaaactg gcgtgccttc acgctttagc 600ggctctggca gtggtacaga cttcactttt
accatctctt cacttcagcc ggaggacatc 660gccacatatt actgtcaaca gtacggaaac
ttgcccttta cttttggagg cggcaccaaa 720gttgaaatca aaagggccgc tgccctggat
aacgaaaaga gcaatgggac tataatacat 780gttaaaggaa aacacctgtg tccatctccc
ctgttccctg gaccgtcaaa gccattttgg 840gtgctcgtgg ttgtcggtgg cgttctcgcc
tgttatagct tgctggtgac agtagccttc 900attatctttt gggtgagatc caaaagaagc
cgcctgctcc atagcgatta catgaatatg 960actccacgcc gccctggccc cacaaggaaa
cactaccagc cttacgcacc acctagagat 1020ttcgctgcct atcggagcag ggtgaagttt
tccagatctg cagatgcacc agcgtatcag 1080cagggccaga accaactgta taacgagctc
aacctgggac gcagggaaga gtatgacgtt 1140ttggacaagc gcagaggacg ggaccctgag
atgggtggca aaccaagacg aaaaaacccc 1200caggagggtc tctataatga gctgcagaag
gataagatgg ctgaagccta ttctgaaata 1260ggcatgaaag gagagcggag aaggggaaaa
gggcacgacg gtttgtacca gggactcagc 1320actgctacga aggatactta tgacgctctc
cacatgcaag ccctgccacc tagg 137428458PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
28Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu1
5 10 15Thr Leu Ser Leu Thr Cys
Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr 20 25
30Tyr Trp Ser Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu
Glu Trp Ile 35 40 45Gly Tyr Ile
Tyr Tyr Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys 50
55 60Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn
Gln Phe Ser Leu65 70 75
80Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Val
85 90 95Ser Leu Val Tyr Cys Gly
Gly Asp Cys Tyr Ser Gly Phe Asp Tyr Trp 100
105 110Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly
Gly Gly Ser Gly 115 120 125Gly Gly
Gly Ser Gly Gly Gly Gly Ser Asp Ile Gln Leu Thr Gln Ser 130
135 140Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg
Val Ser Phe Thr Cys145 150 155
160Gln Ala Ser Gln Asp Ile Asn Asn Phe Leu Asn Trp Tyr Gln Gln Lys
165 170 175Pro Gly Lys Ala
Pro Lys Leu Leu Ile Tyr Asp Ala Ser Asn Leu Glu 180
185 190Thr Gly Val Pro Ser Arg Phe Ser Gly Ser Gly
Ser Gly Thr Asp Phe 195 200 205Thr
Phe Thr Ile Ser Ser Leu Gln Pro Glu Asp Ile Ala Thr Tyr Tyr 210
215 220Cys Gln Gln Tyr Gly Asn Leu Pro Phe Thr
Phe Gly Gly Gly Thr Lys225 230 235
240Val Glu Ile Lys Arg Ala Ala Ala Leu Asp Asn Glu Lys Ser Asn
Gly 245 250 255Thr Ile Ile
His Val Lys Gly Lys His Leu Cys Pro Ser Pro Leu Phe 260
265 270Pro Gly Pro Ser Lys Pro Phe Trp Val Leu
Val Val Val Gly Gly Val 275 280
285Leu Ala Cys Tyr Ser Leu Leu Val Thr Val Ala Phe Ile Ile Phe Trp 290
295 300Val Arg Ser Lys Arg Ser Arg Leu
Leu His Ser Asp Tyr Met Asn Met305 310
315 320Thr Pro Arg Arg Pro Gly Pro Thr Arg Lys His Tyr
Gln Pro Tyr Ala 325 330
335Pro Pro Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser Arg
340 345 350Ser Ala Asp Ala Pro Ala
Tyr Gln Gln Gly Gln Asn Gln Leu Tyr Asn 355 360
365Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr Asp Val Leu Asp
Lys Arg 370 375 380Arg Gly Arg Asp Pro
Glu Met Gly Gly Lys Pro Arg Arg Lys Asn Pro385 390
395 400Gln Glu Gly Leu Tyr Asn Glu Leu Gln Lys
Asp Lys Met Ala Glu Ala 405 410
415Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His
420 425 430Asp Gly Leu Tyr Gln
Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp 435
440 445Ala Leu His Met Gln Ala Leu Pro Pro Arg 450
455291467DNAArtificial SequenceDescription of Artificial
Sequence Synthetic polynucleotide 29atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccgcaggtgc agctgcagga atccggaccg
gggctggtga agcccagcga gactctgagt 120ctcacgtgta cagtttctgg aggtagcatt
agctcctact attggtcatg gataaggcag 180ccccccggga agggattgga atggatcggc
tatatttact acagtgggag caccaattac 240aacccctcac tgaagtctag agttacaatc
agcgttgaca cctcaaagaa tcagttcagt 300ttgaaattgt ctagcgtcac agcagctgat
acagccgtct attattgtgt ttctctggtc 360tattgcggtg gggattgtta cagtggcttt
gactattggg ggcagggtac tctggttaca 420gtttcttccg gggggggagg ctctgggggc
ggaggctcag gtggtggagg cagcgacatc 480cagttgacac agagcccgag ttccttgtcc
gcctccgtcg gggatagagt gtcatttacc 540tgtcaggcct ctcaggatat taataacttt
ctgaattggt atcagcaaaa gcccggaaag 600gcacccaagc tgttgattta cgacgccagt
aacctggaga caggcgtgcc ctcccggttt 660agtggtagcg gaagcggtac ggattttacc
tttactatca gctctctcca acccgaagac 720attgcaacct actattgtca acaatatgga
aacctgcctt ttacatttgg cggcggcacc 780aaggtggaga ttaagcgggc ggcagctatt
gaggtgatgt atccaccgcc ttacctggat 840aacgaaaaga gtaacggtac catcattcac
gtgaaaggta aacacctgtg tccttctccc 900ctcttccccg ggccatcaaa gcccttctgg
gttcttgtgg tcgtgggagg cgtgcttgct 960tgttattctc tgctcgttac cgtggcgttt
atcatttttt gggttagatc caaaagaagc 1020cgcctgctcc atagcgatta catgaatatg
actccacgcc gccctggccc cacaaggaaa 1080cactaccagc cttacgcacc acctagagat
ttcgctgcct atcggagcag ggtgaagttt 1140tccagatctg cagatgcacc agcgtatcag
cagggccaga accaactgta taacgagctc 1200aacctgggac gcagggaaga gtatgacgtt
ttggacaagc gcagaggacg ggaccctgag 1260atgggtggca aaccaagacg aaaaaacccc
caggagggtc tctataatga gctgcagaag 1320gataagatgg ctgaagccta ttctgaaata
ggcatgaaag gagagcggag aaggggaaaa 1380gggcacgacg gtttgtacca gggactcagc
actgctacga aggatactta tgacgctctc 1440cacatgcaag ccctgccacc taggtaa
146730488PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
30Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Gln
Val Gln Leu Gln Glu Ser Gly Pro Gly Leu 20 25
30Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Thr Val
Ser Gly Gly 35 40 45Ser Ile Ser
Ser Tyr Tyr Trp Ser Trp Ile Arg Gln Pro Pro Gly Lys 50
55 60Gly Leu Glu Trp Ile Gly Tyr Ile Tyr Tyr Ser Gly
Ser Thr Asn Tyr65 70 75
80Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys
85 90 95Asn Gln Phe Ser Leu Lys
Leu Ser Ser Val Thr Ala Ala Asp Thr Ala 100
105 110Val Tyr Tyr Cys Val Ser Leu Val Tyr Cys Gly Gly
Asp Cys Tyr Ser 115 120 125Gly Phe
Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly 130
135 140Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly
Gly Gly Ser Asp Ile145 150 155
160Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg
165 170 175Val Ser Phe Thr
Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe Leu Asn 180
185 190Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Leu Leu Ile Tyr Asp 195 200 205Ala
Ser Asn Leu Glu Thr Gly Val Pro Ser Arg Phe Ser Gly Ser Gly 210
215 220Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser
Ser Leu Gln Pro Glu Asp225 230 235
240Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Gly Asn Leu Pro Phe Thr
Phe 245 250 255Gly Gly Gly
Thr Lys Val Glu Ile Lys Arg Ala Ala Ala Ile Glu Val 260
265 270Met Tyr Pro Pro Pro Tyr Leu Asp Asn Glu
Lys Ser Asn Gly Thr Ile 275 280
285Ile His Val Lys Gly Lys His Leu Cys Pro Ser Pro Leu Phe Pro Gly 290
295 300Pro Ser Lys Pro Phe Trp Val Leu
Val Val Val Gly Gly Val Leu Ala305 310
315 320Cys Tyr Ser Leu Leu Val Thr Val Ala Phe Ile Ile
Phe Trp Val Arg 325 330
335Ser Lys Arg Ser Arg Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro
340 345 350Arg Arg Pro Gly Pro Thr
Arg Lys His Tyr Gln Pro Tyr Ala Pro Pro 355 360
365Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser Arg
Ser Ala 370 375 380Asp Ala Pro Ala Tyr
Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu385 390
395 400Asn Leu Gly Arg Arg Glu Glu Tyr Asp Val
Leu Asp Lys Arg Arg Gly 405 410
415Arg Asp Pro Glu Met Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu
420 425 430Gly Leu Tyr Asn Glu
Leu Gln Lys Asp Lys Met Ala Glu Ala Tyr Ser 435
440 445Glu Ile Gly Met Lys Gly Glu Arg Arg Arg Gly Lys
Gly His Asp Gly 450 455 460Leu Tyr Gln
Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu465
470 475 480His Met Gln Ala Leu Pro Pro
Arg 485311401DNAArtificial SequenceDescription of
Artificial Sequence Synthetic polynucleotide 31caggtgcagc tgcaggaatc
cggaccgggg ctggtgaagc ccagcgagac tctgagtctc 60acgtgtacag tttctggagg
tagcattagc tcctactatt ggtcatggat aaggcagccc 120cccgggaagg gattggaatg
gatcggctat atttactaca gtgggagcac caattacaac 180ccctcactga agtctagagt
tacaatcagc gttgacacct caaagaatca gttcagtttg 240aaattgtcta gcgtcacagc
agctgataca gccgtctatt attgtgtttc tctggtctat 300tgcggtgggg attgttacag
tggctttgac tattgggggc agggtactct ggttacagtt 360tcttccgggg ggggaggctc
tgggggcgga ggctcaggtg gtggaggcag cgacatccag 420ttgacacaga gcccgagttc
cttgtccgcc tccgtcgggg atagagtgtc atttacctgt 480caggcctctc aggatattaa
taactttctg aattggtatc agcaaaagcc cggaaaggca 540cccaagctgt tgatttacga
cgccagtaac ctggagacag gcgtgccctc ccggtttagt 600ggtagcggaa gcggtacgga
ttttaccttt actatcagct ctctccaacc cgaagacatt 660gcaacctact attgtcaaca
atatggaaac ctgcctttta catttggcgg cggcaccaag 720gtggagatta agcgggcggc
agctattgag gtgatgtatc caccgcctta cctggataac 780gaaaagagta acggtaccat
cattcacgtg aaaggtaaac acctgtgtcc ttctcccctc 840ttccccgggc catcaaagcc
cttctgggtt cttgtggtcg tgggaggcgt gcttgcttgt 900tattctctgc tcgttaccgt
ggcgtttatc attttttggg ttagatccaa aagaagccgc 960ctgctccata gcgattacat
gaatatgact ccacgccgcc ctggccccac aaggaaacac 1020taccagcctt acgcaccacc
tagagatttc gctgcctatc ggagcagggt gaagttttcc 1080agatctgcag atgcaccagc
gtatcagcag ggccagaacc aactgtataa cgagctcaac 1140ctgggacgca gggaagagta
tgacgttttg gacaagcgca gaggacggga ccctgagatg 1200ggtggcaaac caagacgaaa
aaacccccag gagggtctct ataatgagct gcagaaggat 1260aagatggctg aagcctattc
tgaaataggc atgaaaggag agcggagaag gggaaaaggg 1320cacgacggtt tgtaccaggg
actcagcact gctacgaagg atacttatga cgctctccac 1380atgcaagccc tgccacctag g
140132467PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
32Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu1
5 10 15Thr Leu Ser Leu Thr Cys
Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr 20 25
30Tyr Trp Ser Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu
Glu Trp Ile 35 40 45Gly Tyr Ile
Tyr Tyr Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys 50
55 60Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn
Gln Phe Ser Leu65 70 75
80Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Val
85 90 95Ser Leu Val Tyr Cys Gly
Gly Asp Cys Tyr Ser Gly Phe Asp Tyr Trp 100
105 110Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly
Gly Gly Ser Gly 115 120 125Gly Gly
Gly Ser Gly Gly Gly Gly Ser Asp Ile Gln Leu Thr Gln Ser 130
135 140Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg
Val Ser Phe Thr Cys145 150 155
160Gln Ala Ser Gln Asp Ile Asn Asn Phe Leu Asn Trp Tyr Gln Gln Lys
165 170 175Pro Gly Lys Ala
Pro Lys Leu Leu Ile Tyr Asp Ala Ser Asn Leu Glu 180
185 190Thr Gly Val Pro Ser Arg Phe Ser Gly Ser Gly
Ser Gly Thr Asp Phe 195 200 205Thr
Phe Thr Ile Ser Ser Leu Gln Pro Glu Asp Ile Ala Thr Tyr Tyr 210
215 220Cys Gln Gln Tyr Gly Asn Leu Pro Phe Thr
Phe Gly Gly Gly Thr Lys225 230 235
240Val Glu Ile Lys Arg Ala Ala Ala Ile Glu Val Met Tyr Pro Pro
Pro 245 250 255Tyr Leu Asp
Asn Glu Lys Ser Asn Gly Thr Ile Ile His Val Lys Gly 260
265 270Lys His Leu Cys Pro Ser Pro Leu Phe Pro
Gly Pro Ser Lys Pro Phe 275 280
285Trp Val Leu Val Val Val Gly Gly Val Leu Ala Cys Tyr Ser Leu Leu 290
295 300Val Thr Val Ala Phe Ile Ile Phe
Trp Val Arg Ser Lys Arg Ser Arg305 310
315 320Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg
Arg Pro Gly Pro 325 330
335Thr Arg Lys His Tyr Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala
340 345 350Tyr Arg Ser Arg Val Lys
Phe Ser Arg Ser Ala Asp Ala Pro Ala Tyr 355 360
365Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly
Arg Arg 370 375 380Glu Glu Tyr Asp Val
Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu Met385 390
395 400Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln
Glu Gly Leu Tyr Asn Glu 405 410
415Leu Gln Lys Asp Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys
420 425 430Gly Glu Arg Arg Arg
Gly Lys Gly His Asp Gly Leu Tyr Gln Gly Leu 435
440 445Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu His
Met Gln Ala Leu 450 455 460Pro Pro
Arg465331548DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 33atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccgcaggtgc aattgcaaga gtccggcccc
ggactcgtta aacccagtga gacgcttagc 120ctgacctgta ccgtctcagg gggcagcatc
tcctcttatt actggagctg gatcaggcag 180cctccaggaa aaggccttga atggattggg
tacatctact actctggctc aacaaattat 240aatccatccc tgaagtcccg cgtgactatc
tctgtggaca ccagcaagaa tcagttttca 300ctgaagttgt ctagtgttac cgcggccgac
accgccgtat actactgtgt gtctcttgtg 360tactgtggcg gcgactgcta ttccgggttc
gactactggg gccaagggac tctggtaacc 420gtgtcctcag gcggcggcgg gtcaggagga
ggcggcagtg gaggtggcgg ctccgacatc 480cagctgacac aatcaccatc ttccctttca
gcttcagtcg gggacagagt gtccttcaca 540tgccaggcca gccaggatat caataacttc
ctgaactggt accaacagaa acccggaaag 600gctccaaagc tcctgatcta tgatgcttcc
aacctggaga ccggcgtgcc ctccaggttc 660agtggttcag gatcaggcac tgactttacg
ttcaccatat ccagtcttca gcccgaagac 720attgcaacct attactgcca acaatacggg
aaccttccct ttacattcgg aggcggcacc 780aaggtggaaa tcaaaagggc tgcagcattg
agcaactcaa taatgtattt tagtcacttt 840gtaccagtgt tcttgccggc taagcctact
accacacccg ctccacggcc acctacccca 900gctcctacca tcgcttcaca gcctctgtcc
ctgcgcccag aggcttgccg accggccgca 960gggggcgctg ttcataccag aggactggat
ttcgcctgcg atatctatat ctgggcaccc 1020ctggccggaa cctgcggcgt actcctgctg
tccctggtca tcacgctcta ttgtaatcac 1080aggaacagat ccaaaagaag ccgcctgctc
catagcgatt acatgaatat gactccacgc 1140cgccctggcc ccacaaggaa acactaccag
ccttacgcac cacctagaga tttcgctgcc 1200tatcggagca gggtgaagtt ttccagatct
gcagatgcac cagcgtatca gcagggccag 1260aaccaactgt ataacgagct caacctggga
cgcagggaag agtatgacgt tttggacaag 1320cgcagaggac gggaccctga gatgggtggc
aaaccaagac gaaaaaaccc ccaggagggt 1380ctctataatg agctgcagaa ggataagatg
gctgaagcct attctgaaat aggcatgaaa 1440ggagagcgga gaaggggaaa agggcacgac
ggtttgtacc agggactcag cactgctacg 1500aaggatactt atgacgctct ccacatgcaa
gccctgccac ctaggtaa 154834515PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
34Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Gln
Val Gln Leu Gln Glu Ser Gly Pro Gly Leu 20 25
30Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Thr Val
Ser Gly Gly 35 40 45Ser Ile Ser
Ser Tyr Tyr Trp Ser Trp Ile Arg Gln Pro Pro Gly Lys 50
55 60Gly Leu Glu Trp Ile Gly Tyr Ile Tyr Tyr Ser Gly
Ser Thr Asn Tyr65 70 75
80Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys
85 90 95Asn Gln Phe Ser Leu Lys
Leu Ser Ser Val Thr Ala Ala Asp Thr Ala 100
105 110Val Tyr Tyr Cys Val Ser Leu Val Tyr Cys Gly Gly
Asp Cys Tyr Ser 115 120 125Gly Phe
Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly 130
135 140Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly
Gly Gly Ser Asp Ile145 150 155
160Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg
165 170 175Val Ser Phe Thr
Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe Leu Asn 180
185 190Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Leu Leu Ile Tyr Asp 195 200 205Ala
Ser Asn Leu Glu Thr Gly Val Pro Ser Arg Phe Ser Gly Ser Gly 210
215 220Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser
Ser Leu Gln Pro Glu Asp225 230 235
240Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Gly Asn Leu Pro Phe Thr
Phe 245 250 255Gly Gly Gly
Thr Lys Val Glu Ile Lys Arg Ala Ala Ala Leu Ser Asn 260
265 270Ser Ile Met Tyr Phe Ser His Phe Val Pro
Val Phe Leu Pro Ala Lys 275 280
285Pro Thr Thr Thr Pro Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile 290
295 300Ala Ser Gln Pro Leu Ser Leu Arg
Pro Glu Ala Cys Arg Pro Ala Ala305 310
315 320Gly Gly Ala Val His Thr Arg Gly Leu Asp Phe Ala
Cys Asp Ile Tyr 325 330
335Ile Trp Ala Pro Leu Ala Gly Thr Cys Gly Val Leu Leu Leu Ser Leu
340 345 350Val Ile Thr Leu Tyr Cys
Asn His Arg Asn Arg Ser Lys Arg Ser Arg 355 360
365Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro
Gly Pro 370 375 380Thr Arg Lys His Tyr
Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala385 390
395 400Tyr Arg Ser Arg Val Lys Phe Ser Arg Ser
Ala Asp Ala Pro Ala Tyr 405 410
415Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg
420 425 430Glu Glu Tyr Asp Val
Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu Met 435
440 445Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly
Leu Tyr Asn Glu 450 455 460Leu Gln Lys
Asp Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys465
470 475 480Gly Glu Arg Arg Arg Gly Lys
Gly His Asp Gly Leu Tyr Gln Gly Leu 485
490 495Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu His
Met Gln Ala Leu 500 505 510Pro
Pro Arg 515351482DNAArtificial SequenceDescription of Artificial
Sequence Synthetic polynucleotide 35caggtgcaat tgcaagagtc cggccccgga
ctcgttaaac ccagtgagac gcttagcctg 60acctgtaccg tctcaggggg cagcatctcc
tcttattact ggagctggat caggcagcct 120ccaggaaaag gccttgaatg gattgggtac
atctactact ctggctcaac aaattataat 180ccatccctga agtcccgcgt gactatctct
gtggacacca gcaagaatca gttttcactg 240aagttgtcta gtgttaccgc ggccgacacc
gccgtatact actgtgtgtc tcttgtgtac 300tgtggcggcg actgctattc cgggttcgac
tactggggcc aagggactct ggtaaccgtg 360tcctcaggcg gcggcgggtc aggaggaggc
ggcagtggag gtggcggctc cgacatccag 420ctgacacaat caccatcttc cctttcagct
tcagtcgggg acagagtgtc cttcacatgc 480caggccagcc aggatatcaa taacttcctg
aactggtacc aacagaaacc cggaaaggct 540ccaaagctcc tgatctatga tgcttccaac
ctggagaccg gcgtgccctc caggttcagt 600ggttcaggat caggcactga ctttacgttc
accatatcca gtcttcagcc cgaagacatt 660gcaacctatt actgccaaca atacgggaac
cttcccttta cattcggagg cggcaccaag 720gtggaaatca aaagggctgc agcattgagc
aactcaataa tgtattttag tcactttgta 780ccagtgttct tgccggctaa gcctactacc
acacccgctc cacggccacc taccccagct 840cctaccatcg cttcacagcc tctgtccctg
cgcccagagg cttgccgacc ggccgcaggg 900ggcgctgttc ataccagagg actggatttc
gcctgcgata tctatatctg ggcacccctg 960gccggaacct gcggcgtact cctgctgtcc
ctggtcatca cgctctattg taatcacagg 1020aacagatcca aaagaagccg cctgctccat
agcgattaca tgaatatgac tccacgccgc 1080cctggcccca caaggaaaca ctaccagcct
tacgcaccac ctagagattt cgctgcctat 1140cggagcaggg tgaagttttc cagatctgca
gatgcaccag cgtatcagca gggccagaac 1200caactgtata acgagctcaa cctgggacgc
agggaagagt atgacgtttt ggacaagcgc 1260agaggacggg accctgagat gggtggcaaa
ccaagacgaa aaaaccccca ggagggtctc 1320tataatgagc tgcagaagga taagatggct
gaagcctatt ctgaaatagg catgaaagga 1380gagcggagaa ggggaaaagg gcacgacggt
ttgtaccagg gactcagcac tgctacgaag 1440gatacttatg acgctctcca catgcaagcc
ctgccaccta gg 148236494PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
36Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu1
5 10 15Thr Leu Ser Leu Thr Cys
Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr 20 25
30Tyr Trp Ser Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu
Glu Trp Ile 35 40 45Gly Tyr Ile
Tyr Tyr Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys 50
55 60Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn
Gln Phe Ser Leu65 70 75
80Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Val
85 90 95Ser Leu Val Tyr Cys Gly
Gly Asp Cys Tyr Ser Gly Phe Asp Tyr Trp 100
105 110Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly
Gly Gly Ser Gly 115 120 125Gly Gly
Gly Ser Gly Gly Gly Gly Ser Asp Ile Gln Leu Thr Gln Ser 130
135 140Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg
Val Ser Phe Thr Cys145 150 155
160Gln Ala Ser Gln Asp Ile Asn Asn Phe Leu Asn Trp Tyr Gln Gln Lys
165 170 175Pro Gly Lys Ala
Pro Lys Leu Leu Ile Tyr Asp Ala Ser Asn Leu Glu 180
185 190Thr Gly Val Pro Ser Arg Phe Ser Gly Ser Gly
Ser Gly Thr Asp Phe 195 200 205Thr
Phe Thr Ile Ser Ser Leu Gln Pro Glu Asp Ile Ala Thr Tyr Tyr 210
215 220Cys Gln Gln Tyr Gly Asn Leu Pro Phe Thr
Phe Gly Gly Gly Thr Lys225 230 235
240Val Glu Ile Lys Arg Ala Ala Ala Leu Ser Asn Ser Ile Met Tyr
Phe 245 250 255Ser His Phe
Val Pro Val Phe Leu Pro Ala Lys Pro Thr Thr Thr Pro 260
265 270Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr
Ile Ala Ser Gln Pro Leu 275 280
285Ser Leu Arg Pro Glu Ala Cys Arg Pro Ala Ala Gly Gly Ala Val His 290
295 300Thr Arg Gly Leu Asp Phe Ala Cys
Asp Ile Tyr Ile Trp Ala Pro Leu305 310
315 320Ala Gly Thr Cys Gly Val Leu Leu Leu Ser Leu Val
Ile Thr Leu Tyr 325 330
335Cys Asn His Arg Asn Arg Ser Lys Arg Ser Arg Leu Leu His Ser Asp
340 345 350Tyr Met Asn Met Thr Pro
Arg Arg Pro Gly Pro Thr Arg Lys His Tyr 355 360
365Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala Tyr Arg Ser
Arg Val 370 375 380Lys Phe Ser Arg Ser
Ala Asp Ala Pro Ala Tyr Gln Gln Gly Gln Asn385 390
395 400Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg
Arg Glu Glu Tyr Asp Val 405 410
415Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu Met Gly Gly Lys Pro Arg
420 425 430Arg Lys Asn Pro Gln
Glu Gly Leu Tyr Asn Glu Leu Gln Lys Asp Lys 435
440 445Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly
Glu Arg Arg Arg 450 455 460Gly Lys Gly
His Asp Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys465
470 475 480Asp Thr Tyr Asp Ala Leu His
Met Gln Ala Leu Pro Pro Arg 485
490371440DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 37atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccggatatcc agctcacgca atccccctca
agcttgagtg cctccgtggg cgaccgggtg 120tccttcacat gtcaggcaag ccaagacata
aataatttcc tgaattggta ccaacaaaaa 180cccggcaagg ctcccaaact cctgatttat
gatgcctcca atctggagac cggggtccct 240tctagattca gcggaagtgg cagcggcaca
gactttacat ttactatctc ttctctgcaa 300ccagaggaca tcgccacata ctattgccag
caatacggca atctgccctt caccttcgga 360ggcggaacca aggtagaaat taaaaggggc
ggtggaggct ccggaggggg gggctctggc 420ggagggggct cccaagtaca attgcaggag
tcagggcctg gactcgtgaa gccttcagaa 480actttgtcac tgacatgtac agtgtccggc
ggaagcattt ccagttacta ttggtcctgg 540attagacagc cacccggcaa aggactggaa
tggattggat atatctacta ctctggatct 600acaaactata atcccagcct caaatccagg
gtcactatta gtgtggatac atcaaagaat 660cagttctcct tgaagctgag ctcagtcact
gctgccgaca ccgcagtgta ctattgtgtg 720agcctggtct actgcggcgg agattgctac
agcggtttcg attactgggg ccagggcacc 780ctggttaccg ttagttccgc ggctgctctt
gataacgaga agtccaacgg tacgattatc 840cacgttaagg gtaagcacct ttgccctagc
ccgctgttcc caggccccag taagcccttt 900tgggtcctcg ttgtggtagg tggggtactc
gcctgctact ccctgctcgt cactgtcgca 960ttcatcatct tctgggtcag atccaaaaga
agccgcctgc tccatagcga ttacatgaat 1020atgactccac gccgccctgg ccccacaagg
aaacactacc agccttacgc accacctaga 1080gatttcgctg cctatcggag cagggtgaag
ttttccagat ctgcagatgc accagcgtat 1140cagcagggcc agaaccaact gtataacgag
ctcaacctgg gacgcaggga agagtatgac 1200gttttggaca agcgcagagg acgggaccct
gagatgggtg gcaaaccaag acgaaaaaac 1260ccccaggagg gtctctataa tgagctgcag
aaggataaga tggctgaagc ctattctgaa 1320ataggcatga aaggagagcg gagaagggga
aaagggcacg acggtttgta ccagggactc 1380agcactgcta cgaaggatac ttatgacgct
ctccacatgc aagccctgcc acctaggtaa 144038458PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
38Asp Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1
5 10 15Asp Arg Val Ser Phe Thr
Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe 20 25
30Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Leu Leu Ile 35 40 45Tyr Asp Ala
Ser Asn Leu Glu Thr Gly Val Pro Ser Arg Phe Ser Gly 50
55 60Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser
Ser Leu Gln Pro65 70 75
80Glu Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Gly Asn Leu Pro Phe
85 90 95Thr Phe Gly Gly Gly Thr
Lys Val Glu Ile Lys Arg Gly Gly Gly Gly 100
105 110Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gln
Val Gln Leu Gln 115 120 125Glu Ser
Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr 130
135 140Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr
Tyr Trp Ser Trp Ile145 150 155
160Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Tyr Ile Tyr Tyr
165 170 175Ser Gly Ser Thr
Asn Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile 180
185 190Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu
Lys Leu Ser Ser Val 195 200 205Thr
Ala Ala Asp Thr Ala Val Tyr Tyr Cys Val Ser Leu Val Tyr Cys 210
215 220Gly Gly Asp Cys Tyr Ser Gly Phe Asp Tyr
Trp Gly Gln Gly Thr Leu225 230 235
240Val Thr Val Ser Ser Ala Ala Ala Leu Asp Asn Glu Lys Ser Asn
Gly 245 250 255Thr Ile Ile
His Val Lys Gly Lys His Leu Cys Pro Ser Pro Leu Phe 260
265 270Pro Gly Pro Ser Lys Pro Phe Trp Val Leu
Val Val Val Gly Gly Val 275 280
285Leu Ala Cys Tyr Ser Leu Leu Val Thr Val Ala Phe Ile Ile Phe Trp 290
295 300Val Arg Ser Lys Arg Ser Arg Leu
Leu His Ser Asp Tyr Met Asn Met305 310
315 320Thr Pro Arg Arg Pro Gly Pro Thr Arg Lys His Tyr
Gln Pro Tyr Ala 325 330
335Pro Pro Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser Arg
340 345 350Ser Ala Asp Ala Pro Ala
Tyr Gln Gln Gly Gln Asn Gln Leu Tyr Asn 355 360
365Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr Asp Val Leu Asp
Lys Arg 370 375 380Arg Gly Arg Asp Pro
Glu Met Gly Gly Lys Pro Arg Arg Lys Asn Pro385 390
395 400Gln Glu Gly Leu Tyr Asn Glu Leu Gln Lys
Asp Lys Met Ala Glu Ala 405 410
415Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His
420 425 430Asp Gly Leu Tyr Gln
Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp 435
440 445Ala Leu His Met Gln Ala Leu Pro Pro Arg 450
455391374DNAArtificial SequenceDescription of Artificial
Sequence Synthetic polynucleotide 39gatatccagc tcacgcaatc cccctcaagc
ttgagtgcct ccgtgggcga ccgggtgtcc 60ttcacatgtc aggcaagcca agacataaat
aatttcctga attggtacca acaaaaaccc 120ggcaaggctc ccaaactcct gatttatgat
gcctccaatc tggagaccgg ggtcccttct 180agattcagcg gaagtggcag cggcacagac
tttacattta ctatctcttc tctgcaacca 240gaggacatcg ccacatacta ttgccagcaa
tacggcaatc tgcccttcac cttcggaggc 300ggaaccaagg tagaaattaa aaggggcggt
ggaggctccg gagggggggg ctctggcgga 360gggggctccc aagtacaatt gcaggagtca
gggcctggac tcgtgaagcc ttcagaaact 420ttgtcactga catgtacagt gtccggcgga
agcatttcca gttactattg gtcctggatt 480agacagccac ccggcaaagg actggaatgg
attggatata tctactactc tggatctaca 540aactataatc ccagcctcaa atccagggtc
actattagtg tggatacatc aaagaatcag 600ttctccttga agctgagctc agtcactgct
gccgacaccg cagtgtacta ttgtgtgagc 660ctggtctact gcggcggaga ttgctacagc
ggtttcgatt actggggcca gggcaccctg 720gttaccgtta gttccgcggc tgctcttgat
aacgagaagt ccaacggtac gattatccac 780gttaagggta agcacctttg ccctagcccg
ctgttcccag gccccagtaa gcccttttgg 840gtcctcgttg tggtaggtgg ggtactcgcc
tgctactccc tgctcgtcac tgtcgcattc 900atcatcttct gggtcagatc caaaagaagc
cgcctgctcc atagcgatta catgaatatg 960actccacgcc gccctggccc cacaaggaaa
cactaccagc cttacgcacc acctagagat 1020ttcgctgcct atcggagcag ggtgaagttt
tccagatctg cagatgcacc agcgtatcag 1080cagggccaga accaactgta taacgagctc
aacctgggac gcagggaaga gtatgacgtt 1140ttggacaagc gcagaggacg ggaccctgag
atgggtggca aaccaagacg aaaaaacccc 1200caggagggtc tctataatga gctgcagaag
gataagatgg ctgaagccta ttctgaaata 1260ggcatgaaag gagagcggag aaggggaaaa
gggcacgacg gtttgtacca gggactcagc 1320actgctacga aggatactta tgacgctctc
cacatgcaag ccctgccacc tagg 137440458PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
40Asp Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1
5 10 15Asp Arg Val Ser Phe Thr
Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe 20 25
30Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Leu Leu Ile 35 40 45Tyr Asp Ala
Ser Asn Leu Glu Thr Gly Val Pro Ser Arg Phe Ser Gly 50
55 60Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser
Ser Leu Gln Pro65 70 75
80Glu Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Gly Asn Leu Pro Phe
85 90 95Thr Phe Gly Gly Gly Thr
Lys Val Glu Ile Lys Arg Gly Gly Gly Gly 100
105 110Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gln
Val Gln Leu Gln 115 120 125Glu Ser
Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr 130
135 140Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr
Tyr Trp Ser Trp Ile145 150 155
160Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Tyr Ile Tyr Tyr
165 170 175Ser Gly Ser Thr
Asn Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile 180
185 190Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu
Lys Leu Ser Ser Val 195 200 205Thr
Ala Ala Asp Thr Ala Val Tyr Tyr Cys Val Ser Leu Val Tyr Cys 210
215 220Gly Gly Asp Cys Tyr Ser Gly Phe Asp Tyr
Trp Gly Gln Gly Thr Leu225 230 235
240Val Thr Val Ser Ser Ala Ala Ala Leu Asp Asn Glu Lys Ser Asn
Gly 245 250 255Thr Ile Ile
His Val Lys Gly Lys His Leu Cys Pro Ser Pro Leu Phe 260
265 270Pro Gly Pro Ser Lys Pro Phe Trp Val Leu
Val Val Val Gly Gly Val 275 280
285Leu Ala Cys Tyr Ser Leu Leu Val Thr Val Ala Phe Ile Ile Phe Trp 290
295 300Val Arg Ser Lys Arg Ser Arg Leu
Leu His Ser Asp Tyr Met Asn Met305 310
315 320Thr Pro Arg Arg Pro Gly Pro Thr Arg Lys His Tyr
Gln Pro Tyr Ala 325 330
335Pro Pro Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser Arg
340 345 350Ser Ala Asp Ala Pro Ala
Tyr Gln Gln Gly Gln Asn Gln Leu Tyr Asn 355 360
365Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr Asp Val Leu Asp
Lys Arg 370 375 380Arg Gly Arg Asp Pro
Glu Met Gly Gly Lys Pro Arg Arg Lys Asn Pro385 390
395 400Gln Glu Gly Leu Tyr Asn Glu Leu Gln Lys
Asp Lys Met Ala Glu Ala 405 410
415Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His
420 425 430Asp Gly Leu Tyr Gln
Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp 435
440 445Ala Leu His Met Gln Ala Leu Pro Pro Arg 450
455411467DNAArtificial SequenceDescription of Artificial
Sequence Synthetic polynucleotide 41atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccggatatcc agctgaccca gtctccatcc
tctttgagtg cctccgtggg tgaccgcgtc 120tctttcactt gccaagccag ccaagacatc
aacaactttc tgaattggta ccagcagaaa 180ccaggcaaag caccaaagct cctcatctac
gacgcctcca acctggaaac cggggtgccc 240agcaggttta gcgggagcgg ttctggcacg
gattttacgt tcaccatctc ctctctgcag 300cccgaggata tagctactta ttactgtcag
cagtacggga atctgccatt tacttttggg 360ggtggaacta aggtggaaat caaaaggggc
ggcgggggaa gcgggggcgg gggctcaggt 420ggcggaggga gccaggtgca actccaggaa
agtggcccag gattggtgaa gcccagcgag 480accctttccc ttacttgtac tgttagcgga
ggcagcataa gcagctacta ttggtcctgg 540atcagacagc caccagggaa agggcttgaa
tggattggct acatttacta ttccgggtcc 600accaactaca acccatccct caagtcccgc
gtgacaattt ccgtcgacac aagcaagaac 660cagttctccc tgaaacttag tagcgtcact
gctgcagata cagcagtgta ctattgtgtc 720agccttgtct actgtggcgg cgactgctac
agtggctttg attactgggg acagggcacg 780ctcgtgacag tgtccagcgc tgcggctatc
gaggtaatgt atccgccacc gtatctggac 840aacgagaagt ctaatgggac aatcattcac
gtgaagggga agcacctgtg tccatccccc 900ctgtttccgg gtcccagtaa acccttctgg
gtgcttgttg tcgttggcgg ggtgctggcc 960tgctattccc tgctggtgac cgtcgcgttt
attattttct gggttagatc caaaagaagc 1020cgcctgctcc atagcgatta catgaatatg
actccacgcc gccctggccc cacaaggaaa 1080cactaccagc cttacgcacc acctagagat
ttcgctgcct atcggagcag ggtgaagttt 1140tccagatctg cagatgcacc agcgtatcag
cagggccaga accaactgta taacgagctc 1200aacctgggac gcagggaaga gtatgacgtt
ttggacaagc gcagaggacg ggaccctgag 1260atgggtggca aaccaagacg aaaaaacccc
caggagggtc tctataatga gctgcagaag 1320gataagatgg ctgaagccta ttctgaaata
ggcatgaaag gagagcggag aaggggaaaa 1380gggcacgacg gtttgtacca gggactcagc
actgctacga aggatactta tgacgctctc 1440cacatgcaag ccctgccacc taggtaa
146742488PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
42Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Asp
Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu 20 25
30Ser Ala Ser Val Gly Asp Arg Val Ser Phe Thr Cys Gln
Ala Ser Gln 35 40 45Asp Ile Asn
Asn Phe Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala 50
55 60Pro Lys Leu Leu Ile Tyr Asp Ala Ser Asn Leu Glu
Thr Gly Val Pro65 70 75
80Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile
85 90 95Ser Ser Leu Gln Pro Glu
Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr 100
105 110Gly Asn Leu Pro Phe Thr Phe Gly Gly Gly Thr Lys
Val Glu Ile Lys 115 120 125Arg Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 130
135 140Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu
Val Lys Pro Ser Glu145 150 155
160Thr Leu Ser Leu Thr Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr
165 170 175Tyr Trp Ser Trp
Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile 180
185 190Gly Tyr Ile Tyr Tyr Ser Gly Ser Thr Asn Tyr
Asn Pro Ser Leu Lys 195 200 205Ser
Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu 210
215 220Lys Leu Ser Ser Val Thr Ala Ala Asp Thr
Ala Val Tyr Tyr Cys Val225 230 235
240Ser Leu Val Tyr Cys Gly Gly Asp Cys Tyr Ser Gly Phe Asp Tyr
Trp 245 250 255Gly Gln Gly
Thr Leu Val Thr Val Ser Ser Ala Ala Ala Ile Glu Val 260
265 270Met Tyr Pro Pro Pro Tyr Leu Asp Asn Glu
Lys Ser Asn Gly Thr Ile 275 280
285Ile His Val Lys Gly Lys His Leu Cys Pro Ser Pro Leu Phe Pro Gly 290
295 300Pro Ser Lys Pro Phe Trp Val Leu
Val Val Val Gly Gly Val Leu Ala305 310
315 320Cys Tyr Ser Leu Leu Val Thr Val Ala Phe Ile Ile
Phe Trp Val Arg 325 330
335Ser Lys Arg Ser Arg Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro
340 345 350Arg Arg Pro Gly Pro Thr
Arg Lys His Tyr Gln Pro Tyr Ala Pro Pro 355 360
365Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser Arg
Ser Ala 370 375 380Asp Ala Pro Ala Tyr
Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu385 390
395 400Asn Leu Gly Arg Arg Glu Glu Tyr Asp Val
Leu Asp Lys Arg Arg Gly 405 410
415Arg Asp Pro Glu Met Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu
420 425 430Gly Leu Tyr Asn Glu
Leu Gln Lys Asp Lys Met Ala Glu Ala Tyr Ser 435
440 445Glu Ile Gly Met Lys Gly Glu Arg Arg Arg Gly Lys
Gly His Asp Gly 450 455 460Leu Tyr Gln
Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu465
470 475 480His Met Gln Ala Leu Pro Pro
Arg 485431401DNAArtificial SequenceDescription of
Artificial Sequence Synthetic polynucleotide 43gatatccagc tgacccagtc
tccatcctct ttgagtgcct ccgtgggtga ccgcgtctct 60ttcacttgcc aagccagcca
agacatcaac aactttctga attggtacca gcagaaacca 120ggcaaagcac caaagctcct
catctacgac gcctccaacc tggaaaccgg ggtgcccagc 180aggtttagcg ggagcggttc
tggcacggat tttacgttca ccatctcctc tctgcagccc 240gaggatatag ctacttatta
ctgtcagcag tacgggaatc tgccatttac ttttgggggt 300ggaactaagg tggaaatcaa
aaggggcggc gggggaagcg ggggcggggg ctcaggtggc 360ggagggagcc aggtgcaact
ccaggaaagt ggcccaggat tggtgaagcc cagcgagacc 420ctttccctta cttgtactgt
tagcggaggc agcataagca gctactattg gtcctggatc 480agacagccac cagggaaagg
gcttgaatgg attggctaca tttactattc cgggtccacc 540aactacaacc catccctcaa
gtcccgcgtg acaatttccg tcgacacaag caagaaccag 600ttctccctga aacttagtag
cgtcactgct gcagatacag cagtgtacta ttgtgtcagc 660cttgtctact gtggcggcga
ctgctacagt ggctttgatt actggggaca gggcacgctc 720gtgacagtgt ccagcgctgc
ggctatcgag gtaatgtatc cgccaccgta tctggacaac 780gagaagtcta atgggacaat
cattcacgtg aaggggaagc acctgtgtcc atcccccctg 840tttccgggtc ccagtaaacc
cttctgggtg cttgttgtcg ttggcggggt gctggcctgc 900tattccctgc tggtgaccgt
cgcgtttatt attttctggg ttagatccaa aagaagccgc 960ctgctccata gcgattacat
gaatatgact ccacgccgcc ctggccccac aaggaaacac 1020taccagcctt acgcaccacc
tagagatttc gctgcctatc ggagcagggt gaagttttcc 1080agatctgcag atgcaccagc
gtatcagcag ggccagaacc aactgtataa cgagctcaac 1140ctgggacgca gggaagagta
tgacgttttg gacaagcgca gaggacggga ccctgagatg 1200ggtggcaaac caagacgaaa
aaacccccag gagggtctct ataatgagct gcagaaggat 1260aagatggctg aagcctattc
tgaaataggc atgaaaggag agcggagaag gggaaaaggg 1320cacgacggtt tgtaccaggg
actcagcact gctacgaagg atacttatga cgctctccac 1380atgcaagccc tgccacctag g
140144467PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
44Asp Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1
5 10 15Asp Arg Val Ser Phe Thr
Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe 20 25
30Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Leu Leu Ile 35 40 45Tyr Asp Ala
Ser Asn Leu Glu Thr Gly Val Pro Ser Arg Phe Ser Gly 50
55 60Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser
Ser Leu Gln Pro65 70 75
80Glu Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Gly Asn Leu Pro Phe
85 90 95Thr Phe Gly Gly Gly Thr
Lys Val Glu Ile Lys Arg Gly Gly Gly Gly 100
105 110Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gln
Val Gln Leu Gln 115 120 125Glu Ser
Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr 130
135 140Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr
Tyr Trp Ser Trp Ile145 150 155
160Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Tyr Ile Tyr Tyr
165 170 175Ser Gly Ser Thr
Asn Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile 180
185 190Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu
Lys Leu Ser Ser Val 195 200 205Thr
Ala Ala Asp Thr Ala Val Tyr Tyr Cys Val Ser Leu Val Tyr Cys 210
215 220Gly Gly Asp Cys Tyr Ser Gly Phe Asp Tyr
Trp Gly Gln Gly Thr Leu225 230 235
240Val Thr Val Ser Ser Ala Ala Ala Ile Glu Val Met Tyr Pro Pro
Pro 245 250 255Tyr Leu Asp
Asn Glu Lys Ser Asn Gly Thr Ile Ile His Val Lys Gly 260
265 270Lys His Leu Cys Pro Ser Pro Leu Phe Pro
Gly Pro Ser Lys Pro Phe 275 280
285Trp Val Leu Val Val Val Gly Gly Val Leu Ala Cys Tyr Ser Leu Leu 290
295 300Val Thr Val Ala Phe Ile Ile Phe
Trp Val Arg Ser Lys Arg Ser Arg305 310
315 320Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg
Arg Pro Gly Pro 325 330
335Thr Arg Lys His Tyr Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala
340 345 350Tyr Arg Ser Arg Val Lys
Phe Ser Arg Ser Ala Asp Ala Pro Ala Tyr 355 360
365Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly
Arg Arg 370 375 380Glu Glu Tyr Asp Val
Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu Met385 390
395 400Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln
Glu Gly Leu Tyr Asn Glu 405 410
415Leu Gln Lys Asp Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys
420 425 430Gly Glu Arg Arg Arg
Gly Lys Gly His Asp Gly Leu Tyr Gln Gly Leu 435
440 445Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu His
Met Gln Ala Leu 450 455 460Pro Pro
Arg465451548DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 45atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccggacattc aattgaccca gtcccctagc
agtctctcag caagtgtggg agatagggtg 120tcattcacct gtcaggcttc acaggacatc
aacaacttcc tcaattggta tcagcagaag 180ccagggaagg caccaaagct gctcatatat
gacgcttcaa accttgaaac cggagtacct 240agccgcttca gcggaagcgg atcagggact
gacttcactt ttaccatctc ttcactgcag 300cccgaagaca tcgccacata ctactgccag
cagtacggaa acttgccttt tacatttggg 360ggcggcacca aagtggagat taagcgaggg
ggaggcggct caggaggcgg tggctccgga 420ggcgggggtt cccaggtcca gctccaggaa
tccggcccag gtctggttaa gcccagtgaa 480actttgtccc tcacgtgtac tgtgagcggt
ggttcaatct cctcatacta ttggtcttgg 540atacggcaac ctcctggaaa gggcctcgag
tggatcggct atatctacta tagtggctcc 600actaattaca acccttccct caagtccaga
gtcaccattt ccgtggacac atctaagaac 660cagttcagtc tgaagttgtc cagcgttaca
gccgcagaca cagccgttta ttactgtgtg 720tctcttgttt actgcggggg agactgttat
agcggcttcg attactgggg ccagggcacc 780ttggtcacag tctcttccgc ggccgccctc
tctaacagta ttatgtactt ttctcatttt 840gtacccgtgt tccttcccgc taagccaact
actaccccgg ccccacggcc gcctacccct 900gcacccacaa tagccagtca gcctttgagc
ctgagacctg aggcttgtcg gccggctgct 960gggggtgcag tgcacacacg aggtcttgat
tttgcttgcg acatatacat ctgggcccct 1020ctggccggga cctgtggggt gctgcttctg
agcttggtca tcacgctcta ttgcaaccat 1080cgcaacagat ccaaaagaag ccgcctgctc
catagcgatt acatgaatat gactccacgc 1140cgccctggcc ccacaaggaa acactaccag
ccttacgcac cacctagaga tttcgctgcc 1200tatcggagca gggtgaagtt ttccagatct
gcagatgcac cagcgtatca gcagggccag 1260aaccaactgt ataacgagct caacctggga
cgcagggaag agtatgacgt tttggacaag 1320cgcagaggac gggaccctga gatgggtggc
aaaccaagac gaaaaaaccc ccaggagggt 1380ctctataatg agctgcagaa ggataagatg
gctgaagcct attctgaaat aggcatgaaa 1440ggagagcgga gaaggggaaa agggcacgac
ggtttgtacc agggactcag cactgctacg 1500aaggatactt atgacgctct ccacatgcaa
gccctgccac ctaggtaa 154846515PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
46Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Asp
Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu 20 25
30Ser Ala Ser Val Gly Asp Arg Val Ser Phe Thr Cys Gln
Ala Ser Gln 35 40 45Asp Ile Asn
Asn Phe Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala 50
55 60Pro Lys Leu Leu Ile Tyr Asp Ala Ser Asn Leu Glu
Thr Gly Val Pro65 70 75
80Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile
85 90 95Ser Ser Leu Gln Pro Glu
Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr 100
105 110Gly Asn Leu Pro Phe Thr Phe Gly Gly Gly Thr Lys
Val Glu Ile Lys 115 120 125Arg Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 130
135 140Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu
Val Lys Pro Ser Glu145 150 155
160Thr Leu Ser Leu Thr Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr
165 170 175Tyr Trp Ser Trp
Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile 180
185 190Gly Tyr Ile Tyr Tyr Ser Gly Ser Thr Asn Tyr
Asn Pro Ser Leu Lys 195 200 205Ser
Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu 210
215 220Lys Leu Ser Ser Val Thr Ala Ala Asp Thr
Ala Val Tyr Tyr Cys Val225 230 235
240Ser Leu Val Tyr Cys Gly Gly Asp Cys Tyr Ser Gly Phe Asp Tyr
Trp 245 250 255Gly Gln Gly
Thr Leu Val Thr Val Ser Ser Ala Ala Ala Leu Ser Asn 260
265 270Ser Ile Met Tyr Phe Ser His Phe Val Pro
Val Phe Leu Pro Ala Lys 275 280
285Pro Thr Thr Thr Pro Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile 290
295 300Ala Ser Gln Pro Leu Ser Leu Arg
Pro Glu Ala Cys Arg Pro Ala Ala305 310
315 320Gly Gly Ala Val His Thr Arg Gly Leu Asp Phe Ala
Cys Asp Ile Tyr 325 330
335Ile Trp Ala Pro Leu Ala Gly Thr Cys Gly Val Leu Leu Leu Ser Leu
340 345 350Val Ile Thr Leu Tyr Cys
Asn His Arg Asn Arg Ser Lys Arg Ser Arg 355 360
365Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro
Gly Pro 370 375 380Thr Arg Lys His Tyr
Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala385 390
395 400Tyr Arg Ser Arg Val Lys Phe Ser Arg Ser
Ala Asp Ala Pro Ala Tyr 405 410
415Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg
420 425 430Glu Glu Tyr Asp Val
Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu Met 435
440 445Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly
Leu Tyr Asn Glu 450 455 460Leu Gln Lys
Asp Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys465
470 475 480Gly Glu Arg Arg Arg Gly Lys
Gly His Asp Gly Leu Tyr Gln Gly Leu 485
490 495Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu His
Met Gln Ala Leu 500 505 510Pro
Pro Arg 515471482DNAArtificial SequenceDescription of Artificial
Sequence Synthetic polynucleotide 47gacattcaat tgacccagtc ccctagcagt
ctctcagcaa gtgtgggaga tagggtgtca 60ttcacctgtc aggcttcaca ggacatcaac
aacttcctca attggtatca gcagaagcca 120gggaaggcac caaagctgct catatatgac
gcttcaaacc ttgaaaccgg agtacctagc 180cgcttcagcg gaagcggatc agggactgac
ttcactttta ccatctcttc actgcagccc 240gaagacatcg ccacatacta ctgccagcag
tacggaaact tgccttttac atttgggggc 300ggcaccaaag tggagattaa gcgaggggga
ggcggctcag gaggcggtgg ctccggaggc 360gggggttccc aggtccagct ccaggaatcc
ggcccaggtc tggttaagcc cagtgaaact 420ttgtccctca cgtgtactgt gagcggtggt
tcaatctcct catactattg gtcttggata 480cggcaacctc ctggaaaggg cctcgagtgg
atcggctata tctactatag tggctccact 540aattacaacc cttccctcaa gtccagagtc
accatttccg tggacacatc taagaaccag 600ttcagtctga agttgtccag cgttacagcc
gcagacacag ccgtttatta ctgtgtgtct 660cttgtttact gcgggggaga ctgttatagc
ggcttcgatt actggggcca gggcaccttg 720gtcacagtct cttccgcggc cgccctctct
aacagtatta tgtacttttc tcattttgta 780cccgtgttcc ttcccgctaa gccaactact
accccggccc cacggccgcc tacccctgca 840cccacaatag ccagtcagcc tttgagcctg
agacctgagg cttgtcggcc ggctgctggg 900ggtgcagtgc acacacgagg tcttgatttt
gcttgcgaca tatacatctg ggcccctctg 960gccgggacct gtggggtgct gcttctgagc
ttggtcatca cgctctattg caaccatcgc 1020aacagatcca aaagaagccg cctgctccat
agcgattaca tgaatatgac tccacgccgc 1080cctggcccca caaggaaaca ctaccagcct
tacgcaccac ctagagattt cgctgcctat 1140cggagcaggg tgaagttttc cagatctgca
gatgcaccag cgtatcagca gggccagaac 1200caactgtata acgagctcaa cctgggacgc
agggaagagt atgacgtttt ggacaagcgc 1260agaggacggg accctgagat gggtggcaaa
ccaagacgaa aaaaccccca ggagggtctc 1320tataatgagc tgcagaagga taagatggct
gaagcctatt ctgaaatagg catgaaagga 1380gagcggagaa ggggaaaagg gcacgacggt
ttgtaccagg gactcagcac tgctacgaag 1440gatacttatg acgctctcca catgcaagcc
ctgccaccta gg 148248494PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
48Asp Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1
5 10 15Asp Arg Val Ser Phe Thr
Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe 20 25
30Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys
Leu Leu Ile 35 40 45Tyr Asp Ala
Ser Asn Leu Glu Thr Gly Val Pro Ser Arg Phe Ser Gly 50
55 60Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser
Ser Leu Gln Pro65 70 75
80Glu Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Gly Asn Leu Pro Phe
85 90 95Thr Phe Gly Gly Gly Thr
Lys Val Glu Ile Lys Arg Gly Gly Gly Gly 100
105 110Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gln
Val Gln Leu Gln 115 120 125Glu Ser
Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr 130
135 140Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Tyr
Tyr Trp Ser Trp Ile145 150 155
160Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Tyr Ile Tyr Tyr
165 170 175Ser Gly Ser Thr
Asn Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile 180
185 190Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu
Lys Leu Ser Ser Val 195 200 205Thr
Ala Ala Asp Thr Ala Val Tyr Tyr Cys Val Ser Leu Val Tyr Cys 210
215 220Gly Gly Asp Cys Tyr Ser Gly Phe Asp Tyr
Trp Gly Gln Gly Thr Leu225 230 235
240Val Thr Val Ser Ser Ala Ala Ala Leu Ser Asn Ser Ile Met Tyr
Phe 245 250 255Ser His Phe
Val Pro Val Phe Leu Pro Ala Lys Pro Thr Thr Thr Pro 260
265 270Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr
Ile Ala Ser Gln Pro Leu 275 280
285Ser Leu Arg Pro Glu Ala Cys Arg Pro Ala Ala Gly Gly Ala Val His 290
295 300Thr Arg Gly Leu Asp Phe Ala Cys
Asp Ile Tyr Ile Trp Ala Pro Leu305 310
315 320Ala Gly Thr Cys Gly Val Leu Leu Leu Ser Leu Val
Ile Thr Leu Tyr 325 330
335Cys Asn His Arg Asn Arg Ser Lys Arg Ser Arg Leu Leu His Ser Asp
340 345 350Tyr Met Asn Met Thr Pro
Arg Arg Pro Gly Pro Thr Arg Lys His Tyr 355 360
365Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala Tyr Arg Ser
Arg Val 370 375 380Lys Phe Ser Arg Ser
Ala Asp Ala Pro Ala Tyr Gln Gln Gly Gln Asn385 390
395 400Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg
Arg Glu Glu Tyr Asp Val 405 410
415Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu Met Gly Gly Lys Pro Arg
420 425 430Arg Lys Asn Pro Gln
Glu Gly Leu Tyr Asn Glu Leu Gln Lys Asp Lys 435
440 445Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly
Glu Arg Arg Arg 450 455 460Gly Lys Gly
His Asp Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys465
470 475 480Asp Thr Tyr Asp Ala Leu His
Met Gln Ala Leu Pro Pro Arg 485
49049372DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 49caggtacagc tgcaggaatc tgggcccgga
cttgtcaagc caagtcagac actttctctt 60acatgtaccg tgagcggcgg aagtataagc
agtggaggct tttactggtc ttggatacgg 120cagcacccag gcaaaggctt ggagtggatt
ggatacattc atcattcagg atctacacac 180tataatccat cccttaagtc ccgggtcacc
attagcattg atacgtctaa gaatctgttc 240agtctcaggc tgtcctccgt cactgctgcc
gacacagccg tgtactactg cgcctccttg 300gtttactgcg gaggcgactg ttatagcggc
tttgattatt gggggcaggg gaccctcgta 360accgtgagct ct
37250124PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
50Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Gln1
5 10 15Thr Leu Ser Leu Thr Cys
Thr Val Ser Gly Gly Ser Ile Ser Ser Gly 20 25
30Gly Phe Tyr Trp Ser Trp Ile Arg Gln His Pro Gly Lys
Gly Leu Glu 35 40 45Trp Ile Gly
Tyr Ile His His Ser Gly Ser Thr His Tyr Asn Pro Ser 50
55 60Leu Lys Ser Arg Val Thr Ile Ser Ile Asp Thr Ser
Lys Asn Leu Phe65 70 75
80Ser Leu Arg Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr
85 90 95Cys Ala Ser Leu Val Tyr
Cys Gly Gly Asp Cys Tyr Ser Gly Phe Asp 100
105 110Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser
115 120519PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 51Gly Gly Ser Ile Ser Ser Gly
Gly Phe1 5525PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 52His His Ser Gly Ser1
55314PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 53Leu Val Tyr Cys Gly Gly Asp Cys Tyr Ser Gly Phe
Asp Tyr1 5 1054324DNAArtificial
SequenceDescription of Artificial Sequence Synthetic polynucleotide
54gatatccagc tcactcaaag cccctctagt ctctctgcct cagtggggga tcgggtcagt
60tttacttgtc aagcttcaca ggatatcaac aacttcctta attggtatca gcagaagcca
120ggaaaagcac ccaagctgct catctatgat gcctcaaatt tggagacggg tgttcccagt
180cgattctctg ggtcagggtc cgggaccgac tttacgttta cgatctcctc tctgcagccc
240gaagacatcg ccacatacta ttgtcaacag tacggcaact tgcctttcac atttgggggc
300gggactaagg ttgaaatcaa gagg
32455108PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 55Asp Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu
Ser Ala Ser Val Gly1 5 10
15Asp Arg Val Ser Phe Thr Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe
20 25 30Leu Asn Trp Tyr Gln Gln Lys
Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40
45Tyr Asp Ala Ser Asn Leu Glu Thr Gly Val Pro Ser Arg Phe Ser
Gly 50 55 60Ser Gly Ser Gly Thr Asp
Phe Thr Phe Thr Ile Ser Ser Leu Gln Pro65 70
75 80Glu Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr
Gly Asn Leu Pro Phe 85 90
95Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys Arg 100
1055611PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 56Gln Ala Ser Gln Asp Ile Asn Asn Phe Leu Asn1
5 10577PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 57Asp Ala Ser Asn Leu Glu Thr1
5589PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 58Gln Gln Tyr Gly Asn Leu Pro Phe Thr1
5591446DNAArtificial SequenceDescription of Artificial
Sequence Synthetic polynucleotide 59atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccgcaggtac agctgcagga atctgggccc
ggacttgtca agccaagtca gacactttct 120cttacatgta ccgtgagcgg cggaagtata
agcagtggag gcttttactg gtcttggata 180cggcagcacc caggcaaagg cttggagtgg
attggataca ttcatcattc aggatctaca 240cactataatc catcccttaa gtcccgggtc
accattagca ttgatacgtc taagaatctg 300ttcagtctca ggctgtcctc cgtcactgct
gccgacacag ccgtgtacta ctgcgcctcc 360ttggtttact gcggaggcga ctgttatagc
ggctttgatt attgggggca ggggaccctc 420gtaaccgtga gctctggagg gggtgggagc
gggggaggag gttcaggggg gggcggctcc 480gatatccagc tcactcaaag cccctctagt
ctctctgcct cagtggggga tcgggtcagt 540tttacttgtc aagcttcaca ggatatcaac
aacttcctta attggtatca gcagaagcca 600ggaaaagcac ccaagctgct catctatgat
gcctcaaatt tggagacggg tgttcccagt 660cgattctctg ggtcagggtc cgggaccgac
tttacgttta cgatctcctc tctgcagccc 720gaagacatcg ccacatacta ttgtcaacag
tacggcaact tgcctttcac atttgggggc 780gggactaagg ttgaaatcaa gagggccgct
gcactggaca atgagaagtc caacggcacc 840atcatccacg tgaagggcaa gcacctgtgc
cctagtcctc tgttcccagg cccatccaaa 900cctttttggg ttcttgttgt ggtcgggggg
gtgctggcct gctattctct gctggtcacg 960gtggccttca taattttctg ggttagatcc
aaaagaagcc gcctgctcca tagcgattac 1020atgaatatga ctccacgccg ccctggcccc
acaaggaaac actaccagcc ttacgcacca 1080cctagagatt tcgctgccta tcggagcagg
gtgaagtttt ccagatctgc agatgcacca 1140gcgtatcagc agggccagaa ccaactgtat
aacgagctca acctgggacg cagggaagag 1200tatgacgttt tggacaagcg cagaggacgg
gaccctgaga tgggtggcaa accaagacga 1260aaaaaccccc aggagggtct ctataatgag
ctgcagaagg ataagatggc tgaagcctat 1320tctgaaatag gcatgaaagg agagcggaga
aggggaaaag ggcacgacgg tttgtaccag 1380ggactcagca ctgctacgaa ggatacttat
gacgctctcc acatgcaagc cctgccacct 1440aggtaa
144660481PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
60Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Gln
Val Gln Leu Gln Glu Ser Gly Pro Gly Leu 20 25
30Val Lys Pro Ser Gln Thr Leu Ser Leu Thr Cys Thr Val
Ser Gly Gly 35 40 45Ser Ile Ser
Ser Gly Gly Phe Tyr Trp Ser Trp Ile Arg Gln His Pro 50
55 60Gly Lys Gly Leu Glu Trp Ile Gly Tyr Ile His His
Ser Gly Ser Thr65 70 75
80His Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Ile Asp Thr
85 90 95Ser Lys Asn Leu Phe Ser
Leu Arg Leu Ser Ser Val Thr Ala Ala Asp 100
105 110Thr Ala Val Tyr Tyr Cys Ala Ser Leu Val Tyr Cys
Gly Gly Asp Cys 115 120 125Tyr Ser
Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser 130
135 140Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
Gly Gly Gly Gly Ser145 150 155
160Asp Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly
165 170 175Asp Arg Val Ser
Phe Thr Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe 180
185 190Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala
Pro Lys Leu Leu Ile 195 200 205Tyr
Asp Ala Ser Asn Leu Glu Thr Gly Val Pro Ser Arg Phe Ser Gly 210
215 220Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr
Ile Ser Ser Leu Gln Pro225 230 235
240Glu Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Gly Asn Leu Pro
Phe 245 250 255Thr Phe Gly
Gly Gly Thr Lys Val Glu Ile Lys Arg Ala Ala Ala Leu 260
265 270Asp Asn Glu Lys Ser Asn Gly Thr Ile Ile
His Val Lys Gly Lys His 275 280
285Leu Cys Pro Ser Pro Leu Phe Pro Gly Pro Ser Lys Pro Phe Trp Val 290
295 300Leu Val Val Val Gly Gly Val Leu
Ala Cys Tyr Ser Leu Leu Val Thr305 310
315 320Val Ala Phe Ile Ile Phe Trp Val Arg Ser Lys Arg
Ser Arg Leu Leu 325 330
335His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro Gly Pro Thr Arg
340 345 350Lys His Tyr Gln Pro Tyr
Ala Pro Pro Arg Asp Phe Ala Ala Tyr Arg 355 360
365Ser Arg Val Lys Phe Ser Arg Ser Ala Asp Ala Pro Ala Tyr
Gln Gln 370 375 380Gly Gln Asn Gln Leu
Tyr Asn Glu Leu Asn Leu Gly Arg Arg Glu Glu385 390
395 400Tyr Asp Val Leu Asp Lys Arg Arg Gly Arg
Asp Pro Glu Met Gly Gly 405 410
415Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu Tyr Asn Glu Leu Gln
420 425 430Lys Asp Lys Met Ala
Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly Glu 435
440 445Arg Arg Arg Gly Lys Gly His Asp Gly Leu Tyr Gln
Gly Leu Ser Thr 450 455 460Ala Thr Lys
Asp Thr Tyr Asp Ala Leu His Met Gln Ala Leu Pro Pro465
470 475 480Arg611380DNAArtificial
SequenceDescription of Artificial Sequence Synthetic polynucleotide
61caggtacagc tgcaggaatc tgggcccgga cttgtcaagc caagtcagac actttctctt
60acatgtaccg tgagcggcgg aagtataagc agtggaggct tttactggtc ttggatacgg
120cagcacccag gcaaaggctt ggagtggatt ggatacattc atcattcagg atctacacac
180tataatccat cccttaagtc ccgggtcacc attagcattg atacgtctaa gaatctgttc
240agtctcaggc tgtcctccgt cactgctgcc gacacagccg tgtactactg cgcctccttg
300gtttactgcg gaggcgactg ttatagcggc tttgattatt gggggcaggg gaccctcgta
360accgtgagct ctggaggggg tgggagcggg ggaggaggtt cagggggggg cggctccgat
420atccagctca ctcaaagccc ctctagtctc tctgcctcag tgggggatcg ggtcagtttt
480acttgtcaag cttcacagga tatcaacaac ttccttaatt ggtatcagca gaagccagga
540aaagcaccca agctgctcat ctatgatgcc tcaaatttgg agacgggtgt tcccagtcga
600ttctctgggt cagggtccgg gaccgacttt acgtttacga tctcctctct gcagcccgaa
660gacatcgcca catactattg tcaacagtac ggcaacttgc ctttcacatt tgggggcggg
720actaaggttg aaatcaagag ggccgctgca ctggacaatg agaagtccaa cggcaccatc
780atccacgtga agggcaagca cctgtgccct agtcctctgt tcccaggccc atccaaacct
840ttttgggttc ttgttgtggt cgggggggtg ctggcctgct attctctgct ggtcacggtg
900gccttcataa ttttctgggt tagatccaaa agaagccgcc tgctccatag cgattacatg
960aatatgactc cacgccgccc tggccccaca aggaaacact accagcctta cgcaccacct
1020agagatttcg ctgcctatcg gagcagggtg aagttttcca gatctgcaga tgcaccagcg
1080tatcagcagg gccagaacca actgtataac gagctcaacc tgggacgcag ggaagagtat
1140gacgttttgg acaagcgcag aggacgggac cctgagatgg gtggcaaacc aagacgaaaa
1200aacccccagg agggtctcta taatgagctg cagaaggata agatggctga agcctattct
1260gaaataggca tgaaaggaga gcggagaagg ggaaaagggc acgacggttt gtaccaggga
1320ctcagcactg ctacgaagga tacttatgac gctctccaca tgcaagccct gccacctagg
138062460PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 62Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu
Val Lys Pro Ser Gln1 5 10
15Thr Leu Ser Leu Thr Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Gly
20 25 30Gly Phe Tyr Trp Ser Trp Ile
Arg Gln His Pro Gly Lys Gly Leu Glu 35 40
45Trp Ile Gly Tyr Ile His His Ser Gly Ser Thr His Tyr Asn Pro
Ser 50 55 60Leu Lys Ser Arg Val Thr
Ile Ser Ile Asp Thr Ser Lys Asn Leu Phe65 70
75 80Ser Leu Arg Leu Ser Ser Val Thr Ala Ala Asp
Thr Ala Val Tyr Tyr 85 90
95Cys Ala Ser Leu Val Tyr Cys Gly Gly Asp Cys Tyr Ser Gly Phe Asp
100 105 110Tyr Trp Gly Gln Gly Thr
Leu Val Thr Val Ser Ser Gly Gly Gly Gly 115 120
125Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Asp Ile Gln
Leu Thr 130 135 140Gln Ser Pro Ser Ser
Leu Ser Ala Ser Val Gly Asp Arg Val Ser Phe145 150
155 160Thr Cys Gln Ala Ser Gln Asp Ile Asn Asn
Phe Leu Asn Trp Tyr Gln 165 170
175Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Asp Ala Ser Asn
180 185 190Leu Glu Thr Gly Val
Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr 195
200 205Asp Phe Thr Phe Thr Ile Ser Ser Leu Gln Pro Glu
Asp Ile Ala Thr 210 215 220Tyr Tyr Cys
Gln Gln Tyr Gly Asn Leu Pro Phe Thr Phe Gly Gly Gly225
230 235 240Thr Lys Val Glu Ile Lys Arg
Ala Ala Ala Leu Asp Asn Glu Lys Ser 245
250 255Asn Gly Thr Ile Ile His Val Lys Gly Lys His Leu
Cys Pro Ser Pro 260 265 270Leu
Phe Pro Gly Pro Ser Lys Pro Phe Trp Val Leu Val Val Val Gly 275
280 285Gly Val Leu Ala Cys Tyr Ser Leu Leu
Val Thr Val Ala Phe Ile Ile 290 295
300Phe Trp Val Arg Ser Lys Arg Ser Arg Leu Leu His Ser Asp Tyr Met305
310 315 320Asn Met Thr Pro
Arg Arg Pro Gly Pro Thr Arg Lys His Tyr Gln Pro 325
330 335Tyr Ala Pro Pro Arg Asp Phe Ala Ala Tyr
Arg Ser Arg Val Lys Phe 340 345
350Ser Arg Ser Ala Asp Ala Pro Ala Tyr Gln Gln Gly Gln Asn Gln Leu
355 360 365Tyr Asn Glu Leu Asn Leu Gly
Arg Arg Glu Glu Tyr Asp Val Leu Asp 370 375
380Lys Arg Arg Gly Arg Asp Pro Glu Met Gly Gly Lys Pro Arg Arg
Lys385 390 395 400Asn Pro
Gln Glu Gly Leu Tyr Asn Glu Leu Gln Lys Asp Lys Met Ala
405 410 415Glu Ala Tyr Ser Glu Ile Gly
Met Lys Gly Glu Arg Arg Arg Gly Lys 420 425
430Gly His Asp Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys
Asp Thr 435 440 445Tyr Asp Ala Leu
His Met Gln Ala Leu Pro Pro Arg 450 455
460631473DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 63atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccgcaggtgc agctgcagga aagcggtccg
ggacttgtca agccgtccca aacgctgagt 120ctgacgtgta ctgtctctgg tggctctatt
tcttccgggg gcttttattg gtcttggatc 180agacaacacc ctggcaaagg gctggagtgg
atagggtata ttcaccactc tgggtccact 240cactacaacc catcattgaa atccagagtg
actatctcaa tcgacacatc caagaacctt 300ttcagcctga ggttgtcatc agttaccgcc
gctgacaccg cggtgtatta ttgcgcctct 360ctcgtgtact gcggtggcga ttgttatagt
ggctttgact actgggggca ggggacattg 420gttaccgttt caagtggagg cggtgggtct
ggcgggggcg gtagcggagg tggggggagc 480gacatacagc ttacgcagag cccctccagc
ctttcagcct ccgtggggga tagggtgtcc 540tttacctgcc aggcttccca ggacataaac
aacttcctca attggtatca gcaaaagccc 600gggaaagcac caaagctgct catctacgat
gccagcaacc tggaaaccgg agtgccgtct 660cgcttctctg gaagtggcag tgggaccgat
ttcactttta caatctcaag tttgcagcca 720gaagacattg caacatacta ctgtcaacag
tacggcaatc tcccctttac atttgggggg 780ggaactaaag tggagattaa gcgcgctgca
gccattgaag ttatgtatcc gcccccgtat 840ctggataacg agaaatctaa tggtaccata
atacatgtga aggggaagca cctctgtcca 900tcaccgctgt tccccggccc ttcaaaacct
ttctgggtac tcgttgtcgt gggtggagtt 960ctggcctgct atagtctgct ggtgaccgtg
gcgtttatca tcttctgggt aagatccaaa 1020agaagccgcc tgctccatag cgattacatg
aatatgactc cacgccgccc tggccccaca 1080aggaaacact accagcctta cgcaccacct
agagatttcg ctgcctatcg gagcagggtg 1140aagttttcca gatctgcaga tgcaccagcg
tatcagcagg gccagaacca actgtataac 1200gagctcaacc tgggacgcag ggaagagtat
gacgttttgg acaagcgcag aggacgggac 1260cctgagatgg gtggcaaacc aagacgaaaa
aacccccagg agggtctcta taatgagctg 1320cagaaggata agatggctga agcctattct
gaaataggca tgaaaggaga gcggagaagg 1380ggaaaagggc acgacggttt gtaccaggga
ctcagcactg ctacgaagga tacttatgac 1440gctctccaca tgcaagccct gccacctagg
taa 147364490PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
64Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Gln
Val Gln Leu Gln Glu Ser Gly Pro Gly Leu 20 25
30Val Lys Pro Ser Gln Thr Leu Ser Leu Thr Cys Thr Val
Ser Gly Gly 35 40 45Ser Ile Ser
Ser Gly Gly Phe Tyr Trp Ser Trp Ile Arg Gln His Pro 50
55 60Gly Lys Gly Leu Glu Trp Ile Gly Tyr Ile His His
Ser Gly Ser Thr65 70 75
80His Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Ile Asp Thr
85 90 95Ser Lys Asn Leu Phe Ser
Leu Arg Leu Ser Ser Val Thr Ala Ala Asp 100
105 110Thr Ala Val Tyr Tyr Cys Ala Ser Leu Val Tyr Cys
Gly Gly Asp Cys 115 120 125Tyr Ser
Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser 130
135 140Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
Gly Gly Gly Gly Ser145 150 155
160Asp Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly
165 170 175Asp Arg Val Ser
Phe Thr Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe 180
185 190Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala
Pro Lys Leu Leu Ile 195 200 205Tyr
Asp Ala Ser Asn Leu Glu Thr Gly Val Pro Ser Arg Phe Ser Gly 210
215 220Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr
Ile Ser Ser Leu Gln Pro225 230 235
240Glu Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Gly Asn Leu Pro
Phe 245 250 255Thr Phe Gly
Gly Gly Thr Lys Val Glu Ile Lys Arg Ala Ala Ala Ile 260
265 270Glu Val Met Tyr Pro Pro Pro Tyr Leu Asp
Asn Glu Lys Ser Asn Gly 275 280
285Thr Ile Ile His Val Lys Gly Lys His Leu Cys Pro Ser Pro Leu Phe 290
295 300Pro Gly Pro Ser Lys Pro Phe Trp
Val Leu Val Val Val Gly Gly Val305 310
315 320Leu Ala Cys Tyr Ser Leu Leu Val Thr Val Ala Phe
Ile Ile Phe Trp 325 330
335Val Arg Ser Lys Arg Ser Arg Leu Leu His Ser Asp Tyr Met Asn Met
340 345 350Thr Pro Arg Arg Pro Gly
Pro Thr Arg Lys His Tyr Gln Pro Tyr Ala 355 360
365Pro Pro Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val Lys Phe
Ser Arg 370 375 380Ser Ala Asp Ala Pro
Ala Tyr Gln Gln Gly Gln Asn Gln Leu Tyr Asn385 390
395 400Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr
Asp Val Leu Asp Lys Arg 405 410
415Arg Gly Arg Asp Pro Glu Met Gly Gly Lys Pro Arg Arg Lys Asn Pro
420 425 430Gln Glu Gly Leu Tyr
Asn Glu Leu Gln Lys Asp Lys Met Ala Glu Ala 435
440 445Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg Arg Arg
Gly Lys Gly His 450 455 460Asp Gly Leu
Tyr Gln Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp465
470 475 480Ala Leu His Met Gln Ala Leu
Pro Pro Arg 485 490651407DNAArtificial
SequenceDescription of Artificial Sequence Synthetic polynucleotide
65caggtgcagc tgcaggaaag cggtccggga cttgtcaagc cgtcccaaac gctgagtctg
60acgtgtactg tctctggtgg ctctatttct tccgggggct tttattggtc ttggatcaga
120caacaccctg gcaaagggct ggagtggata gggtatattc accactctgg gtccactcac
180tacaacccat cattgaaatc cagagtgact atctcaatcg acacatccaa gaaccttttc
240agcctgaggt tgtcatcagt taccgccgct gacaccgcgg tgtattattg cgcctctctc
300gtgtactgcg gtggcgattg ttatagtggc tttgactact gggggcaggg gacattggtt
360accgtttcaa gtggaggcgg tgggtctggc gggggcggta gcggaggtgg ggggagcgac
420atacagctta cgcagagccc ctccagcctt tcagcctccg tgggggatag ggtgtccttt
480acctgccagg cttcccagga cataaacaac ttcctcaatt ggtatcagca aaagcccggg
540aaagcaccaa agctgctcat ctacgatgcc agcaacctgg aaaccggagt gccgtctcgc
600ttctctggaa gtggcagtgg gaccgatttc acttttacaa tctcaagttt gcagccagaa
660gacattgcaa catactactg tcaacagtac ggcaatctcc cctttacatt tgggggggga
720actaaagtgg agattaagcg cgctgcagcc attgaagtta tgtatccgcc cccgtatctg
780gataacgaga aatctaatgg taccataata catgtgaagg ggaagcacct ctgtccatca
840ccgctgttcc ccggcccttc aaaacctttc tgggtactcg ttgtcgtggg tggagttctg
900gcctgctata gtctgctggt gaccgtggcg tttatcatct tctgggtaag atccaaaaga
960agccgcctgc tccatagcga ttacatgaat atgactccac gccgccctgg ccccacaagg
1020aaacactacc agccttacgc accacctaga gatttcgctg cctatcggag cagggtgaag
1080ttttccagat ctgcagatgc accagcgtat cagcagggcc agaaccaact gtataacgag
1140ctcaacctgg gacgcaggga agagtatgac gttttggaca agcgcagagg acgggaccct
1200gagatgggtg gcaaaccaag acgaaaaaac ccccaggagg gtctctataa tgagctgcag
1260aaggataaga tggctgaagc ctattctgaa ataggcatga aaggagagcg gagaagggga
1320aaagggcacg acggtttgta ccagggactc agcactgcta cgaaggatac ttatgacgct
1380ctccacatgc aagccctgcc acctagg
140766469PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 66Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu
Val Lys Pro Ser Gln1 5 10
15Thr Leu Ser Leu Thr Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Gly
20 25 30Gly Phe Tyr Trp Ser Trp Ile
Arg Gln His Pro Gly Lys Gly Leu Glu 35 40
45Trp Ile Gly Tyr Ile His His Ser Gly Ser Thr His Tyr Asn Pro
Ser 50 55 60Leu Lys Ser Arg Val Thr
Ile Ser Ile Asp Thr Ser Lys Asn Leu Phe65 70
75 80Ser Leu Arg Leu Ser Ser Val Thr Ala Ala Asp
Thr Ala Val Tyr Tyr 85 90
95Cys Ala Ser Leu Val Tyr Cys Gly Gly Asp Cys Tyr Ser Gly Phe Asp
100 105 110Tyr Trp Gly Gln Gly Thr
Leu Val Thr Val Ser Ser Gly Gly Gly Gly 115 120
125Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Asp Ile Gln
Leu Thr 130 135 140Gln Ser Pro Ser Ser
Leu Ser Ala Ser Val Gly Asp Arg Val Ser Phe145 150
155 160Thr Cys Gln Ala Ser Gln Asp Ile Asn Asn
Phe Leu Asn Trp Tyr Gln 165 170
175Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Asp Ala Ser Asn
180 185 190Leu Glu Thr Gly Val
Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr 195
200 205Asp Phe Thr Phe Thr Ile Ser Ser Leu Gln Pro Glu
Asp Ile Ala Thr 210 215 220Tyr Tyr Cys
Gln Gln Tyr Gly Asn Leu Pro Phe Thr Phe Gly Gly Gly225
230 235 240Thr Lys Val Glu Ile Lys Arg
Ala Ala Ala Ile Glu Val Met Tyr Pro 245
250 255Pro Pro Tyr Leu Asp Asn Glu Lys Ser Asn Gly Thr
Ile Ile His Val 260 265 270Lys
Gly Lys His Leu Cys Pro Ser Pro Leu Phe Pro Gly Pro Ser Lys 275
280 285Pro Phe Trp Val Leu Val Val Val Gly
Gly Val Leu Ala Cys Tyr Ser 290 295
300Leu Leu Val Thr Val Ala Phe Ile Ile Phe Trp Val Arg Ser Lys Arg305
310 315 320Ser Arg Leu Leu
His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro 325
330 335Gly Pro Thr Arg Lys His Tyr Gln Pro Tyr
Ala Pro Pro Arg Asp Phe 340 345
350Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser Arg Ser Ala Asp Ala Pro
355 360 365Ala Tyr Gln Gln Gly Gln Asn
Gln Leu Tyr Asn Glu Leu Asn Leu Gly 370 375
380Arg Arg Glu Glu Tyr Asp Val Leu Asp Lys Arg Arg Gly Arg Asp
Pro385 390 395 400Glu Met
Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu Tyr
405 410 415Asn Glu Leu Gln Lys Asp Lys
Met Ala Glu Ala Tyr Ser Glu Ile Gly 420 425
430Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His Asp Gly Leu
Tyr Gln 435 440 445Gly Leu Ser Thr
Ala Thr Lys Asp Thr Tyr Asp Ala Leu His Met Gln 450
455 460Ala Leu Pro Pro Arg465671554DNAArtificial
SequenceDescription of Artificial Sequence Synthetic polynucleotide
67atggcactcc ccgtaactgc tctgctgctg ccgttggcat tgctcctgca cgccgcacgc
60ccgcaggtgc agttgcagga aagcgggcct ggccttgtga aaccaagcca gacactgagc
120ctgacatgca ctgtgtccgg cgggtccata tcttccgggg gtttttattg gtcctggata
180cgccagcatc ccgggaaagg acttgaatgg attggatata tccaccattc cggaagcacc
240cactacaatc caagccttaa atcccgggtg acaatctcca tcgacacctc aaagaatctt
300ttttccctgc ggttgtcttc agtaactgcc gccgataccg ctgtgtacta ctgtgccagc
360ctcgtctatt gcggcggaga ttgttattct gggttcgatt attggggtca aggcacactg
420gtaactgtca gcagcggagg cggcggttcc gggggcgggg gcagtggagg gggcggatct
480gacattcagc ttacgcagtc cccatcttca cttagcgcca gcgttggcga tcgggtcagc
540ttcacgtgtc aagcaagtca ggatatcaac aactttctta actggtacca gcagaagcca
600ggcaaggcac ccaagttgct gatttacgat gcttctaacc tcgagacggg agtgcctagc
660cgcttctccg ggagcggcag cggcacagac tttaccttta cgatttccag tctgcagcca
720gaggatatag caacttatta ctgtcagcag tatggcaacc tcccttttac cttcggtggt
780ggcacaaagg tcgagattaa aagagccgca gcgttgtcca actccataat gtatttttct
840cattttgtgc ccgtctttct gcctgccaaa cctaccacca cccccgcccc acgaccacct
900actccagccc ccaccatcgc ctcccagccc ctcagcctga ggccagaggc ttgtcgccct
960gctgcggggg gcgctgtcca taccagagga ctcgacttcg cctgcgatat ttatatatgg
1020gcccccctcg ccggcacctg cggagtcttg ctcctgagcc ttgtgatcac gctttattgt
1080aaccatcgga atagatccaa aagaagccgc ctgctccata gcgattacat gaatatgact
1140ccacgccgcc ctggccccac aaggaaacac taccagcctt acgcaccacc tagagatttc
1200gctgcctatc ggagcagggt gaagttttcc agatctgcag atgcaccagc gtatcagcag
1260ggccagaacc aactgtataa cgagctcaac ctgggacgca gggaagagta tgacgttttg
1320gacaagcgca gaggacggga ccctgagatg ggtggcaaac caagacgaaa aaacccccag
1380gagggtctct ataatgagct gcagaaggat aagatggctg aagcctattc tgaaataggc
1440atgaaaggag agcggagaag gggaaaaggg cacgacggtt tgtaccaggg actcagcact
1500gctacgaagg atacttatga cgctctccac atgcaagccc tgccacctag gtaa
155468517PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 68Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro
Leu Ala Leu Leu Leu1 5 10
15His Ala Ala Arg Pro Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu
20 25 30Val Lys Pro Ser Gln Thr Leu
Ser Leu Thr Cys Thr Val Ser Gly Gly 35 40
45Ser Ile Ser Ser Gly Gly Phe Tyr Trp Ser Trp Ile Arg Gln His
Pro 50 55 60Gly Lys Gly Leu Glu Trp
Ile Gly Tyr Ile His His Ser Gly Ser Thr65 70
75 80His Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr
Ile Ser Ile Asp Thr 85 90
95Ser Lys Asn Leu Phe Ser Leu Arg Leu Ser Ser Val Thr Ala Ala Asp
100 105 110Thr Ala Val Tyr Tyr Cys
Ala Ser Leu Val Tyr Cys Gly Gly Asp Cys 115 120
125Tyr Ser Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr
Val Ser 130 135 140Ser Gly Gly Gly Gly
Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser145 150
155 160Asp Ile Gln Leu Thr Gln Ser Pro Ser Ser
Leu Ser Ala Ser Val Gly 165 170
175Asp Arg Val Ser Phe Thr Cys Gln Ala Ser Gln Asp Ile Asn Asn Phe
180 185 190Leu Asn Trp Tyr Gln
Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 195
200 205Tyr Asp Ala Ser Asn Leu Glu Thr Gly Val Pro Ser
Arg Phe Ser Gly 210 215 220Ser Gly Ser
Gly Thr Asp Phe Thr Phe Thr Ile Ser Ser Leu Gln Pro225
230 235 240Glu Asp Ile Ala Thr Tyr Tyr
Cys Gln Gln Tyr Gly Asn Leu Pro Phe 245
250 255Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys Arg
Ala Ala Ala Leu 260 265 270Ser
Asn Ser Ile Met Tyr Phe Ser His Phe Val Pro Val Phe Leu Pro 275
280 285Ala Lys Pro Thr Thr Thr Pro Ala Pro
Arg Pro Pro Thr Pro Ala Pro 290 295
300Thr Ile Ala Ser Gln Pro Leu Ser Leu Arg Pro Glu Ala Cys Arg Pro305
310 315 320Ala Ala Gly Gly
Ala Val His Thr Arg Gly Leu Asp Phe Ala Cys Asp 325
330 335Ile Tyr Ile Trp Ala Pro Leu Ala Gly Thr
Cys Gly Val Leu Leu Leu 340 345
350Ser Leu Val Ile Thr Leu Tyr Cys Asn His Arg Asn Arg Ser Lys Arg
355 360 365Ser Arg Leu Leu His Ser Asp
Tyr Met Asn Met Thr Pro Arg Arg Pro 370 375
380Gly Pro Thr Arg Lys His Tyr Gln Pro Tyr Ala Pro Pro Arg Asp
Phe385 390 395 400Ala Ala
Tyr Arg Ser Arg Val Lys Phe Ser Arg Ser Ala Asp Ala Pro
405 410 415Ala Tyr Gln Gln Gly Gln Asn
Gln Leu Tyr Asn Glu Leu Asn Leu Gly 420 425
430Arg Arg Glu Glu Tyr Asp Val Leu Asp Lys Arg Arg Gly Arg
Asp Pro 435 440 445Glu Met Gly Gly
Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu Tyr 450
455 460Asn Glu Leu Gln Lys Asp Lys Met Ala Glu Ala Tyr
Ser Glu Ile Gly465 470 475
480Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His Asp Gly Leu Tyr Gln
485 490 495Gly Leu Ser Thr Ala
Thr Lys Asp Thr Tyr Asp Ala Leu His Met Gln 500
505 510Ala Leu Pro Pro Arg 515691488DNAArtificial
SequenceDescription of Artificial Sequence Synthetic polynucleotide
69caggtgcagt tgcaggaaag cgggcctggc cttgtgaaac caagccagac actgagcctg
60acatgcactg tgtccggcgg gtccatatct tccgggggtt tttattggtc ctggatacgc
120cagcatcccg ggaaaggact tgaatggatt ggatatatcc accattccgg aagcacccac
180tacaatccaa gccttaaatc ccgggtgaca atctccatcg acacctcaaa gaatcttttt
240tccctgcggt tgtcttcagt aactgccgcc gataccgctg tgtactactg tgccagcctc
300gtctattgcg gcggagattg ttattctggg ttcgattatt ggggtcaagg cacactggta
360actgtcagca gcggaggcgg cggttccggg ggcgggggca gtggaggggg cggatctgac
420attcagctta cgcagtcccc atcttcactt agcgccagcg ttggcgatcg ggtcagcttc
480acgtgtcaag caagtcagga tatcaacaac tttcttaact ggtaccagca gaagccaggc
540aaggcaccca agttgctgat ttacgatgct tctaacctcg agacgggagt gcctagccgc
600ttctccggga gcggcagcgg cacagacttt acctttacga tttccagtct gcagccagag
660gatatagcaa cttattactg tcagcagtat ggcaacctcc cttttacctt cggtggtggc
720acaaaggtcg agattaaaag agccgcagcg ttgtccaact ccataatgta tttttctcat
780tttgtgcccg tctttctgcc tgccaaacct accaccaccc ccgccccacg accacctact
840ccagccccca ccatcgcctc ccagcccctc agcctgaggc cagaggcttg tcgccctgct
900gcggggggcg ctgtccatac cagaggactc gacttcgcct gcgatattta tatatgggcc
960cccctcgccg gcacctgcgg agtcttgctc ctgagccttg tgatcacgct ttattgtaac
1020catcggaata gatccaaaag aagccgcctg ctccatagcg attacatgaa tatgactcca
1080cgccgccctg gccccacaag gaaacactac cagccttacg caccacctag agatttcgct
1140gcctatcgga gcagggtgaa gttttccaga tctgcagatg caccagcgta tcagcagggc
1200cagaaccaac tgtataacga gctcaacctg ggacgcaggg aagagtatga cgttttggac
1260aagcgcagag gacgggaccc tgagatgggt ggcaaaccaa gacgaaaaaa cccccaggag
1320ggtctctata atgagctgca gaaggataag atggctgaag cctattctga aataggcatg
1380aaaggagagc ggagaagggg aaaagggcac gacggtttgt accagggact cagcactgct
1440acgaaggata cttatgacgc tctccacatg caagccctgc cacctagg
148870496PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 70Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu
Val Lys Pro Ser Gln1 5 10
15Thr Leu Ser Leu Thr Cys Thr Val Ser Gly Gly Ser Ile Ser Ser Gly
20 25 30Gly Phe Tyr Trp Ser Trp Ile
Arg Gln His Pro Gly Lys Gly Leu Glu 35 40
45Trp Ile Gly Tyr Ile His His Ser Gly Ser Thr His Tyr Asn Pro
Ser 50 55 60Leu Lys Ser Arg Val Thr
Ile Ser Ile Asp Thr Ser Lys Asn Leu Phe65 70
75 80Ser Leu Arg Leu Ser Ser Val Thr Ala Ala Asp
Thr Ala Val Tyr Tyr 85 90
95Cys Ala Ser Leu Val Tyr Cys Gly Gly Asp Cys Tyr Ser Gly Phe Asp
100 105 110Tyr Trp Gly Gln Gly Thr
Leu Val Thr Val Ser Ser Gly Gly Gly Gly 115 120
125Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Asp Ile Gln
Leu Thr 130 135 140Gln Ser Pro Ser Ser
Leu Ser Ala Ser Val Gly Asp Arg Val Ser Phe145 150
155 160Thr Cys Gln Ala Ser Gln Asp Ile Asn Asn
Phe Leu Asn Trp Tyr Gln 165 170
175Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Asp Ala Ser Asn
180 185 190Leu Glu Thr Gly Val
Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr 195
200 205Asp Phe Thr Phe Thr Ile Ser Ser Leu Gln Pro Glu
Asp Ile Ala Thr 210 215 220Tyr Tyr Cys
Gln Gln Tyr Gly Asn Leu Pro Phe Thr Phe Gly Gly Gly225
230 235 240Thr Lys Val Glu Ile Lys Arg
Ala Ala Ala Leu Ser Asn Ser Ile Met 245
250 255Tyr Phe Ser His Phe Val Pro Val Phe Leu Pro Ala
Lys Pro Thr Thr 260 265 270Thr
Pro Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile Ala Ser Gln 275
280 285Pro Leu Ser Leu Arg Pro Glu Ala Cys
Arg Pro Ala Ala Gly Gly Ala 290 295
300Val His Thr Arg Gly Leu Asp Phe Ala Cys Asp Ile Tyr Ile Trp Ala305
310 315 320Pro Leu Ala Gly
Thr Cys Gly Val Leu Leu Leu Ser Leu Val Ile Thr 325
330 335Leu Tyr Cys Asn His Arg Asn Arg Ser Lys
Arg Ser Arg Leu Leu His 340 345
350Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro Gly Pro Thr Arg Lys
355 360 365His Tyr Gln Pro Tyr Ala Pro
Pro Arg Asp Phe Ala Ala Tyr Arg Ser 370 375
380Arg Val Lys Phe Ser Arg Ser Ala Asp Ala Pro Ala Tyr Gln Gln
Gly385 390 395 400Gln Asn
Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr
405 410 415Asp Val Leu Asp Lys Arg Arg
Gly Arg Asp Pro Glu Met Gly Gly Lys 420 425
430Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu Tyr Asn Glu Leu
Gln Lys 435 440 445Asp Lys Met Ala
Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg 450
455 460Arg Arg Gly Lys Gly His Asp Gly Leu Tyr Gln Gly
Leu Ser Thr Ala465 470 475
480Thr Lys Asp Thr Tyr Asp Ala Leu His Met Gln Ala Leu Pro Pro Arg
485 490 49571363DNAArtificial
SequenceDescription of Artificial Sequence Synthetic polynucleotide
71caggtccaac tggtgcagtc cggagccgaa gtcaagaaac caggtgcctc cgttaaagtg
60agttgcaaag tctctggata cactctgacc gagctctcta tgcactgggt ccggcaggcc
120cccggcaagg gattggaatg gatgggcggg ttcgatcctg aggacggaga gactatctac
180gctcaaaaat tccagggacg agtgactgtg accgaagaca ctagtaccga cactgcctac
240atggaacttt cctctctgcg atcagaagat accgcagtgt actactgtgc tactgaatct
300aggggcattg gatggcccta cttcgattac tggggtcagg gaactctggt gactgtctcc
360agc
36372121PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 72Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val
Lys Lys Pro Gly Ala1 5 10
15Ser Val Lys Val Ser Cys Lys Val Ser Gly Tyr Thr Leu Thr Glu Leu
20 25 30Ser Met His Trp Val Arg Gln
Ala Pro Gly Lys Gly Leu Glu Trp Met 35 40
45Gly Gly Phe Asp Pro Glu Asp Gly Glu Thr Ile Tyr Ala Gln Lys
Phe 50 55 60Gln Gly Arg Val Thr Val
Thr Glu Asp Thr Ser Thr Asp Thr Ala Tyr65 70
75 80Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr
Ala Val Tyr Tyr Cys 85 90
95Ala Thr Glu Ser Arg Gly Ile Gly Trp Pro Tyr Phe Asp Tyr Trp Gly
100 105 110Gln Gly Thr Leu Val Thr
Val Ser Ser 115 120737PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 73Gly
Tyr Thr Leu Thr Glu Leu1 5746PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 74Asp
Pro Glu Asp Gly Glu1 57512PRTArtificial SequenceDescription
of Artificial Sequence Synthetic peptide 75Glu Ser Arg Gly Ile Gly
Trp Pro Tyr Phe Asp Tyr1 5
1076324DNAArtificial SequenceDescription of Artificial Sequence Synthetic
polynucleotide 76gatattcaga tgactcaatc tccttcttct ctgtccgctt
ccgtgggcga tagagtgacc 60attacttgta gggcgtccca gtcaatctcc agttatttga
attggtatca gcagaagccc 120gggaaagcac ctaagctgtt gatcagcggg gcttctagcc
tgaagagtgg ggtaccttca 180cggttcagcg gaagcggaag cggaaccgat ttcaccctga
ctatcagcag cctgccacct 240gaggactttg caacttacta ctgccaacag tcatacagca
ctccgatcac tttcggccag 300ggcacccggc tcgaaatcaa gcgc
32477108PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 77Asp Ile Gln Met Thr Gln
Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5
10 15Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ser
Ile Ser Ser Tyr 20 25 30Leu
Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35
40 45Ser Gly Ala Ser Ser Leu Lys Ser Gly
Val Pro Ser Arg Phe Ser Gly 50 55
60Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Pro Pro65
70 75 80Glu Asp Phe Ala Thr
Tyr Tyr Cys Gln Gln Ser Tyr Ser Thr Pro Ile 85
90 95Thr Phe Gly Gln Gly Thr Arg Leu Glu Ile Lys
Arg 100 1057811PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 78Arg
Ala Ser Gln Ser Ile Ser Ser Tyr Leu Asn1 5
10797PRTArtificial SequenceDescription of Artificial Sequence Synthetic
peptide 79Gly Ala Ser Ser Leu Lys Ser1
5809PRTArtificial SequenceDescription of Artificial Sequence Synthetic
peptide 80Gln Gln Ser Tyr Ser Thr Pro Ile Thr1
5811437DNAArtificial SequenceDescription of Artificial Sequence Synthetic
polynucleotide 81atggcactcc ccgtaactgc tctgctgctg ccgttggcat
tgctcctgca cgccgcacgc 60ccgcaggtcc aactggtgca gtccggagcc gaagtcaaga
aaccaggtgc ctccgttaaa 120gtgagttgca aagtctctgg atacactctg accgagctct
ctatgcactg ggtccggcag 180gcccccggca agggattgga atggatgggc gggttcgatc
ctgaggacgg agagactatc 240tacgctcaaa aattccaggg acgagtgact gtgaccgaag
acactagtac cgacactgcc 300tacatggaac tttcctctct gcgatcagaa gataccgcag
tgtactactg tgctactgaa 360tctaggggca ttggatggcc ctacttcgat tactggggtc
agggaactct ggtgactgtc 420tccagcggtg gaggtggcag cggtggtggc ggaagcgggg
ggggcggctc tgatattcag 480atgactcaat ctccttcttc tctgtccgct tccgtgggcg
atagagtgac cattacttgt 540agggcgtccc agtcaatctc cagttatttg aattggtatc
agcagaagcc cgggaaagca 600cctaagctgt tgatcagcgg ggcttctagc ctgaagagtg
gggtaccttc acggttcagc 660ggaagcggaa gcggaaccga tttcaccctg actatcagca
gcctgccacc tgaggacttt 720gcaacttact actgccaaca gtcatacagc actccgatca
ctttcggcca gggcacccgg 780ctcgaaatca agcgcgctgc tgctttggac aatgagaagt
caaacggcac catcatacat 840gttaaaggta aacatctgtg tccctccccg ctgttccccg
gcccttccaa accgttctgg 900gttctggtgg tggtcggagg cgtactcgct tgctatagtc
tgctggtaac tgtcgccttc 960atcatctttt gggtgagatc caaaagaagc cgcctgctcc
atagcgatta catgaatatg 1020actccacgcc gccctggccc cacaaggaaa cactaccagc
cttacgcacc acctagagat 1080ttcgctgcct atcggagcag ggtgaagttt tccagatctg
cagatgcacc agcgtatcag 1140cagggccaga accaactgta taacgagctc aacctgggac
gcagggaaga gtatgacgtt 1200ttggacaagc gcagaggacg ggaccctgag atgggtggca
aaccaagacg aaaaaacccc 1260caggagggtc tctataatga gctgcagaag gataagatgg
ctgaagccta ttctgaaata 1320ggcatgaaag gagagcggag aaggggaaaa gggcacgacg
gtttgtacca gggactcagc 1380actgctacga aggatactta tgacgctctc cacatgcaag
ccctgccacc taggtaa 143782478PRTArtificial SequenceDescription of
Artificial Sequence Synthetic polypeptide 82Met Ala Leu Pro Val Thr
Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1 5
10 15His Ala Ala Arg Pro Gln Val Gln Leu Val Gln Ser
Gly Ala Glu Val 20 25 30Lys
Lys Pro Gly Ala Ser Val Lys Val Ser Cys Lys Val Ser Gly Tyr 35
40 45Thr Leu Thr Glu Leu Ser Met His Trp
Val Arg Gln Ala Pro Gly Lys 50 55
60Gly Leu Glu Trp Met Gly Gly Phe Asp Pro Glu Asp Gly Glu Thr Ile65
70 75 80Tyr Ala Gln Lys Phe
Gln Gly Arg Val Thr Val Thr Glu Asp Thr Ser 85
90 95Thr Asp Thr Ala Tyr Met Glu Leu Ser Ser Leu
Arg Ser Glu Asp Thr 100 105
110Ala Val Tyr Tyr Cys Ala Thr Glu Ser Arg Gly Ile Gly Trp Pro Tyr
115 120 125Phe Asp Tyr Trp Gly Gln Gly
Thr Leu Val Thr Val Ser Ser Gly Gly 130 135
140Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Asp Ile
Gln145 150 155 160Met Thr
Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val
165 170 175Thr Ile Thr Cys Arg Ala Ser
Gln Ser Ile Ser Ser Tyr Leu Asn Trp 180 185
190Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Ser
Gly Ala 195 200 205Ser Ser Leu Lys
Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser 210
215 220Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Pro
Pro Glu Asp Phe225 230 235
240Ala Thr Tyr Tyr Cys Gln Gln Ser Tyr Ser Thr Pro Ile Thr Phe Gly
245 250 255Gln Gly Thr Arg Leu
Glu Ile Lys Arg Ala Ala Ala Leu Asp Asn Glu 260
265 270Lys Ser Asn Gly Thr Ile Ile His Val Lys Gly Lys
His Leu Cys Pro 275 280 285Ser Pro
Leu Phe Pro Gly Pro Ser Lys Pro Phe Trp Val Leu Val Val 290
295 300Val Gly Gly Val Leu Ala Cys Tyr Ser Leu Leu
Val Thr Val Ala Phe305 310 315
320Ile Ile Phe Trp Val Arg Ser Lys Arg Ser Arg Leu Leu His Ser Asp
325 330 335Tyr Met Asn Met
Thr Pro Arg Arg Pro Gly Pro Thr Arg Lys His Tyr 340
345 350Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala
Tyr Arg Ser Arg Val 355 360 365Lys
Phe Ser Arg Ser Ala Asp Ala Pro Ala Tyr Gln Gln Gly Gln Asn 370
375 380Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg
Arg Glu Glu Tyr Asp Val385 390 395
400Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu Met Gly Gly Lys Pro
Arg 405 410 415Arg Lys Asn
Pro Gln Glu Gly Leu Tyr Asn Glu Leu Gln Lys Asp Lys 420
425 430Met Ala Glu Ala Tyr Ser Glu Ile Gly Met
Lys Gly Glu Arg Arg Arg 435 440
445Gly Lys Gly His Asp Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys 450
455 460Asp Thr Tyr Asp Ala Leu His Met
Gln Ala Leu Pro Pro Arg465 470
475831371DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 83caggtccaac tggtgcagtc cggagccgaa
gtcaagaaac caggtgcctc cgttaaagtg 60agttgcaaag tctctggata cactctgacc
gagctctcta tgcactgggt ccggcaggcc 120cccggcaagg gattggaatg gatgggcggg
ttcgatcctg aggacggaga gactatctac 180gctcaaaaat tccagggacg agtgactgtg
accgaagaca ctagtaccga cactgcctac 240atggaacttt cctctctgcg atcagaagat
accgcagtgt actactgtgc tactgaatct 300aggggcattg gatggcccta cttcgattac
tggggtcagg gaactctggt gactgtctcc 360agcggtggag gtggcagcgg tggtggcgga
agcggggggg gcggctctga tattcagatg 420actcaatctc cttcttctct gtccgcttcc
gtgggcgata gagtgaccat tacttgtagg 480gcgtcccagt caatctccag ttatttgaat
tggtatcagc agaagcccgg gaaagcacct 540aagctgttga tcagcggggc ttctagcctg
aagagtgggg taccttcacg gttcagcgga 600agcggaagcg gaaccgattt caccctgact
atcagcagcc tgccacctga ggactttgca 660acttactact gccaacagtc atacagcact
ccgatcactt tcggccaggg cacccggctc 720gaaatcaagc gcgctgctgc tttggacaat
gagaagtcaa acggcaccat catacatgtt 780aaaggtaaac atctgtgtcc ctccccgctg
ttccccggcc cttccaaacc gttctgggtt 840ctggtggtgg tcggaggcgt actcgcttgc
tatagtctgc tggtaactgt cgccttcatc 900atcttttggg tgagatccaa aagaagccgc
ctgctccata gcgattacat gaatatgact 960ccacgccgcc ctggccccac aaggaaacac
taccagcctt acgcaccacc tagagatttc 1020gctgcctatc ggagcagggt gaagttttcc
agatctgcag atgcaccagc gtatcagcag 1080ggccagaacc aactgtataa cgagctcaac
ctgggacgca gggaagagta tgacgttttg 1140gacaagcgca gaggacggga ccctgagatg
ggtggcaaac caagacgaaa aaacccccag 1200gagggtctct ataatgagct gcagaaggat
aagatggctg aagcctattc tgaaataggc 1260atgaaaggag agcggagaag gggaaaaggg
cacgacggtt tgtaccaggg actcagcact 1320gctacgaagg atacttatga cgctctccac
atgcaagccc tgccacctag g 137184457PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
84Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala1
5 10 15Ser Val Lys Val Ser Cys
Lys Val Ser Gly Tyr Thr Leu Thr Glu Leu 20 25
30Ser Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Met 35 40 45Gly Gly Phe
Asp Pro Glu Asp Gly Glu Thr Ile Tyr Ala Gln Lys Phe 50
55 60Gln Gly Arg Val Thr Val Thr Glu Asp Thr Ser Thr
Asp Thr Ala Tyr65 70 75
80Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95Ala Thr Glu Ser Arg Gly
Ile Gly Trp Pro Tyr Phe Asp Tyr Trp Gly 100
105 110Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Gly
Gly Ser Gly Gly 115 120 125Gly Gly
Ser Gly Gly Gly Gly Ser Asp Ile Gln Met Thr Gln Ser Pro 130
135 140Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val
Thr Ile Thr Cys Arg145 150 155
160Ala Ser Gln Ser Ile Ser Ser Tyr Leu Asn Trp Tyr Gln Gln Lys Pro
165 170 175Gly Lys Ala Pro
Lys Leu Leu Ile Ser Gly Ala Ser Ser Leu Lys Ser 180
185 190Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser
Gly Thr Asp Phe Thr 195 200 205Leu
Thr Ile Ser Ser Leu Pro Pro Glu Asp Phe Ala Thr Tyr Tyr Cys 210
215 220Gln Gln Ser Tyr Ser Thr Pro Ile Thr Phe
Gly Gln Gly Thr Arg Leu225 230 235
240Glu Ile Lys Arg Ala Ala Ala Leu Asp Asn Glu Lys Ser Asn Gly
Thr 245 250 255Ile Ile His
Val Lys Gly Lys His Leu Cys Pro Ser Pro Leu Phe Pro 260
265 270Gly Pro Ser Lys Pro Phe Trp Val Leu Val
Val Val Gly Gly Val Leu 275 280
285Ala Cys Tyr Ser Leu Leu Val Thr Val Ala Phe Ile Ile Phe Trp Val 290
295 300Arg Ser Lys Arg Ser Arg Leu Leu
His Ser Asp Tyr Met Asn Met Thr305 310
315 320Pro Arg Arg Pro Gly Pro Thr Arg Lys His Tyr Gln
Pro Tyr Ala Pro 325 330
335Pro Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser Arg Ser
340 345 350Ala Asp Ala Pro Ala Tyr
Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu 355 360
365Leu Asn Leu Gly Arg Arg Glu Glu Tyr Asp Val Leu Asp Lys
Arg Arg 370 375 380Gly Arg Asp Pro Glu
Met Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln385 390
395 400Glu Gly Leu Tyr Asn Glu Leu Gln Lys Asp
Lys Met Ala Glu Ala Tyr 405 410
415Ser Glu Ile Gly Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His Asp
420 425 430Gly Leu Tyr Gln Gly
Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala 435
440 445Leu His Met Gln Ala Leu Pro Pro Arg 450
455851464DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 85atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccgcaggtgc agcttgtgca gagcggggcc
gaggtgaaga agcccggggc cagcgtcaaa 120gtgtcctgta aggtcagcgg ttacaccctc
accgagctga gcatgcactg ggtacggcag 180gctcccggca aaggtcttga gtggatgggt
ggatttgatc cagaagatgg agagactatc 240tacgcccaga agttccaggg ccgggtcacc
gtaacagaag acacctcaac tgacaccgct 300tacatggagc tgagttcact gcggtccgag
gacacggccg tgtattattg tgccaccgag 360agccgcggaa tcggatggcc ttacttcgac
tactggggac agggtacact tgttacagta 420tcatccgggg gtggcggctc tggtgggggc
ggctccggag ggggtggatc agatatccaa 480atgactcaaa gtccaagttc cctgtctgcc
tcagtcggag atagagtcac cataacctgc 540agggcaagtc agtccatctc ctcctatctg
aactggtacc aacagaaacc tggaaaggcg 600cctaagctcc tgatctccgg agcctcatct
ttgaaatccg gtgtcccatc tcgcttcagt 660ggctctggaa gcggtacaga ttttactttg
accattagca gcctcccacc ggaagacttt 720gctacatatt actgccagca gtcttactca
accccaatca ccttcgggca aggcaccaga 780ctcgaaataa aaagagcagc tgctatcgag
gttatgtacc caccgccgta cttggataac 840gaaaaaagca atgggaccat cattcatgtg
aagggtaagc acctttgccc tagcccactg 900tttcctggcc cgagtaaacc cttttgggta
cttgtggtcg tcggcggcgt gctggcctgc 960tactcactcc tggttaccgt cgcattcatc
atcttttggg tgagatccaa aagaagccgc 1020ctgctccata gcgattacat gaatatgact
ccacgccgcc ctggccccac aaggaaacac 1080taccagcctt acgcaccacc tagagatttc
gctgcctatc ggagcagggt gaagttttcc 1140agatctgcag atgcaccagc gtatcagcag
ggccagaacc aactgtataa cgagctcaac 1200ctgggacgca gggaagagta tgacgttttg
gacaagcgca gaggacggga ccctgagatg 1260ggtggcaaac caagacgaaa aaacccccag
gagggtctct ataatgagct gcagaaggat 1320aagatggctg aagcctattc tgaaataggc
atgaaaggag agcggagaag gggaaaaggg 1380cacgacggtt tgtaccaggg actcagcact
gctacgaagg atacttatga cgctctccac 1440atgcaagccc tgccacctag gtaa
146486487PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
86Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Gln
Val Gln Leu Val Gln Ser Gly Ala Glu Val 20 25
30Lys Lys Pro Gly Ala Ser Val Lys Val Ser Cys Lys Val
Ser Gly Tyr 35 40 45Thr Leu Thr
Glu Leu Ser Met His Trp Val Arg Gln Ala Pro Gly Lys 50
55 60Gly Leu Glu Trp Met Gly Gly Phe Asp Pro Glu Asp
Gly Glu Thr Ile65 70 75
80Tyr Ala Gln Lys Phe Gln Gly Arg Val Thr Val Thr Glu Asp Thr Ser
85 90 95Thr Asp Thr Ala Tyr Met
Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr 100
105 110Ala Val Tyr Tyr Cys Ala Thr Glu Ser Arg Gly Ile
Gly Trp Pro Tyr 115 120 125Phe Asp
Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly 130
135 140Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly
Gly Ser Asp Ile Gln145 150 155
160Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val
165 170 175Thr Ile Thr Cys
Arg Ala Ser Gln Ser Ile Ser Ser Tyr Leu Asn Trp 180
185 190Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu
Leu Ile Ser Gly Ala 195 200 205Ser
Ser Leu Lys Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser 210
215 220Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser
Leu Pro Pro Glu Asp Phe225 230 235
240Ala Thr Tyr Tyr Cys Gln Gln Ser Tyr Ser Thr Pro Ile Thr Phe
Gly 245 250 255Gln Gly Thr
Arg Leu Glu Ile Lys Arg Ala Ala Ala Ile Glu Val Met 260
265 270Tyr Pro Pro Pro Tyr Leu Asp Asn Glu Lys
Ser Asn Gly Thr Ile Ile 275 280
285His Val Lys Gly Lys His Leu Cys Pro Ser Pro Leu Phe Pro Gly Pro 290
295 300Ser Lys Pro Phe Trp Val Leu Val
Val Val Gly Gly Val Leu Ala Cys305 310
315 320Tyr Ser Leu Leu Val Thr Val Ala Phe Ile Ile Phe
Trp Val Arg Ser 325 330
335Lys Arg Ser Arg Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg
340 345 350Arg Pro Gly Pro Thr Arg
Lys His Tyr Gln Pro Tyr Ala Pro Pro Arg 355 360
365Asp Phe Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser Arg Ser
Ala Asp 370 375 380Ala Pro Ala Tyr Gln
Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn385 390
395 400Leu Gly Arg Arg Glu Glu Tyr Asp Val Leu
Asp Lys Arg Arg Gly Arg 405 410
415Asp Pro Glu Met Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly
420 425 430Leu Tyr Asn Glu Leu
Gln Lys Asp Lys Met Ala Glu Ala Tyr Ser Glu 435
440 445Ile Gly Met Lys Gly Glu Arg Arg Arg Gly Lys Gly
His Asp Gly Leu 450 455 460Tyr Gln Gly
Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu His465
470 475 480Met Gln Ala Leu Pro Pro Arg
485871398DNAArtificial SequenceDescription of Artificial
Sequence Synthetic polynucleotide 87caggtgcagc ttgtgcagag cggggccgag
gtgaagaagc ccggggccag cgtcaaagtg 60tcctgtaagg tcagcggtta caccctcacc
gagctgagca tgcactgggt acggcaggct 120cccggcaaag gtcttgagtg gatgggtgga
tttgatccag aagatggaga gactatctac 180gcccagaagt tccagggccg ggtcaccgta
acagaagaca cctcaactga caccgcttac 240atggagctga gttcactgcg gtccgaggac
acggccgtgt attattgtgc caccgagagc 300cgcggaatcg gatggcctta cttcgactac
tggggacagg gtacacttgt tacagtatca 360tccgggggtg gcggctctgg tgggggcggc
tccggagggg gtggatcaga tatccaaatg 420actcaaagtc caagttccct gtctgcctca
gtcggagata gagtcaccat aacctgcagg 480gcaagtcagt ccatctcctc ctatctgaac
tggtaccaac agaaacctgg aaaggcgcct 540aagctcctga tctccggagc ctcatctttg
aaatccggtg tcccatctcg cttcagtggc 600tctggaagcg gtacagattt tactttgacc
attagcagcc tcccaccgga agactttgct 660acatattact gccagcagtc ttactcaacc
ccaatcacct tcgggcaagg caccagactc 720gaaataaaaa gagcagctgc tatcgaggtt
atgtacccac cgccgtactt ggataacgaa 780aaaagcaatg ggaccatcat tcatgtgaag
ggtaagcacc tttgccctag cccactgttt 840cctggcccga gtaaaccctt ttgggtactt
gtggtcgtcg gcggcgtgct ggcctgctac 900tcactcctgg ttaccgtcgc attcatcatc
ttttgggtga gatccaaaag aagccgcctg 960ctccatagcg attacatgaa tatgactcca
cgccgccctg gccccacaag gaaacactac 1020cagccttacg caccacctag agatttcgct
gcctatcgga gcagggtgaa gttttccaga 1080tctgcagatg caccagcgta tcagcagggc
cagaaccaac tgtataacga gctcaacctg 1140ggacgcaggg aagagtatga cgttttggac
aagcgcagag gacgggaccc tgagatgggt 1200ggcaaaccaa gacgaaaaaa cccccaggag
ggtctctata atgagctgca gaaggataag 1260atggctgaag cctattctga aataggcatg
aaaggagagc ggagaagggg aaaagggcac 1320gacggtttgt accagggact cagcactgct
acgaaggata cttatgacgc tctccacatg 1380caagccctgc cacctagg
139888466PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
88Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala1
5 10 15Ser Val Lys Val Ser Cys
Lys Val Ser Gly Tyr Thr Leu Thr Glu Leu 20 25
30Ser Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Met 35 40 45Gly Gly Phe
Asp Pro Glu Asp Gly Glu Thr Ile Tyr Ala Gln Lys Phe 50
55 60Gln Gly Arg Val Thr Val Thr Glu Asp Thr Ser Thr
Asp Thr Ala Tyr65 70 75
80Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95Ala Thr Glu Ser Arg Gly
Ile Gly Trp Pro Tyr Phe Asp Tyr Trp Gly 100
105 110Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Gly
Gly Ser Gly Gly 115 120 125Gly Gly
Ser Gly Gly Gly Gly Ser Asp Ile Gln Met Thr Gln Ser Pro 130
135 140Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val
Thr Ile Thr Cys Arg145 150 155
160Ala Ser Gln Ser Ile Ser Ser Tyr Leu Asn Trp Tyr Gln Gln Lys Pro
165 170 175Gly Lys Ala Pro
Lys Leu Leu Ile Ser Gly Ala Ser Ser Leu Lys Ser 180
185 190Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser
Gly Thr Asp Phe Thr 195 200 205Leu
Thr Ile Ser Ser Leu Pro Pro Glu Asp Phe Ala Thr Tyr Tyr Cys 210
215 220Gln Gln Ser Tyr Ser Thr Pro Ile Thr Phe
Gly Gln Gly Thr Arg Leu225 230 235
240Glu Ile Lys Arg Ala Ala Ala Ile Glu Val Met Tyr Pro Pro Pro
Tyr 245 250 255Leu Asp Asn
Glu Lys Ser Asn Gly Thr Ile Ile His Val Lys Gly Lys 260
265 270His Leu Cys Pro Ser Pro Leu Phe Pro Gly
Pro Ser Lys Pro Phe Trp 275 280
285Val Leu Val Val Val Gly Gly Val Leu Ala Cys Tyr Ser Leu Leu Val 290
295 300Thr Val Ala Phe Ile Ile Phe Trp
Val Arg Ser Lys Arg Ser Arg Leu305 310
315 320Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg
Pro Gly Pro Thr 325 330
335Arg Lys His Tyr Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala Tyr
340 345 350Arg Ser Arg Val Lys Phe
Ser Arg Ser Ala Asp Ala Pro Ala Tyr Gln 355 360
365Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg
Arg Glu 370 375 380Glu Tyr Asp Val Leu
Asp Lys Arg Arg Gly Arg Asp Pro Glu Met Gly385 390
395 400Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu
Gly Leu Tyr Asn Glu Leu 405 410
415Gln Lys Asp Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly
420 425 430Glu Arg Arg Arg Gly
Lys Gly His Asp Gly Leu Tyr Gln Gly Leu Ser 435
440 445Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu His Met
Gln Ala Leu Pro 450 455 460Pro
Arg465891545DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 89atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccgcaggtgc agttggtgca aagcggcgca
gaagttaaga aacctggggc gtcagttaag 120gtgtcttgca aagtatctgg ctataccctc
actgagctgt ccatgcattg ggtaaggcag 180gctcctggaa aggggctcga atggatggga
ggatttgacc ctgaagacgg agagaccatc 240tacgcccaga aattccaggg tagagtaaca
gtgactgagg acactagcac tgacacagcg 300tacatggagc tgagttctct gagaagtgag
gacacagccg tttactactg cgctaccgag 360tccagaggta ttggctggcc atacttcgac
tattggggtc agggcaccct ggttacagtg 420agttcaggag gcgggggctc tggggggggc
ggttccggag gggggggctc agatatacag 480atgacgcaga gtccatcaag tctctcagcc
agcgtgggag atcgcgtgac tattacttgc 540cgcgccagcc agagtattag ctcctatctg
aattggtacc agcaaaagcc cgggaaggcc 600cctaagcttc tgatttctgg cgcctcctct
ttgaagtcag gtgtgccaag cagatttagc 660gggtctggaa gtggcactga ctttacactt
actatctcca gcctgccccc agaggatttt 720gccacatatt actgtcagca aagctactct
actccaatca ctttcggcca gggcacaaga 780ttggagatta agagggctgc cgcactttca
aattccatca tgtatttcag ccattttgtg 840cctgtttttc ttccggccaa acctacaacc
actcccgccc cacgcccacc tactcccgcc 900cctaccattg cctcccagcc tctgtctctt
agacctgagg cttgtagacc tgctgccggc 960ggagccgtgc acactcgcgg tctggacttc
gcctgcgaca tctatatctg ggcccctctg 1020gccggcacct gcggcgttct ccttctctca
ctcgtaatca cactctattg caatcacagg 1080aacagatcca aaagaagccg cctgctccat
agcgattaca tgaatatgac tccacgccgc 1140cctggcccca caaggaaaca ctaccagcct
tacgcaccac ctagagattt cgctgcctat 1200cggagcaggg tgaagttttc cagatctgca
gatgcaccag cgtatcagca gggccagaac 1260caactgtata acgagctcaa cctgggacgc
agggaagagt atgacgtttt ggacaagcgc 1320agaggacggg accctgagat gggtggcaaa
ccaagacgaa aaaaccccca ggagggtctc 1380tataatgagc tgcagaagga taagatggct
gaagcctatt ctgaaatagg catgaaagga 1440gagcggagaa ggggaaaagg gcacgacggt
ttgtaccagg gactcagcac tgctacgaag 1500gatacttatg acgctctcca catgcaagcc
ctgccaccta ggtaa 154590514PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
90Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Gln
Val Gln Leu Val Gln Ser Gly Ala Glu Val 20 25
30Lys Lys Pro Gly Ala Ser Val Lys Val Ser Cys Lys Val
Ser Gly Tyr 35 40 45Thr Leu Thr
Glu Leu Ser Met His Trp Val Arg Gln Ala Pro Gly Lys 50
55 60Gly Leu Glu Trp Met Gly Gly Phe Asp Pro Glu Asp
Gly Glu Thr Ile65 70 75
80Tyr Ala Gln Lys Phe Gln Gly Arg Val Thr Val Thr Glu Asp Thr Ser
85 90 95Thr Asp Thr Ala Tyr Met
Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr 100
105 110Ala Val Tyr Tyr Cys Ala Thr Glu Ser Arg Gly Ile
Gly Trp Pro Tyr 115 120 125Phe Asp
Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly 130
135 140Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly
Gly Ser Asp Ile Gln145 150 155
160Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val
165 170 175Thr Ile Thr Cys
Arg Ala Ser Gln Ser Ile Ser Ser Tyr Leu Asn Trp 180
185 190Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu
Leu Ile Ser Gly Ala 195 200 205Ser
Ser Leu Lys Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser 210
215 220Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser
Leu Pro Pro Glu Asp Phe225 230 235
240Ala Thr Tyr Tyr Cys Gln Gln Ser Tyr Ser Thr Pro Ile Thr Phe
Gly 245 250 255Gln Gly Thr
Arg Leu Glu Ile Lys Arg Ala Ala Ala Leu Ser Asn Ser 260
265 270Ile Met Tyr Phe Ser His Phe Val Pro Val
Phe Leu Pro Ala Lys Pro 275 280
285Thr Thr Thr Pro Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile Ala 290
295 300Ser Gln Pro Leu Ser Leu Arg Pro
Glu Ala Cys Arg Pro Ala Ala Gly305 310
315 320Gly Ala Val His Thr Arg Gly Leu Asp Phe Ala Cys
Asp Ile Tyr Ile 325 330
335Trp Ala Pro Leu Ala Gly Thr Cys Gly Val Leu Leu Leu Ser Leu Val
340 345 350Ile Thr Leu Tyr Cys Asn
His Arg Asn Arg Ser Lys Arg Ser Arg Leu 355 360
365Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro Gly
Pro Thr 370 375 380Arg Lys His Tyr Gln
Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala Tyr385 390
395 400Arg Ser Arg Val Lys Phe Ser Arg Ser Ala
Asp Ala Pro Ala Tyr Gln 405 410
415Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg Glu
420 425 430Glu Tyr Asp Val Leu
Asp Lys Arg Arg Gly Arg Asp Pro Glu Met Gly 435
440 445Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu
Tyr Asn Glu Leu 450 455 460Gln Lys Asp
Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly465
470 475 480Glu Arg Arg Arg Gly Lys Gly
His Asp Gly Leu Tyr Gln Gly Leu Ser 485
490 495Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu His Met
Gln Ala Leu Pro 500 505 510Pro
Arg911479DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 91caggtgcagt tggtgcaaag cggcgcagaa
gttaagaaac ctggggcgtc agttaaggtg 60tcttgcaaag tatctggcta taccctcact
gagctgtcca tgcattgggt aaggcaggct 120cctggaaagg ggctcgaatg gatgggagga
tttgaccctg aagacggaga gaccatctac 180gcccagaaat tccagggtag agtaacagtg
actgaggaca ctagcactga cacagcgtac 240atggagctga gttctctgag aagtgaggac
acagccgttt actactgcgc taccgagtcc 300agaggtattg gctggccata cttcgactat
tggggtcagg gcaccctggt tacagtgagt 360tcaggaggcg ggggctctgg ggggggcggt
tccggagggg ggggctcaga tatacagatg 420acgcagagtc catcaagtct ctcagccagc
gtgggagatc gcgtgactat tacttgccgc 480gccagccaga gtattagctc ctatctgaat
tggtaccagc aaaagcccgg gaaggcccct 540aagcttctga tttctggcgc ctcctctttg
aagtcaggtg tgccaagcag atttagcggg 600tctggaagtg gcactgactt tacacttact
atctccagcc tgcccccaga ggattttgcc 660acatattact gtcagcaaag ctactctact
ccaatcactt tcggccaggg cacaagattg 720gagattaaga gggctgccgc actttcaaat
tccatcatgt atttcagcca ttttgtgcct 780gtttttcttc cggccaaacc tacaaccact
cccgccccac gcccacctac tcccgcccct 840accattgcct cccagcctct gtctcttaga
cctgaggctt gtagacctgc tgccggcgga 900gccgtgcaca ctcgcggtct ggacttcgcc
tgcgacatct atatctgggc ccctctggcc 960ggcacctgcg gcgttctcct tctctcactc
gtaatcacac tctattgcaa tcacaggaac 1020agatccaaaa gaagccgcct gctccatagc
gattacatga atatgactcc acgccgccct 1080ggccccacaa ggaaacacta ccagccttac
gcaccaccta gagatttcgc tgcctatcgg 1140agcagggtga agttttccag atctgcagat
gcaccagcgt atcagcaggg ccagaaccaa 1200ctgtataacg agctcaacct gggacgcagg
gaagagtatg acgttttgga caagcgcaga 1260ggacgggacc ctgagatggg tggcaaacca
agacgaaaaa acccccagga gggtctctat 1320aatgagctgc agaaggataa gatggctgaa
gcctattctg aaataggcat gaaaggagag 1380cggagaaggg gaaaagggca cgacggtttg
taccagggac tcagcactgc tacgaaggat 1440acttatgacg ctctccacat gcaagccctg
ccacctagg 147992493PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
92Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala1
5 10 15Ser Val Lys Val Ser Cys
Lys Val Ser Gly Tyr Thr Leu Thr Glu Leu 20 25
30Ser Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Met 35 40 45Gly Gly Phe
Asp Pro Glu Asp Gly Glu Thr Ile Tyr Ala Gln Lys Phe 50
55 60Gln Gly Arg Val Thr Val Thr Glu Asp Thr Ser Thr
Asp Thr Ala Tyr65 70 75
80Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95Ala Thr Glu Ser Arg Gly
Ile Gly Trp Pro Tyr Phe Asp Tyr Trp Gly 100
105 110Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Gly
Gly Ser Gly Gly 115 120 125Gly Gly
Ser Gly Gly Gly Gly Ser Asp Ile Gln Met Thr Gln Ser Pro 130
135 140Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val
Thr Ile Thr Cys Arg145 150 155
160Ala Ser Gln Ser Ile Ser Ser Tyr Leu Asn Trp Tyr Gln Gln Lys Pro
165 170 175Gly Lys Ala Pro
Lys Leu Leu Ile Ser Gly Ala Ser Ser Leu Lys Ser 180
185 190Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser
Gly Thr Asp Phe Thr 195 200 205Leu
Thr Ile Ser Ser Leu Pro Pro Glu Asp Phe Ala Thr Tyr Tyr Cys 210
215 220Gln Gln Ser Tyr Ser Thr Pro Ile Thr Phe
Gly Gln Gly Thr Arg Leu225 230 235
240Glu Ile Lys Arg Ala Ala Ala Leu Ser Asn Ser Ile Met Tyr Phe
Ser 245 250 255His Phe Val
Pro Val Phe Leu Pro Ala Lys Pro Thr Thr Thr Pro Ala 260
265 270Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile
Ala Ser Gln Pro Leu Ser 275 280
285Leu Arg Pro Glu Ala Cys Arg Pro Ala Ala Gly Gly Ala Val His Thr 290
295 300Arg Gly Leu Asp Phe Ala Cys Asp
Ile Tyr Ile Trp Ala Pro Leu Ala305 310
315 320Gly Thr Cys Gly Val Leu Leu Leu Ser Leu Val Ile
Thr Leu Tyr Cys 325 330
335Asn His Arg Asn Arg Ser Lys Arg Ser Arg Leu Leu His Ser Asp Tyr
340 345 350Met Asn Met Thr Pro Arg
Arg Pro Gly Pro Thr Arg Lys His Tyr Gln 355 360
365Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala Tyr Arg Ser Arg
Val Lys 370 375 380Phe Ser Arg Ser Ala
Asp Ala Pro Ala Tyr Gln Gln Gly Gln Asn Gln385 390
395 400Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg
Glu Glu Tyr Asp Val Leu 405 410
415Asp Lys Arg Arg Gly Arg Asp Pro Glu Met Gly Gly Lys Pro Arg Arg
420 425 430Lys Asn Pro Gln Glu
Gly Leu Tyr Asn Glu Leu Gln Lys Asp Lys Met 435
440 445Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly Glu
Arg Arg Arg Gly 450 455 460Lys Gly His
Asp Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys Asp465
470 475 480Thr Tyr Asp Ala Leu His Met
Gln Ala Leu Pro Pro Arg 485
49093351DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 93caggtccagt tggtcgaaag tggcggtggt
gtagtgcagc cgggccgcag tttgaggctt 60tcctgtgcgg cttcaggctt tactttttcc
agctatggaa tgcactgggt gcggcaggcc 120cccggcaaag gacttgagtg ggtggccgtc
atttcttatg acggatcaga taagtactac 180gtggacagcg tcaagggcag attcaccatc
tctagggaca acagtaaaaa tagactctac 240ctccagatga atagcctcag agctgaagac
acggccgtct actattgtgc tcgggagcgg 300tatagtggca gagactactg ggggcagggc
acactcgtta cagtgagtag c 35194117PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
94Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1
5 10 15Ser Leu Arg Leu Ser Cys
Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25
30Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45Ala Val Ile
Ser Tyr Asp Gly Ser Asp Lys Tyr Tyr Val Asp Ser Val 50
55 60Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys
Asn Arg Leu Tyr65 70 75
80Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95Ala Arg Glu Arg Tyr Ser
Gly Arg Asp Tyr Trp Gly Gln Gly Thr Leu 100
105 110Val Thr Val Ser Ser 115957PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 95Gly
Phe Thr Phe Ser Ser Tyr1 5966PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 96Ser
Tyr Asp Gly Ser Asp1 5978PRTArtificial SequenceDescription
of Artificial Sequence Synthetic peptide 97Glu Arg Tyr Ser Gly Arg
Asp Tyr1 598324DNAArtificial SequenceDescription of
Artificial Sequence Synthetic polynucleotide 98gagattgtta tgacccagag
tcctgcgacc ctctcagtca gccccgggga gcgcgcaact 60ttgtcttgca gagctagtca
gtccgtgtcc tctcttctga catggtacca gcaaaagccc 120gggcaggctc cgcgcctttt
gatctttggg gcttcaacaa gagccactgg gattcccgca 180cgattctctg gctccgggag
cggtactggt ttcaccctga cgattagcag tctccagagc 240gaggacttcg ccgtatacta
ctgccagcag tacgatacgt ggccattcac ttttggacca 300gggactaaag tggattttaa
gcgc 32499108PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
99Glu Ile Val Met Thr Gln Ser Pro Ala Thr Leu Ser Val Ser Pro Gly1
5 10 15Glu Arg Ala Thr Leu Ser
Cys Arg Ala Ser Gln Ser Val Ser Ser Leu 20 25
30Leu Thr Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg
Leu Leu Ile 35 40 45Phe Gly Ala
Ser Thr Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50
55 60Ser Gly Ser Gly Thr Gly Phe Thr Leu Thr Ile Ser
Ser Leu Gln Ser65 70 75
80Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asp Thr Trp Pro Phe
85 90 95Thr Phe Gly Pro Gly Thr
Lys Val Asp Phe Lys Arg 100
10510011PRTArtificial SequenceDescription of Artificial Sequence
Synthetic peptide 100Arg Ala Ser Gln Ser Val Ser Ser Leu Leu Thr1
5 101017PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 101Gly Ala Ser Thr Arg Ala
Thr1 51029PRTArtificial SequenceDescription of Artificial
Sequence Synthetic peptide 102Gln Gln Tyr Asp Thr Trp Pro Phe Thr1
51031425DNAArtificial SequenceDescription of Artificial
Sequence Synthetic polynucleotide 103atggcactcc ccgtaactgc
tctgctgctg ccgttggcat tgctcctgca cgccgcacgc 60ccgcaggtcc agttggtcga
aagtggcggt ggtgtagtgc agccgggccg cagtttgagg 120ctttcctgtg cggcttcagg
ctttactttt tccagctatg gaatgcactg ggtgcggcag 180gcccccggca aaggacttga
gtgggtggcc gtcatttctt atgacggatc agataagtac 240tacgtggaca gcgtcaaggg
cagattcacc atctctaggg acaacagtaa aaatagactc 300tacctccaga tgaatagcct
cagagctgaa gacacggccg tctactattg tgctcgggag 360cggtatagtg gcagagacta
ctgggggcag ggcacactcg ttacagtgag tagcggcgga 420ggagggagtg ggggcggtgg
ctccggtgga ggaggttctg agattgttat gacccagagt 480cctgcgaccc tctcagtcag
ccccggggag cgcgcaactt tgtcttgcag agctagtcag 540tccgtgtcct ctcttctgac
atggtaccag caaaagcccg ggcaggctcc gcgccttttg 600atctttgggg cttcaacaag
agccactggg attcccgcac gattctctgg ctccgggagc 660ggtactggtt tcaccctgac
gattagcagt ctccagagcg aggacttcgc cgtatactac 720tgccagcagt acgatacgtg
gccattcact tttggaccag ggactaaagt ggattttaag 780cgcgccgccg ctctcgataa
cgaaaagtca aatggcacca taatccacgt caaaggcaag 840cacctgtgcc cttccccgct
cttccccgga cccagtaaac cattttgggt gctggttgtt 900gtggggggcg tgctggcctg
ctatagcctt ttggtcactg tagccttcat tattttttgg 960gtcagatcca aaagaagccg
cctgctccat agcgattaca tgaatatgac tccacgccgc 1020cctggcccca caaggaaaca
ctaccagcct tacgcaccac ctagagattt cgctgcctat 1080cggagcaggg tgaagttttc
cagatctgca gatgcaccag cgtatcagca gggccagaac 1140caactgtata acgagctcaa
cctgggacgc agggaagagt atgacgtttt ggacaagcgc 1200agaggacggg accctgagat
gggtggcaaa ccaagacgaa aaaaccccca ggagggtctc 1260tataatgagc tgcagaagga
taagatggct gaagcctatt ctgaaatagg catgaaagga 1320gagcggagaa ggggaaaagg
gcacgacggt ttgtaccagg gactcagcac tgctacgaag 1380gatacttatg acgctctcca
catgcaagcc ctgccaccta ggtaa 1425104474PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
104Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Gln
Val Gln Leu Val Glu Ser Gly Gly Gly Val 20 25
30Val Gln Pro Gly Arg Ser Leu Arg Leu Ser Cys Ala Ala
Ser Gly Phe 35 40 45Thr Phe Ser
Ser Tyr Gly Met His Trp Val Arg Gln Ala Pro Gly Lys 50
55 60Gly Leu Glu Trp Val Ala Val Ile Ser Tyr Asp Gly
Ser Asp Lys Tyr65 70 75
80Tyr Val Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser
85 90 95Lys Asn Arg Leu Tyr Leu
Gln Met Asn Ser Leu Arg Ala Glu Asp Thr 100
105 110Ala Val Tyr Tyr Cys Ala Arg Glu Arg Tyr Ser Gly
Arg Asp Tyr Trp 115 120 125Gly Gln
Gly Thr Leu Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly 130
135 140Gly Gly Gly Ser Gly Gly Gly Gly Ser Glu Ile
Val Met Thr Gln Ser145 150 155
160Pro Ala Thr Leu Ser Val Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys
165 170 175Arg Ala Ser Gln
Ser Val Ser Ser Leu Leu Thr Trp Tyr Gln Gln Lys 180
185 190Pro Gly Gln Ala Pro Arg Leu Leu Ile Phe Gly
Ala Ser Thr Arg Ala 195 200 205Thr
Gly Ile Pro Ala Arg Phe Ser Gly Ser Gly Ser Gly Thr Gly Phe 210
215 220Thr Leu Thr Ile Ser Ser Leu Gln Ser Glu
Asp Phe Ala Val Tyr Tyr225 230 235
240Cys Gln Gln Tyr Asp Thr Trp Pro Phe Thr Phe Gly Pro Gly Thr
Lys 245 250 255Val Asp Phe
Lys Arg Ala Ala Ala Leu Asp Asn Glu Lys Ser Asn Gly 260
265 270Thr Ile Ile His Val Lys Gly Lys His Leu
Cys Pro Ser Pro Leu Phe 275 280
285Pro Gly Pro Ser Lys Pro Phe Trp Val Leu Val Val Val Gly Gly Val 290
295 300Leu Ala Cys Tyr Ser Leu Leu Val
Thr Val Ala Phe Ile Ile Phe Trp305 310
315 320Val Arg Ser Lys Arg Ser Arg Leu Leu His Ser Asp
Tyr Met Asn Met 325 330
335Thr Pro Arg Arg Pro Gly Pro Thr Arg Lys His Tyr Gln Pro Tyr Ala
340 345 350Pro Pro Arg Asp Phe Ala
Ala Tyr Arg Ser Arg Val Lys Phe Ser Arg 355 360
365Ser Ala Asp Ala Pro Ala Tyr Gln Gln Gly Gln Asn Gln Leu
Tyr Asn 370 375 380Glu Leu Asn Leu Gly
Arg Arg Glu Glu Tyr Asp Val Leu Asp Lys Arg385 390
395 400Arg Gly Arg Asp Pro Glu Met Gly Gly Lys
Pro Arg Arg Lys Asn Pro 405 410
415Gln Glu Gly Leu Tyr Asn Glu Leu Gln Lys Asp Lys Met Ala Glu Ala
420 425 430Tyr Ser Glu Ile Gly
Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His 435
440 445Asp Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys
Asp Thr Tyr Asp 450 455 460Ala Leu His
Met Gln Ala Leu Pro Pro Arg465 4701051359DNAArtificial
SequenceDescription of Artificial Sequence Synthetic polynucleotide
105caggtccagt tggtcgaaag tggcggtggt gtagtgcagc cgggccgcag tttgaggctt
60tcctgtgcgg cttcaggctt tactttttcc agctatggaa tgcactgggt gcggcaggcc
120cccggcaaag gacttgagtg ggtggccgtc atttcttatg acggatcaga taagtactac
180gtggacagcg tcaagggcag attcaccatc tctagggaca acagtaaaaa tagactctac
240ctccagatga atagcctcag agctgaagac acggccgtct actattgtgc tcgggagcgg
300tatagtggca gagactactg ggggcagggc acactcgtta cagtgagtag cggcggagga
360gggagtgggg gcggtggctc cggtggagga ggttctgaga ttgttatgac ccagagtcct
420gcgaccctct cagtcagccc cggggagcgc gcaactttgt cttgcagagc tagtcagtcc
480gtgtcctctc ttctgacatg gtaccagcaa aagcccgggc aggctccgcg ccttttgatc
540tttggggctt caacaagagc cactgggatt cccgcacgat tctctggctc cgggagcggt
600actggtttca ccctgacgat tagcagtctc cagagcgagg acttcgccgt atactactgc
660cagcagtacg atacgtggcc attcactttt ggaccaggga ctaaagtgga ttttaagcgc
720gccgccgctc tcgataacga aaagtcaaat ggcaccataa tccacgtcaa aggcaagcac
780ctgtgccctt ccccgctctt ccccggaccc agtaaaccat tttgggtgct ggttgttgtg
840gggggcgtgc tggcctgcta tagccttttg gtcactgtag ccttcattat tttttgggtc
900agatccaaaa gaagccgcct gctccatagc gattacatga atatgactcc acgccgccct
960ggccccacaa ggaaacacta ccagccttac gcaccaccta gagatttcgc tgcctatcgg
1020agcagggtga agttttccag atctgcagat gcaccagcgt atcagcaggg ccagaaccaa
1080ctgtataacg agctcaacct gggacgcagg gaagagtatg acgttttgga caagcgcaga
1140ggacgggacc ctgagatggg tggcaaacca agacgaaaaa acccccagga gggtctctat
1200aatgagctgc agaaggataa gatggctgaa gcctattctg aaataggcat gaaaggagag
1260cggagaaggg gaaaagggca cgacggtttg taccagggac tcagcactgc tacgaaggat
1320acttatgacg ctctccacat gcaagccctg ccacctagg
1359106453PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 106Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val
Val Gln Pro Gly Arg1 5 10
15Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr
20 25 30Gly Met His Trp Val Arg Gln
Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40
45Ala Val Ile Ser Tyr Asp Gly Ser Asp Lys Tyr Tyr Val Asp Ser
Val 50 55 60Lys Gly Arg Phe Thr Ile
Ser Arg Asp Asn Ser Lys Asn Arg Leu Tyr65 70
75 80Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr
Ala Val Tyr Tyr Cys 85 90
95Ala Arg Glu Arg Tyr Ser Gly Arg Asp Tyr Trp Gly Gln Gly Thr Leu
100 105 110Val Thr Val Ser Ser Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly 115 120
125Gly Gly Gly Ser Glu Ile Val Met Thr Gln Ser Pro Ala Thr
Leu Ser 130 135 140Val Ser Pro Gly Glu
Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser145 150
155 160Val Ser Ser Leu Leu Thr Trp Tyr Gln Gln
Lys Pro Gly Gln Ala Pro 165 170
175Arg Leu Leu Ile Phe Gly Ala Ser Thr Arg Ala Thr Gly Ile Pro Ala
180 185 190Arg Phe Ser Gly Ser
Gly Ser Gly Thr Gly Phe Thr Leu Thr Ile Ser 195
200 205Ser Leu Gln Ser Glu Asp Phe Ala Val Tyr Tyr Cys
Gln Gln Tyr Asp 210 215 220Thr Trp Pro
Phe Thr Phe Gly Pro Gly Thr Lys Val Asp Phe Lys Arg225
230 235 240Ala Ala Ala Leu Asp Asn Glu
Lys Ser Asn Gly Thr Ile Ile His Val 245
250 255Lys Gly Lys His Leu Cys Pro Ser Pro Leu Phe Pro
Gly Pro Ser Lys 260 265 270Pro
Phe Trp Val Leu Val Val Val Gly Gly Val Leu Ala Cys Tyr Ser 275
280 285Leu Leu Val Thr Val Ala Phe Ile Ile
Phe Trp Val Arg Ser Lys Arg 290 295
300Ser Arg Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro305
310 315 320Gly Pro Thr Arg
Lys His Tyr Gln Pro Tyr Ala Pro Pro Arg Asp Phe 325
330 335Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser
Arg Ser Ala Asp Ala Pro 340 345
350Ala Tyr Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly
355 360 365Arg Arg Glu Glu Tyr Asp Val
Leu Asp Lys Arg Arg Gly Arg Asp Pro 370 375
380Glu Met Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu
Tyr385 390 395 400Asn Glu
Leu Gln Lys Asp Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly
405 410 415Met Lys Gly Glu Arg Arg Arg
Gly Lys Gly His Asp Gly Leu Tyr Gln 420 425
430Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu His
Met Gln 435 440 445Ala Leu Pro Pro
Arg 4501071452DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 107atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccgcaggtgc agctcgtgga gtctggcggc
ggcgtggtcc agcccggccg gtccctgcgc 120ctgtcctgcg ccgccagcgg gtttactttt
tcctcctacg gcatgcactg ggtgcgccag 180gctcccggca agggcctcga gtgggtcgcc
gtgatctcat acgatgggtc agacaaatac 240tatgtcgatt ctgttaaagg gcggtttacc
atttcaagag ataactctaa gaataggctg 300tatttgcaga tgaacagcct gagggctgaa
gataccgcag tgtactattg cgctagggag 360cggtatagtg gccgcgatta ctggggacag
ggtacactgg tgaccgtgag ctctgggggt 420ggcggaagcg ggggtggcgg aagcggcgga
gggggtagtg aaattgtgat gacccagtct 480ccggctacac tttcagtctc ccctggggag
agagctacac tgtcatgcag agcgtcccag 540tccgtctctt ctctccttac ctggtatcag
cagaagcccg gccaggctcc tcgactgctg 600atcttcggtg cctccacaag ggcgaccggg
attccagccc gcttctcagg ttctgggagc 660ggaactggtt tcactttgac aatcagttca
ctgcagtcag aggatttcgc cgtgtactac 720tgccagcaat acgacacatg gccattcact
ttcggacccg gtaccaaagt cgatttcaag 780agagccgcgg ccatcgaggt tatgtaccca
ccaccatatc tggacaatga aaaaagcaat 840ggaaccatta tccatgtgaa gggtaaacac
ctctgcccta gcccactttt ccctggccca 900tcaaagccct tctgggtctt ggtggtcgtg
gggggtgtgc tggcctgtta cagccttctg 960gtgacggttg ctttcattat cttctgggtt
agatccaaaa gaagccgcct gctccatagc 1020gattacatga atatgactcc acgccgccct
ggccccacaa ggaaacacta ccagccttac 1080gcaccaccta gagatttcgc tgcctatcgg
agcagggtga agttttccag atctgcagat 1140gcaccagcgt atcagcaggg ccagaaccaa
ctgtataacg agctcaacct gggacgcagg 1200gaagagtatg acgttttgga caagcgcaga
ggacgggacc ctgagatggg tggcaaacca 1260agacgaaaaa acccccagga gggtctctat
aatgagctgc agaaggataa gatggctgaa 1320gcctattctg aaataggcat gaaaggagag
cggagaaggg gaaaagggca cgacggtttg 1380taccagggac tcagcactgc tacgaaggat
acttatgacg ctctccacat gcaagccctg 1440ccacctaggt aa
1452108483PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
108Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Gln
Val Gln Leu Val Glu Ser Gly Gly Gly Val 20 25
30Val Gln Pro Gly Arg Ser Leu Arg Leu Ser Cys Ala Ala
Ser Gly Phe 35 40 45Thr Phe Ser
Ser Tyr Gly Met His Trp Val Arg Gln Ala Pro Gly Lys 50
55 60Gly Leu Glu Trp Val Ala Val Ile Ser Tyr Asp Gly
Ser Asp Lys Tyr65 70 75
80Tyr Val Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser
85 90 95Lys Asn Arg Leu Tyr Leu
Gln Met Asn Ser Leu Arg Ala Glu Asp Thr 100
105 110Ala Val Tyr Tyr Cys Ala Arg Glu Arg Tyr Ser Gly
Arg Asp Tyr Trp 115 120 125Gly Gln
Gly Thr Leu Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly 130
135 140Gly Gly Gly Ser Gly Gly Gly Gly Ser Glu Ile
Val Met Thr Gln Ser145 150 155
160Pro Ala Thr Leu Ser Val Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys
165 170 175Arg Ala Ser Gln
Ser Val Ser Ser Leu Leu Thr Trp Tyr Gln Gln Lys 180
185 190Pro Gly Gln Ala Pro Arg Leu Leu Ile Phe Gly
Ala Ser Thr Arg Ala 195 200 205Thr
Gly Ile Pro Ala Arg Phe Ser Gly Ser Gly Ser Gly Thr Gly Phe 210
215 220Thr Leu Thr Ile Ser Ser Leu Gln Ser Glu
Asp Phe Ala Val Tyr Tyr225 230 235
240Cys Gln Gln Tyr Asp Thr Trp Pro Phe Thr Phe Gly Pro Gly Thr
Lys 245 250 255Val Asp Phe
Lys Arg Ala Ala Ala Ile Glu Val Met Tyr Pro Pro Pro 260
265 270Tyr Leu Asp Asn Glu Lys Ser Asn Gly Thr
Ile Ile His Val Lys Gly 275 280
285Lys His Leu Cys Pro Ser Pro Leu Phe Pro Gly Pro Ser Lys Pro Phe 290
295 300Trp Val Leu Val Val Val Gly Gly
Val Leu Ala Cys Tyr Ser Leu Leu305 310
315 320Val Thr Val Ala Phe Ile Ile Phe Trp Val Arg Ser
Lys Arg Ser Arg 325 330
335Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro Gly Pro
340 345 350Thr Arg Lys His Tyr Gln
Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala 355 360
365Tyr Arg Ser Arg Val Lys Phe Ser Arg Ser Ala Asp Ala Pro
Ala Tyr 370 375 380Gln Gln Gly Gln Asn
Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg385 390
395 400Glu Glu Tyr Asp Val Leu Asp Lys Arg Arg
Gly Arg Asp Pro Glu Met 405 410
415Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu Tyr Asn Glu
420 425 430Leu Gln Lys Asp Lys
Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys 435
440 445Gly Glu Arg Arg Arg Gly Lys Gly His Asp Gly Leu
Tyr Gln Gly Leu 450 455 460Ser Thr Ala
Thr Lys Asp Thr Tyr Asp Ala Leu His Met Gln Ala Leu465
470 475 480Pro Pro
Arg1091386DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 109caggtgcagc tcgtggagtc tggcggcggc
gtggtccagc ccggccggtc cctgcgcctg 60tcctgcgccg ccagcgggtt tactttttcc
tcctacggca tgcactgggt gcgccaggct 120cccggcaagg gcctcgagtg ggtcgccgtg
atctcatacg atgggtcaga caaatactat 180gtcgattctg ttaaagggcg gtttaccatt
tcaagagata actctaagaa taggctgtat 240ttgcagatga acagcctgag ggctgaagat
accgcagtgt actattgcgc tagggagcgg 300tatagtggcc gcgattactg gggacagggt
acactggtga ccgtgagctc tgggggtggc 360ggaagcgggg gtggcggaag cggcggaggg
ggtagtgaaa ttgtgatgac ccagtctccg 420gctacacttt cagtctcccc tggggagaga
gctacactgt catgcagagc gtcccagtcc 480gtctcttctc tccttacctg gtatcagcag
aagcccggcc aggctcctcg actgctgatc 540ttcggtgcct ccacaagggc gaccgggatt
ccagcccgct tctcaggttc tgggagcgga 600actggtttca ctttgacaat cagttcactg
cagtcagagg atttcgccgt gtactactgc 660cagcaatacg acacatggcc attcactttc
ggacccggta ccaaagtcga tttcaagaga 720gccgcggcca tcgaggttat gtacccacca
ccatatctgg acaatgaaaa aagcaatgga 780accattatcc atgtgaaggg taaacacctc
tgccctagcc cacttttccc tggcccatca 840aagcccttct gggtcttggt ggtcgtgggg
ggtgtgctgg cctgttacag ccttctggtg 900acggttgctt tcattatctt ctgggttaga
tccaaaagaa gccgcctgct ccatagcgat 960tacatgaata tgactccacg ccgccctggc
cccacaagga aacactacca gccttacgca 1020ccacctagag atttcgctgc ctatcggagc
agggtgaagt tttccagatc tgcagatgca 1080ccagcgtatc agcagggcca gaaccaactg
tataacgagc tcaacctggg acgcagggaa 1140gagtatgacg ttttggacaa gcgcagagga
cgggaccctg agatgggtgg caaaccaaga 1200cgaaaaaacc cccaggaggg tctctataat
gagctgcaga aggataagat ggctgaagcc 1260tattctgaaa taggcatgaa aggagagcgg
agaaggggaa aagggcacga cggtttgtac 1320cagggactca gcactgctac gaaggatact
tatgacgctc tccacatgca agccctgcca 1380cctagg
1386110462PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
110Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1
5 10 15Ser Leu Arg Leu Ser Cys
Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25
30Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu
Glu Trp Val 35 40 45Ala Val Ile
Ser Tyr Asp Gly Ser Asp Lys Tyr Tyr Val Asp Ser Val 50
55 60Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys
Asn Arg Leu Tyr65 70 75
80Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95Ala Arg Glu Arg Tyr Ser
Gly Arg Asp Tyr Trp Gly Gln Gly Thr Leu 100
105 110Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly
Gly Gly Ser Gly 115 120 125Gly Gly
Gly Ser Glu Ile Val Met Thr Gln Ser Pro Ala Thr Leu Ser 130
135 140Val Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys
Arg Ala Ser Gln Ser145 150 155
160Val Ser Ser Leu Leu Thr Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro
165 170 175Arg Leu Leu Ile
Phe Gly Ala Ser Thr Arg Ala Thr Gly Ile Pro Ala 180
185 190Arg Phe Ser Gly Ser Gly Ser Gly Thr Gly Phe
Thr Leu Thr Ile Ser 195 200 205Ser
Leu Gln Ser Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asp 210
215 220Thr Trp Pro Phe Thr Phe Gly Pro Gly Thr
Lys Val Asp Phe Lys Arg225 230 235
240Ala Ala Ala Ile Glu Val Met Tyr Pro Pro Pro Tyr Leu Asp Asn
Glu 245 250 255Lys Ser Asn
Gly Thr Ile Ile His Val Lys Gly Lys His Leu Cys Pro 260
265 270Ser Pro Leu Phe Pro Gly Pro Ser Lys Pro
Phe Trp Val Leu Val Val 275 280
285Val Gly Gly Val Leu Ala Cys Tyr Ser Leu Leu Val Thr Val Ala Phe 290
295 300Ile Ile Phe Trp Val Arg Ser Lys
Arg Ser Arg Leu Leu His Ser Asp305 310
315 320Tyr Met Asn Met Thr Pro Arg Arg Pro Gly Pro Thr
Arg Lys His Tyr 325 330
335Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val
340 345 350Lys Phe Ser Arg Ser Ala
Asp Ala Pro Ala Tyr Gln Gln Gly Gln Asn 355 360
365Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr
Asp Val 370 375 380Leu Asp Lys Arg Arg
Gly Arg Asp Pro Glu Met Gly Gly Lys Pro Arg385 390
395 400Arg Lys Asn Pro Gln Glu Gly Leu Tyr Asn
Glu Leu Gln Lys Asp Lys 405 410
415Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg Arg Arg
420 425 430Gly Lys Gly His Asp
Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys 435
440 445Asp Thr Tyr Asp Ala Leu His Met Gln Ala Leu Pro
Pro Arg 450 455
4601111533DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 111atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccgcaggtgc agttggttga atcaggaggg
ggtgtggtgc aacccggtcg gtcactgcgc 120ctcagttgtg ctgcttccgg gtttactttc
agctcatatg ggatgcactg ggtacggcag 180gctccaggta aaggcttgga atgggtggcg
gtgatcagct atgacggctc tgacaaatat 240tatgtggact ccgtgaaagg cagattcacc
atcagtcgag acaactcaaa gaatagactc 300tacttgcaga tgaatagcct ccgggccgaa
gatactgcag tctattattg cgcccgggag 360cgctacagtg gaagagacta ttgggggcaa
ggaactcttg tcacagtctc atctggcggc 420ggcggcagcg gtgggggcgg atctggcggg
ggcggcagcg aaatcgttat gactcagagt 480cctgccacac tgagcgttag ccctggtgag
agagcaacac ttagctgcag agctagtcag 540agtgtttcca gtcttttgac atggtaccaa
cagaagcccg gtcaagctcc acgactgctc 600atcttcggtg catccacccg cgcaaccggg
atacccgccc ggttttccgg ttctggaagt 660ggcacaggat tcacgctcac catttcttct
ctgcagtctg aagactttgc cgtgtattac 720tgccagcagt acgatacctg gccctttacc
tttggcccag gtactaaagt ggattttaaa 780cgagctgctg cactttccaa tagtattatg
tacttttcac attttgtgcc cgtgttcctg 840cctgcgaagc ctacgacaac cccagcccct
aggccgccca caccggcccc aactattgcc 900tcccagccat tgtctctgag acccgaagct
tgcagacctg ctgctggagg cgccgttcac 960acccgaggat tggatttcgc atgtgacatt
tacatctggg cccctttggc cggaacctgc 1020ggtgtgctgc tgctgtcact cgtgattaca
ctttactgca accaccgaaa cagatccaaa 1080agaagccgcc tgctccatag cgattacatg
aatatgactc cacgccgccc tggccccaca 1140aggaaacact accagcctta cgcaccacct
agagatttcg ctgcctatcg gagcagggtg 1200aagttttcca gatctgcaga tgcaccagcg
tatcagcagg gccagaacca actgtataac 1260gagctcaacc tgggacgcag ggaagagtat
gacgttttgg acaagcgcag aggacgggac 1320cctgagatgg gtggcaaacc aagacgaaaa
aacccccagg agggtctcta taatgagctg 1380cagaaggata agatggctga agcctattct
gaaataggca tgaaaggaga gcggagaagg 1440ggaaaagggc acgacggttt gtaccaggga
ctcagcactg ctacgaagga tacttatgac 1500gctctccaca tgcaagccct gccacctagg
taa 1533112509PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
112Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Gln
Val Gln Leu Val Glu Ser Gly Gly Gly Val 20 25
30Val Gln Pro Gly Arg Ser Leu Arg Leu Ser Cys Ala Ala
Ser Gly Phe 35 40 45Thr Phe Ser
Ser Tyr Gly Met His Trp Val Arg Gln Ala Pro Gly Lys 50
55 60Gly Leu Glu Trp Val Ala Val Ile Ser Tyr Asp Gly
Ser Asp Lys Tyr65 70 75
80Tyr Val Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser
85 90 95Lys Asn Arg Leu Tyr Leu
Gln Met Asn Ser Leu Arg Ala Glu Asp Thr 100
105 110Ala Val Tyr Tyr Cys Ala Arg Glu Arg Tyr Ser Gly
Arg Asp Tyr Trp 115 120 125Gly Gly
Thr Leu Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly 130
135 140Gly Gly Ser Gly Gly Gly Gly Ser Glu Ile Val
Met Thr Gln Ser Pro145 150 155
160Ala Thr Leu Ser Val Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg
165 170 175Ala Ser Gln Ser
Val Ser Ser Leu Leu Thr Trp Tyr Gln Gln Lys Pro 180
185 190Gly Gln Ala Pro Arg Leu Leu Ile Phe Gly Ala
Ser Thr Arg Ala Thr 195 200 205Gly
Ile Pro Ala Arg Phe Ser Gly Ser Gly Ser Gly Thr Gly Phe Thr 210
215 220Leu Thr Ile Ser Ser Leu Gln Ser Glu Asp
Phe Ala Val Tyr Tyr Cys225 230 235
240Gln Gln Tyr Asp Thr Trp Pro Phe Thr Phe Gly Pro Gly Thr Lys
Val 245 250 255Asp Phe Lys
Arg Ala Ala Ala Leu Ser Asn Ser Ile Met Tyr Phe Ser 260
265 270His Phe Val Pro Val Phe Leu Pro Ala Lys
Pro Thr Thr Thr Pro Ala 275 280
285Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile Ala Ser Gln Pro Leu Ser 290
295 300Leu Arg Pro Glu Ala Cys Arg Pro
Ala Ala Gly Gly Ala Val His Thr305 310
315 320Arg Gly Leu Asp Phe Ala Cys Asp Ile Tyr Ile Trp
Ala Pro Leu Ala 325 330
335Gly Thr Cys Gly Val Leu Leu Leu Ser Leu Val Ile Thr Leu Tyr Cys
340 345 350Asn His Arg Asn Arg Ser
Lys Arg Ser Arg Leu Leu His Ser Asp Tyr 355 360
365Met Asn Met Thr Pro Arg Arg Pro Gly Pro Thr Arg Lys His
Tyr Gln 370 375 380Pro Tyr Ala Pro Pro
Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val Lys385 390
395 400Phe Ser Arg Ser Ala Asp Ala Pro Ala Tyr
Gln Gln Gly Gln Asn Gln 405 410
415Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr Asp Val Leu
420 425 430Asp Lys Arg Arg Gly
Arg Asp Pro Glu Met Gly Gly Lys Pro Arg Arg 435
440 445Lys Asn Pro Gln Glu Gly Leu Tyr Asn Glu Leu Gln
Lys Asp Lys Met 450 455 460Ala Glu Ala
Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg Arg Arg Gly465
470 475 480Lys Gly His Asp Gly Leu Tyr
Gln Gly Leu Ser Thr Ala Thr Lys Asp 485
490 495Thr Tyr Asp Ala Leu His Met Gln Ala Leu Pro Pro
Arg 500 5051131467DNAArtificial
SequenceDescription of Artificial Sequence Synthetic polynucleotide
113caggtgcagt tggttgaatc aggagggggt gtggtgcaac ccggtcggtc actgcgcctc
60agttgtgctg cttccgggtt tactttcagc tcatatggga tgcactgggt acggcaggct
120ccaggtaaag gcttggaatg ggtggcggtg atcagctatg acggctctga caaatattat
180gtggactccg tgaaaggcag attcaccatc agtcgagaca actcaaagaa tagactctac
240ttgcagatga atagcctccg ggccgaagat actgcagtct attattgcgc ccgggagcgc
300tacagtggaa gagactattg ggggcaagga actcttgtca cagtctcatc tggcggcggc
360ggcagcggtg ggggcggatc tggcgggggc ggcagcgaaa tcgttatgac tcagagtcct
420gccacactga gcgttagccc tggtgagaga gcaacactta gctgcagagc tagtcagagt
480gtttccagtc ttttgacatg gtaccaacag aagcccggtc aagctccacg actgctcatc
540ttcggtgcat ccacccgcgc aaccgggata cccgcccggt tttccggttc tggaagtggc
600acaggattca cgctcaccat ttcttctctg cagtctgaag actttgccgt gtattactgc
660cagcagtacg atacctggcc ctttaccttt ggcccaggta ctaaagtgga ttttaaacga
720gctgctgcac tttccaatag tattatgtac ttttcacatt ttgtgcccgt gttcctgcct
780gcgaagccta cgacaacccc agcccctagg ccgcccacac cggccccaac tattgcctcc
840cagccattgt ctctgagacc cgaagcttgc agacctgctg ctggaggcgc cgttcacacc
900cgaggattgg atttcgcatg tgacatttac atctgggccc ctttggccgg aacctgcggt
960gtgctgctgc tgtcactcgt gattacactt tactgcaacc accgaaacag atccaaaaga
1020agccgcctgc tccatagcga ttacatgaat atgactccac gccgccctgg ccccacaagg
1080aaacactacc agccttacgc accacctaga gatttcgctg cctatcggag cagggtgaag
1140ttttccagat ctgcagatgc accagcgtat cagcagggcc agaaccaact gtataacgag
1200ctcaacctgg gacgcaggga agagtatgac gttttggaca agcgcagagg acgggaccct
1260gagatgggtg gcaaaccaag acgaaaaaac ccccaggagg gtctctataa tgagctgcag
1320aaggataaga tggctgaagc ctattctgaa ataggcatga aaggagagcg gagaagggga
1380aaagggcacg acggtttgta ccagggactc agcactgcta cgaaggatac ttatgacgct
1440ctccacatgc aagccctgcc acctagg
1467114489PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 114Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val
Val Gln Pro Gly Arg1 5 10
15Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr
20 25 30Gly Met His Trp Val Arg Gln
Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40
45Ala Val Ile Ser Tyr Asp Gly Ser Asp Lys Tyr Tyr Val Asp Ser
Val 50 55 60Lys Gly Arg Phe Thr Ile
Ser Arg Asp Asn Ser Lys Asn Arg Leu Tyr65 70
75 80Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr
Ala Val Tyr Tyr Cys 85 90
95Ala Arg Glu Arg Tyr Ser Gly Arg Asp Tyr Trp Gly Gln Gly Thr Leu
100 105 110Val Thr Val Ser Ser Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly 115 120
125Gly Gly Gly Ser Glu Ile Val Met Thr Gln Ser Pro Ala Thr
Leu Ser 130 135 140Val Ser Pro Gly Glu
Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser145 150
155 160Val Ser Ser Leu Leu Thr Trp Tyr Gln Gln
Lys Pro Gly Gln Ala Pro 165 170
175Arg Leu Leu Ile Phe Gly Ala Ser Thr Arg Ala Thr Gly Ile Pro Ala
180 185 190Arg Phe Ser Gly Ser
Gly Ser Gly Thr Gly Phe Thr Leu Thr Ile Ser 195
200 205Ser Leu Gln Ser Glu Asp Phe Ala Val Tyr Tyr Cys
Gln Gln Tyr Asp 210 215 220Thr Trp Pro
Phe Thr Phe Gly Pro Gly Thr Lys Val Asp Phe Lys Arg225
230 235 240Ala Ala Ala Leu Ser Asn Ser
Ile Met Tyr Phe Ser His Phe Val Pro 245
250 255Val Phe Leu Pro Ala Lys Pro Thr Thr Thr Pro Ala
Pro Arg Pro Pro 260 265 270Thr
Pro Ala Pro Thr Ile Ala Ser Gln Pro Leu Ser Leu Arg Pro Glu 275
280 285Ala Cys Arg Pro Ala Ala Gly Gly Ala
Val His Thr Arg Gly Leu Asp 290 295
300Phe Ala Cys Asp Ile Tyr Ile Trp Ala Pro Leu Ala Gly Thr Cys Gly305
310 315 320Val Leu Leu Leu
Ser Leu Val Ile Thr Leu Tyr Cys Asn His Arg Asn 325
330 335Arg Ser Lys Arg Ser Arg Leu Leu His Ser
Asp Tyr Met Asn Met Thr 340 345
350Pro Arg Arg Pro Gly Pro Thr Arg Lys His Tyr Gln Pro Tyr Ala Pro
355 360 365Pro Arg Asp Phe Ala Ala Tyr
Arg Ser Arg Val Lys Phe Ser Arg Ser 370 375
380Ala Asp Ala Pro Ala Tyr Gln Gln Gly Gln Asn Gln Leu Tyr Asn
Glu385 390 395 400Leu Asn
Leu Gly Arg Arg Glu Glu Tyr Asp Val Leu Asp Lys Arg Arg
405 410 415Gly Arg Asp Pro Glu Met Gly
Gly Lys Pro Arg Arg Lys Asn Pro Gln 420 425
430Glu Gly Leu Tyr Asn Glu Leu Gln Lys Asp Lys Met Ala Glu
Ala Tyr 435 440 445Ser Glu Ile Gly
Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His Asp 450
455 460Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys Asp
Thr Tyr Asp Ala465 470 475
480Leu His Met Gln Ala Leu Pro Pro Arg
4851151425DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 115atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccggagattg tgatgaccca gtcccctgct
accctgtccg tcagtccggg cgagagagcc 120accttgtcat gccgggccag ccagtccgtc
agcagtctcc tgacttggta tcagcaaaaa 180ccagggcagg caccgcggct tttgattttt
ggtgcaagca cacgcgccac tggcattcca 240gctaggtttt ctggaagtgg atctgggaca
ggcttcactc tgacaatcag tagcctgcag 300agtgaggact ttgctgttta ctactgtcaa
cagtacgaca cctggccatt cacattcggg 360cccggcacca aggtcgactt caagaggggc
ggtggaggtt caggtggtgg cgggtcaggc 420ggcggtgggt ctcaggttca actggtggaa
tcaggtggcg gcgttgtcca accggggcga 480tcacttcgac tttcctgtgc tgcctcaggc
tttacttttt catcctatgg gatgcactgg 540gttcggcagg ctcccggaaa aggactcgag
tgggttgcag tgatctctta cgatggctca 600gacaagtatt atgtggactc agtcaagggg
agattcacaa taagccgaga caactccaaa 660aaccggcttt atctccagat gaacagcctt
agagcggaag ataccgcggt atactactgt 720gcccgcgaga ggtattccgg cagagactac
tggggacagg gcacactggt caccgtgagt 780tctgccgcag cgctcgataa cgaaaagagc
aacggaacca ttatccacgt taagggcaag 840cacctgtgcc ccagtcccct cttcccagga
ccatctaaac ccttctgggt tctggtagta 900gttggagggg tccttgcatg ttactccctt
ttggtcaccg tcgccttcat tattttctgg 960gtgagatcca aaagaagccg cctgctccat
agcgattaca tgaatatgac tccacgccgc 1020cctggcccca caaggaaaca ctaccagcct
tacgcaccac ctagagattt cgctgcctat 1080cggagcaggg tgaagttttc cagatctgca
gatgcaccag cgtatcagca gggccagaac 1140caactgtata acgagctcaa cctgggacgc
agggaagagt atgacgtttt ggacaagcgc 1200agaggacggg accctgagat gggtggcaaa
ccaagacgaa aaaaccccca ggagggtctc 1260tataatgagc tgcagaagga taagatggct
gaagcctatt ctgaaatagg catgaaagga 1320gagcggagaa ggggaaaagg gcacgacggt
ttgtaccagg gactcagcac tgctacgaag 1380gatacttatg acgctctcca catgcaagcc
ctgccaccta ggtaa 1425116474PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
116Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Glu
Ile Val Met Thr Gln Ser Pro Ala Thr Leu 20 25
30Ser Val Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg
Ala Ser Gln 35 40 45Ser Val Ser
Ser Leu Leu Thr Trp Tyr Gln Gln Lys Pro Gly Gln Ala 50
55 60Pro Arg Leu Leu Ile Phe Gly Ala Ser Thr Arg Ala
Thr Gly Ile Pro65 70 75
80Ala Arg Phe Ser Gly Ser Gly Ser Gly Thr Gly Phe Thr Leu Thr Ile
85 90 95Ser Ser Leu Gln Ser Glu
Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr 100
105 110Asp Thr Trp Pro Phe Thr Phe Gly Pro Gly Thr Lys
Val Asp Phe Lys 115 120 125Arg Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 130
135 140Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val
Val Gln Pro Gly Arg145 150 155
160Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr
165 170 175Gly Met His Trp
Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 180
185 190Ala Val Ile Ser Tyr Asp Gly Ser Asp Lys Tyr
Tyr Val Asp Ser Val 195 200 205Lys
Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Arg Leu Tyr 210
215 220Leu Gln Met Asn Ser Leu Arg Ala Glu Asp
Thr Ala Val Tyr Tyr Cys225 230 235
240Ala Arg Glu Arg Tyr Ser Gly Arg Asp Tyr Trp Gly Gln Gly Thr
Leu 245 250 255Val Thr Val
Ser Ser Ala Ala Ala Leu Asp Asn Glu Lys Ser Asn Gly 260
265 270Thr Ile Ile His Val Lys Gly Lys His Leu
Cys Pro Ser Pro Leu Phe 275 280
285Pro Gly Pro Ser Lys Pro Phe Trp Val Leu Val Val Val Gly Gly Val 290
295 300Leu Ala Cys Tyr Ser Leu Leu Val
Thr Val Ala Phe Ile Ile Phe Trp305 310
315 320Val Arg Ser Lys Arg Ser Arg Leu Leu His Ser Asp
Tyr Met Asn Met 325 330
335Thr Pro Arg Arg Pro Gly Pro Thr Arg Lys His Tyr Gln Pro Tyr Ala
340 345 350Pro Pro Arg Asp Phe Ala
Ala Tyr Arg Ser Arg Val Lys Phe Ser Arg 355 360
365Ser Ala Asp Ala Pro Ala Tyr Gln Gln Gly Gln Asn Gln Leu
Tyr Asn 370 375 380Glu Leu Asn Leu Gly
Arg Arg Glu Glu Tyr Asp Val Leu Asp Lys Arg385 390
395 400Arg Gly Arg Asp Pro Glu Met Gly Gly Lys
Pro Arg Arg Lys Asn Pro 405 410
415Gln Glu Gly Leu Tyr Asn Glu Leu Gln Lys Asp Lys Met Ala Glu Ala
420 425 430Tyr Ser Glu Ile Gly
Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His 435
440 445Asp Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys
Asp Thr Tyr Asp 450 455 460Ala Leu His
Met Gln Ala Leu Pro Pro Arg465 4701171359DNAArtificial
SequenceDescription of Artificial Sequence Synthetic polynucleotide
117gagattgtga tgacccagtc ccctgctacc ctgtccgtca gtccgggcga gagagccacc
60ttgtcatgcc gggccagcca gtccgtcagc agtctcctga cttggtatca gcaaaaacca
120gggcaggcac cgcggctttt gatttttggt gcaagcacac gcgccactgg cattccagct
180aggttttctg gaagtggatc tgggacaggc ttcactctga caatcagtag cctgcagagt
240gaggactttg ctgtttacta ctgtcaacag tacgacacct ggccattcac attcgggccc
300ggcaccaagg tcgacttcaa gaggggcggt ggaggttcag gtggtggcgg gtcaggcggc
360ggtgggtctc aggttcaact ggtggaatca ggtggcggcg ttgtccaacc ggggcgatca
420cttcgacttt cctgtgctgc ctcaggcttt actttttcat cctatgggat gcactgggtt
480cggcaggctc ccggaaaagg actcgagtgg gttgcagtga tctcttacga tggctcagac
540aagtattatg tggactcagt caaggggaga ttcacaataa gccgagacaa ctccaaaaac
600cggctttatc tccagatgaa cagccttaga gcggaagata ccgcggtata ctactgtgcc
660cgcgagaggt attccggcag agactactgg ggacagggca cactggtcac cgtgagttct
720gccgcagcgc tcgataacga aaagagcaac ggaaccatta tccacgttaa gggcaagcac
780ctgtgcccca gtcccctctt cccaggacca tctaaaccct tctgggttct ggtagtagtt
840ggaggggtcc ttgcatgtta ctcccttttg gtcaccgtcg ccttcattat tttctgggtg
900agatccaaaa gaagccgcct gctccatagc gattacatga atatgactcc acgccgccct
960ggccccacaa ggaaacacta ccagccttac gcaccaccta gagatttcgc tgcctatcgg
1020agcagggtga agttttccag atctgcagat gcaccagcgt atcagcaggg ccagaaccaa
1080ctgtataacg agctcaacct gggacgcagg gaagagtatg acgttttgga caagcgcaga
1140ggacgggacc ctgagatggg tggcaaacca agacgaaaaa acccccagga gggtctctat
1200aatgagctgc agaaggataa gatggctgaa gcctattctg aaataggcat gaaaggagag
1260cggagaaggg gaaaagggca cgacggtttg taccagggac tcagcactgc tacgaaggat
1320acttatgacg ctctccacat gcaagccctg ccacctagg
1359118453PRTArtificial SequenceDescription of Artificial Sequence
Synthetic polypeptide 118Glu Ile Val Met Thr Gln Ser Pro Ala Thr Leu
Ser Val Ser Pro Gly1 5 10
15Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Leu
20 25 30Leu Thr Trp Tyr Gln Gln Lys
Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40
45Phe Gly Ala Ser Thr Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser
Gly 50 55 60Ser Gly Ser Gly Thr Gly
Phe Thr Leu Thr Ile Ser Ser Leu Gln Ser65 70
75 80Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr
Asp Thr Trp Pro Phe 85 90
95Thr Phe Gly Pro Gly Thr Lys Val Asp Phe Lys Arg Gly Gly Gly Gly
100 105 110Ser Gly Gly Gly Gly Ser
Gly Gly Gly Gly Ser Gln Val Gln Leu Val 115 120
125Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg Ser Leu Arg
Leu Ser 130 135 140Cys Ala Ala Ser Gly
Phe Thr Phe Ser Ser Tyr Gly Met His Trp Val145 150
155 160Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp
Val Ala Val Ile Ser Tyr 165 170
175Asp Gly Ser Asp Lys Tyr Tyr Val Asp Ser Val Lys Gly Arg Phe Thr
180 185 190Ile Ser Arg Asp Asn
Ser Lys Asn Arg Leu Tyr Leu Gln Met Asn Ser 195
200 205Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala
Arg Glu Arg Tyr 210 215 220Ser Gly Arg
Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser225
230 235 240Ala Ala Ala Leu Asp Asn Glu
Lys Ser Asn Gly Thr Ile Ile His Val 245
250 255Lys Gly Lys His Leu Cys Pro Ser Pro Leu Phe Pro
Gly Pro Ser Lys 260 265 270Pro
Phe Trp Val Leu Val Val Val Gly Gly Val Leu Ala Cys Tyr Ser 275
280 285Leu Leu Val Thr Val Ala Phe Ile Ile
Phe Trp Val Arg Ser Lys Arg 290 295
300Ser Arg Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro305
310 315 320Gly Pro Thr Arg
Lys His Tyr Gln Pro Tyr Ala Pro Pro Arg Asp Phe 325
330 335Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser
Arg Ser Ala Asp Ala Pro 340 345
350Ala Tyr Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly
355 360 365Arg Arg Glu Glu Tyr Asp Val
Leu Asp Lys Arg Arg Gly Arg Asp Pro 370 375
380Glu Met Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu
Tyr385 390 395 400Asn Glu
Leu Gln Lys Asp Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly
405 410 415Met Lys Gly Glu Arg Arg Arg
Gly Lys Gly His Asp Gly Leu Tyr Gln 420 425
430Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu His
Met Gln 435 440 445Ala Leu Pro Pro
Arg 4501191452DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 119atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccggagatcg tcatgacaca gagtccagct
accctgagcg tgtcccctgg agagagagcc 120accctgtcct gtagggctag tcagagtgtg
tccagcctcc tcacctggta tcaacagaag 180cctggtcaag ctccccggct gcttatcttc
ggggccagca cgcgagccac aggcatcccg 240gccagattct ctggctctgg cagtggcacc
gggttcactc tcacgatctc atccctgcag 300tcagaggatt tcgctgtgta ttactgtcag
cagtacgata catggccctt caccttcggc 360ccgggcacaa aagtagattt caagcgcggc
ggcgggggta gtgggggcgg gggatcagga 420ggagggggct cccaagtaca gctggttgag
agcggcggcg gggtggttca gcccgggcgc 480agcctcaggc tgagttgcgc agcatcagga
ttcacattca gttcttatgg aatgcattgg 540gtcagacagg ctcccgggaa gggccttgaa
tgggtggcag tcattagcta cgacggaagc 600gataagtact atgtggactc agttaaaggg
agatttacta tcagccgcga caattccaaa 660aacagattgt atttgcagat gaactccctc
agggcggagg acactgctgt atattactgc 720gcacgagaga gatactccgg ccgagactat
tggggccaag gaacattggt aactgtgagc 780tccgccgcag ctattgaggt catgtacccc
ccaccttatc tcgataatga gaagagtaat 840gggactataa ttcacgtaaa gggcaaacac
ctgtgccctt ccccgctgtt tccaggtcca 900agtaagccgt tctgggtcct ggttgtggtg
ggaggggtgc tggcctgcta ttctctgttg 960gttaccgtgg cctttatcat tttctgggtg
agatccaaaa gaagccgcct gctccatagc 1020gattacatga atatgactcc acgccgccct
ggccccacaa ggaaacacta ccagccttac 1080gcaccaccta gagatttcgc tgcctatcgg
agcagggtga agttttccag atctgcagat 1140gcaccagcgt atcagcaggg ccagaaccaa
ctgtataacg agctcaacct gggacgcagg 1200gaagagtatg acgttttgga caagcgcaga
ggacgggacc ctgagatggg tggcaaacca 1260agacgaaaaa acccccagga gggtctctat
aatgagctgc agaaggataa gatggctgaa 1320gcctattctg aaataggcat gaaaggagag
cggagaaggg gaaaagggca cgacggtttg 1380taccagggac tcagcactgc tacgaaggat
acttatgacg ctctccacat gcaagccctg 1440ccacctaggt aa
1452120483PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
120Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Glu
Ile Val Met Thr Gln Ser Pro Ala Thr Leu 20 25
30Ser Val Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg
Ala Ser Gln 35 40 45Ser Val Ser
Ser Leu Leu Thr Trp Tyr Gln Gln Lys Pro Gly Gln Ala 50
55 60Pro Arg Leu Leu Ile Phe Gly Ala Ser Thr Arg Ala
Thr Gly Ile Pro65 70 75
80Ala Arg Phe Ser Gly Ser Gly Ser Gly Thr Gly Phe Thr Leu Thr Ile
85 90 95Ser Ser Leu Gln Ser Glu
Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr 100
105 110Asp Thr Trp Pro Phe Thr Phe Gly Pro Gly Thr Lys
Val Asp Phe Lys 115 120 125Arg Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 130
135 140Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val
Val Gln Pro Gly Arg145 150 155
160Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr
165 170 175Gly Met His Trp
Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 180
185 190Ala Val Ile Ser Tyr Asp Gly Ser Asp Lys Tyr
Tyr Val Asp Ser Val 195 200 205Lys
Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Arg Leu Tyr 210
215 220Leu Gln Met Asn Ser Leu Arg Ala Glu Asp
Thr Ala Val Tyr Tyr Cys225 230 235
240Ala Arg Glu Arg Tyr Ser Gly Arg Asp Tyr Trp Gly Gln Gly Thr
Leu 245 250 255Val Thr Val
Ser Ser Ala Ala Ala Ile Glu Val Met Tyr Pro Pro Pro 260
265 270Tyr Leu Asp Asn Glu Lys Ser Asn Gly Thr
Ile Ile His Val Lys Gly 275 280
285Lys His Leu Cys Pro Ser Pro Leu Phe Pro Gly Pro Ser Lys Pro Phe 290
295 300Trp Val Leu Val Val Val Gly Gly
Val Leu Ala Cys Tyr Ser Leu Leu305 310
315 320Val Thr Val Ala Phe Ile Ile Phe Trp Val Arg Ser
Lys Arg Ser Arg 325 330
335Leu Leu His Ser Asp Tyr Met Asn Met Thr Pro Arg Arg Pro Gly Pro
340 345 350Thr Arg Lys His Tyr Gln
Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala 355 360
365Tyr Arg Ser Arg Val Lys Phe Ser Arg Ser Ala Asp Ala Pro
Ala Tyr 370 375 380Gln Gln Gly Gln Asn
Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg385 390
395 400Glu Glu Tyr Asp Val Leu Asp Lys Arg Arg
Gly Arg Asp Pro Glu Met 405 410
415Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu Tyr Asn Glu
420 425 430Leu Gln Lys Asp Lys
Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys 435
440 445Gly Glu Arg Arg Arg Gly Lys Gly His Asp Gly Leu
Tyr Gln Gly Leu 450 455 460Ser Thr Ala
Thr Lys Asp Thr Tyr Asp Ala Leu His Met Gln Ala Leu465
470 475 480Pro Pro
Arg1211386DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 121gagatcgtca tgacacagag tccagctacc
ctgagcgtgt cccctggaga gagagccacc 60ctgtcctgta gggctagtca gagtgtgtcc
agcctcctca cctggtatca acagaagcct 120ggtcaagctc cccggctgct tatcttcggg
gccagcacgc gagccacagg catcccggcc 180agattctctg gctctggcag tggcaccggg
ttcactctca cgatctcatc cctgcagtca 240gaggatttcg ctgtgtatta ctgtcagcag
tacgatacat ggcccttcac cttcggcccg 300ggcacaaaag tagatttcaa gcgcggcggc
gggggtagtg ggggcggggg atcaggagga 360gggggctccc aagtacagct ggttgagagc
ggcggcgggg tggttcagcc cgggcgcagc 420ctcaggctga gttgcgcagc atcaggattc
acattcagtt cttatggaat gcattgggtc 480agacaggctc ccgggaaggg ccttgaatgg
gtggcagtca ttagctacga cggaagcgat 540aagtactatg tggactcagt taaagggaga
tttactatca gccgcgacaa ttccaaaaac 600agattgtatt tgcagatgaa ctccctcagg
gcggaggaca ctgctgtata ttactgcgca 660cgagagagat actccggccg agactattgg
ggccaaggaa cattggtaac tgtgagctcc 720gccgcagcta ttgaggtcat gtacccccca
ccttatctcg ataatgagaa gagtaatggg 780actataattc acgtaaaggg caaacacctg
tgcccttccc cgctgtttcc aggtccaagt 840aagccgttct gggtcctggt tgtggtggga
ggggtgctgg cctgctattc tctgttggtt 900accgtggcct ttatcatttt ctgggtgaga
tccaaaagaa gccgcctgct ccatagcgat 960tacatgaata tgactccacg ccgccctggc
cccacaagga aacactacca gccttacgca 1020ccacctagag atttcgctgc ctatcggagc
agggtgaagt tttccagatc tgcagatgca 1080ccagcgtatc agcagggcca gaaccaactg
tataacgagc tcaacctggg acgcagggaa 1140gagtatgacg ttttggacaa gcgcagagga
cgggaccctg agatgggtgg caaaccaaga 1200cgaaaaaacc cccaggaggg tctctataat
gagctgcaga aggataagat ggctgaagcc 1260tattctgaaa taggcatgaa aggagagcgg
agaaggggaa aagggcacga cggtttgtac 1320cagggactca gcactgctac gaaggatact
tatgacgctc tccacatgca agccctgcca 1380cctagg
1386122462PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
122Glu Ile Val Met Thr Gln Ser Pro Ala Thr Leu Ser Val Ser Pro Gly1
5 10 15Glu Arg Ala Thr Leu Ser
Cys Arg Ala Ser Gln Ser Val Ser Ser Leu 20 25
30Leu Thr Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg
Leu Leu Ile 35 40 45Phe Gly Ala
Ser Thr Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50
55 60Ser Gly Ser Gly Thr Gly Phe Thr Leu Thr Ile Ser
Ser Leu Gln Ser65 70 75
80Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asp Thr Trp Pro Phe
85 90 95Thr Phe Gly Pro Gly Thr
Lys Val Asp Phe Lys Arg Gly Gly Gly Gly 100
105 110Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gln
Val Gln Leu Val 115 120 125Glu Ser
Gly Gly Gly Val Val Gln Pro Gly Arg Ser Leu Arg Leu Ser 130
135 140Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr
Gly Met His Trp Val145 150 155
160Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ala Val Ile Ser Tyr
165 170 175Asp Gly Ser Asp
Lys Tyr Tyr Val Asp Ser Val Lys Gly Arg Phe Thr 180
185 190Ile Ser Arg Asp Asn Ser Lys Asn Arg Leu Tyr
Leu Gln Met Asn Ser 195 200 205Leu
Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg Glu Arg Tyr 210
215 220Ser Gly Arg Asp Tyr Trp Gly Gln Gly Thr
Leu Val Thr Val Ser Ser225 230 235
240Ala Ala Ala Ile Glu Val Met Tyr Pro Pro Pro Tyr Leu Asp Asn
Glu 245 250 255Lys Ser Asn
Gly Thr Ile Ile His Val Lys Gly Lys His Leu Cys Pro 260
265 270Ser Pro Leu Phe Pro Gly Pro Ser Lys Pro
Phe Trp Val Leu Val Val 275 280
285Val Gly Gly Val Leu Ala Cys Tyr Ser Leu Leu Val Thr Val Ala Phe 290
295 300Ile Ile Phe Trp Val Arg Ser Lys
Arg Ser Arg Leu Leu His Ser Asp305 310
315 320Tyr Met Asn Met Thr Pro Arg Arg Pro Gly Pro Thr
Arg Lys His Tyr 325 330
335Gln Pro Tyr Ala Pro Pro Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val
340 345 350Lys Phe Ser Arg Ser Ala
Asp Ala Pro Ala Tyr Gln Gln Gly Gln Asn 355 360
365Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr
Asp Val 370 375 380Leu Asp Lys Arg Arg
Gly Arg Asp Pro Glu Met Gly Gly Lys Pro Arg385 390
395 400Arg Lys Asn Pro Gln Glu Gly Leu Tyr Asn
Glu Leu Gln Lys Asp Lys 405 410
415Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg Arg Arg
420 425 430Gly Lys Gly His Asp
Gly Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys 435
440 445Asp Thr Tyr Asp Ala Leu His Met Gln Ala Leu Pro
Pro Arg 450 455
4601231533DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 123atggcactcc ccgtaactgc tctgctgctg
ccgttggcat tgctcctgca cgccgcacgc 60ccggaaatag tgatgactca gtccccggcc
accctcagcg tgtcccccgg ggagcgagcg 120accctgtcat gcagggcttc ccagagtgtc
agctccctgc tcacttggta tcagcaaaag 180ccggggcagg ctccccgcct cctcatcttc
ggggcatcaa ctagggccac cggcattcct 240gcaagatttt ccgggtctgg cagcggcacc
ggcttcaccc ttaccattag ctctctgcag 300tctgaggact tcgccgttta ctattgtcag
cagtatgata cttggccctt taccttcggt 360cccggaacta aggtggactt caagcgcggg
gggggtggat ctggaggtgg tggctccggg 420ggcggtggaa gccaggtcca gttggttgag
agcggcggcg gagtggtgca gcccgggagg 480tccttgcggc tgagctgtgc agcctccggt
tttacttttt ctagctatgg aatgcattgg 540gtaagacagg ctcccggaaa aggcctcgag
tgggtggcgg tcattagcta tgatggatct 600gataaatact atgtggactc agttaagggg
cgcttcacaa tctcaagaga caatagcaaa 660aatagactgt acctgcagat gaatagtctg
cgcgccgagg acactgccgt gtactactgc 720gcccgcgaga gatacagcgg acgggattac
tggggccagg gtaccctcgt aacggtgtcc 780tccgctgccg cccttagcaa cagcattatg
tacttttctc atttcgtgcc agtctttctc 840ccagcaaagc ccaccactac cccggccccc
aggccgccta ctcctgcccc cactatcgcg 900tctcagcctc tctccttgcg gcccgaggcc
tgccggccag ccgcaggggg cgccgtacat 960actcggggtt tggatttcgc ttgcgacata
tatatttggg cccccctcgc cggcacatgt 1020ggagtgctgc tcctgagtct cgttataacc
ctctattgca accatagaaa cagatccaaa 1080agaagccgcc tgctccatag cgattacatg
aatatgactc cacgccgccc tggccccaca 1140aggaaacact accagcctta cgcaccacct
agagatttcg ctgcctatcg gagcagggtg 1200aagttttcca gatctgcaga tgcaccagcg
tatcagcagg gccagaacca actgtataac 1260gagctcaacc tgggacgcag ggaagagtat
gacgttttgg acaagcgcag aggacgggac 1320cctgagatgg gtggcaaacc aagacgaaaa
aacccccagg agggtctcta taatgagctg 1380cagaaggata agatggctga agcctattct
gaaataggca tgaaaggaga gcggagaagg 1440ggaaaagggc acgacggttt gtaccaggga
ctcagcactg ctacgaagga tacttatgac 1500gctctccaca tgcaagccct gccacctagg
taa 1533124510PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
124Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro Glu
Ile Val Met Thr Gln Ser Pro Ala Thr Leu 20 25
30Ser Val Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg
Ala Ser Gln 35 40 45Ser Val Ser
Ser Leu Leu Thr Trp Tyr Gln Gln Lys Pro Gly Gln Ala 50
55 60Pro Arg Leu Leu Ile Phe Gly Ala Ser Thr Arg Ala
Thr Gly Ile Pro65 70 75
80Ala Arg Phe Ser Gly Ser Gly Ser Gly Thr Gly Phe Thr Leu Thr Ile
85 90 95Ser Ser Leu Gln Ser Glu
Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr 100
105 110Asp Thr Trp Pro Phe Thr Phe Gly Pro Gly Thr Lys
Val Asp Phe Lys 115 120 125Arg Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 130
135 140Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val
Val Gln Pro Gly Arg145 150 155
160Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr
165 170 175Gly Met His Trp
Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 180
185 190Ala Val Ile Ser Tyr Asp Gly Ser Asp Lys Tyr
Tyr Val Asp Ser Val 195 200 205Lys
Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Arg Leu Tyr 210
215 220Leu Gln Met Asn Ser Leu Arg Ala Glu Asp
Thr Ala Val Tyr Tyr Cys225 230 235
240Ala Arg Glu Arg Tyr Ser Gly Arg Asp Tyr Trp Gly Gln Gly Thr
Leu 245 250 255Val Thr Val
Ser Ser Ala Ala Ala Leu Ser Asn Ser Ile Met Tyr Phe 260
265 270Ser His Phe Val Pro Val Phe Leu Pro Ala
Lys Pro Thr Thr Thr Pro 275 280
285Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile Ala Ser Gln Pro Leu 290
295 300Ser Leu Arg Pro Glu Ala Cys Arg
Pro Ala Ala Gly Gly Ala Val His305 310
315 320Thr Arg Gly Leu Asp Phe Ala Cys Asp Ile Tyr Ile
Trp Ala Pro Leu 325 330
335Ala Gly Thr Cys Gly Val Leu Leu Leu Ser Leu Val Ile Thr Leu Tyr
340 345 350Cys Asn His Arg Asn Arg
Ser Lys Arg Ser Arg Leu Leu His Ser Asp 355 360
365Tyr Met Asn Met Thr Pro Arg Arg Pro Gly Pro Thr Arg Lys
His Tyr 370 375 380Gln Pro Tyr Ala Pro
Pro Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val385 390
395 400Lys Phe Ser Arg Ser Ala Asp Ala Pro Ala
Tyr Gln Gln Gly Gln Asn 405 410
415Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr Asp Val
420 425 430Leu Asp Lys Arg Arg
Gly Arg Asp Pro Glu Met Gly Gly Lys Pro Arg 435
440 445Arg Lys Asn Pro Gln Glu Gly Leu Tyr Asn Glu Leu
Gln Lys Asp Lys 450 455 460Met Ala Glu
Ala Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg Arg Arg465
470 475 480Gly Lys Gly His Asp Gly Leu
Tyr Gln Gly Leu Ser Thr Ala Thr Lys 485
490 495Asp Thr Tyr Asp Ala Leu His Met Gln Ala Leu Pro
Pro Arg 500 505
5101251467DNAArtificial SequenceDescription of Artificial Sequence
Synthetic polynucleotide 125gaaatagtga tgactcagtc cccggccacc
ctcagcgtgt cccccgggga gcgagcgacc 60ctgtcatgca gggcttccca gagtgtcagc
tccctgctca cttggtatca gcaaaagccg 120gggcaggctc cccgcctcct catcttcggg
gcatcaacta gggccaccgg cattcctgca 180agattttccg ggtctggcag cggcaccggc
ttcaccctta ccattagctc tctgcagtct 240gaggacttcg ccgtttacta ttgtcagcag
tatgatactt ggccctttac cttcggtccc 300ggaactaagg tggacttcaa gcgcgggggg
ggtggatctg gaggtggtgg ctccgggggc 360ggtggaagcc aggtccagtt ggttgagagc
ggcggcggag tggtgcagcc cgggaggtcc 420ttgcggctga gctgtgcagc ctccggtttt
actttttcta gctatggaat gcattgggta 480agacaggctc ccggaaaagg cctcgagtgg
gtggcggtca ttagctatga tggatctgat 540aaatactatg tggactcagt taaggggcgc
ttcacaatct caagagacaa tagcaaaaat 600agactgtacc tgcagatgaa tagtctgcgc
gccgaggaca ctgccgtgta ctactgcgcc 660cgcgagagat acagcggacg ggattactgg
ggccagggta ccctcgtaac ggtgtcctcc 720gctgccgccc ttagcaacag cattatgtac
ttttctcatt tcgtgccagt ctttctccca 780gcaaagccca ccactacccc ggcccccagg
ccgcctactc ctgcccccac tatcgcgtct 840cagcctctct ccttgcggcc cgaggcctgc
cggccagccg cagggggcgc cgtacatact 900cggggtttgg atttcgcttg cgacatatat
atttgggccc ccctcgccgg cacatgtgga 960gtgctgctcc tgagtctcgt tataaccctc
tattgcaacc atagaaacag atccaaaaga 1020agccgcctgc tccatagcga ttacatgaat
atgactccac gccgccctgg ccccacaagg 1080aaacactacc agccttacgc accacctaga
gatttcgctg cctatcggag cagggtgaag 1140ttttccagat ctgcagatgc accagcgtat
cagcagggcc agaaccaact gtataacgag 1200ctcaacctgg gacgcaggga agagtatgac
gttttggaca agcgcagagg acgggaccct 1260gagatgggtg gcaaaccaag acgaaaaaac
ccccaggagg gtctctataa tgagctgcag 1320aaggataaga tggctgaagc ctattctgaa
ataggcatga aaggagagcg gagaagggga 1380aaagggcacg acggtttgta ccagggactc
agcactgcta cgaaggatac ttatgacgct 1440ctccacatgc aagccctgcc acctagg
1467126489PRTArtificial
SequenceDescription of Artificial Sequence Synthetic polypeptide
126Glu Ile Val Met Thr Gln Ser Pro Ala Thr Leu Ser Val Ser Pro Gly1
5 10 15Glu Arg Ala Thr Leu Ser
Cys Arg Ala Ser Gln Ser Val Ser Ser Leu 20 25
30Leu Thr Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg
Leu Leu Ile 35 40 45Phe Gly Ala
Ser Thr Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50
55 60Ser Gly Ser Gly Thr Gly Phe Thr Leu Thr Ile Ser
Ser Leu Gln Ser65 70 75
80Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asp Thr Trp Pro Phe
85 90 95Thr Phe Gly Pro Gly Thr
Lys Val Asp Phe Lys Arg Gly Gly Gly Gly 100
105 110Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gln
Val Gln Leu Val 115 120 125Glu Ser
Gly Gly Gly Val Val Gln Pro Gly Arg Ser Leu Arg Leu Ser 130
135 140Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr
Gly Met His Trp Val145 150 155
160Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ala Val Ile Ser Tyr
165 170 175Asp Gly Ser Asp
Lys Tyr Tyr Val Asp Ser Val Lys Gly Arg Phe Thr 180
185 190Ile Ser Arg Asp Asn Ser Lys Asn Arg Leu Tyr
Leu Gln Met Asn Ser 195 200 205Leu
Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg Glu Arg Tyr 210
215 220Ser Gly Arg Asp Tyr Trp Gly Gln Gly Thr
Leu Val Thr Val Ser Ser225 230 235
240Ala Ala Ala Leu Ser Asn Ser Ile Met Tyr Phe Ser His Phe Val
Pro 245 250 255Val Phe Leu
Pro Ala Lys Pro Thr Thr Thr Pro Ala Pro Arg Pro Pro 260
265 270Thr Pro Ala Pro Thr Ile Ala Ser Gln Pro
Leu Ser Leu Arg Pro Glu 275 280
285Ala Cys Arg Pro Ala Ala Gly Gly Ala Val His Thr Arg Gly Leu Asp 290
295 300Phe Ala Cys Asp Ile Tyr Ile Trp
Ala Pro Leu Ala Gly Thr Cys Gly305 310
315 320Val Leu Leu Leu Ser Leu Val Ile Thr Leu Tyr Cys
Asn His Arg Asn 325 330
335Arg Ser Lys Arg Ser Arg Leu Leu His Ser Asp Tyr Met Asn Met Thr
340 345 350Pro Arg Arg Pro Gly Pro
Thr Arg Lys His Tyr Gln Pro Tyr Ala Pro 355 360
365Pro Arg Asp Phe Ala Ala Tyr Arg Ser Arg Val Lys Phe Ser
Arg Ser 370 375 380Ala Asp Ala Pro Ala
Tyr Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu385 390
395 400Leu Asn Leu Gly Arg Arg Glu Glu Tyr Asp
Val Leu Asp Lys Arg Arg 405 410
415Gly Arg Asp Pro Glu Met Gly Gly Lys Pro Arg Arg Lys Asn Pro Gln
420 425 430Glu Gly Leu Tyr Asn
Glu Leu Gln Lys Asp Lys Met Ala Glu Ala Tyr 435
440 445Ser Glu Ile Gly Met Lys Gly Glu Arg Arg Arg Gly
Lys Gly His Asp 450 455 460Gly Leu Tyr
Gln Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala465
470 475 480Leu His Met Gln Ala Leu Pro
Pro Arg 48512763DNAHomo sapiens 127atggcactcc ccgtaactgc
tctgctgctg ccgttggcat tgctcctgca cgccgcacgc 60ccg
6312821PRTHomo sapiens
128Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu1
5 10 15His Ala Ala Arg Pro
2012945DNAArtificial SequenceDescription of Artificial Sequence
Synthetic oligonucleotide 129ggcggtggag gctccggagg ggggggctct
ggcggagggg gctcc 4513015PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 130Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser1 5
10 1513154DNAArtificial
SequenceDescription of Artificial Sequence Synthetic oligonucleotide
131gggtctacat ccggctccgg gaagcccgga agtggcgaag gtagtacaaa gggg
5413218PRTArtificial SequenceDescription of Artificial Sequence Synthetic
peptide 132Gly Ser Thr Ser Gly Ser Gly Lys Pro Gly Ser Gly Glu Gly
Ser Thr1 5 10 15Lys
Gly133152PRTHomo sapiens 133Met Leu Arg Leu Leu Leu Ala Leu Asn Leu Phe
Pro Ser Ile Gln Val1 5 10
15Thr Gly Asn Lys Ile Leu Val Lys Gln Ser Pro Met Leu Val Ala Tyr
20 25 30Asp Asn Ala Val Asn Leu Ser
Cys Lys Tyr Ser Tyr Asn Leu Phe Ser 35 40
45Arg Glu Phe Arg Ala Ser Leu His Lys Gly Leu Asp Ser Ala Val
Glu 50 55 60Val Cys Val Val Tyr Gly
Asn Tyr Ser Gln Gln Leu Gln Val Tyr Ser65 70
75 80Lys Thr Gly Phe Asn Cys Asp Gly Lys Leu Gly
Asn Glu Ser Val Thr 85 90
95Phe Tyr Leu Gln Asn Leu Tyr Val Asn Gln Thr Asp Ile Tyr Phe Cys
100 105 110Lys Ile Glu Val Met Tyr
Pro Pro Pro Tyr Leu Asp Asn Glu Lys Ser 115 120
125Asn Gly Thr Ile Ile His Val Lys Gly Lys His Leu Cys Pro
Ser Pro 130 135 140Leu Phe Pro Gly Pro
Ser Lys Pro145 1501349PRTArtificial SequenceDescription
of Artificial Sequence Synthetic peptideMOD_RES(2)..(2)Gly, Phe or
TyrMOD_RES(3)..(3)Ser or ThrMOD_RES(4)..(4)Ile, Phe or
LeuMOD_RES(5)..(5)Ser or ThrMOD_RES(6)..(7)May or may not be
presentMOD_RES(8)..(8)Glu, Gly or absentMOD_RES(9)..(9)Phe, Leu or Tyr
134Gly Xaa Xaa Xaa Xaa Ser Gly Xaa Xaa1 51356PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
peptideMOD_RES(1)..(1)Asp, His, Ser or TyrMOD_RES(2)..(2)His, Pro or
TyrMOD_RES(3)..(3)Asp, Glu or SerMOD_RES(4)..(4)Asp or
GlyMOD_RES(5)..(5)Gly or SerMOD_RES(6)..(6)Asp, Glu or absent 135Xaa Xaa
Xaa Xaa Xaa Xaa1 513614PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptideMOD_RES(1)..(1)Glu or
LeuMOD_RES(2)..(2)Arg, Ser or ValMOD_RES(3)..(3)Arg or
TyrMOD_RES(4)..(4)Cys, Gly or SerMOD_RES(5)..(5)Gly, Ile or
absentMOD_RES(6)..(8)May or may not be presentMOD_RES(9)..(9)Trp, Tyr or
absentMOD_RES(10)..(10)Pro, Ser or absentMOD_RES(11)..(11)Gly, Tyr or
absentMOD_RES(12)..(12)Phe or Arg 136Xaa Xaa Xaa Xaa Xaa Gly Asp Cys Xaa
Xaa Xaa Xaa Asp Tyr1 5
1013711PRTArtificial SequenceDescription of Artificial Sequence Synthetic
peptideMOD_RES(1)..(1)Gln or ArgMOD_RES(5)..(5)Asp or
SerMOD_RES(6)..(6)Ile or ValMOD_RES(7)..(8)Asn or SerMOD_RES(9)..(9)Phe,
Leu or TyrMOD_RES(11)..(11)Asn or Thr 137Xaa Ala Ser Gln Xaa Xaa Xaa Xaa
Xaa Leu Xaa1 5 101387PRTArtificial
SequenceDescription of Artificial Sequence Synthetic
peptideMOD_RES(1)..(1)Asp or GlyMOD_RES(4)..(4)Asn, Ser or
ThrMOD_RES(5)..(5)Leu or ArgMOD_RES(6)..(6)Ala, Glu or
LysMOD_RES(7)..(7)Ser or Thr 138Xaa Ala Ser Xaa Xaa Xaa Xaa1
51399PRTArtificial SequenceDescription of Artificial Sequence Synthetic
peptideMOD_RES(3)..(3)Ser or TyrMOD_RES(4)..(4)Asp, Gly or
TyrMOD_RES(5)..(5)Asn, Ser or ThrMOD_RES(6)..(6)Leu, Thr or
TyrMOD_RES(8)..(8)Phe or Ile 139Gln Gln Xaa Xaa Xaa Xaa Pro Xaa Thr1
5140265PRTHomo sapiens 140Met Ser Glu Glu Val Thr Tyr Ala Asp
Leu Gln Phe Gln Asn Ser Ser1 5 10
15Glu Met Glu Lys Ile Pro Glu Ile Gly Lys Phe Gly Glu Lys Ala
Pro 20 25 30Pro Ala Pro Ser
His Val Trp Arg Pro Ala Ala Leu Phe Leu Thr Leu 35
40 45Leu Cys Leu Leu Leu Leu Ile Gly Leu Gly Val Leu
Ala Ser Met Phe 50 55 60His Val Thr
Leu Lys Ile Glu Met Lys Lys Met Asn Lys Leu Gln Asn65 70
75 80Ile Ser Glu Glu Leu Gln Arg Asn
Ile Ser Leu Gln Leu Met Ser Asn 85 90
95Met Asn Ile Ser Asn Lys Ile Arg Asn Leu Ser Thr Thr Leu
Gln Thr 100 105 110Ile Ala Thr
Lys Leu Cys Arg Glu Leu Tyr Ser Lys Glu Gln Glu His 115
120 125Lys Cys Lys Pro Cys Pro Arg Arg Trp Ile Trp
His Lys Asp Ser Cys 130 135 140Tyr Phe
Leu Ser Asp Asp Val Gln Thr Trp Gln Glu Ser Lys Met Ala145
150 155 160Cys Ala Ala Gln Asn Ala Ser
Leu Leu Lys Ile Asn Asn Lys Asn Ala 165
170 175Leu Glu Phe Ile Lys Ser Gln Ser Arg Ser Tyr Asp
Tyr Trp Leu Gly 180 185 190Leu
Ser Pro Glu Glu Asp Ser Thr Arg Gly Met Arg Val Asp Asn Ile 195
200 205Ile Asn Ser Ser Ala Trp Val Ile Arg
Asn Ala Pro Asp Leu Asn Asn 210 215
220Met Tyr Cys Gly Tyr Ile Asn Arg Leu Tyr Val Gln Tyr Tyr His Cys225
230 235 240Thr Tyr Lys Lys
Arg Met Ile Cys Glu Lys Met Ala Asn Pro Val Gln 245
250 255Leu Gly Ser Thr Tyr Phe Arg Glu Ala
260 265141126DNAHomo sapiens 141aagcgcggca
ggaagaagct cctctacatt tttaagcagc cttttatgag gcccgtacag 60acaacacagg
aggaagatgg ctgtagctgc agatttcccg aggaggagga aggtgggtgc 120gagctg
12614242PRTHomo
sapiens 142Lys Arg Gly Arg Lys Lys Leu Leu Tyr Ile Phe Lys Gln Pro Phe
Met1 5 10 15Arg Pro Val
Gln Thr Thr Gln Glu Glu Asp Gly Cys Ser Cys Arg Phe 20
25 30Pro Glu Glu Glu Glu Gly Gly Cys Glu Leu
35 4014337PRTHomo sapiens 143Arg Arg Asp Gln Arg Leu
Pro Pro Asp Ala His Lys Pro Pro Gly Gly1 5
10 15Gly Ser Phe Arg Thr Pro Ile Gln Glu Glu Gln Ala
Asp Ala His Ser 20 25 30Thr
Leu Ala Lys Ile 3514421PRTArtificial SequenceDescription of
Artificial Sequence Synthetic peptide 144Met Ala Leu Pro Val Thr Ala
Leu Leu Leu Pro Leu Ala Leu Leu Leu1 5 10
15His Ala Ala Arg Pro 2014520PRTArtificial
SequenceDescription of Artificial Sequence Synthetic peptide 145Gly
Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly1
5 10 15Gly Gly Gly Ser
20146112PRTHomo sapiens 146Arg Val Lys Phe Ser Arg Ser Ala Asp Ala Pro
Ala Tyr Lys Gln Gly1 5 10
15Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr
20 25 30Asp Val Leu Asp Lys Arg Arg
Gly Arg Asp Pro Glu Met Gly Gly Lys 35 40
45Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu Tyr Asn Glu Leu Gln
Lys 50 55 60Asp Lys Met Ala Glu Ala
Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg65 70
75 80Arg Arg Gly Lys Gly His Asp Gly Leu Tyr Gln
Gly Leu Ser Thr Ala 85 90
95Thr Lys Asp Thr Tyr Asp Ala Leu His Met Gln Ala Leu Pro Pro Arg
100 105 1101476762DNALentivirus
147ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
60ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
120ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
180ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
240ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
300gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
360tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
420ttaacgcgaa ttttaacaaa atattaacgc ttacaatttg ccattcgcca ttcaggctgc
480gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
540ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
600gtaaaacgac ggccagtgaa ttgtaatacg actcactata gggcgacccg gggatggcgc
660gccagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
720aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
780taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
840agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
900cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
960tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatgctga
1020tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
1080gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc
1140caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg
1200aggtctatat aagcagagct ggtttagtga accggggtct ctctggttag accagatctg
1260agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
1320ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
1380cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag ggacttgaaa
1440gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga agcgcgcacg
1500gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag cggaggctag
1560aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag atcgcgatgg
1620gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca tatagtatgg
1680gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac atcagaaggc
1740tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga agaacttaga
1800tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga gataaaagac
1860accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac caccgcacag
1920caagccgccg ctgatcttca gacctggagg aggagatatg agggacaatt ggagaagtga
1980attatataaa tataaagtag taaaaattga accattagga gtagcaccca ccaaggcaaa
2040gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt tccttgggtt
2100cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg tacaggccag
2160acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta ttgaggcgca
2220acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa gaatcctggc
2280tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct ctggaaaact
2340catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc tggaacagat
2400ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca caagcttaat
2460acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag aattattgga
2520attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc tgtggtatat
2580aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt ttgctgtact
2640ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga cccacctccc
2700aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag agagagacag
2760agacagatcc attcgattag tgaacggatc tcgacggtat cggttaactt ttaaaagaaa
2820aggggggatt ggggggtaca gtgcagggga aagaatagta gacataatag caacagacat
2880acaaactaaa gaattacaaa aacaaattac aaaattcaaa attttatcgc gatcgcggaa
2940tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat
3000ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca
3060gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
3120acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt
3180ccagggtgcc ccaaggacct gaaaatgacc ctgtgcctta tttgaactaa ccaatcagtt
3240cgcttctcgc ttctgttcgc gcgcttctgc tccccgagct caataaaaga gcccacaacc
3300cctcactcgg cgcgccagtc cttcgaagta gatctttgtc gatcctacca tccactcgac
3360acacccgcca gcggccgctg ccaagcttcc gagctctcga attaattcac ggtacccacc
3420atggcctagg gagactagtc gaatcgatat caacctctgg attacaaaat ttgtgaaaga
3480ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg
3540cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc
3600tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc
3660actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt
3720tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt
3780gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg
3840aagctgacgt ccttttcatg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg
3900tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg
3960ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt
4020tgggccgcct ccccgcctgg ttaattaaag tacctttaag accaatgact tacaaggcag
4080ctgtagatct tagccacttt ttaaaagaaa aggggggact ggaagggcga attcactccc
4140aacgaagaca agatctgctt tttgcttgta ctgggtctct ctggttagac cagatctgag
4200cctgggagct ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt
4260gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca
4320gaccctttta gtcagtgtgg aaaatctcta gcaggcatgc cagacatgat aagatacatt
4380gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt
4440tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac
4500aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttggcgcgcc
4560atcgtcgagg ttccctttag tgagggttaa ttgcgagctt ggcgtaatca tggtcatagc
4620tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
4680taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
4740cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
4800gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
4860tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
4920tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
4980ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
5040agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
5100accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
5160ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
5220gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
5280ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
5340gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
5400taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
5460tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
5520gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
5580cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
5640agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
5700cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
5760cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
5820ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
5880taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
5940tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
6000ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
6060atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
6120gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
6180tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
6240cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
6300taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
6360ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
6420ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
6480cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
6540ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
6600gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
6660gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
6720aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc ac
6762
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