STEM CELL FACTOR INHIBITOR

Abstract

Provided herein are methods, compositions, and uses relating to inhibitors of stem cell factor. For example, provided herein are antibodies targeting stem cell factor and methods for treating fibrotic and tissue remodeling diseases.

Description

[0001] This application is a continuation of U.S. patent application Ser. No. 15/782,503, filed Oct. 12, 2017, which is a continuation of U.S. patent application Ser. No. 15/058,918, filed Mar. 2, 2016, now U.S. Pat. No. 9,790,272, issued Oct. 17, 2017, which is a divisional of U.S. patent application Ser. No. 13/937,852, filed Jul. 9, 2013, now U.S. Pat. No. 9,353,178, issued on May 31, 2016, which is a divisional of U.S. patent application Ser. No. 13/347,459, filed on Jan. 10, 2012, now U.S. Pat. No. 8,911,729, issued on Dec. 16, 2014, which claims priority to U.S. Patent Application Ser. No. 61/431,246 filed on Jan. 10, 2011, each of which is incorporated herein by reference in its entirety for all purposes.

FIELD OF INVENTION

[0003] Provided herein are methods, compositions, and uses relating to inhibitors of stem cell factor. For example, provided herein are antibodies targeting stem cell factor and methods for treating fibrotic and tissue remodeling diseases.

BACKGROUND

[0004] Diseases involving tissue remodeling and fibrosis are a leading cause of death worldwide. Nearly 45 percent of all natural deaths in the western world are attributable to some type of chronic fibroproliferative disease and the associated health care costs are in the billions of dollars. Tissue remodeling is the reorganization or renovation of existing tissues, which can either change the characteristics of a tissue (e.g., blood vessel remodeling) or participate in establishing the dynamic equilibrium of a tissue (e.g., bone remodeling). Fibrosis is the formation or development of excess fibrous connective tissue in an organ or tissue as a reparative or reactive process, as opposed to formation of fibrous tissue as a normal constituent of an organ or tissue. Fibrosis affects nearly all tissues and organ systems, and fibrotic tissue remodeling can influence cancer metastasis and accelerate chronic graft rejection in transplant recipients. Diseases in which fibrosis is a major cause of morbidity and mortality include the interstitial lung diseases, liver cirrhosis, kidney disease, heart disease, and systemic sclerosis, among others.

[0005] Stem cell factor (SCF) and its receptor c-Kit have been implicated in fibrotic and tissue remodeling diseases (El-Koraie, et al., Kidney Int. 60: 167 (2001); Powell, et al., Am. J. Physiol. 289: G2 (2005); El Kossi, et al., Am. J. Kidney Dis. 41: 785 (2003); Powell, et al., Am. J. Physiol. 277: C183 (1999)). c-Kit is a type III receptor-tyrosine kinase that is present in many cell types (Orr-Urtreger et al., Development 109: 911 (1990)). It is also expressed in the early stages of differentiation (Andre et al., Oncogene 4: 1047 (1989)) and certain tumors exhibit elevated expression of c-kit. SCF is a ligand specific for the c-Kit receptor kinase. Binding causes dimerization of c-Kit and activation of its kinase activity. SCF was first isolated from the supernatant of murine fibroblasts. At the time, SCF was called mast cell growth factor (MGF) (Williams et al., Cell 63: 167 (1990)) or hematopoietic growth factor KL (Kit ligand) (Huang et al., Cell 63: 225 (1990)). A homologue was subsequently isolated from rat liver cells and designated stem cell factor (SCF) (Zsebo et al., Cell 63: 195 (1990)). The corresponding human protein is designated variously as SCF, MGF, or Steel Factor (SF) (Cell 63: 203 (1990)).

[0006] Previous studies have suggested that an inhibitor of c-Kit receptor tyrosine kinase can significantly inhibit aberrant tissue fibrosis (see, e.g., Aono, Am. J. Respir. Crit. Care Med. 171: 1279 (2005); Vuorinen, et al., Exp. Lung Res. 33: 357 (2007); Vittal, et al., J. Pharmacol. Exp. Ther. 321: 35 (2007); Distler, et al., Arthritis Rheum 56: 311 (2007)). However, this inhibitor has several disadvantages. It needs to be given systemically by oral administration, it has some toxicity associated with its use, and the compound must be delivered intracellularly for efficacy. Consequently, alternative therapies are needed.

SUMMARY

[0007] Provided herein are methods, compositions, and uses relating to inhibitors of stem cell factor. For example, provided herein are antibodies targeting stem cell factor and methods for treating fibrotic and tissue remodeling diseases as well as for research and diagnostic uses.

[0008] In some embodiments, the compositions, methods, and uses herein provide therapies relating to inhibiting stem cell factor (SCF). Some embodiments provide an isolated antibody that targets SCF. In some embodiments, inhibiting SCF affects the activity of c-Kit. The compositions, methods, and uses provided herein find use in treating fibrotic diseases and maladies associated with tissue remodeling. Unlike some other therapies that produce undesirable side effects due to interfering with general intracellular signaling pathways, the embodiments provided herein eliminate or minimize such side effects by modulating the activity of SCF. Consequently, toxicity is minimized. Moreover, targeting an extracellular ligand removes the need to deliver a composition into a cell to interact with an intracellular target. In some embodiments, the compositions are delivered into the airway, thus providing an advantage over previous technologies that require oral administration and, as such, resulting in systemic bioavailability.

[0009] Provided herein are embodiments of methods for treating a fibrotic or tissue remodeling disease comprising administering a therapeutically effective amount of a stem cell factor inhibitor to a subject with or at risk for a fibrotic or tissue remodeling disease. For example, in some embodiments, provided herein are methods comprising providing an inhibitor of stem cell factor and administering a therapeutically effective amount of the inhibitor to a subject. In some embodiments the inhibitor is an isolated antibody (e.g., a monoclonal or polyclonal antibody) or an antigen-binding fragment thereof (e.g., Fab, Fab', F(ab').sub.2, and Fv fragments, etc.). In some embodiments the inhibitor is a small interfering RNA. In more specific embodiments, the antibody is a monoclonal antibody or a polyclonal antibody. Some embodiments provide that the antibody or antigen-binding fragment thereof specifically binds to stem cell factor. Some embodiments provide that the antibody or antigen-binding fragment thereof specifically binds to a peptide comprising amino acid sequence SEQ ID NO: 1 or SEQ ID NO: 8.

[0010] In some embodiments of the methods provided herein, the subject has a disease. Accordingly, some embodiments provide that administering the inhibitor prevents or reduces the severity of at least one sign or symptom of the disease. In some embodiments, the subject has an abnormal activity of stem cell factor or the subject has abnormal collagen production. In some embodiments, the subject has a disease including, but not limited to, fibrosis or a remodeling disease. In additional embodiments, the disease is a pulmonary disease. Some embodiments provide that a subject has a pulmonary disease including, but not limited to, idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, acute respiratory distress syndrome, cystic fibrosis, peribronchial fibrosis, hypersensitivity pneumonitis, or asthma. In addition, some embodiments provide that a subject has a disease including, but not limited to, sclerodoma, inflammation, liver cirrhosis, renal fibrosis, parenchymal fibrosis, endomyocardial fibrosis, mediatinal fibrosis, nodular subepidermal fibrosis, fibrous histiocytoma, fibrothorax, hepatic fibrosis, fibromyalgia, gingival fibrosis, or radiation-induced fibrosis.

[0011] While not limited in the mode of administration, in some embodiments of the method, the antibody is delivered into an airway of the subject, e.g., by intranasal administration.

[0012] In some embodiments, administering the inhibitor reduces the activity of a receptor. Some embodiments provide that administering the inhibitor reduces an interaction of stem cell factor with a receptor. In more specific embodiments, the receptor is a receptor tyrosine kinase, and in yet more specific embodiments, the receptor is c-Kit. Importantly, the methods are not limited in the location of the targeted receptor or the origin of stem cell factor. For example, in some embodiments the receptor is found on a hematopoietic progenitor cell, a melanocyte, a germ cell, an eosinophil, a lymphocyte, a fibroblast, a myofibroblast, or a mast cell. Additionally, in some embodiments, stem cell factor originates from a bone marrow cell, a liver cell, an epithelial cell, a smooth muscle cell, or a fibroblast. In some embodiments, administering the inhibitor to a subject results in a direct inhibition of fibroblast activation.

[0013] Some embodiments provide a composition comprising an isolated antibody (e.g., a monoclonal or a polyclonal antibody) or antigen-binding fragment thereof that specifically binds to stem cell factor (e.g., a protein or a peptide fragment thereof (e.g., an epitope)). For example, some embodiments provide a composition comprising an isolated antibody or antigen-binding fragment thereof that specifically binds to a peptide of amino acid sequence SEQ ID NO: 1 or SEQ ID NO: 8. Additional embodiments provide an antibody or antigen-binding fragment than binds to the SCF isoform b precursor (e.g., a protein or peptide fragment of the sequence available at GenBank accession number NP_000890 (SEQ ID NO: 4)), or a variant or modified form thereof, or to the SCF isoform a precursor (e.g., a protein or peptide fragment of the sequence available at GenBank accession number NP_003985 (SEQ ID NO: 6)), or a variant or modified form thereof. Some embodiments provide an antibody or antigen-binding fragment that binds to a protein or peptide, or variants or modified forms thereof, that is a translation product of the NCBI Reference Gene Sequence for SCF (e.g., accession number NG_012098 (SEQ ID NO: 7)) or variants or fragments thereof. Some embodiments provide an antibody or antigen-binding fragment that binds to a peptide comprising the first 11 amino acids of the mature form of SCF (e.g., EGICRNRVTNN (SEQ ID NO: 8)).

[0014] Some embodiments provide an antibody or antigen-binding fragment than binds to the translation product (e.g., a protein or peptide), or a variant or modified form thereof, of a nucleic acid encoding SCF, or a variant or a modified form thereof. For example, embodiments provide an antibody or antigen-binding fragment than binds to the translation product (e.g., a protein or peptide), or a variant or modified form thereof, of the nucleic acids having sequences comprising a sequence as defined by GenBank accession numbers NM_000899 (SEQ ID NO: 3), NM_003994 (SEQ ID NO: 5), and NG_012098 (SEQ ID NO: 7), or fragments or variants thereof (e.g., mutants, cDNAs, expression-optimized variants, operably linked to a regulatory element (e.g., promoter, enhancer, polymerase binding site, etc.), etc.). In some embodiments, the antibody or antigen-binding fragment binds to a protein or peptide, or a variant or modified form thereof, that is the translation product of a nucleotide sequence that encodes the peptide sequence EGICRNRVTNN (SEQ ID NO: 8). The peptides and proteins (and fragments and variants thereof) and the nucleic acids (and fragments and variants thereof) that encode the peptides and proteins (and fragments and variants thereof) are used in some embodiments to raise antibodies. Also contemplated are vectors, plasmids, expression constructs, cells, cell lines, hybridomas, and organisms used to produce the antibodies as provided herein.

[0015] Some embodiments provide a monoclonal antibody and some embodiments provide a humanized antibody. In some embodiments, the composition is used for a medicament or is used for the manufacture of a medicament. In some embodiments, the medicament is used to treat disease. Use of the composition as a medicament is not limited in the disease that can be treated. For example, in some embodiments, the disease is idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, acute respiratory distress syndrome, cystic fibrosis, peribronchial fibrosis, hypersensitivity pneumonitis, asthma, sclerodoma, inflammation, liver cirrhosis, renal fibrosis, parenchymal fibrosis, endomyocardial fibrosis, mediatinal fibrosis, nodular subepidermal fibrosis, fibrous histiocytoma, fibrothorax, hepatic fibrosis, fibromyalgia, gingival fibrosis, or radiation-induced fibrosis. In some embodiments, the composition is used to study disease in vitro or in a model system (e.g., in vivo).

[0016] Embodiments provide herein a method of preparing an antibody (e.g., a monoclonal antibody) targeting stem cell factor comprising the steps of providing a peptide comprising or consisting of an immunogenic portion of SCF (e.g., as provided by SEQ ID NO: 1 or 8), immunizing a host with the peptide, isolating an immune cell from the host, preparing a hybridoma using the immune cell, and isolating the antibody or antigen-binding fragment thereof. Some embodiments provide a method of preparing an antibody (e.g., a monoclonal antibody) targeting stem cell factor, wherein the antibody or antigen-binding fragment thereof specifically binds to stem cell factor (e.g., a protein or a peptide fragment thereof (e.g., an epitope)). For example, some embodiments provide a method of preparing an isolated antibody or antigen-binding fragment thereof that specifically binds to a peptide of amino acid sequence SEQ ID NO: 1. Additional embodiments provide a method of preparing an antibody or antigen-binding fragment than binds to the SCF isoform b precursor (e.g., a protein or peptide fragment of the sequence available at GenBank accession number NP_000890 (SEQ ID NO: 4)), or a variant or modified form thereof, or to the SCF isoform a precursor (e.g., a protein or peptide fragment of the sequence available at GenBank accession number NP_003985 (SEQ ID NO: 6)), or a variant or modified form thereof. Some embodiments provide a method of preparing an antibody or antigen-binding fragment that binds to a protein or peptide, or variants or modified forms thereof, that is a translation product of the NCBI Reference Gene Sequence for SCF (e.g., accession number NG_012098 (SEQ ID NO: 7)) or variants or fragments thereof. Some embodiments provide a method of preparing an antibody or antigen-binding fragment that binds to a peptide comprising the first 11 amino acids of the mature form of SCF (e.g., EGICRNRVTNN (SEQ ID NO: 8)).

[0017] Some embodiments provide a method of preparing an antibody or antigen-binding fragment than binds to the translation product (e.g., a protein or peptide), or a variant or modified form thereof, of a nucleic acid encoding SCF, or a variant or a modified form thereof. For example, embodiments provide a method of preparing an antibody or antigen-binding fragment than binds to the translation product (e.g., a protein or peptide), or a variant or modified form thereof, of the nucleic acids having sequences comprising a sequence as defined by GenBank accession numbers NM_000899 (SEQ ID NO: 3), NM_003994 (SEQ ID NO: 5), and NG_012098 (SEQ ID NO: 7), or fragments or variants thereof (e.g., mutants, cDNAs, expression-optimized variants, operably linked to a regulatory element (e.g., promoter, enhancer, polymerase binding site, etc.), etc.). In some embodiments, the antibody or antigen-binding fragment binds to a protein or peptide, or a variant or modified form thereof, that is the translation product of a nucleotide sequence that encodes the peptide sequence EGICRNRVTNN (SEQ ID NO: 8). The peptides, proteins, and fragments and variants thereof; and nucleic acids, and fragments and variants thereof, that encode the peptides, proteins, and fragments and variants thereof, find use in some embodiments in a method of preparing antibodies as provided by the technology provided. Also contemplated are methods of producing vectors, plasmids, expression constructs, cells, cell lines, hybridomas, and organisms that find use in producing the antibodies as provided herein.

[0018] Some embodiments provide a method comprising the steps of providing an inhibitor of stem cell factor and administering the inhibitor to a cell or tissue.

[0019] In addition, some embodiments provide a kit comprising a composition comprising an isolated antibody or antigen-binding fragment thereof that specifically binds to stem cell factor, a means for administering the composition to a subject, and/or instructions for use.

[0020] Additional embodiments will be apparent to persons skilled in the relevant art based on the teachings contained herein.

BRIEF DESCRIPTION OF THE DRAWINGS

[0021] These and other features, aspects, and advantages of the present technology will become better understood with regard to the following drawings:

[0022] FIG. 1 shows a series of plots demonstrating that inhibiting SCF with an antibody reduces the expression of tissue remodeling mediators. FIG. 1A shows a plot demonstrating that an anti-SCF antibody reduces the amount of hydroxyproline in bleomycin treated lung;

[0023] FIG. 1B shows a plot demonstrating that an anti-SCF antibody reduces the amount of IL-25 mRNA; FIG. 1C shows a plot demonstrating that an anti-SCF antibody reduces the amount of IL-13 mRNA; FIG. 1D shows a plot demonstrating that an anti-SCF antibody reduces the amount of soluble SCF present in plasma. FIG. 1E shows a plot demonstrating that an anti-SCF antibody reduces the amount of IL-25 receptor.

[0024] FIG. 2 shows a plot demonstrating that IL-4 stimulates c-kit expression in human fibroblasts.

[0025] FIG. 3 shows an amino acid sequence and the corresponding nucleotide sequence of an immunogenic peptide used to produce antibodies specific for SCF.

[0026] FIG. 4 shows a plot demonstrating that a monoclonal antibody specific for SCF inhibits the activation of HMC-1 cells for MCP-1 production.

[0027] FIG. 5 shows a plot demonstrating that a lower amount of hydroxyproline is detected in a mouse deficient in SCF production after bleomycin injury.

DETAILED DESCRIPTION

[0028] Provided herein are methods, compositions, and uses relating to inhibitors of stem cell factor. For example, provided herein are antibodies targeting stem cell factor, methods of producing antibodies targeting stem cell factor, and methods for treating fibrotic and tissue remodeling diseases as well as for research and diagnostic uses. In some embodiments, the compositions, methods, and uses herein provide therapies relating to inhibiting stem cell factor (SCF). Some embodiments provide an isolated antibody that targets SCF. In some embodiments, inhibiting SCF affects the activity of c-Kit. The compositions, methods, and uses provided herein find use in treating fibrotic diseases and maladies associated with tissue remodeling.

Definitions

[0029] To facilitate an understanding of embodiments of the present technology, a number of terms and phrases are defined below. Additional definitions are set forth throughout the detailed description.

[0030] Throughout the specification and claims, the following terms take the meanings explicitly associated herein, unless the context clearly dictates otherwise. The phrase "in one embodiment" as used herein does not necessarily refer to the same embodiment, though it may. Furthermore, the phrase "in another embodiment" as used herein does not necessarily refer to a different embodiment, although it may. Thus, as described below, various embodiments of the invention may be readily combined, without departing from the scope or spirit of the invention.

[0031] In addition, as used herein, the term "or" is an inclusive "or" operator and is equivalent to the term "and/or" unless the context clearly dictates otherwise. The term "based on" is not exclusive and allows for being based on additional factors not described, unless the context clearly dictates otherwise. In addition, throughout the specification, the meaning of "a", "an", and "the" include plural references. The meaning of "in" includes "in" and "on."

[0032] The terms "protein" and "polypeptide" refer to compounds comprising amino acids joined via peptide bonds and are used interchangeably. A "protein" or "polypeptide" encoded by a gene is not limited to the amino acid sequence encoded by the gene, but includes post-translational modifications of the protein.

[0033] Where the term "amino acid sequence" is recited herein to refer to an amino acid sequence of a protein molecule, "amino acid sequence" and like terms, such as "polypeptide" or "protein" are not meant to limit the amino acid sequence to the complete, native amino acid sequence associated with the recited protein molecule. Furthermore, an "amino acid sequence" can be deduced from the nucleic acid sequence encoding the protein.

[0034] The term "nascent" when used in reference to a protein refers to a newly synthesized protein, which has not been subject to post-translational modifications, which includes but is not limited to glycosylation and polypeptide shortening. The term "mature" when used in reference to a protein refers to a protein which has been subject to post-translational processing and/or which is in a cellular location (such as within a membrane or a multi-molecular complex) from which it can perform a particular function which it could not if it were not in the location.

[0035] The term "portion" when used in reference to a protein (as in "a portion of a given protein") refers to fragments of that protein. The fragments may range in size from four amino acid residues to the entire amino sequence minus one amino acid (for example, the range in size includes 4, 5, 6, 7, 8, 9, 10, or 11 . . . amino acids up to the entire amino acid sequence minus one amino acid).

[0036] The term "homolog" or "homologous" when used in reference to a polypeptide refers to a high degree of sequence identity between two polypeptides, or to a high degree of similarity between the three-dimensional structure or to a high degree of similarity between the active site and the mechanism of action. In a preferred embodiment, a homolog has a greater than 60% sequence identity, and more preferably greater than 75% sequence identity, and still more preferably greater than 90% sequence identity, with a reference sequence.

[0037] The terms "variant" and "mutant" when used in reference to a polypeptide refer to an amino acid sequence that differs by one or more amino acids from another, usually related polypeptide. The variant may have "conservative" changes, wherein a substituted amino acid has similar structural or chemical properties. One type of conservative amino acid substitutions refers to the interchangeability of residues having similar side chains. For example, a group of amino acids having aliphatic side chains is glycine, alanine, valine, leucine, and isoleucine; a group of amino acids having aliphatic-hydroxyl side chains is serine and threonine; a group of amino acids having amide-containing side chains is asparagine and glutamine; a group of amino acids having aromatic side chains is phenylalanine, tyrosine, and tryptophan; a group of amino acids having basic side chains is lysine, arginine, and histidine; and a group of amino acids having sulfur-containing side chains is cysteine and methionine. Preferred conservative amino acids substitution groups are: valine-leucine-isoleucine, phenylalanine-tyrosine, lysine-arginine, alanine-valine, and asparagine-glutamine. More rarely, a variant may have "non-conservative" changes (e.g., replacement of a glycine with a tryptophan). Similar minor variations may also include amino acid deletions or insertions (i.e., additions), or both. Guidance in determining which and how many amino acid residues may be substituted, inserted or deleted without abolishing biological activity may be found using computer programs well known in the art, for example, DNAStar software. Variants can be tested in functional assays. Preferred variants have less than 10%, and preferably less than 5%, and still more preferably less than 2% changes (whether substitutions, deletions, and so on).

[0038] The term "domain" when used in reference to a polypeptide refers to a subsection of the polypeptide which possesses a unique structural and/or functional characteristic; typically, this characteristic is similar across diverse polypeptides. The subsection typically comprises contiguous amino acids, although it may also comprise amino acids which act in concert or which are in close proximity due to folding or other configurations. Examples of a protein domain include the transmembrane domains, and the glycosylation sites.

[0039] The term "gene" refers to a nucleic acid (e.g., DNA or RNA) sequence that comprises coding sequences necessary for the production of an RNA, or a polypeptide or its precursor (e.g., proinsulin). A functional polypeptide can be encoded by a full length coding sequence or by any portion of the coding sequence as long as the desired activity or functional properties (e.g., enzymatic activity, ligand binding, signal transduction, etc.) of the polypeptide are retained. The term "portion" when used in reference to a gene refers to fragments of that gene. The fragments may range in size from a few nucleotides to the entire gene sequence minus one nucleotide. Thus, "a nucleotide comprising at least a portion of a gene" may comprise fragments of the gene or the entire gene.

[0040] The term "gene" also encompasses the coding regions of a structural gene and includes sequences located adjacent to the coding region on both the 5' and 3' ends for a distance of about 1 kb on either end such that the gene corresponds to the length of the full-length mRNA. The sequences which are located 5' of the coding region and which are present on the mRNA are referred to as 5' non-translated sequences. The sequences which are located 3' or downstream of the coding region and which are present on the mRNA are referred to as 3' non-translated sequences. The term "gene" encompasses both cDNA and genomic forms of a gene. A genomic form or clone of a gene contains the coding region interrupted with non-coding sequences termed "introns" or "intervening regions" or "intervening sequences." Introns are segments of a gene which are transcribed into nuclear RNA (hnRNA); introns may contain regulatory elements such as enhancers. Introns are removed or "spliced out" from the nuclear or primary transcript; introns therefore are absent in the messenger RNA (mRNA) transcript. The mRNA functions during translation to specify the sequence or order of amino acids in a nascent polypeptide.

[0041] In addition to containing introns, genomic forms of a gene may also include sequences located on both the 5' and 3' end of the sequences which are present on the RNA transcript. These sequences are referred to as "flanking" sequences or regions (these flanking sequences are located 5' or 3' to the non-translated sequences present on the mRNA transcript). The 5' flanking region may contain regulatory sequences such as promoters and enhancers which control or influence the transcription of the gene. The 3' flanking region may contain sequences which direct the termination of transcription, posttranscriptional cleavage and polyadenylation.

[0042] The terms "oligonucleotide" or "polynucleotide" or "nucleotide" or "nucleic acid" refer to a molecule comprised of two or more deoxyribonucleotides or ribonucleotides, preferably more than three, and usually more than ten. The exact size will depend on many factors, which in turn depends on the ultimate function or use of the oligonucleotide. The oligonucleotide may be generated in any manner, including chemical synthesis, DNA replication, reverse transcription, or a combination thereof.

[0043] The terms "an oligonucleotide having a nucleotide sequence encoding a gene" or "a nucleic acid sequence encoding" a specified polypeptide refer to a nucleic acid sequence comprising the coding region of a gene or in other words the nucleic acid sequence which encodes a gene product. The coding region may be present in either a cDNA, genomic DNA or RNA form. When present in a DNA form, the oligonucleotide may be single-stranded (i.e., the sense strand) or double-stranded. Suitable control elements such as enhancers/promoters, splice junctions, polyadenylation signals, etc. may be placed in close proximity to the coding region of the gene if needed to permit proper initiation of transcription and/or correct processing of the primary RNA transcript. Alternatively, the coding region utilized in the expression vectors of the present invention may contain endogenous enhancers/promoters, splice junctions, intervening sequences, polyadenylation signals, etc. or a combination of both endogenous and exogenous control elements.

[0044] The term "recombinant" when made in reference to a nucleic acid molecule refers to a nucleic acid molecule which is comprised of segments of nucleic acid joined together by means of molecular biological techniques. The term "recombinant" when made in reference to a protein or a polypeptide refers to a protein molecule which is expressed using a recombinant nucleic acid molecule.

[0045] The terms "complementary" and "complementarity" refer to polynucleotides (i.e., a sequence of nucleotides) related by the base-pairing rules. For example, for the sequence "5'-A-G-T-3'," is complementary to the sequence "3'-T-C-A-S'." Complementarity may be "partial," in which only some of the nucleic acids' bases are matched according to the base pairing rules. Or, there may be "complete" or "total" complementarity between the nucleic acids. The degree of complementarity between nucleic acid strands has significant effects on the efficiency and strength of hybridization between nucleic acid strands. This is of particular importance in amplification reactions, as well as detection methods which depend upon binding between nucleic acids.

[0046] The term "wild-type" when made in reference to a gene refers to a gene that has the characteristics of a gene isolated from a naturally occurring source. The term "wild-type" when made in reference to a gene product refers to a gene product that has the characteristics of a gene product isolated from a naturally occurring source. The term "naturally-occurring" as applied to an object refers to the fact that an object can be found in nature. For example, a polypeptide or polynucleotide sequence that is present in an organism (including viruses) that can be isolated from a source in nature and which has not been intentionally modified by man in the laboratory is naturally-occurring. A wild-type gene is frequently that gene which is most frequently observed in a population and is thus arbitrarily designated the "normal" or "wild-type" form of the gene. In contrast, the term "modified" or "mutant" when made in reference to a gene or to a gene product refers, respectively, to a gene or to a gene product which displays modifications in sequence and/or functional properties (i.e., altered characteristics) when compared to the wild-type gene or gene product. It is noted that naturally-occurring mutants can be isolated; these are identified by the fact that they have altered characteristics when compared to the wild-type gene or gene product.

[0047] The term "allele" refers to different variations in a gene; the variations include but are not limited to variants and mutants, polymorphic loci and single nucleotide polymorphic loci, frameshift and splice mutations. An allele may occur naturally in a population, or it might arise during the lifetime of any particular individual of the population.

[0048] Thus, the terms "variant" and "mutant" when used in reference to a nucleotide sequence refer to an nucleic acid sequence that differs by one or more nucleotides from another, usually related nucleotide acid sequence. A "variation" is a difference between two different nucleotide sequences; typically, one sequence is a reference sequence.

[0049] The term "antisense" refers to a deoxyribonucleotide sequence whose sequence of deoxyribonucleotide residues is in reverse 5' to 3' orientation in relation to the sequence of deoxyribonucleotide residues in a sense strand of a DNA duplex. A "sense strand" of a DNA duplex refers to a strand in a DNA duplex which is transcribed by a cell in its natural state into a "sense mRNA." Thus an "antisense" sequence is a sequence having the same sequence as the non-coding strand in a DNA duplex. The term "antisense RNA" refers to a RNA transcript that is complementary to all or part of a target primary transcript or mRNA and that blocks the expression of a target gene by interfering with the processing, transport and/or translation of its primary transcript or mRNA. The complementarity of an antisense RNA may be with any part of the specific gene transcript, i.e., at the 5' non-coding sequence, 3' non-coding sequence, introns, or the coding sequence. In addition, as used herein, antisense RNA may contain regions of ribozyme sequences that increase the efficacy of antisense RNA to block gene expression. "Ribozyme" refers to a catalytic RNA and includes sequence-specific endoribonucleases. "Antisense inhibition" refers to the production of antisense RNA transcripts capable of preventing the expression of the target protein.

[0050] The term "primer" refers to an oligonucleotide, whether occurring naturally as in a purified restriction digest or produced synthetically, which is capable of acting as a point of initiation of synthesis when placed under conditions in which synthesis of a primer extension product which is complementary to a nucleic acid strand is induced, (e.g., in the presence of nucleotides and an inducing agent such as DNA polymerase and at a suitable temperature and pH). The primer is preferably single stranded for maximum efficiency in amplification, but may alternatively be double stranded. If double stranded, the primer is first treated to separate its strands before being used to prepare extension products. Preferably, the primer is an oligodeoxyribonucleotide. The primer must be sufficiently long to prime the synthesis of extension products in the presence of the inducing agent. The exact lengths of the primers will depend on many factors, including temperature, source of primer and the use of the method.

[0051] The term "probe" refers to an oligonucleotide (i.e., a sequence of nucleotides), whether occurring naturally as in a purified restriction digest or produced synthetically, recombinantly or by PCR amplification, that is capable of hybridizing to another oligonucleotide of interest. A probe may be single-stranded or double-stranded. Probes are useful in the detection, identification and isolation of particular gene sequences. It is contemplated that any probe used in the present invention will be labeled with any "reporter molecule," so that is detectable in any detection system, including, but not limited to enzyme (e.g., ELISA, as well as enzyme-based histochemical assays), fluorescent, radioactive, and luminescent systems. It is not intended that the present invention be limited to any particular detection system or label.

[0052] The term "isolated" when used in relation to a nucleic acid, as in "an isolated oligonucleotide" refers to a nucleic acid sequence that is identified and separated from at least one contaminant nucleic acid with which it is ordinarily associated in its natural source. Isolated nucleic acid is present in a form or setting that is different from that in which it is found in nature. In contrast, non-isolated nucleic acids, such as DNA and RNA, are found in the state they exist in nature. Examples of non-isolated nucleic acids include: a given DNA sequence (e.g., a gene) found on the host cell chromosome in proximity to neighboring genes; RNA sequences, such as a specific mRNA sequence encoding a specific protein, found in the cell as a mixture with numerous other mRNAs which encode a multitude of proteins. However, isolated nucleic acid encoding a particular protein includes, by way of example, such nucleic acid in cells ordinarily expressing the protein, where the nucleic acid is in a chromosomal location different from that of natural cells, or is otherwise flanked by a different nucleic acid sequence than that found in nature. The isolated nucleic acid or oligonucleotide may be present in single-stranded or double-stranded form. When an isolated nucleic acid or oligonucleotide is to be utilized to express a protein, the oligonucleotide will contain at a minimum the sense or coding strand (i.e., the oligonucleotide may single-stranded), but may contain both the sense and anti-sense strands (i.e., the oligonucleotide may be double-stranded).

[0053] The term "purified" refers to molecules, either nucleic or amino acid sequences, that are removed from their natural environment, isolated or separated. An "isolated nucleic acid sequence" may therefore be a purified nucleic acid sequence. "Substantially purified" molecules are at least 60% free, preferably at least 75% free, and more preferably at least 90% free from other components with which they are naturally associated. As used herein, the term "purified" or "to purify" also refer to the removal of contaminants from a sample. The removal of contaminating proteins results in an increase in the percent of polypeptide of interest in the sample. In another example, recombinant polypeptides are expressed in plant, bacterial, yeast, or mammalian host cells and the polypeptides are purified by the removal of host cell proteins; the percent of recombinant polypeptides is thereby increased in the sample.

[0054] The term "composition comprising" a given polynucleotide sequence or polypeptide refers broadly to any composition containing the given polynucleotide sequence or polypeptide. The composition may comprise an aqueous solution. Compositions comprising polynucleotide sequences or fragments thereof may be employed as hybridization probes. In some embodiments, polynucleotide sequences are employed in an aqueous solution containing salts (e.g., NaCl), detergents (e.g., SDS), and other components (e.g., Denhardt's solution, dry milk, salmon sperm DNA, etc.).

[0055] The term "test compound" refers to any chemical entity, pharmaceutical, drug, and the like that can be used to treat or prevent a disease, illness, sickness, or disorder of bodily function, or otherwise alter the physiological or cellular status of a sample. Test compounds comprise both known and potential therapeutic compounds. A test compound can be determined to be therapeutic by screening using the screening methods of the present invention. A "known therapeutic compound" refers to a therapeutic compound that has been shown (e.g., through animal trials or prior experience with administration to humans) to be effective in such treatment or prevention.

[0056] As used herein, the term "antibody" is used in its broadest sense to refer to whole antibodies, monoclonal antibodies (including human, humanized, or chimeric antibodies), polyclonal antibodies, and antibody fragments that can bind antigen (e.g., Fab', F' (ab).sub.2, Fv, single chain antibodies), comprising complementarity determining regions (CDRs) of the foregoing as long as they exhibit the desired biological activity.

[0057] As used herein, "antibody fragments" comprise a portion of an intact antibody, preferably the antigen binding or variable region of the intact antibody. Examples of antibody fragments include Fab, Fab', F(ab').sub.2, and Fv fragments; diabodies; linear antibodies (Zapata et al., Protein Eng. 8(10): 1057-1062 (1995)); single-chain antibody molecules; and multispecific antibodies formed from antibody fragments.

[0058] An antibody that "specifically binds to" or is "specific for" a particular polypeptide or an epitope on a particular polypeptide is one that binds to that particular polypeptide or epitope on a particular polypeptide without substantially binding to any other polypeptide or polypeptide epitope.

[0059] As used herein, "active" or "activity" refers to native or naturally occurring biological and/or immunological activity.

[0060] As used herein the term, "in vitro" refers to an artificial environment and to processes or reactions that occur within an artificial environment. In vitro environments may include, but are not limited to, test tubes and cell cultures. The term "in vivo" refers to the natural environment (e.g., an animal or a cell) and to processes or reactions that occur within a natural environment.

[0061] As used herein, "inhibitor" refers to a molecule which eliminates, minimizes, or decreases the activity, e.g., the biological, enzymatic, chemical, or immunological activity, of a target.

[0062] As used herein the term "disease" refers to a deviation from the condition regarded as normal or average for members of a species, and which is detrimental to an affected individual under conditions that are not inimical to the majority of individuals of that species (e.g., diarrhea, nausea, fever, pain, inflammation, etc.).

[0063] As used herein, the term "administration" refers to the act of giving a drug, prodrug, antibody, or other agent, or therapeutic treatment to a physiological system (e.g., a subject or in vivo, in vitro, or ex vivo cells, tissues, and organs). Exemplary routes of administration to the human body can be through the eyes (ophthalmic), mouth (oral), skin (transdermal), nose (nasal), lungs (inhalant), oral mucosa (buccal), ear, by injection (e.g., intravenously, subcutaneously, intratumorally, intraperitoneally, etc.) and the like. "Coadministration" refers to administration of more than one chemical agent or therapeutic treatment (e.g., radiation therapy) to a physiological system (e.g., a subject or in vivo, in vitro, or ex vivo cells, tissues, and organs). As used herein, administration "in combination with" one or more further therapeutic agents includes simultaneous (concurrent) and consecutive administration in any order. "Coadministration" of therapeutic treatments may be concurrent, or in any temporal order or physical combination.

[0064] As used herein, the term "treating" includes reducing or alleviating at least one adverse effect or symptom of a disease or disorder through introducing in any way a therapeutic composition of the present technology into or onto the body of a subject. "Treatment" refers to both therapeutic treatment and prophylactic or preventative measures, wherein the object is to prevent or slow down (lessen) the targeted pathologic condition or disorder. Those in need of treatment include those already with the disorder as well as those prone to have the disorder or those in whom the disorder is to be prevented.

[0065] As used herein, "therapeutically effective dose" refers to an amount of a therapeutic agent sufficient to bring about a beneficial or desired clinical effect. Said dose can be administered in one or more administrations. However, the precise determination of what would be considered an effective dose may be based on factors individual to each patient, including, but not limited to, the patient's age, size, type or extent of disease, stage of the disease, route of administration, the type or extent of supplemental therapy used, ongoing disease process, and type of treatment desired (e.g., aggressive vs. conventional treatment).

[0066] As used herein, the term "effective amount" refers to the amount of a composition sufficient to effect beneficial or desired results. An effective amount can be administered in one or more administrations, applications, or dosages and is not intended to be limited to a particular formulation or administration route.

[0067] As used herein, the term "pharmaceutical composition" refers to the combination of an active agent with, as desired, a carrier, inert or active, making the composition especially suitable for diagnostic or therapeutic use in vitro, in vivo, or ex vivo.

[0068] As used herein, the terms "pharmaceutically acceptable" or "pharmacologically acceptable" refer to compositions that do not substantially produce adverse reactions, e.g., toxic, allergic, or immunological reactions, when administered to a subject.

[0069] As used herein, "carriers" include pharmaceutically acceptable carriers, excipients, or stabilizers which are nontoxic to the cell or mammal being exposed thereto at the dosages and concentrations employed. Often the physiologically acceptable carrier is an aqueous pH-buffered solution. Examples of physiologically acceptable carriers include buffers such as phosphate, citrate, and other organic acids; antioxidants including ascorbic acid; low molecular weight (less than about 10 residues) polypeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrins; chelating agents such as EDTA; sugar alcohols such as mannitol or sorbitol; salt-forming counterions such as sodium; and/or nonionic surfactants.

[0070] As used herein, the terms "patient" or "subject" refer to organisms to be treated by the compositions of the present technology or to be subject to various tests provided by the technology. The term "subject" includes animals, preferably mammals, including humans. In a preferred embodiment, the subject is a primate. In an even more preferred embodiment, the subject is a human.

[0071] As used herein, the term "sample" is used in its broadest sense. In one sense it can refer to animal cells or tissues. In another sense, it is meant to include a specimen or culture obtained from any source, such as biological and environmental samples. Biological samples may be obtained from plants or animals (including humans) and encompass fluids, solids, tissues, and gases. Environmental samples include environmental material such as surface matter, soil, water, and industrial samples. These examples are not to be construed as limiting the sample types applicable to the present technology.

Embodiments of the Technology

[0072] Although the disclosure herein refers to certain illustrated embodiments, it is to be understood that these embodiments are presented by way of example and not by way of limitation.

[0073] 1. Inhibitors of SCF

[0074] Stem cell factor (SCF) is a ligand that is specific for the c-Kit receptor kinase. Binding of SCF to c-Kit causes dimerization of c-Kit and activation of its kinase activity, which is important for hemopoiesis, melanogenesis, and fertility. Through c-Kit, SCF acts to promote cell survival, proliferation, differentiation, adhesion, and functional activation. Aberrant activation of c-Kit can result in disease, including fibrosis and tissue remodeling defects. In particular, there are multiple pulmonary diseases with known remodeling defects as well as other chronic tissue remodeling diseases affecting other organs and tissues. Specific examples of diseases involving fibrosis or tissue remodeling defects are idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, acute respiratory distress syndrome, cystic fibrosis, peribronchial fibrosis, hypersensitivity pneumonitis, asthma, sclerodoma, inflammation, liver cirrhosis, renal fibrosis, parenchymal fibrosis, endomyocardial fibrosis, mediatinal fibrosis, nodular subepidermal fibrosis, fibrous histiocytoma, fibrothorax, hepatic fibrosis, fibromyalgia, gingival fibrosis, and radiation-induced fibrosis.

[0075] Accordingly, interfering with the interaction between SCF and c-Kit can be used to treat or study diseases involving aberrant activation of c-Kit that causes fibrosis and tissue remodeling defects. The c-Kit receptor is found on hematopoietic progenitor cells, melanocytes, germ cells, eosinophils, lymphocytes, and mast cells. Thus, preventing SCF interaction with c-Kit can alter the activation of several disease-associated cell populations that have been implicated in fibrosis and tissue remodeling disease phenotypes.

[0076] Additionally, SCF induces key mediators in the fibrotic response, IL-25 and IL-13. Data suggest that IL-25 can drive IL-13 expression in a T-cell and antigen-independent manner. Therefore, these processes can progress without an antigen-specific response and consequently chronically perpetuate remodeling and fibrotic disease. It is contemplated that a complex cascade is established in which SCF induces IL-25, which in turn induces production of IL-13, myofibroblast differentiation, and collagen production. IL-4 has also been identified as a fibrosis-associated cytokine.

[0077] 2. Antibodies

[0078] In some embodiments, inhibiting the ability of SCF to interact with c-Kit is accomplished by means of an antibody that recognizes SCF. The antibody can be a monoclonal antibody or a polyclonal antibody, and may be, for example, a human, humanized, or chimeric antibody. Monoclonal antibodies against target antigens are produced by a variety of techniques including conventional monoclonal antibody methodologies such as the somatic cell hybridization techniques of Kohler and Milstein (Nature, 256:495 (1975)). Although in some embodiments, somatic cell hybridization procedures are preferred, other techniques for producing monoclonal antibodies are contemplated as well (e.g., viral or oncogenic transformation of B lymphocytes).

[0079] It is contemplated that antibodies against SCF find use in the experimental, diagnostic, and therapeutic methods described herein. In certain embodiments, the antibodies provided herein are used to detect the expression of SCF in biological samples. For example, a sample comprising a tissue biopsy can be sectioned and protein detected using, for example, immunofluorescence or immunohistochemistry. Alternatively, individual cells from a sample can be isolated, and protein expression detected on fixed or live cells by FACS analysis. Furthermore, the antibodies can be used on protein arrays to detect expression of SCF. In other embodiments, the antibodies provided herein are used to decrease the activity of cells expressing c-Kit by inhibiting SCF either in an in vitro cell-based assay or in an in vivo animal model. In some embodiments, antibodies are used to treat a human patient by administering a therapeutically effective amount of an antibody against SCF.

[0080] For the production of antibodies, various host animals can be immunized by injection with the peptide corresponding to the desired epitope (e.g., a fragment of SCF, e.g., a fragment comprising the sequence provided by SEQ ID NO: 1 or 8 or immunogenic portions thereof) including, but not limited to, rabbits, mice, rats, sheep, goats, etc. Antibodies to SCF can be raised by immunizing (e.g., by injection) with an antigen comprising a peptide, a portion, or the full protein of the SCF isoform b precursor (e.g., a protein or peptide fragment of the sequence available at GenBank accession number NP_000890 (SEQ ID NO: 4)), or a variant or modified version thereof, or a peptide, a portion, or the full protein of the SCF isoform a precursor (e.g., a protein or peptide fragment of the sequence available at GenBank accession number NP_003985 (SEQ ID NO: 6)), or a variant or modified version thereof. Antibodies can also be raised by immunization with a translation product of the NCBI Reference Gene Sequence for SCF (e.g., accession number NG_012098 (SEQ ID NO: 7)) or variants or fragments thereof.

[0081] In some embodiments, the peptide is conjugated to an immunogenic carrier (e.g., diphtheria toxoid, bovine serum albumin (BSA), or keyhole limpet hemocyanin (KLH)). Various adjuvants are used to increase the immunological response, depending on the host species, including, but not limited to, Freund's (complete and incomplete), mineral gels such as aluminum hydroxide, surface active substances such as lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, keyhole limpet hemocyanins, dinitrophenol, and potentially useful human adjuvants such as BCG (Bacille Calmette-Guerin) and Corynebacterium parvum.

[0082] Polyclonal antibodies can be prepared by any known method. Polyclonal antibodies can be raised by immunizing an animal (e.g., a rabbit, rat, mouse, donkey, etc) by multiple subcutaneous or intraperitoneal injections of the relevant antigen (a purified peptide fragment, full-length recombinant protein, fusion protein, etc.) optionally conjugated to KLH, serum albumin, etc., diluted in sterile saline, and combined with an adjuvant to form a stable emulsion. The polyclonal antibody is then recovered from blood, ascites, and the like, of an animal so immunized. Collected blood is clotted, and the serum decanted, clarified by centrifugation, and assayed for antibody titer. The polyclonal antibodies can be purified from serum or ascites according to standard methods in the art including affinity chromatography, ion-exchange chromatography, gel electrophoresis, dialysis, etc.

[0083] For preparation of monoclonal antibodies, any technique that provides for the production of antibody molecules by continuous cell lines in culture may be used (see e.g., Harlow and Lane, Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.). These include, but are not limited to, the hybridoma technique originally developed by Kohler and Milstein and the trioma technique, the human B-cell hybridoma technique (See, e.g., Kozbor et al., Immunol. Today, 4:72 (1983)), and the EBV-hybridoma technique to produce human monoclonal antibodies (Cole et al., in Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc., pp. 77-96 (1985)).

[0084] In some embodiments provided herein, the antibodies are prepared from a hybridoma. Using the hybridoma method, a mouse, hamster, or other appropriate host animal, is immunized as described above to elicit the production by lymphocytes of antibodies that will specifically bind to an immunizing antigen. Alternatively, lymphocytes can be immunized in vitro. Following immunization, the lymphocytes are isolated and fused with a suitable myeloma cell line using, for example, polyethylene glycol, to form hybridoma cells that can then be selected away from unfused lymphocytes and myeloma cells. Hybridomas that produce monoclonal antibodies directed specifically against a chosen antigen as determined by immunoprecipitation, immunoblotting, or by an in vitro binding assay such as radioimmunoassay (RIA) or enzyme-linked immunosorbent assay (ELISA) can then be propagated in vitro (e.g., in culture) using standard methods (Goding, Monoclonal Antibodies: Principles and Practice, Academic Press, 1986) or in vivo as ascites tumors in an animal. The monoclonal antibodies can then be purified from the culture medium or ascites fluid as described for polyclonal antibodies above.

[0085] The preferred animal system for preparing hybridomas is the murine system. Hybridoma production in the mouse is a well-established procedure. Immunization protocols and techniques for isolation of immunized splenocytes for fusion are known in the art. Fusion partners (e.g., murine myeloma cells) and fusion procedures are also known. Embodiments of the technology herein provide antibodies (e.g., monoclonal antibodies) produced from a hybridoma prepared by immunizing mice with a peptide that is a portion or fragment of the SCF protein. For example, some embodiments provide an antibody or antigen-binding fragment than binds to SCF by immunizing with, e.g., a protein or peptide fragment of the sequence available at GenBank accession number NP_000890 (SEQ ID NO: 4)), or a variant or modified version thereof, or by immunizing with, e.g., a protein or peptide fragment of the sequence available at GenBank accession number NP_003985 (SEQ ID NO: 6)), or a variant or modified version thereof. Some embodiments provide an antibody or antigen-binding fragment that binds to a protein or peptide, or variants or modified versions thereof, that is a translation product of the NCBI Reference Gene Sequence for SCF (e.g., accession number NG_012098 (SEQ ID NO: 7)) or variants or fragments thereof.

[0086] For example, embodiments of the technology herein provide monoclonal antibodies produced from a hybridoma prepared by immunizing mice with a peptide of amino acid sequence SEQ ID NO: 1 or 8. Also contemplated are methods and compositions related to antibodies prepared using a variant of SEQ ID NO: 1 or 8 comprising one or more substitutions, deletions, insertions, or other changes, as long as said variant produces an antibody specific for SCF. Producing polypeptides of SEQ ID NO: 1 or 8 and similar sequences thereto can be accomplished according to various techniques well known in the art. For example, a polypeptide of SEQ ID NO: 1 or 8 or a variant thereof can be produced using a bacterial expression system and a nucleic acid encoding a polypeptide of SEQ ID NO: 1 or 8 or a variant thereof. As an example, a polypeptide according to SEQ ID NO: 1 can be produced using the nucleotide sequence according to SEQ ID NO: 2.

[0087] Moreover, human monoclonal antibodies directed against human proteins can be generated using transgenic mice carrying the complete human immune system rather than the mouse system. Splenocytes from the transgenic mice are immunized with the antigen of interest, which are used to produce hybridomas that secrete human monoclonal antibodies with specific affinities for epitopes from a human protein.

[0088] Monoclonal antibodies can also be generated by other methods known to those skilled in the art of recombinant DNA technology. For instance, combinatorial antibody display has can be utilized to produce monoclonal antibodies (see, e.g., Sastry et al., Proc. Nat. Acad. Sci. USA, 86: 5728 (1989); Huse et al., Science, 246: 1275 (1989); Orlandi et al., Proc. Nat. Acad. Sci. USA, 86:3833 (1989)). After immunizing an animal with an immunogen as described above, the antibody repertoire of the resulting B-cell pool is cloned. Methods are generally known for obtaining the DNA sequence of the variable regions of a diverse population of immunoglobulin molecules by using a mixture of oligomer primers and PCR. For instance, mixed oligonucleotide primers corresponding to the 5' leader (signal peptide) sequences and/or framework 1 (FR1) sequences, as well as primers to a conserved 3' region can be used to amplify and isolate the heavy and light chain variable regions from a number of murine antibodies (see. e.g., Larrick et al., Biotechniques, 11: 152 (1991)). A similar strategy can also been used to amplify human heavy and light chain variable regions from human antibodies (see, e.g., Larrick et al., Methods: Companion to Methods in Enzymology, 2: 106 (1991)).

[0089] Alternatively, monoclonal antibodies can also be made using recombinant DNA methods as described in U.S. Pat. No. 4,816,567. The polynucleotides encoding a monoclonal antibody are isolated (e.g., from mature B-cells or hybridoma cells), by, e.g., RT-PCR using oligonucleotide primers that specifically amplify the genes encoding the heavy and light chains of the antibody, and their sequences are determined using conventional procedures. The isolated polynucleotides encoding the heavy and light chains are then cloned into suitable expression vectors, which, when transfected into host cells such as E. coli cells, simian COS cells, Chinese hamster ovary (CHO) cells, or myeloma cells that do not otherwise produce immunoglobulin protein, cause monoclonal antibodies to be generated by the host cells. Also, recombinant monoclonal antibodies or fragments thereof of the desired species can be isolated from phage display libraries as described (McCafferty et al., 1990, Nature, 348:552-554; Clackson et al., 1991, Nature, 352:624-628; and Marks et al., 1991, J. Mol. Biol., 222:581-597).

[0090] The polynucleotide encoding a monoclonal antibody can further be modified in a number of different manners using recombinant DNA technology to generate alternative antibodies. In one embodiment, the constant domains of the light and heavy chains of, for example, a mouse monoclonal antibody can be substituted 1) for those regions of, for example, a human antibody to generate a chimeric antibody or 2) for a non-immunoglobulin polypeptide to generate a fusion antibody. In other embodiments, the constant regions are truncated or removed to generate the desired antibody fragment of a monoclonal antibody. Furthermore, site-directed or high-density mutagenesis of the variable region can be used to optimize specificity, affinity, etc. of a monoclonal antibody.

[0091] For example, also contemplated are chimeric mouse-human monoclonal antibodies, which can be produced by recombinant DNA techniques known in the art. For example, a gene encoding the constant region of a murine (or other species) monoclonal antibody molecule is digested with restriction enzymes to remove the region encoding the murine constant region, and the equivalent portion of a gene encoding a human constant region is substituted (see, e.g., Robinson et al., PCT/US86/02269; European Patent Application 184,187; European Patent Application 171,496; European Patent Application 173,494; WO 86/01533; U.S. Pat. No. 4,816,567; European Patent Application 125,023 (each of which is herein incorporated by reference in its entirety); Better et al., Science, 240:1041-1043 (1988); Liu et al., Proc. Nat. Acad. Sci. USA, 84:3439-3443 (1987); Liu et al., J. Immunol., 139:3521-3526 (1987); Sun et al., Proc. Nat. Acad. Sci. USA, 84:214-218 (1987); Nishimura et al., Canc. Res., 47:999-1005 (1987); Wood et al., Nature, 314:446-449 (1985); and Shaw et al., J. Natl. Cancer Inst., 80:1553-1559 (1988)).

[0092] The chimeric antibody can be further humanized by replacing sequences of the variable region that are not directly involved in antigen binding with equivalent sequences from human variable regions. General reviews of humanized chimeric antibodies are provided by S. L. Morrison, Science, 229:1202-1207 (1985) and by Oi et al., Bio Techniques, 4:214 (1986). Those methods include isolating, manipulating, and expressing the nucleic acid sequences that encode all or part of immunoglobulin variable regions from at least one of a heavy or light chain. Sources of such nucleic acid are well known to those skilled in the art. The recombinant DNA encoding the chimeric antibody, or fragment thereof, can then be cloned into an appropriate expression vector.

[0093] Suitable humanized antibodies can alternatively be produced by CDR substitution (see, e.g., U.S. Pat. No. 5,225,539; Jones et al., Nature, 321:552-525 (1986); Verhoeyan et al., Science, 239:1534 (1988); and Beidler et al., J. Immunol., 141:4053 (1988)). All of the CDRs of a particular human antibody may be replaced with at least a portion of a non-human CDR or only some of the CDRs may be replaced with non-human CDRs. It is only necessary to replace the number of CDRs important for binding of the humanized antibody to the Fc receptor.

[0094] An antibody can be humanized by any method that is capable of replacing at least a portion of a CDR of a human antibody with a CDR derived from a non-human antibody. The human CDRs may be replaced with non-human CDRs using oligonucleotide site-directed mutagenesis.

[0095] Also contemplated are chimeric and humanized antibodies in which specific amino acids have been substituted, deleted, or added. In particular, preferred humanized antibodies have amino acid substitutions in the framework region, such as to improve binding to the antigen. For example, in a humanized antibody having mouse CDRs, amino acids located in the human framework region can be replaced with the amino acids located at the corresponding positions in the mouse antibody. Such substitutions are known to improve binding of humanized antibodies to the antigen in some instances.

[0096] In certain embodiments provided herein, it is desirable to use an antibody fragment. Various techniques are known for the production of antibody fragments. Traditionally, these fragments are derived via proteolytic digestion of intact antibodies (for example Morimoto et al., 1993, Journal of Biochemical and Biophysical Methods 24:107-117 and Brennan et al., 1985, Science, 229:81). For example, papain digestion of antibodies produces two identical antigen-binding fragments, called Fab fragments, each with a single antigen-binding site, and a residual Fc fragment. Pepsin treatment yields an F(ab').sub.2 fragment that has two antigen-combining sites and is still capable of cross-linking antigen.

[0097] However, these fragments are now typically produced directly by recombinant host cells as described above. Thus Fab, Fv, and scFv antibody fragments can all be expressed in and secreted from E. coli or other host cells, thus allowing the production of large amounts of these fragments. Alternatively, such antibody fragments can be isolated from the antibody phage libraries discussed above. The antibody fragment can also be linear antibodies as described in U.S. Pat. No. 5,641,870, for example, and can be monospecific or bispecific. Other techniques for the production of antibody fragments will be apparent to the skilled practitioner.

[0098] Fv is the minimum antibody fragment which contains a complete antigen-recognition and antigen-binding site. This region consists of a dimer of one heavy-chain and one light-chain variable domain in tight, non-covalent association. It is in this configuration that the three CDRs of each variable domain interact to define an antigen-binding site on the surface of the V.sub.H-V.sub.L dimer. Collectively, the six CDRs confer antigen-binding specificity to the antibody. However, even a single variable domain (or half of an Fv comprising only three CDRs specific for an antigen) has the ability to recognize and bind antigen, although at a lower affinity than the entire binding site.

[0099] The Fab fragment also contains the constant domain of the light chain and the first constant domain (CH1) of the heavy chain. Fab fragments differ from Fab' fragments by the addition of a few residues at the carboxy terminus of the heavy chain CH1 domain including one or more cysteines from the antibody hinge region. F(ab').sub.2 antibody fragments originally were produced as pairs of Fab' fragments which have hinge cysteines between them. Other chemical couplings of antibody fragments are also known to the skilled artisan.

[0100] The technology herein provided also contemplates modifying an antibody to increase its serum half-life. This can be achieved, for example, by incorporating a salvage receptor binding epitope into the antibody fragment by mutation of the appropriate region in the antibody fragment or by incorporating the epitope into a peptide tag that is then fused to the antibody fragment at either end or in the middle (e.g., by DNA or peptide synthesis).

[0101] The technology embraces variants and equivalents which are substantially homologous to the chimeric, humanized, and human antibodies, or antibody fragments thereof, provided herein. These can contain, for example, conservative substitution mutations, i.e. the substitution of one or more amino acids by similar amino acids. For example, conservative substitution refers to the substitution of an amino acid with another within the same general class such as, for example, one acidic amino acid with another acidic amino acid, one basic amino acid with another basic amino acid, or one neutral amino acid by another neutral amino acid. What is intended by a conservative amino acid substitution is well known in the art.

[0102] An additional embodiment utilizes the techniques known in the art for the construction of Fab expression libraries (Huse et al., Science, 246:1275-1281 (1989)) to allow rapid and easy identification of monoclonal Fab fragments with the desired specificity.

[0103] Also, this technology encompasses bispecific antibodies that specifically recognize SCF. Bispecific antibodies are antibodies that are capable of specifically recognizing and binding at least two different epitopes. Bispecific antibodies can be intact antibodies or antibody fragments. Techniques for making bispecific antibodies are common in the art (Millstein et al., 1983, Nature 305:537-539; Brennan et al., 1985, Science 229:81; Suresh et al, 1986, Methods in Enzymol. 121:120; Traunecker et al., 1991, EMBO J. 10:3655-3659; Shalaby et al., 1992, J. Exp. Med. 175:217-225; Kostelny et al., 1992, J. Immunol. 148:1547-1553; Gruber et al., 1994, J. Immunol. 152:5368; and U.S. Pat. No. 5,731,168).

[0104] Techniques described for the production of single chain antibodies (U.S. Pat. No. 4,946,778; herein incorporated by reference) can be adapted to produce specific single chain antibodies as desired. Single-chain Fv antibody fragments comprise the V.sub.H and V.sub.L domains of an antibody, wherein these domains are present in a single polypeptide chain. Preferably, the Fv polypeptide further comprises a polypeptide linker between the V.sub.H and V.sub.L domains that enables the single-chain Fv antibody fragments to form the desired structure for antigen binding. For a review of single-chain Fv antibody fragments, see Pluckthun in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds., Springer-Verlag, New York, pp. 269-315 (1994).

[0105] 3. Other SCF Inhibitors

[0106] It is also contemplated that inhibiting SCF can be accomplished by a variety of other types of inhibitors. For example, in some embodiments a small interfering RNA (siRNA) can be designed to target and degrade SCF mRNA. siRNAs are double-stranded RNA molecules of 20-25 nucleotides in length. While not limited in their features, typically an siRNA is 21 nucleotides long and has 2-nt 3' overhangs on both ends. Each strand has a 5' phosphate group and a 3' hydroxyl group. In vivo, this structure is the result of processing by dicer, an enzyme that converts either long dsRNAs or small hairpin RNAs into siRNAs. However, siRNAs can also be synthesized and exogenously introduced into cells to bring about the specific knockdown of a gene of interest. Essentially any gene of which the sequence is known can be targeted based on sequence complementarity with an appropriately tailored siRNA. For example, those of ordinary skill in the art can synthesize an siRNA (see, e.g., Elbashir, et al., Nature 411: 494 (2001); Elbashir, et al. Genes Dev 15:188 (2001); Tuschl T, et al., Genes Dev 13:3191 (1999)).

[0107] In some embodiments, RNAi is utilized to inhibit SCF. RNAi represents an evolutionarily conserved cellular defense for controlling the expression of foreign genes in most eukaryotes, including humans. RNAi is typically triggered by double-stranded RNA (dsRNA) and causes sequence-specific degradation of single-stranded target RNAs (e.g., an mRNA). The mediators of mRNA degradation are small interfering RNAs (siRNAs), which are normally produced from long dsRNA by enzymatic cleavage in the cell. siRNAs are generally approximately twenty-one nucleotides in length (e.g. 21-23 nucleotides in length) and have a base-paired structure characterized by two nucleotide 3' overhangs. Following the introduction of a small RNA, or RNAi, into the cell, it is believed the sequence is delivered to an enzyme complex called RISC (RNA-induced silencing complex). RISC recognizes the target and cleaves it with an endonuclease. It is noted that if larger RNA sequences are delivered to a cell, an RNase III enzyme (e.g., Dicer) converts the longer dsRNA into 21-23 nt double-stranded siRNA fragments. In some embodiments, RNAi oligonucleotides are designed to target the junction region of fusion proteins. Chemically synthesized siRNAs have become powerful reagents for genome-wide analysis of mammalian gene function in cultured somatic cells. Beyond their value for validation of gene function, siRNAs also hold great potential as gene-specific therapeutic agents (see, e.g., Tuschl and Borkhardt, Molecular Intervent. 2002; 2(3): 158-67, herein incorporated by reference).

[0108] The transfection of siRNAs into animal cells results in the potent, long-lasting post-transcriptional silencing of specific genes (Caplen et al, Proc Natl Acad Sci U.S.A. 2001; 98: 9742-47; Elbashir et al., Nature. 2001; 411:4 94-98; Elbashir et al., Genes Dev. 2001; 15: 188-200; and Elbashir et al., EMBO J. 2001; 20: 6877-88, all of which are herein incorporated by reference). Methods and compositions for performing RNAi with siRNAs are described, for example, in U.S. Pat. No. 6,506,559, herein incorporated by reference.

[0109] siRNAs are extraordinarily effective at lowering the amounts of targeted RNA and their protein products, frequently to undetectable levels. The silencing effect can last several months, and is extraordinarily specific--a one-nucleotide mismatch between the target RNA and the central region of the siRNA is frequently sufficient to prevent silencing (Brummelkamp et al, Science 2002; 296: 550-53; and Holen et al, Nucleic Acids Res. 2002; 30: 1757-66, both of which are herein incorporated by reference).

[0110] An important factor in the design of siRNAs is the presence of accessible sites for siRNA binding. Bahoia et al., (J. Biol. Chem., 2003; 278: 15991-97; herein incorporated by reference) describe the use of a type of DNA array called a scanning array to find accessible sites in mRNAs for designing effective siRNAs. These arrays comprise oligonucleotides ranging in size from monomers to a certain maximum, usually Co-mers, synthesized using a physical barrier (mask) by stepwise addition of each base in the sequence. Thus the arrays represent a full oligonucleotide complement of a region of the target gene. Hybridization of the target mRNA to these arrays provides an exhaustive accessibility profile of this region of the target mRNA. Such data are useful in the design of antisense oligonucleotides (ranging from 7mers to 25mers), where it is important to achieve a compromise between oligonucleotide length and binding affinity, e.g., to retain efficacy and target specificity (Sohail et al, Nucleic Acids Res., 2001; 29(10): 2041-45). Additional methods and concerns for selecting siRNAs are described, for example, in WO 05054270, WO05038054A1, WO03070966A2, J Mol Biol. 2005 May 13; 348(4):883-93, J Mol Biol. 2005 May 13; 348(4):871-81, and Nucleic Acids Res. 2003 Aug. 1; 31(15):4417-24, each of which is herein incorporated by reference in its entirety. In addition, software (e.g., the MWG online siMAX siRNA design tool) is commercially or publicly available for use in the selection and design of siRNAs and RNAi reagents.

[0111] In some embodiments, the present invention utilizes siRNA including blunt ends (See e.g., US20080200420, herein incorporated by reference in its entirety), overhangs (See e.g., US20080269147A1, herein incorporated by reference in its entirety), locked nucleic acids (See e.g., WO2008/006369, WO2008/043753, and WO2008/051306, each of which is herein incorporated by reference in its entirety). In some embodiments, siRNAs are delivered via gene expression or using bacteria (See e.g., Xiang et al., Nature 24: 6 (2006) and WO06066048, each of which is herein incorporated by reference in its entirety).

[0112] In other embodiments, shRNA techniques (See e.g., 20080025958, herein incorporated by reference in its entirety) are utilized. A small hairpin RNA or short hairpin RNA (shRNA) is a sequence of RNA that makes a tight hairpin turn that can be used to silence gene expression via RNA interference. shRNA uses a vector introduced into cells and utilizes the U6 promoter to ensure that the shRNA is always expressed. This vector is usually passed on to daughter cells, allowing the gene silencing to be inherited. The shRNA hairpin structure is cleaved by the cellular machinery into siRNA, which is then bound to the RNA-induced silencing complex (RISC). This complex binds to and cleaves mRNAs which match the siRNA that is bound to it. shRNA is transcribed by RNA polymerase III.

[0113] The present invention also includes pharmaceutical compositions and formulations that include the RNAi compounds of the present invention as described below.

[0114] SCF exists in both transmembrane and soluble forms. Upon cleavage of the SCF soluble domain from the transmembrane form, SCF is released from the cell surface to function as the ligand of c-Kit. Thus, it is contemplated that SCF activity can be altered by inhibiting the release of soluble SCF from the membrane-bound form, for example, by inhibiting or otherwise reducing the activity of a protease that cleaves the soluble domain from the membrane-bound form.

[0115] In addition, it is contemplated that SCF can be inhibited by chemicals (e.g., a small molecule, e.g., a pharmacological agent) or other biological agents that bind or modify SCF. For example, one of ordinary skill in the art can design and produce RNA aptamers or other nucleic acids that specifically recognize and bind to SCF, for instance by using SELEX or other in vitro evolution methods known in the art. Furthermore, SCF activity can be inhibited by specifically degrading SCF or inducing an altered conformation of SCF such that it is less effective in interacting with c-Kit. In some embodiments, the SCF inhibitor is a "designed ankyrin repeat protein" (DARPin) (see, e.g., Stumpp M T & Amstutz P, "DARPins: a true alternative to antibodies", Curr Opin Drug Discov Devel 2007, 10(2): 153-59, incorporated herein in its entirety for all purposes). In some embodiments, SCF is inhibited by a small molecule, e.g., a small molecule that binds to SCF and blocks its function (e.g., inhibits its binding and/or other interaction (e.g., an activating interaction) with the c-Kit receptor).

[0116] It is contemplated that altering SCF activity can be effected by inhibiting the expression of SCF, for instance, by inhibiting the transcription of SCF, by inhibiting the translation of SCF, by inhibiting the processing of the SCF mRNA, by inhibiting the processing of the SCF polypeptide, by inhibiting the folding of the SCF polypeptide, by inhibiting trafficking of SCF within a cell, or by inhibiting the insertion of SCF into the plasma membrane. SCF activity can be altered by changes in chromatin structure or other means of epigenetic regulation of SCF (e.g., changes in DNA methylation). Also, SCF activity may be altered by specifically sequestering SCF in a vesicle or other cellular compartment that hinders its action upon c-Kit.

[0117] 4. Therapies Using Inhibitors of SCF

[0118] Inhibiting SCF finds use in therapies to treat disease. Accordingly, provided herein are therapies comprising inhibiting SCF to benefit individuals suffering from disease. In particular, as shown herein, disease states involving fibrosis and tissue remodeling demonstrate aberrant SCF activity. For example, fibroblasts isolated from diseased individuals with fibrotic or tissue remodeling phenotypes directly respond to SCF, which results in the generation of a more severe phenotype that includes increased collagen production. As such, as shown herein, inhibiting SCF can significantly affect the generation of severe disease consequences including inflammation and remodeling of target tissue. Also contemplated are therapies targeting SCF during the generation of fibrosis associated with acute and chronic disorders that have either a dynamic disease course or a more predictable disease course. Indications that can benefit from therapy inhibiting SCF include, but are not limited to, idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, acute respiratory distress syndrome, cystic fibrosis, peribronchial fibrosis, hypersensitivity pneumonitis, asthma, sclerodoma, inflammation, liver cirrhosis, renal fibrosis, parenchymal fibrosis, endomyocardial fibrosis, mediatinal fibrosis, nodular subepidermal fibrosis, fibrous histiocytoma, fibrothorax, hepatic fibrosis, fibromyalgia, gingival fibrosis, and radiation-induced fibrosis.

[0119] Importantly, therapies targeting SCF reduce or eliminate toxic effects associated with other similar therapies, for example those targeting c-Kit. These undesirable toxic effects are associated with targeting an intracellular, rather than extracellular, target, and the more widespread and general changes in cell signaling that result. While the therapies are not limited in their route of administration, embodiments of the technology provided herein deliver the SCF inhibitor via the airway by intranasal administration. Such administration allows direct delivery of the therapeutic agent to target tissues in pulmonary diseases involving fibrosis and tissue remodeling, rather than relying on systemic delivery via an orally administered composition.

[0120] In certain embodiments, a physiologically appropriate solution containing an effective concentration of an antibody specific for SCF can be administered topically, intraocularly, parenterally, orally, intranasally, intravenously, intramuscularly, subcutaneously, or by any other effective means. In particular, the antibody may delivered into an airway of a subject by intranasal administration. Alternatively, a tissue can receive a physiologically appropriate composition (e.g., a solution such as a saline or phosphate buffer, a suspension, or an emulsion, which is sterile) containing an effective concentration of an antibody specific for SCF via direct injection with a needle or via a catheter or other delivery tube. Any effective imaging device such as X-ray, sonogram, or fiber-optic visualization system may be used to locate the target tissue and guide the admistration. In another alternative, a physiologically appropriate solution containing an effective concentration of an antibody specific for SCF can be administered systemically into the blood circulation to treat tissue that cannot be directly reached or anatomically isolated. Such manipulations have in common the goal of placing an effective concentration of an antibody specific for SCF in sufficient contact with the target tissue to permit the antibody specific for SCF to contact the tissue.

[0121] With respect to administration of a SCF inhibitor (e.g., an antibody specific for SCF) to a subject, it is contemplated that the SCF inhibitor be administered in a pharmaceutically effective amount. One of ordinary skill recognizes that a pharmaceutically effective amount varies depending on the therapeutic agent used, the subject's age, condition, and sex, and on the extent of the disease in the subject. Generally, the dosage should not be so large as to cause adverse side effects, such as hyperviscosity syndromes, pulmonary edema, congestive heart failure, and the like. The dosage can also be adjusted by the individual physician or veterinarian to achieve the desired therapeutic goal.

[0122] As used herein, the actual amount encompassed by the term "pharmaceutically effective amount" will depend on the route of administration, the type of subject being treated, and the physical characteristics of the specific subject under consideration. These factors and their relationship to determining this amount are well known to skilled practitioners in the medical, veterinary, and other related arts. This amount and the method of administration can be tailored to achieve optimal efficacy but will depend on such factors as weight, diet, concurrent medication, and other factors that those skilled in the art will recognize.

[0123] In some embodiments, a SCF inhibitor (e.g., an antibody specific for SCF) according to the technology provided herein is administered in a pharmaceutically effective amount. In some embodiments, a SCF inhibitor (e.g., an antibody specific for SCF) is administered in a therapeutically effective dose. The dosage amount and frequency are selected to create an effective level of the SCF inhibitor without substantially harmful effects. When administered, the dosage of a SCF inhibitor (e.g., an antibody specific for SCF) will generally range from 0.001 to 10,000 mg/kg/day or dose (e.g., 0.01 to 1000 mg/kg/day or dose; 0.1 to 100 mg/kg/day or dose).

[0124] Pharmaceutical compositions preferably comprise one or more compounds of the present invention associated with one or more pharmaceutically acceptable carriers, diluents, or excipients. Pharmaceutically acceptable carriers are known in the art such as those described in, for example, Remingtons Pharmaceutical Sciences, Mack Publishing Co. (A. R. Gennaro ed., 1985).

[0125] In some embodiments, a single dose of a SCF inhibitor (e.g., an antibody specific for SCF) according to the technology provided herein is administered to a subject. In other embodiments, multiple doses are administered over two or more time points, separated by hours, days, weeks, etc. In some embodiments, compounds are administered over a long period of time (e.g., chronically), for example, for a period of months or years (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or more months or years; e.g., for the lifetime of the subject). In such embodiments, compounds may be taken on a regular scheduled basis (e.g., daily, weekly, etc.) for the duration of the extended period.

[0126] In some embodiments, a SCF inhibitor (e.g., an antibody specific for SCF) according to the technology provided herein is co-administered with another compound or more than one other compound (e.g., 2 or 3 or more other compounds).

[0127] 5. Kits

[0128] Some embodiments provide herein kits for the treatment of a subject. In some embodiments, the kits include an inhibitor of SCF and appropriate solutions and buffers. Embodiments include all controls and instructions for use.

Examples

Materials and Methods

SCF Nucleotide Sequences and Proteins

[0129] The human gene encoding Stem Cell Factor (SCF) is also known as kit ligand and has the official symbol KITLG and HGNC number HGNC:6343. SCF is also known as SF; MGF; SCF; FPH2; KL-1; Kit1; SHEP7; and kit-ligand. Two transcript variants encoding different isoforms have been found for this gene. The SCF (kit ligand) isoform b precursor is available at GenBank accession numbers NM_000899 (mRNA transcript; SEQ ID NO: 3) and NP_000890 (protein sequence; SEQ ID NO: 4). The SCF (kit ligand) isoform a precursor is available at GenBank accession numbers NM_003994 (mRNA transcript; SEQ ID NO: 5) and NP_003985 (protein sequence; SEQ ID NO: 6). The NCBI Reference Gene Sequence has accession number NG_012098 (SEQ ID NO: 7). For both isoforms, the first 25 amino acids comprise the signal peptide and the mature form begins at amino acid 26. The first 11 amino acids of the mature form are EGICRNRVTNN (SEQ ID NO: 8).

Bleomycin Model

[0130] Interstitial pulmonary fibrosis was induced in specific pathogen-free (SPF) female, CBA/J mice (6-8 weeks old; The Jackson Laboratory, Bar Harbor, Me.) by the i.t. injection of 0.003 U of bleomycin (Blenoxane, sterile bleomycin sulfate; Bristol-Meyers Pharmaceuticals, Evansville, Ind.; 0.15 U/Kg of mouse body weight) dissolved in 60 .mu.l of phosphate-buffered saline (PBS). Controls received 60 .mu.l of PBS by the same route. All procedures were conducted in a sterile environment and were approved by the institutional animal care and use committee.

Whole Lung Histology

[0131] Following anesthesia-induced euthanasia, whole lungs from bleomycin-challenged mice were fully inflated with 10% formalin, dissected, and placed in fresh formalin for 24 hours. Routine histological techniques were used to embed the entire lung in paraffin, and 5-.mu.m sections of whole lung were stained with hematoxylin and eosin.

Production and Administration of Anti-SCF Polyclonal Antibodies

[0132] Anti-SCF antibodies were generated by immunizing rabbits with recombinant (whole protein) SCF and generating polyclonal SCF-specific antibodies. Polyclonal antibodies were isolated from the serum using a protein G column. The isolated IgG portion was quantified and used at the specified concentrations suspended in saline. IgG from pre-immune serum was isolated in a similar fashion for use as a control. Briefly, 100, 150 or 200 .mu.g of control or anti-SCF was given to mice by intranasal administration 7 days after treatment with bleomycin. This treatment was repeated on a daily basis until 12 days after bleomycin administration. Thus, the treatment protocol is considered therapeutic.

Generation of Mouse Anti-Human Monoclonal Antibodies

[0133] After identifying an immunogenic human peptide (e.g., SEQ ID NO: 1 or 8), mice were immunized with a standard protocol. The determination of high titer serum antibodies indicated the appropriate immunization and fusion hybridomas were made. Culture supernatants were analyzed from individual clones for SCF-specific antibody and chosen based upon specificity. Five hybridomas producing specific monoclonal antibodies against the peptide were propagated and the monoclonal with the highest titer was subsequently tested in biologically relevant cultures. In some embodiments, a peptide having the sequence EGICRNRVTNN (SEQ ID NO: 8) was used to generate an antibody (e.g., a monoclonal antibody). In some embodiments, any peptide fragment (e.g., an antigenic fragment) of the SCF protein sequence (e.g., as provided by SEQ ID NO: 4 and/or SEQ ID NO: 6) is used to generate antibodies. In some embodiments, mutant or variant forms (e.g., comprising one or more amino acid substitutions with respect to the sequences provided by SEQ ID NO: 4 and SEQ ID NO: 6) of SCF are used to provide a peptide for generating antibodies. It is to be understood that these embodiments comprise additions, deletions, substitutions, post-translational modifications (e.g., glycosylation, cyclization, N- and C-terminal modification, etc.) and other variations of proteins and peptides that are known in the art of molecular biology as applied to provide a peptide for antibody generation.

Testing Mouse Anti-Human Monoclonal Antibodies

[0134] To demonstrate that monoclonal antibodies inhibit SCF, mast cell lines that are sensitive to SCF were tested. The HMC-1 cell line, a mastocytoma cell line that expresses c-Kit and responds to SCF was first used. In brief, HMC-1 cells were cultured in specific growth media and plated in 24-well tissue culture plates at a concentration of 1.times.10.sup.6 cells/ml. Recombinant human SCF (1-100 ng/ml) was mixed with monoclonal anti-SCF antibody (12 .mu.g/ml) and incubated at 37.degree. C. for 30 minutes. After incubation, the antibody/SCF or SCF alone was added to the HMC-1 cells. After 1 hour or 24 hours, the cultured HMC-1 cells were harvested and mRNA and protein levels were measured as an indication of SCF inhibition by the monoclonal antibodies.

Analysis of mRNA Expression by Quantitative PCR

[0135] Cells or tissue to be tested were dispersed in 1 ml of Trizol reagent (Invitrogen). RNA was isolated as described (Invitrogen), and 5 .mu.g of mRNA was reverse-transcribed to assess gene expression. Detection of cytokine mRNA was determined using previously available primer/probe sets (PE Biosystems, Foster City, Calif.) and analyzed using an ABI Prism 7500 Sequence Detection System (Applied Biosystems, Foster City, Calif.). GAPDH mRNA was measured as a control for normalizing mRNA expression. Changes in gene expression were calculated relative to gene expression in unchallenged mice.

Determination of Cytokine Production

[0136] Protein levels of cytokines were quantified using a Bio-Plex bead-based cytokine assay purchased from Bio-Rad Laboratories (Hercules, Calif.). Using standard protocols, the level of cytokines can be quickly and consistently assessed with this methodology.

Statistical Analysis

[0137] Data were analyzed using Prism GraphPad software. Unless otherwise specified, data shown are representative of two or more experiments. Statistical significance in all experiments was determined by one-way ANOVA, followed by a Newman-Keuls post test. Significant differences were regarded as p<0.05.

Isolation and Propagation of Pulmonary Fibroblasts from Patient Populations

[0138] The Institutional Review Board at the University of Michigan Medical School approved this study. All patients underwent clinical evaluation, including chest radiography, lung function measurements, and thin-section computed tomography before fiber optic bronchoscopy. In these patients, interstitial pneumonia was determined from a compilation of symptoms, physiological symptoms, and radiographical findings. Surgical lung biopsies were obtained via the Clinical Core at the University of Michigan Medical School from patients suspected of having interstitial pneumonia between May 2000 and May 2002. Histologically normal lung was obtained from resected specimens in patients undergoing thoracic resection. Each biopsy was processed separately using sterile technique in a laminar flow hood and processed for culturing primary fibroblast lines. Two pathologists who were unaware of any other clinical findings independently reviewed each biopsy and histological classification was based on previously published criteria for idiopathic interstitial pneumonia.

[0139] Interstitial pneumonia and normal biopsies were finely minced and the dispersed tissue pieces were placed into 150-cm.sup.2 cell culture flasks (Corning Inc., Corning, N.Y.) containing Dulbecco's modified Eagle's medium (DMEM, BioWhittaker, Walkersville, Md.) supplemented with 15% fetal bovine serum (DMEM-15, BioWhittaker), 1 mmol/L glutamine (BioWhittaker), 100 U/ml penicillin (BioWhittaker), 100 .mu.g/ml streptomycin (BioWhittaker), and 0.25 .mu.g amphotericin B (Fungizone; BioWhittaker). All primary lung cell lines were maintained in DMEM-15 at 37.degree. C. in a 5% CO.sub.2 incubator and were serially passaged a total of five times to yield pure populations of lung fibroblasts. All primary fibroblast cell lines were used at passages 6 to 10 in the experiments outlined below and all of the experiments were performed under comparable conditions.

[0140] 1. Anti-SCF Antibody Reduces Fibrosis and Inflammation

[0141] Experiments conducted while developing embodiments of the technology demonstrated that anti-SCF antibody reduced fibrosis and inflammation. Pulmonary fibrosis was induced in mice as described. On day 7 following bleomycin injury, mice were subjected to treatment with anti-SCF antibodies delivered into the airway by intranasal administration. Treatment continued until day 12 following bleomycin exposure. Lungs were harvested on day 16 and examined by microscopy and a series of micrographs were taken. Lung histology demonstrated that anti-SCF antibodies reduced overall inflammation. In addition, Masson's trichrome staining, which designates collagen deposition, was reduced.

[0142] 2. Anti-SCF Antibody Reduces Levels of SCF, Hydroxyproline, IL-25, and IL-13

[0143] Levels of hydroxyproline and particular cytokines were monitored while developing embodiments of the technology. Lung tissue sections from the above experiment were examined for the presence of hydroxyproline, a collagen precursor. The data demonstrated that the anti-SCF antibody reduced the production of hydroxyproline and plasma levels of SCF in a dose-dependent manner (FIGS. 1A and D). Also, IL-25 and IL-13 expression, measured as a function of mRNA levels, were reduced, as was expression of IL-25 receptor (FIGS. 1B, C, and E).

[0144] In particular, the experiments tested the effect of anti-SCF antibody treatment in the BLM model (FIG. 1). Mice were treated with saline (FIG. 1, "SAL") or BLM (FIG. 1, "BLM") on day 0. On days 8 and 12, different groups were also treated intratracheally with non-immune (FIG. 1, "IgG") or anti-SCF antibodies (FIG. 1, "aSCF") at the indicated doses. H&E stained lung tissue sections from each treatment group were acquired and examined. Fibrosis was quantified biochemically as lung hydroxyproline content (FIG. 1A). Lungs were then analyzed for IL-13 mRNAs by real time PCR (FIG. 1C). Plasma and lung tissue collected from SAL- or BLM-treated mice were then analyzed for soluble SCF by ELISA (FIG. 1D) or IL-25 mRNA by real time PCR (FIG. 1B). Values represent the means+/- the standard error with an n=7. A single asterisk (*) indicates statistical significance (P<0.05) when compared to the saline control group, while double asterisks (**) indicate significance with respect to the BLM+IgG control group.

[0145] 3. IL-4 Stimulates c-Kit Expression in Human Fibroblasts

[0146] Experiments conducted while developing embodiments of the technology demonstrated that IL-4 stimulated c-kit expression in human fibroblasts. In addition to the mouse model of pulmonary inflammation, SCF receptor is expressed in fibroblast populations from patients diagnosed with hypersensitivity pneumonitis and who thus have a pro-fibrotic environment. Pulmonary fibroblasts were grown from normal areas of lungs from patients (normal) and those diagnosed with hypersensitivity pneumonitis. Expression of c-kit was measured after stimulation with IL-4 at 1 or 10 ng/ml. Individual cell lines (133, 131, 173, 177A, 177B) were assessed using real-time PCR. Compared to lung fibroblasts grown from patients with non-fibrotic disease, fibroblasts from the hypersensitivity pneumonitis patients displayed significant upregulation of c-kit when stimulated with IL-4, a fibrosis-associated cytokine. The data demonstrated that SCF activated fibroblasts from inflammatory lesions, but not those from normal tissue, and promoted the expression of fibrosis-associated genes including collagen (FIG. 2).

[0147] 4. A Mouse Anti-Human Monoclonal Antibody Blocks SCF-Induced HMC Mast Cell Activation.

[0148] Experiments conducted while developing embodiments of the technology demonstrated that the monoclonal antibody specific for SCF inhibited the activation of HMC-1 cells for MCP-1 production. The activation of mast cells is a classic SCF-induced response that can be used to monitor antibody neutralization of SCF-mediated cytokine responses. Previous studies have demonstrated that monocyte chemotactic protein (MCP)-1 is strongly upregulated by SCF in mast cells. A monoclonal antibody was produced against SEQ ID NO: 1 (FIG. 3). The efficacy of this antibody was tested using a human mast cell line, HMC-1, stimulated with 100 ng/ml of SCF. The monoclonal antibody (6 .mu.g/ml) was preincubated with the recombinant SCF for 5 minutes prior to placing the SCF or the SCF plus anti-SCF onto the cultured HMC-1 cells (1.times.10.sup.6 cells/ml). The cells were subsequently incubated for 12 hours, after which the cell-free supernatant was collected and MCP-1 was analyzed by Bio-Plex. The data illustrate that the monoclonal antibody specific for SCF inhibited the activation of HMC-1 cells for MCP-1 production (FIG. 4).

[0149] 5. SCF-Deficient Mice Subjected to BLM-Induced Injury have Reduced Fibrosis.

[0150] During the development of embodiments of the technology provided herein, the effects of SCF deficiency in Kit1.sup.S1/Kitl.sup.S1-d mutant mice were examined (FIG. 5). These mice have a complete deletion of the SCF gene in one allele (S1) and a deletion of the membrane-bound ligand in the other (S1.sup.d), which significantly decreases the expression of soluble SCF. When these mice and their wild type controls (WT) were subjected to BLM-induced lung injury, there was a significantly reduced fibrosis in the mutant mice compared to wild type mice, both morphologically (Masson trichrome stain) and biochemically by hydroxyproline analysis (FIG. 5).

[0151] Wild-type and SCF deficient mice were treated with saline ("SAL") or BLM ("BLM") on day 0 and lungs were harvested 21 days later. Fibrosis was quantified biochemically as lung hydroxyproline content. Values represent the mean+/-standard deviation with an n=3. A single asterisk (*) indicates statistical significance (P<0.05) when compared to the WT saline-treated control mean, while double asterisks (**) indicate significance when compared to the WT BLM-treated group.

[0152] Similar suppression of cytokine expression and telomerase induction was also noted in S1/S1d mice. These data taken together indicated an essential role for the SCF/c-Kit signaling induced pulmonary fibrosis.

[0153] All publications and patents mentioned in the above specification are herein incorporated by reference in their entirety for all purposes. Various modifications and variations of the described compositions, methods, and uses of the technology will be apparent to those skilled in the art without departing from the scope and spirit of the technology as described. Although the technology has been described in connection with specific exemplary embodiments, it should be understood that the invention as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications of the described modes for carrying out the invention that are obvious to those skilled in pharmacology, biochemistry, medical science, or related fields are intended to be within the scope of the following claims.

Sequence CWU 1

1

8115PRTHomo sapiens 1Ser Ser Ser Asn Arg Lys Ala Lys Asn Pro Pro Gly Asp Ser Cys1 5 10 15245DNAHomo sapiens 2tcgtcgtcga accgcaaagc gaaaaacccg ccgggcgatt cgtgc 4535460DNAHomo sapiens 3gggcttcgct cgccgcctcg cgccgagact agaagcgctg cgggaagcag ggacagtgga 60gagggcgctg cgctcgggct acccaatgcg tggactatct gccgccgctg ttcgtgcaat 120atgctggagc tccagaacag ctaaacggag tcgccacacc actgtttgtg ctggatcgca 180gcgctgcctt tccttatgaa gaagacacaa acttggattc tcacttgcat ttatcttcag 240ctgctcctat ttaatcctct cgtcaaaact gaagggatct gcaggaatcg tgtgactaat 300aatgtaaaag acgtcactaa attggtggca aatcttccaa aagactacat gataaccctc 360aaatatgtcc ccgggatgga tgttttgcca agtcattgtt ggataagcga gatggtagta 420caattgtcag acagcttgac tgatcttctg gacaagtttt caaatatttc tgaaggcttg 480agtaattatt ccatcataga caaacttgtg aatatagtgg atgaccttgt ggagtgcgtg 540aaagaaaact catctaagga tctaaaaaaa tcattcaaga gcccagaacc caggctcttt 600actcctgaag aattctttag aatttttaat agatccattg atgccttcaa ggactttgta 660gtggcatctg aaactagtga ttgtgtggtt tcttcaacat taagtcctga gaaagattcc 720agagtcagtg tcacaaaacc atttatgtta ccccctgttg cagccagctc ccttaggaat 780gacagcagta gcagtaatag gaaggccaaa aatccccctg gagactccag cctacactgg 840gcagccatgg cattgccagc attgttttct cttataattg gctttgcttt tggagcctta 900tactggaaga agagacagcc aagtcttaca agggcagttg aaaatataca aattaatgaa 960gaggataatg agataagtat gttgcaagag aaagagagag agtttcaaga agtgtaattg 1020tggcttgtat caacactgtt actttcgtac attggctggt aacagttcat gtttgcttca 1080taaatgaagc agctttaaac aaattcatat tctgtctgga gtgacagacc acatctttat 1140ctgttcttgc tacccatgac tttatatgga tgattcagaa attggaacag aatgttttac 1200tgtgaaactg gcactgaatt aatcatctat aaagaagaac ttgcatggag caggactcta 1260ttttaaggac tgcgggactt gggtctcatt tagaacttgc agctgatgtt ggaagagaaa 1320gcacgtgtct cagactgcat gtaccatttg catggctcca gaaatgtcta aatgctgaaa 1380aaacacctag ctttattctt cagatacaaa ctgcagcctg tagttatcct ggtctctgca 1440agtagatttc agcttggata gtgagggtaa caatttttct caaagggatc tggaaaaaat 1500gtttaaaact cagtagtgtc agccactgta cagtgtagaa agcagtggga actgtgattg 1560gatttggcaa catgtcagct ttatagttgc cgattagtga tatgggtctg atttcgatct 1620cttcctgatg taaaccatgc tcacccatat cccactatac aaatgcaaat ggttgcctgg 1680ttccatttat gcaagggagc cagtactgaa ttatgccttg gcagagggga gactccaaaa 1740gagtcatcgc aggaagaagt taagaacact gaacatcaga acagtctgcc aagaaggaca 1800ttggcatcct gggaaagtcc gccttttccc ttgaccacta tagggtgtat aaatcgtgtt 1860tgcaaaatgt gttatgatgt gtttatattc taaaactatt acagagctat gtaaagggac 1920ttaggagaaa atgctgaatg taagatggtc ccatttcaat ttccaccatg ggagagccta 1980aaaataaatt atgacattta gtatctaagg ttagaaaacc acgcccacat gctaatatgg 2040gtgttgaaaa ctaggttact tataatgcaa ggaatcagga aactttagtt atttatagta 2100taatcaccat tatctgttta aaggatccat ttagttaaaa tcgggcactc tatattcatt 2160aaggtttatg aattaaaaag aaagctttat gtagttatgc atgtcagttt gctatttaaa 2220atgtgtgaca gtgtttgtca tattaagagt gaatttggca ggaattccca agatggacat 2280tgtgctttta aactagaact tgtaagacat tatgtgaata tcccttgcca atttttttta 2340taataagaaa acatctgact aaagtcaaag aatgatttct tatggtttat tttgatgaaa 2400gttcttttaa catgtcttga atgtacacat aaaggaatcc aaagctttcc attctaactt 2460aatctttgtg ataacattat tgccatgttc tacaaccgta agatgacagt tttcaatgta 2520gtgacacaaa agggcatgaa aaactaactg ctagctttcc tttcatttca aaagtccaag 2580aatttctagt atatttggat tttagcttct gttcaaagca aatccagatg caactccagt 2640aagtggcctt tgctcttttt tgtaccaaag agcccagatg attcctacag tccctttctt 2700ctctaacatg ctgtggttcc ttaaatatga gtaatttctc taagatataa cccaggtgct 2760ttgagaagct gcattaaggt gttcaggccc tcagatatca catggtacac ttgattagta 2820ataaaaccag agatcaattt aaattgctga taggtcctgt ctcagtgtgt ggcattgact 2880gttttcagga aaatagatac agattaatat gagttatgcg tgtaggttgt gtatagattg 2940agaagataga tacttctcaa tctagtagtt tgatttattt aaccaatggt ttcagtttgc 3000ttgagcatat gaaaatcctg cttaatgtgc ttaagagtat aataaatgtg tacttttgtc 3060ctcaaaccta gtagctgggt tttaacactc atggacatgg tcttaatcaa tggagttaaa 3120taaacaaatt cagcaagtta ttaaatctga catggtagga gaggggagat gtgtcctgct 3180tattaaatgt gttggtccat tgaaagttac atggattgcc aatttttaaa acactaaagt 3240tgaataaaat gcatgaacaa tagaaaaatg ctgaacatta ttttggatgc tagctgcttg 3300gacattaact gtgttatttc tgctttgaga tgaaaatata tatttatctt tgcttatttt 3360atcccagatg tgttctgaat atccttcttc ataaatcatg gaaaactcac tgctgagata 3420gtaaaccatg aaatcgcctt ttcagttggt gccatgtatc tgacagttcc atcttggaag 3480gtttcaaaat taccttttaa aatgatctca gaagtctgta gattctcaat gatactgaaa 3540gctttgcacc tctttggtag aaaccaggtc tatttagaaa atggctttat gataaatgtt 3600gcctcctgag tgataatgaa gtgttcctgg atattgtatt gtaatttaat gtgcttacca 3660cactgccaca ttttaatgag tcagagaaaa attaattttt cttcaataca ataatagaac 3720aagtagccta ttctcttaaa aagtatgtga aaagaaaatt atgaaaaaat atgcatacct 3780aatgaagtat tggttttagt aagaattaaa tacatttcat tgagctttaa agtactttgg 3840agaaactttg gggcacgttt tcctactcta attcaactaa agttataaat aaagagaaaa 3900actcattcag aaatcatgga ttttaaaaat attttactgc agccaagttt tcatttcaaa 3960atgtaatttc agtttggagc ttttaggcat tatgtatatt taaaaaatat attcttcaaa 4020aatgcatttt ggcatggtgg gatggatgtt gcaaaagata tccggagcct ccagtctgtc 4080attaactgat atggtaaatc acctctcttc tttgggtctc aattttttat ttatctatat 4140ggtaaactca gagatcactc cttaggggtg agtcctattg caatatgacc gacaaagaag 4200acaaaatagc attgaaacta acccatacaa aatatccaac tctggattct gtgaataagt 4260atcttgacca taaaaagtca ttgctgttct tgtttctaat gtaaatagtg tccattagta 4320aaagtgaaat tcagtcttaa gtagggtgaa ttggatcacc atttacacaa gagatggctt 4380tttcctttgc ttgaataaac attttggatc acctccaaag aatgaaaacc agtagtacgt 4440tttagtcata ttagtcagga tgagaaacta taagatgtgt gtaacatttg gaaatgcacc 4500aaagtgagcg tttaaatctt ctcattttat tgaaaactaa gagcagaaaa tgtaaaatgc 4560tcatgaaggt tttgaatgcc aaaagatatt ttagaatcaa tttataaagg ggtaattcat 4620taattacact ttaaaattgg aaagtgggat aagaaatcta aagtaaacca gcttatcttt 4680gaaacaatat tattttgaaa ttggctttaa aataaaacca ttcagattga aattctaatt 4740agctcatttg tggagtttga tcacacaatt cataatgttg ctgctttcca ttaactagtc 4800ttgaaatgcc tttgtttgta aaaataaaat aatggtactt tcattttata acaaggtgtt 4860tttttcaaga aataatccat gctaaaatgg atatttgtga tcctgaaatg tttactaagc 4920attgtaaatt tatttataac tgccatctcc aactacatcc ttatgatgtt tttaacaata 4980aaattaaaac aactgttaaa ctaaaaacca caccgttttc cagtacttga tctctgagct 5040acaatactca ctaaatataa ttttccaatc aaaatattct attctatatt ctaagggtta 5100atatgtgatt atagtgtcca cttgccacca tttttttaaa tcaatggact tgaaaagtat 5160taatttagat ggatgcgcag atataccctc agttcagtca tagattggag tttgcatata 5220ataatgtaaa tgtatgtcga cactattcta aatagttcta ttatgactga aatttaatta 5280aataaaaaag gttgtaaaat gtgatgtgta tgtgtatata ctgtatgtgt actttttaaa 5340ataggtgtat gtcccaaccc ttttttatac aggtttgaat ttaaaattac atgatatata 5400catatacttt attgttctaa ataaagaatt ttatgcactc tcaaaaaaaa aaaaaaaaaa 54604273PRTHomo sapiens 4Met Lys Lys Thr Gln Thr Trp Ile Leu Thr Cys Ile Tyr Leu Gln Leu1 5 10 15Leu Leu Phe Asn Pro Leu Val Lys Thr Glu Gly Ile Cys Arg Asn Arg 20 25 30Val Thr Asn Asn Val Lys Asp Val Thr Lys Leu Val Ala Asn Leu Pro 35 40 45Lys Asp Tyr Met Ile Thr Leu Lys Tyr Val Pro Gly Met Asp Val Leu 50 55 60Pro Ser His Cys Trp Ile Ser Glu Met Val Val Gln Leu Ser Asp Ser65 70 75 80Leu Thr Asp Leu Leu Asp Lys Phe Ser Asn Ile Ser Glu Gly Leu Ser 85 90 95Asn Tyr Ser Ile Ile Asp Lys Leu Val Asn Ile Val Asp Asp Leu Val 100 105 110Glu Cys Val Lys Glu Asn Ser Ser Lys Asp Leu Lys Lys Ser Phe Lys 115 120 125Ser Pro Glu Pro Arg Leu Phe Thr Pro Glu Glu Phe Phe Arg Ile Phe 130 135 140Asn Arg Ser Ile Asp Ala Phe Lys Asp Phe Val Val Ala Ser Glu Thr145 150 155 160Ser Asp Cys Val Val Ser Ser Thr Leu Ser Pro Glu Lys Asp Ser Arg 165 170 175Val Ser Val Thr Lys Pro Phe Met Leu Pro Pro Val Ala Ala Ser Ser 180 185 190Leu Arg Asn Asp Ser Ser Ser Ser Asn Arg Lys Ala Lys Asn Pro Pro 195 200 205Gly Asp Ser Ser Leu His Trp Ala Ala Met Ala Leu Pro Ala Leu Phe 210 215 220Ser Leu Ile Ile Gly Phe Ala Phe Gly Ala Leu Tyr Trp Lys Lys Arg225 230 235 240Gln Pro Ser Leu Thr Arg Ala Val Glu Asn Ile Gln Ile Asn Glu Glu 245 250 255Asp Asn Glu Ile Ser Met Leu Gln Glu Lys Glu Arg Glu Phe Gln Glu 260 265 270Val55376DNAHomo sapiens 5gggcttcgct cgccgcctcg cgccgagact agaagcgctg cgggaagcag ggacagtgga 60gagggcgctg cgctcgggct acccaatgcg tggactatct gccgccgctg ttcgtgcaat 120atgctggagc tccagaacag ctaaacggag tcgccacacc actgtttgtg ctggatcgca 180gcgctgcctt tccttatgaa gaagacacaa acttggattc tcacttgcat ttatcttcag 240ctgctcctat ttaatcctct cgtcaaaact gaagggatct gcaggaatcg tgtgactaat 300aatgtaaaag acgtcactaa attggtggca aatcttccaa aagactacat gataaccctc 360aaatatgtcc ccgggatgga tgttttgcca agtcattgtt ggataagcga gatggtagta 420caattgtcag acagcttgac tgatcttctg gacaagtttt caaatatttc tgaaggcttg 480agtaattatt ccatcataga caaacttgtg aatatagtgg atgaccttgt ggagtgcgtg 540aaagaaaact catctaagga tctaaaaaaa tcattcaaga gcccagaacc caggctcttt 600actcctgaag aattctttag aatttttaat agatccattg atgccttcaa ggactttgta 660gtggcatctg aaactagtga ttgtgtggtt tcttcaacat taagtcctga gaaagggaag 720gccaaaaatc cccctggaga ctccagccta cactgggcag ccatggcatt gccagcattg 780ttttctctta taattggctt tgcttttgga gccttatact ggaagaagag acagccaagt 840cttacaaggg cagttgaaaa tatacaaatt aatgaagagg ataatgagat aagtatgttg 900caagagaaag agagagagtt tcaagaagtg taattgtggc ttgtatcaac actgttactt 960tcgtacattg gctggtaaca gttcatgttt gcttcataaa tgaagcagct ttaaacaaat 1020tcatattctg tctggagtga cagaccacat ctttatctgt tcttgctacc catgacttta 1080tatggatgat tcagaaattg gaacagaatg ttttactgtg aaactggcac tgaattaatc 1140atctataaag aagaacttgc atggagcagg actctatttt aaggactgcg ggacttgggt 1200ctcatttaga acttgcagct gatgttggaa gagaaagcac gtgtctcaga ctgcatgtac 1260catttgcatg gctccagaaa tgtctaaatg ctgaaaaaac acctagcttt attcttcaga 1320tacaaactgc agcctgtagt tatcctggtc tctgcaagta gatttcagct tggatagtga 1380gggtaacaat ttttctcaaa gggatctgga aaaaatgttt aaaactcagt agtgtcagcc 1440actgtacagt gtagaaagca gtgggaactg tgattggatt tggcaacatg tcagctttat 1500agttgccgat tagtgatatg ggtctgattt cgatctcttc ctgatgtaaa ccatgctcac 1560ccatatccca ctatacaaat gcaaatggtt gcctggttcc atttatgcaa gggagccagt 1620actgaattat gccttggcag aggggagact ccaaaagagt catcgcagga agaagttaag 1680aacactgaac atcagaacag tctgccaaga aggacattgg catcctggga aagtccgcct 1740tttcccttga ccactatagg gtgtataaat cgtgtttgca aaatgtgtta tgatgtgttt 1800atattctaaa actattacag agctatgtaa agggacttag gagaaaatgc tgaatgtaag 1860atggtcccat ttcaatttcc accatgggag agcctaaaaa taaattatga catttagtat 1920ctaaggttag aaaaccacgc ccacatgcta atatgggtgt tgaaaactag gttacttata 1980atgcaaggaa tcaggaaact ttagttattt atagtataat caccattatc tgtttaaagg 2040atccatttag ttaaaatcgg gcactctata ttcattaagg tttatgaatt aaaaagaaag 2100ctttatgtag ttatgcatgt cagtttgcta tttaaaatgt gtgacagtgt ttgtcatatt 2160aagagtgaat ttggcaggaa ttcccaagat ggacattgtg cttttaaact agaacttgta 2220agacattatg tgaatatccc ttgccaattt tttttataat aagaaaacat ctgactaaag 2280tcaaagaatg atttcttatg gtttattttg atgaaagttc ttttaacatg tcttgaatgt 2340acacataaag gaatccaaag ctttccattc taacttaatc tttgtgataa cattattgcc 2400atgttctaca accgtaagat gacagttttc aatgtagtga cacaaaaggg catgaaaaac 2460taactgctag ctttcctttc atttcaaaag tccaagaatt tctagtatat ttggatttta 2520gcttctgttc aaagcaaatc cagatgcaac tccagtaagt ggcctttgct cttttttgta 2580ccaaagagcc cagatgattc ctacagtccc tttcttctct aacatgctgt ggttccttaa 2640atatgagtaa tttctctaag atataaccca ggtgctttga gaagctgcat taaggtgttc 2700aggccctcag atatcacatg gtacacttga ttagtaataa aaccagagat caatttaaat 2760tgctgatagg tcctgtctca gtgtgtggca ttgactgttt tcaggaaaat agatacagat 2820taatatgagt tatgcgtgta ggttgtgtat agattgagaa gatagatact tctcaatcta 2880gtagtttgat ttatttaacc aatggtttca gtttgcttga gcatatgaaa atcctgctta 2940atgtgcttaa gagtataata aatgtgtact tttgtcctca aacctagtag ctgggtttta 3000acactcatgg acatggtctt aatcaatgga gttaaataaa caaattcagc aagttattaa 3060atctgacatg gtaggagagg ggagatgtgt cctgcttatt aaatgtgttg gtccattgaa 3120agttacatgg attgccaatt tttaaaacac taaagttgaa taaaatgcat gaacaataga 3180aaaatgctga acattatttt ggatgctagc tgcttggaca ttaactgtgt tatttctgct 3240ttgagatgaa aatatatatt tatctttgct tattttatcc cagatgtgtt ctgaatatcc 3300ttcttcataa atcatggaaa actcactgct gagatagtaa accatgaaat cgccttttca 3360gttggtgcca tgtatctgac agttccatct tggaaggttt caaaattacc ttttaaaatg 3420atctcagaag tctgtagatt ctcaatgata ctgaaagctt tgcacctctt tggtagaaac 3480caggtctatt tagaaaatgg ctttatgata aatgttgcct cctgagtgat aatgaagtgt 3540tcctggatat tgtattgtaa tttaatgtgc ttaccacact gccacatttt aatgagtcag 3600agaaaaatta atttttcttc aatacaataa tagaacaagt agcctattct cttaaaaagt 3660atgtgaaaag aaaattatga aaaaatatgc atacctaatg aagtattggt tttagtaaga 3720attaaataca tttcattgag ctttaaagta ctttggagaa actttggggc acgttttcct 3780actctaattc aactaaagtt ataaataaag agaaaaactc attcagaaat catggatttt 3840aaaaatattt tactgcagcc aagttttcat ttcaaaatgt aatttcagtt tggagctttt 3900aggcattatg tatatttaaa aaatatattc ttcaaaaatg cattttggca tggtgggatg 3960gatgttgcaa aagatatccg gagcctccag tctgtcatta actgatatgg taaatcacct 4020ctcttctttg ggtctcaatt ttttatttat ctatatggta aactcagaga tcactcctta 4080ggggtgagtc ctattgcaat atgaccgaca aagaagacaa aatagcattg aaactaaccc 4140atacaaaata tccaactctg gattctgtga ataagtatct tgaccataaa aagtcattgc 4200tgttcttgtt tctaatgtaa atagtgtcca ttagtaaaag tgaaattcag tcttaagtag 4260ggtgaattgg atcaccattt acacaagaga tggctttttc ctttgcttga ataaacattt 4320tggatcacct ccaaagaatg aaaaccagta gtacgtttta gtcatattag tcaggatgag 4380aaactataag atgtgtgtaa catttggaaa tgcaccaaag tgagcgttta aatcttctca 4440ttttattgaa aactaagagc agaaaatgta aaatgctcat gaaggttttg aatgccaaaa 4500gatattttag aatcaattta taaaggggta attcattaat tacactttaa aattggaaag 4560tgggataaga aatctaaagt aaaccagctt atctttgaaa caatattatt ttgaaattgg 4620ctttaaaata aaaccattca gattgaaatt ctaattagct catttgtgga gtttgatcac 4680acaattcata atgttgctgc tttccattaa ctagtcttga aatgcctttg tttgtaaaaa 4740taaaataatg gtactttcat tttataacaa ggtgtttttt tcaagaaata atccatgcta 4800aaatggatat ttgtgatcct gaaatgttta ctaagcattg taaatttatt tataactgcc 4860atctccaact acatccttat gatgttttta acaataaaat taaaacaact gttaaactaa 4920aaaccacacc gttttccagt acttgatctc tgagctacaa tactcactaa atataatttt 4980ccaatcaaaa tattctattc tatattctaa gggttaatat gtgattatag tgtccacttg 5040ccaccatttt tttaaatcaa tggacttgaa aagtattaat ttagatggat gcgcagatat 5100accctcagtt cagtcataga ttggagtttg catataataa tgtaaatgta tgtcgacact 5160attctaaata gttctattat gactgaaatt taattaaata aaaaaggttg taaaatgtga 5220tgtgtatgtg tatatactgt atgtgtactt tttaaaatag gtgtatgtcc caaccctttt 5280ttatacaggt ttgaatttaa aattacatga tatatacata tactttattg ttctaaataa 5340agaattttat gcactctcaa aaaaaaaaaa aaaaaa 53766245PRTHomo sapiens 6Met Lys Lys Thr Gln Thr Trp Ile Leu Thr Cys Ile Tyr Leu Gln Leu1 5 10 15Leu Leu Phe Asn Pro Leu Val Lys Thr Glu Gly Ile Cys Arg Asn Arg 20 25 30Val Thr Asn Asn Val Lys Asp Val Thr Lys Leu Val Ala Asn Leu Pro 35 40 45Lys Asp Tyr Met Ile Thr Leu Lys Tyr Val Pro Gly Met Asp Val Leu 50 55 60Pro Ser His Cys Trp Ile Ser Glu Met Val Val Gln Leu Ser Asp Ser65 70 75 80Leu Thr Asp Leu Leu Asp Lys Phe Ser Asn Ile Ser Glu Gly Leu Ser 85 90 95Asn Tyr Ser Ile Ile Asp Lys Leu Val Asn Ile Val Asp Asp Leu Val 100 105 110Glu Cys Val Lys Glu Asn Ser Ser Lys Asp Leu Lys Lys Ser Phe Lys 115 120 125Ser Pro Glu Pro Arg Leu Phe Thr Pro Glu Glu Phe Phe Arg Ile Phe 130 135 140Asn Arg Ser Ile Asp Ala Phe Lys Asp Phe Val Val Ala Ser Glu Thr145 150 155 160Ser Asp Cys Val Val Ser Ser Thr Leu Ser Pro Glu Lys Gly Lys Ala 165 170 175Lys Asn Pro Pro Gly Asp Ser Ser Leu His Trp Ala Ala Met Ala Leu 180 185 190Pro Ala Leu Phe Ser Leu Ile Ile Gly Phe Ala Phe Gly Ala Leu Tyr 195 200 205Trp Lys Lys Arg Gln Pro Ser Leu Thr Arg Ala Val Glu Asn Ile Gln 210 215 220Ile Asn Glu Glu Asp Asn Glu Ile Ser Met Leu Gln Glu Lys Glu Arg225 230 235 240Glu Phe Gln Glu Val 245794673DNAHomo sapiens 7agaatactct gtgaacttat tatgtatttc atcattttca aggctggtaa aatcacacgc 60cttcacatag gcaaagagaa cttcacattc ttgtagtttg gaaactccat aggaaggaga 120tactgttccc cattagctca agcacaaaag tcccacagtg gattctgatt ggcccatctc 180atgtgatgta cccattcctc caccaatcag gactggacag atcagggtac taagcctatc 240tccgatagtg gaaggaggtg ggaaaacaga accatgtgaa tgacagtctc tcttgaaaga 300taaggaatag tctgaattct ctaagtgaaa gtgagtgcta ataatggaag aagagtgaca 360ggaaagtgtg ctaagcaaat aaaaaaaaaa aaaatctaag ccgggcatgg tggctcacgc 420atgtaattcc agcactttgg gaggccaagg cgggcagatc acctgaggtc agcagttcga 480gaccagcctg agcaacatgg agaaaccccg tctctactga aaatacaaaa cttagtcagg 540catggtggtg catgtctgta atcccagcta ctcaggaggc tgaggcagga gaatcgcttg 600aacctgggag gcagaggttg cgtgagccaa gatcatgcca ttgcactcca gattgggcaa 660caagagtgaa gctctgtccc cccccgacaa aaaaaaattc ctaaccacag tccatatgtc 720taggttagat catggttttc ccatttatca attgagggat

cttgagcaaa tcaccatact 780tctttgatcc tcagtttcct tatttatcaa gtagaaaagt taaaaattac ctctcttaca 840tgattgtcat aagaagttat tgacagaata taaactaagg gcttagacca attcctggct 900tatatgggca acagcaacct aattgaccaa tatttaaggg attcgctctg ttctttcccc 960tgtgacagat attgacaata aaataatgag caagggcagt cataattact ttaaccaaat 1020agtcaaacaa acaaatacaa aactgcagtt gttttataaa aaataggtaa ttcatgctcg 1080actttatagt aagaggagaa ttgaacaaag caaagagatc agggaaaacc tccctgagga 1140agaaaatata gggaatactg agttgagatc tacatgataa atgcatactt actgggcaca 1200atgatagggt gaaacaaagt ggtccaaaca gaggcagaaa ggagggtatt tgcaaatggc 1260ctgaggtgag aattatgagg acagaagggc cttggccctg gaaaaaaggg agcacataat 1320gaattgacct tggagaagga gggagaggcg agatcctgga ggccttgaag gctttgctaa 1380ggtgctaatt tctttattcc aagagcaatg gcaagccacc aaagagcttt cagcagagga 1440aatttcatga tcaaagaggc attcttctcc ttttccccct cctctagtca ctcaaagaca 1500gatatagaca gtctgtctcc ggtctggctt ggtcaggcaa taagtttcag tttgtgagag 1560acatttttta aaagaaaaat attgaaaaca atatgtgtgt ctaaaagaag atatccacaa 1620aatagtgaat gaacaagaga tatttcacat gcacaaagca agataagact tatttggtaa 1680attacacctg cctcttaaca acaaaaacga taaagctcag gaaaaaagct tttcaaaatt 1740ggattaatca ataatagcat gagtgacgaa tagtattgaa agtattcaag taaaaattag 1800ataatcatga aataactttc tacccagtgt gttaaatgtt gaattccctg agttgtataa 1860tggtagcctg ctatttccat aagaaaacat cccccatccc tcctgtccac ctgggacata 1920gccctgcctc ttgttcagtt taaaggaact ccttggtttt tcttctgcga atcaactgaa 1980tgctggcctg catcccctat caaaagccag tcttaaggag agatctcagt cattctgacc 2040catttgtgac ttccatataa gcctacccaa aatctttcat ttgcatatta gctctttcta 2100atcccttgga tatttgcttc gacattctct tgattcaagt aacattatta cacaacaaaa 2160ctgtaatgca gcagtatgag ggaaacaaaa gatacagtta cgtatgagaa agtcagtgat 2220attaagaaag ctagaaccag ggaatggtgg aggtgggagg aggagacaga actgagaggg 2280aaaaaaaatg gaaagaggga gaaagagaga ggagaatgag aatgcattgt aggagatgaa 2340cgtagcatgt attattgagt tgaatgaaaa gaaattgaac tgaatttctg ttaggaacac 2400tgccttgttt ctttaaacct ggtacaacaa attaggccat tcctttctat tggtctcata 2460aaatccgtat ggttagattt atgattccca ggcttcagta actctcatac cattttgatt 2520atttttgtcg tatctgtcct cctttagtac tacttgttat ttacttaatg ttatacatga 2580aagtgagttt atattgacac taaatatttc ttttcttggg gggaaaatgg tggtaaaggg 2640gtacaaagtt ttagttagac aggatgaata agttctggag atttattgta cagcatgata 2700actacagtta atgtactgta tacagttaac ccttgaacaa cacaggtttg aactgtatgg 2760gtctacttat atgcaatttt tttttcaaac tcagatggaa aatacagtgt tctagagatg 2820taaaacccat gtataagcag ggctgacttt tgtgtctgca cgttctgcag ggttgactgt 2880gggacttgaa tatatgtaga ttttggtata cctgaaaggt ctaggaatca atcccctgaa 2940tatactgagg tactccaaaa ctacaaagag tagatgttaa atactctcag ctcccaaaaa 3000tgatgagtat gtgaggtgat ggatatacta tttagcctga tttaataatt tcacaatgta 3060tacataagca gaacatcaca ttgtacgtgg taaatagata ctatttttgt caaatataac 3120ttaataaagc tggagaaaaa aaataaaatt ctccacttca aaaaagcctt aaaggggccg 3180ggtgcagtgg ctcacgcctg taatcccagc actttggaag gctgaggcag gcggatcacc 3240tgaggtcagg agttcgagac cagcctggcc aaaatggtga aaccccgtct ctactaaaac 3300aacaacaaca acaacaaaaa tacctgggcg tggtggtggg cacctgtaat cccagctact 3360tgtgagactg aggcgggata atcgcttaaa cctgggaggc ggaggctgca ttgagccgag 3420attgcgtcat tgcacttcag cctgggagac agagcgagat tccatctaaa aaataaaaaa 3480taaaaataaa tcttgaagaa aaaaaaaaca atcatttgct atgcctccag tatccaaatt 3540aagctcattt tgatatttga tgttcgatat atttgtacta agagtaccta atgtataaaa 3600atctttctaa gcaactatca acttggaatc atgattgatg tgctgtctat acatattttt 3660tctaacatga gaaagtgaga acgcagtccc ccactaaatt atgcagtcga gtttcccaca 3720tttggggaaa tcacagaagt cagcacatct ggagtgaaat aaacaagctt cgccctgatt 3780acaggtttgg catattcatg tttaacaaac gcaccggaac taattaaagc aaatttggac 3840tcctactcta ttattaaaat aactaattcc taaactcata cattattctt tcttttaaag 3900atcactgatg ttaatgttca gctaatattt tcttttaaaa aattaatgaa atgctttgga 3960atgttctcca gttatacaaa caaaaacaga cacattctgg agaatcaata aaatatttga 4020agtatttttt aagactgtac agctggagct ccaggtttaa attagagctc tgccatttac 4080tagatatggt tggtcaagaa aattaacctt gtatttcatc gtccataaaa tgggaataaa 4140ataataagat ctacactaat agatttgatg aaagaattta atgagataat ctatgcgaag 4200ttcttagtct ggcacctggc acatgaataa gtgcatcaca taatgatgtt agctattatt 4260attgtggttg cattatcttt ccgttcaaaa aagtacaact ttcaaagcct tgctgtgttg 4320gcgacctgta gtttaataat ctatcgcttc tctttcaatc ttaatttcta aagacacagt 4380aacaaaccat tccaaatgaa gcgtctcccc ttgcaatact taaccttgag tcaacaattc 4440ttgtaattat taaatattca ttcactaaag taattgtttt ttttaaaaaa tagaggtgat 4500aggacaatgg cctaactaca gatcgcaatt aaactttatt tctctaaggc aggcagggtg 4560aattctttgc tccgaacctc ccggactcct ctccctctcc gactttgtat aacagaggga 4620gctccgagcc ctctctggcg cgcgaggtat ttcgtctgtc cccgggggtg ccaggtgagc 4680cccagcggat ccgggagggt aagctgggac tcctcgcgag cagtagctgc agggtaccaa 4740gcttcgccct ctgcgtcccc gcgccttcgc ggtctcccgc cagtgcaggt ccggggcccc 4800caggcgagcg gacaaggttg gcctaatctg ccaaacttct ggggcattta ccgtgctctg 4860gccgccctcc cgattcttcc ctccgcgccc ttgcctgctt ctcgcctacc ccgggctccg 4920gaagggaagg aggcgtgtcc ggagcaggcg ggcgggaact gtataaaagc gccggcggct 4980cagcagccgg gcttcgctcg ccgcctcgcg ccgagactag aagcgctgcg ggaagcaggg 5040acagtggaga gggcgctgcg ctcgggctac ccaatgcgtg gactatctgc cgccgctgtt 5100cgtgcaatat gctggagctc cagaacagct aaacggagtc gccacaccac tgtttgtgct 5160ggatcgcagc gctgcctttc cttatgaaga agacacaagt gagtagggcg cgcccgggag 5220ctcccaggct ctccaggaaa aatcgcgccc ggtgccccgg ggaagccggc gctccctggg 5280acttgcagct ggggcgtgca gggctgtgcc tgccgggtga gacaagagga tgcgggggag 5340gccggcgtgg tgtgtgatcc cgagccgagc cgggtgagcc agggagaaaa ggagtgggag 5400tgctgagagg gagccagtgt caagtttgga gcctcagcag ttaagttttg agctgtcagt 5460cggaaaccgt aattcccgtc tggtggaaag attggctttt gggccatgga atgttaagtt 5520atcacgggaa ggcatgtctt tgcataggag acggttttcc cgtggattcc aggagattat 5580gaaacagcac cccacgcgcc accctcacta gctctctcac tcagatttat ttgtgtagcc 5640ccgaggtggg cgccgggcac caggagcggc gcgaacgcgg gtgcagccgc gggagcgaag 5700ccaacgcgga actcctaagc ggcagctctc ggcggggcgg gaacctgccc gcctgcccgg 5760gaaacttggc tctcggcggt gtctgcctag tttgcagctc tgtgtgggct ctaaagagct 5820ctagtacctc cacaatgcac gcaccttttt ccatttcaaa agggataaag gctttttata 5880aaataggcac ctctgaatga tagtgattga gataattccc agtgctgcta aattctgcat 5940tatacataaa gcactgtggg taaagcgctg cctgggggga caattagatt tttttttccc 6000ttgaagaaaa tacctggttc ttcgtttaga aagggccgcc acggtgcccc tatccacagt 6060tctcttcacc attgttgatt gttacctcct ctcactttcc cccttagaaa ttgttatgtg 6120ttatctgccg cggttggcta agggttgccc tttcaccccc cagaggttct gttggagatt 6180gcagttcctg aaatgtgtgg aaggggtgga agtggtgggg aaaaaggaaa ccggggtgtt 6240tgtacacatc gttggtgtag aatgaaggca tcaatgtgct caaaatgctt tatcaaatga 6300cctgcttagc atcatatttt aaacagatat ctaggcctgc aaaaagttgc aagactctag 6360ccccttgtac cggtgtggta aaaggcaggg gaaagggaca ttgcctggct aaaagaggct 6420tgcatagtct ctagagcact ctctaagaga aaagacgcga tgtgcctctc ccctctctta 6480caagcaggtt agctttcagg caaggcattg agcaccgcac ctggaatcac cacagcgtgc 6540tgtcgaaatg caaatgttaa agattcttgc aacaaaagaa aagttttatt gtaatctctt 6600gatttagaca agctttctgc gaaattataa gaagggtaga gacctattcc ttcttatagt 6660agcatctgct cccatggatg gtggtgagat ctgggggaag aagctgctac cttctagttc 6720aggaaaagag cctaaacctt cctatttaaa caagaacaca ggagacagat aaggttaatg 6780tccaacactc caaagccagg ggatgtacta tcaagtcact ttgagaggca gtaggattga 6840ttatccttgc acccttgctt tgcaaaacaa atgtgagctc tgtaatctcc tttctcgctt 6900tgattaaatg gctgtgtaaa tcggattaca tggaataatt ccattgaaaa ctagaaagag 6960gtcaatctag tctgaaacat tgtgactgtg gcattagtga ctcagtgctg ccgaccagag 7020aaatttgaga aagctaagta ttttagatat gtgacctgtt cggtcacaca tcacacaagt 7080taaagtgcag gccctttggg ggaacttgct gatgttaaac ggcttgtagg caaaggaggg 7140cagtaccctt gttgaatgaa ggcattcatt gcctaggatc taatgacctt caaggttttt 7200tttaagaata cttctagcga ttccacagtc actgaaaaag gggacactgg aaatagccca 7260aaatcaataa aataatctgc agaaatggag gaggggatta gatctttcat attgttaaat 7320tttctaatcc aaaaattaca aagacatttt ctgtgcaatg ctgcatggat ggaaggtagc 7380ctatgtaaat tggacattaa gaaataggat cgattgttct gattctaatg ttatcctcac 7440cagggaagaa tatctattaa tatttagtag tttaaatagt catttataaa catggcatca 7500ataccatggt tctattataa agcaacttgt tctgtactct cctactaccc cacatataaa 7560aaaaatttat agtaatattt taaaatgggg tttgatttga attacacagc tctccaagaa 7620tcatggggac tccaggtatt ctggcctttc cgtcaagcta agtaagtgtg aaaattccat 7680caacaaccac ttggggtttg atgaatgatt cattcattta gcatgttggt acacttcagt 7740atttccaata taatgacaca ggcaataaat ttaagcacag ttctacagaa atgaatgtct 7800tttgaatttt tcagatttct gtttgatttt attttattca gattgtgtgg ttttacagtt 7860ttctagaggt gtttgtggct tgggcttaga aaaaacatct ctttcaatgt gtaactattt 7920tttaaacata atattctgat acaccatatt ggataacaat caaggcacca tattttgcaa 7980cactgattaa gatttggtta aaagtgccat tagcttaatg tgcctggtgt tacatttatc 8040aatgttgtgc ttacatttta gtttggggat ggtttgtgag aagtgtcata actcctgtca 8100atttcagaaa tactgagata agaaagattg atcttgcaaa caatgtaaaa aaggccattt 8160taggcatact aactctcaga gaataagcaa atagaatgct aattttcact actccaaaga 8220aaatatctct tttatgtctg tatgaagttt gtcatgtggg tgatgatgca agtttacttt 8280gtagaagaaa attaacatac ttatgaaaag agcatttttt tagtcttgtc cttaagatca 8340ctcaatttgg aatgtatgct ttgaataatt ccaatttatg tagcttttaa ttgatacatt 8400ctgccaagac tttaaaaccc aagtgctaag ttgattacat ttaacataaa acaaggaaaa 8460taaccatatg gaaatctgaa agtctgggcc aaaccaatta aaattgctaa attttattta 8520cgtctaactg ctgatttgaa gatagcaaat tctgggtcac attttttttt taataatagg 8580tgactgaaat gtttccatca ccataaatca cataaaaatt tctcactcaa atagcaagtg 8640tgggtattgg aggtttttta catggattgg caaattccac tacatgtggc cttcgcttta 8700tatctgagaa taaaccatat ctgagcatat atgagaaaca cctaaaatat ggttggtaga 8760aggattctag gttctagctt ctgaagactt agattctaat tcctgtttta ctactcatga 8820gacaaatgac cttaggcaaa tatttgttaa actttttggg atttctgttt tttgtaatct 8880gtatgatgat tataaaagtt ataaagattt ttctcgcatg gtggttttaa ggactaaatg 8940aaattttata tgtgtcttta taccagcata gtagcttact atatgtacag tttgaattta 9000ttgttcctga gaagagagtg cctgctggcc tacttttaca agccagagaa aaactgactt 9060attgcaggta aattatgcct aatgaaccaa gcacagagac caagttttcc taaatctacg 9120tctgtcgatt tggtgactcc catgataggc taatatctgt ttatagacta taattaagac 9180acatcaatcc aggaatatgt gaagacaaag gtgatatgtg ggcttaagag gtgctacgtt 9240tttgaaggtt tttcttttaa agtacaaatg aatgagataa ttagaataga aaatagaaat 9300ggaaaatgga aatattatct ttatcgctga ctcagtttca tatttctaca ctcttgctaa 9360gttctccttg tatgattcta aggaattatt tcattttatt ttattttatt ttggactaaa 9420gagacttggt tagtttccca tccctaatgt tctcgcactt gcttttctga gtgaggccat 9480gcattcattt ctgtggggtg tctttgttgt ctttaccagg caacgtgcag tgacattttt 9540ttttcattat gctagtttga gacttttatt tttcttttct ggattatctt aatgttccct 9600tttgtaaatt atttttattc ccttttgtaa attttattcc cttttgtaaa ttagggcaag 9660gcatttggat ataaatctct gaagagtgat cttttatctt tctttctagc acataaatca 9720tgagcagctg caagaaactg ttcttggtct tacacactgt tcttggtctt agtcacgtgt 9780gtagacagaa caatttacaa aacaagttga gggaacacca agatattttt ataatatatg 9840ctgctttcag ggttgttttg tataggctct gtttcataga aacaattgac tgcagccttt 9900ctgatttata gatgcagcaa gatgtgtttg ccttttcact taattatggg tatactaaaa 9960actgtttaca ggggctggga gtttaagaga attgtcatga tctcatctct acctatttct 10020ttctctatag aaaaaagagg tgggggagtg attttctcat agttggggct catggttctt 10080tctttaaaat cataatgtgt gtcatctaaa ggaagactgt tctttatgta cctgtctttg 10140tcctttggct acattttact atttttgtag gctgccactt tggcttggat attaggaggc 10200tgatttcttt tattaacctg gataataagt agtagtaatt tgggaaatta tttgaactcc 10260agatcttagt ttgctcaact gtaaaatgag taagtgagac taggtgattt ctaagtgcct 10320tctggttcta aaattctcta tttctaaagt actagtaaaa tagatatttg aaccagatgg 10380tatttaagct tccttcagtt ctcagattta tgcagccatt gcttcagtgt gaatgcacat 10440tataaccatt gttttatgat tatgaataag ccaatgggga gaaatgaagg agatttaaat 10500ttagactgtt ctacttttat gcatttcacc tactgcttga tatttctttg taaaaagaat 10560ggtatttaca ctttcctgtt ttagcaaaat ccatagttaa agaggatctt tactttctct 10620gttctaaatt tttaagttct atgaaaaaga attctaagat aaagtcagct aaccagagga 10680gataagctta caaaaggtaa tgcttccctg tatttgcctg ttttttttct gtccctttaa 10740cttccagtac gtgtgtgcat ggagttgaaa tggagaaaga aaaccaagaa gggatctgat 10800gatgtagtag aaagagcact gaggggagaa tcagaggaac tgatcctacc ctttgctctg 10860cctcaagatg tgtgaccttg gactagtctc ccagtcttag tatctctgaa tcatgcccaa 10920gatagttaat cctgagaaga tagtgaagat gagaggctgc agctggagac acggaggtgg 10980aaagaggagg gagcctcagg gaaaggcagt tcctgcagat gatactatgc caaatctctc 11040cttggctggc ttttgtctgg gtccagtacc atccttggaa cagtgcttgc ttacctcctt 11100tccttagctg agagctggag caaacccagc agctagccgt agcccttttc tttttttctt 11160tttttttttt tcttttagca gtcagaagct atggcagtct tttcttgttg acatgagaat 11220attccaagat cagctggacc ctcctcaagc taggagaaca cacatttctc agaaaacagg 11280gccacaacca tgaagataat gttcttctct aatcactggc attgcccact gagctatcca 11340ctaaaacaaa gtttagaatt gaggcaatag tctagaaaat ttaaaaaaaa aaaaaagtag 11400caatggctat tattgggata tttggatgca gaaattcagc acttgacttt gtgcctgtag 11460ctgttgccct ggggctcttg aatgctgctg attagaacat gggtaaattg ggagggagcg 11520gggcaataag gtagcctttt ctgtgttcca ggtacacaga cagcagctca aactttgtct 11580gttgctcaca aagaatgcct ttgctcttta gggtgaccat gtaaaaggac atgaaaggac 11640caccgcaaac tcatcggtgc tcacttgaaa aagcaagcct tgttcacctt tgcagaaagg 11700gcatccccac tgcataattg catccctgtg caaacctggg aagaaagagc acaggttgga 11760ggggggcgtt gtgtttgtct tgtatgccca tactgtgttg ccttttcagg agcagaagtt 11820catctgctga acataagccc tgtgagtgct tagaactttt ccctcttttt caccttgaca 11880taaaatccct aaagctgagg acctttccct ttcaatgtct tcactctgct ccagggctga 11940attgcatctc attattggct tgtttcacct aaatgtgagt atttgagtgg cagtgaacca 12000ggttgcctct aaggaataca actgaataat ggtggtctat tgtgaaggaa cctcttaggg 12060ccctacagtg ctgagttttc tactgtttac ttggagttcc tttctcagtg ccttcccaaa 12120caactgcctt gggaaaacac acagattctt tggcttccaa gaagagtgtg acagagaaag 12180tgtgctttgt tataaaggcc ttgtcaccct cctggatgtg ttatccatat cgatgtactg 12240gctgttaaaa ttcatgttca ctgcagaact ctagatggag gagtggtaag aagggagggt 12300agagatgttt tcatataaaa ttgtttctaa tgtgatctag aaaacccaag gattgcaaca 12360cagacctcag cagtcataca tcattattta tggttctgtg taaatgtaca atttaacgct 12420tctgaaatag taaacacatt agggatttcc ttttttgatg aacaaccaaa agctctgtga 12480ttgctttagt ttggatttct cgcaacttta gaaaactgct gtattccgtg ggccctactc 12540ataaatcttt gatgtgctct gttagtcaga ggactgacct tagcacattt gtcattctgc 12600tgtttgaaag atacattatc acagtatagg agaatggctg gtgctttcta cttccaggtg 12660gtccaaagtg ggaactaaac aaaatgaggg gataataaag ggattaggca acattcatga 12720aaaattcaga ggaaattatt agcataatgt ttttactatt tggatgacat taattgtatg 12780tagaaaagca ctgaactaca aaatagggct tacataattc ttccaatatc tttgtaacaa 12840attatatata tatatatata tatataataa tgtatataat atatatataa tgtacatatt 12900tatatttatt cttagcccct gcccagaaaa gtgtttttaa aactttactt cctgtttgta 12960atgtactgaa gccacagtac tctatgcttt aataatactt taaaaataat gtaccttttt 13020ttaaaatatg aaactagggt ctttaagaaa aaaatgtatt taggcattta ttaggcttgc 13080ttgatttatt tcatatatag atacacatat gactgtagct aattttaatt taaaattata 13140agtagcttct aataatacta gcaaatcctt atataatgct cacaatgtca ggacctgttc 13200taagccgttt gcatttattt atttatattt ttcaataatg agacttttaa atccaagact 13260gatttgacca tttattgttt gcacttctgt acaaatgtca cacataatga ttcaacttga 13320agagatgaaa tttactgtac tatgtgtcaa gtgtttggca tttaaatgtg tagccatttt 13380aaatagcttc tatcattata ttataaaatc aaaccatcct gataatagat catgccaaaa 13440tggcacttag tagaaaacag ttccaagatt aattaattca cagcactttt ccgctgagag 13500gtagttatta tttttttaaa ccccatttca cagataggaa aactaagaca aggtctttct 13560cctgccaatc ccaggtgttt attgcaaaag agactcagct ggcattaacc acagcgcatt 13620agtgtggctt atttaatacc tcaaagatat gcattctgtt tacatgaaca ttgaatcaaa 13680aacaaatgca actcagatgt tcaaaaaagc tgccagtaat agaatttgga caacaaaaat 13740ttcatgattg cagttgcctc aaagagtggc tcctgccaac tgctcagtaa atacacttgg 13800aatgttgtga aatgaatgga gacctaattc aggttgttag atactgatac attttttata 13860agcctttttt ctaatggcaa aattttaaca attgtgctag gtctgttata cacacaaatg 13920tgtttggtgc atgactggca aggctacttt gtttactatt ttacagcatt aattttcagt 13980gaatgcataa catcatttac ttagtgaaac tggagacaat gtgagagaat agcaaaggtt 14040tgaatggata tagatcaaat gtagtattgg ttctgccact taccaacccc agtaagtttt 14100tctctttgaa cttcactttc tcaatattta aaatgggcac agaagtaacc acctagtagt 14160agtattgatg tgataattaa aagagataga ctgtctaaca tcacagcaac tggcacttac 14220ttaatagagc attgtttcct tctctaaaag aatagtaaat cacacagaac attatacaaa 14280tctgtatgct gtaagaaata acttcagtgt tagtaattcc tggctttggc tatagggctt 14340tcataaataa aaaaattaaa accaatcctt agagactaat agcttaactc tcttatttat 14400acgttatcta tcttatttga cactcataaa cacccttagc aaatttttag aaaattttcc 14460caattttatc ttctttactt ttctcccctt tctgttgaaa agaaataaca aagtaggcca 14520aagacaagct gtttaagaag taagagcagc ttactgaata aggaaatata agacaaatta 14580ataagttaat aactgtgaat tactccccat gggtgaaaca ccatgccaca aactgcacaa 14640gtgtatctca ttcgattgta gcaaaatgtt gtaaaataca tattattatt tggcatcatt 14700ttttttaaca aagaaaaata aaaaccctga agttcagaga gattaagaaa acttgcccaa 14760aatgagaatt taaaaagtgc aagtggaaga gtttgtattt gaagccaagc ctatttaaat 14820ctaaaggcca gattccttct aatcactaca atataatgca ccaatataga cacgcatgca 14880tacacataca aactgaaagg gttgaacttc tttgtttttt ttttgttgtt gttgttgttt 14940tttctaattt atttgtggag aagaagatgc aatttaactt tacaccagaa agactgagag 15000caggcaaagc tgagtcctct ggttgagtga tgagttgctt ctgaccccac actcctcagt 15060acttaccaga agtgtaaatg gctgtatgtg agcaacactc ccacaggtca gggagcctgg 15120gtttcctcat caggctttcc cacccatctc cccagcagtc aaatggaaac cccttgcctg 15180actcatctga ctgttgtgag ggaatctcat tgtaaactct aaagatgtcg tcttacttta 15240tggaatcagg aattacttca ttgccaaaaa gctttctgtc tttcaaataa agtggtactt 15300tgttggcaga gggcagattt taaaatattt taaagttatg tttcaaaata acttcttgga 15360aatagaaaga tacttcagaa agtactttta tcaatttgct ctgatcctct ttacatgatg 15420ttttggaaga tggttttaaa tgtctgtgat catattagga aattatttga taataatgtc 15480gtttatgtct ggcttttaaa gctaccagcc atttatattg tggcctttgt tttgttcatt 15540tcttatcaat ccaactgcat taatattaaa aaaaatattt tctgcattaa tatcgcaaga 15600ttttaaatgg gtgattgtca gcacaacctt atcagcagca tcaccttgta acatactcaa 15660tagcagttta gtacaaatga aaatgttggc taaaaagtcc tggcatcatc tcaacacagg 15720taaaggagac aacagaacag agatgtgctt aaccaaactc tgcagtcaga tggtctgctt 15780tggttgccta agaacgaaaa cctcttagcc tcaatttccc

catctgccaa acagaatgag 15840caatggtaca tttatcatag gtagttataa ggataaaatg aaataatcca caaaaaagca 15900cttagtccag tgctggacac caagaaaata ctaaataaac atttactctt gctattacca 15960ttaaacaaat ggtaatgtaa tgcttatcaa tagtgatggc tcctttctta aagaaagtga 16020ctttaactgc aaaatctcca ctaccagagt gaaagggaaa gttgaagcaa tttatccaag 16080taactccagc ctcaaattca cttttgtcct gagtaatgta gttgttttta ttcttttttt 16140aaaacaaaaa ctgaaaaaga aaaatgtgta taattagaat cacaaaagag tttacctaga 16200gcaaagttac ttctttctat catgtgccca agatcgcggc accaatatgt ggtggctggc 16260ttatgaaaca gattcggatt cagagctcgc attcccaagc tctttgcttt acctggaatt 16320agggggacag aacatgagtc attcccattt tgaatcagtg agtgattgcc tacttttccc 16380tatatgagtg aagtcatttt cagcatccat agaaatatag ccacttgtca aaagaaaatg 16440gaaagaacat ggcttttgtg tcaaaagcat atagatttta cctctggctc ttgactctag 16500cttgacatgg gaccttaggc aatttattta tctttcctta accttctctt aaacttctgt 16560aaaatggaag aataatactt tgcagcttcc ttctcttgtt tcctcactct tttttctctc 16620ccttctttcc ttgttctttc ccttcttttt tttaaaaacc tggaaataat gttatatacc 16680taatttgtaa tcacgattca caatcactca aataattgta ctattagaaa acaatgtttt 16740tcagaattaa cacggtttcc tgctctaatt ttaagaagta aattatattt ctcaaagtgt 16800ttggctcatt ggccttgtgt agactggaag aagcatgggg tttggagtta gacagaatga 16860gagttaaatc ttagctccac tattatgatt acaacatgat atttatttcc ttcctctagg 16920ccccaatatc ctcattggtg taatggggtt gtaggggtaa tagtacatag ttggaagaac 16980acaagaatta aatgagatta tagtctgggc atggtggttt atgtctgcaa tccaaacact 17040ttggaagaca gaggcaggag gattgcttga gcccaggagt ttgagaacag cctgggcaac 17100atagtgagac cttgtctcta caaaaatatt ataaataaaa taaataaata aataaataaa 17160taaataaata aataaataaa tgagataata tgtggaaatt gtctgaccca gaagtgatgc 17220tcaataaatt gtatctgtaa gctttaggga atggattatg ccagcagaca catcagtatg 17280tattagactg tctgctctgt gtctgtaagg atgttgacac atgttggcga cctcacaagt 17340tgtgaaagtc aagagagtta attaactgag cagtgctcta catactatac tgtccattca 17400gaggcctagt gttgttgtat tgttctcaga cttgagcaat aaaaaatata aattttccaa 17460gtacatgaaa actttagttt attaatgtct tctaaaatga gcctcaaatt agtttttgta 17520tagtatgatc atgaatagct ttggagactc ataaatctgg aatcatctct tcaaactgta 17580tgactttgag tcacttgatt cttcggataa cattctgatt ttctcacttg taaaatgagg 17640tttataatag tgtctaagtc ctaaggtgac tgtgaggatt aaatgagcta atgcataaga 17700agggcattag gacaggtacc agaaaacctc ggtaaatgtt agctcctttc aaataaaatg 17760cagttttgca ttcaaaggtt gttcgtttgt tgtttttatt ttttttgcat gtgttgaatt 17820taataacttt atagaatcaa cagtttggaa atgtttccat tatgaattca ggttttattt 17880tcttgtcagt gactctgttt agtttaaggg cttggttgtc cagattcatc tttgagtaag 17940ttttgtatca atcaagacag atgcaagtta cttttatcag tccattactt caggtaaaca 18000agcattccat tctttgacag aacattaaac tgttatgttc tttttgatgg agtgctgatt 18060tgttcaaaac acttgtgaaa gagagaaaac agctcctgtt ttataactct tttgtaatac 18120aaagtttgtt tccaaaggtt ttctggaagt ttcttgattg aatctttgat tgtggatacc 18180acagtgatta atttaccaag ccggaaaatg tgggaagtaa ttttgggttc agttcagagt 18240gggatctaac acaacacttt gaaccaatat taaaatgcag aaatctcttt tagatattga 18300aaaagaaaaa caaatatata tatattaaag tccctgcttt taaggcctaa aggagaatgt 18360agtagaaaca ctgacatgtg taatttttgt gaattgctta ggaacagaat tagatttaaa 18420aaggaattcc ttttgcactt ttccctgaag atttgtgtcc tagcatttga ggagtaatac 18480aggaaagtca gaacaagaac ttactgacag cgcaggagat ccatgtccta aattcatgta 18540tgtggtactg tattgctatg cagctttcag gatgtccatt ttattgtcca tcaaaatgta 18600ggggtagatc ctataatttt aaaggttctt tctaatgcac acaatctaga gttattaaca 18660gaagtctgac tgtgtcatta ccctggctaa aatcctctac cgactttaca ttaccctcag 18720gataaagtcc aaaccctcag cacactatac aaggtcctct gtggtttggc tgttgcttat 18780ctccaccact cccgagtggt aagtttcatt ccagcgtcac ctaactatgt acaatgacat 18840accaatattg ctgcacttgg tttccaaggc cctgagggcc tcgcatttgg gtagctttga 18900tggtctctac tccaggaagt cctccctgac ttttgtttgc tctttttcct cagcccctct 18960ccctactatg ttgtttagtg atcttaccct gagctcctgt agcatttaaa aaaacaatca 19020tatcacttat cacattatat tgaagttatt tttttactta cctagctaag ttgtctaaca 19080ctggtatccc tagctagtgg cacagtgcca aaatattgat aaaggttttc tgaactaaac 19140tatgaattta atgccaaatg tctctttgag ggacaacatc cacctgtcag cattctaaga 19200atggatccag acatttgaca attttgtgcc tgctgacctc agcagacaca tgaaactgca 19260tgtaacactg tcctgtataa acatacatac attgttcaaa cagaattttc ctcagagaga 19320gaaatgataa atgagccact ttgggtggta gaagagaatg gagacagtaa attaaaaata 19380aataaataaa taaaggagaa ggtgaggcca ttgcttgcca ttcttaagga aatgacacaa 19440taggctgaat gacaagggcc agttaaaact cgctaatgcc agatatatga agacttaaga 19500accagaaagg attttagggc agaatattgt tgatgatgat gaagaggaaa aatgttgatt 19560tccgatgctt attctctgtg ccaggtcctg agctaaatgt gttacatact ttatgtctcc 19620tgattctccc cacctgtgag ggtggtggta gtgtgattat tccatttgat acaagaggag 19680gaatctgagc tcacagaggt tgagcaaatt tctcagcatc ttacaactct cagtcatcaa 19740acctgatctg tttgttccta atttcttgct tagtattagt actgttgaca ttttattttc 19800tcacctagag attttgtttg tttgtttctt tgtttgttta aatggagtct cgctctgtca 19860ccaggctgga gtgcagtggt gcaatctcag ctcactgcaa cttccgcctc ccaggtttaa 19920gtgattctcc tgcctcagcc tcctgagtag ctgggattac aggcacgtgc caccactccc 19980ggctaatttt tgtattttta gtagagatgt ggtttcacca tgttggccag gatggtctcg 20040atctcttgat ctcgtgatct gcctgcctca acctcccaga gtgctgggat tacaggcctg 20100agccaccgtg cccagccgag attttttttg atacataata attcatgaga ctattacaaa 20160tatggtaaat ggattcttcc ttcccagctc acccacactc aagatgcttc ctttaaaatt 20220ctgttaacaa tttgtgctag ttaataggta gctagtgcaa tactcaagta aaggatgttg 20280tgtagttttg gatactgaga tgtatagtct ttgtctaaaa ggagttgaaa gacagaaata 20340tgacaatatg agagggttta gatgattcta ttagaatgat atttatgtat ttaaaacaga 20400tatggttaag gagttcagca aaaataatgt ctcaatacat aagatcagag attaagcgtg 20460gaggaagaaa gttggagaag gattaacaag aagtgatgtt ttcaaatttc atgtgtggtt 20520agaagccatc atcttagtga ctcaaggcag taatactact gctttacaat acagtgctct 20580aaaataatgt tacagatatt gcatgtactt ttaaaaacat atatcatttt ggggatactc 20640tttgatgcca agtttaaaaa attctgaagt gtttaaaatg atacaaactt agagcatgat 20700ctgtgttcct ttttttttta atcaggtaag cttcttggtt ttgggtgtag atctgaaaag 20760ttaacatttc ttgaaccaat aaaatgtttc cagttttata taattatatc acaaagtcta 20820ccttctgcct tgatttgcaa agaaaagctc caaaaaggac cataaagaag aggagataac 20880acacaaatgg tgagcaatga agtagatgta caaaatggtt tgtgcagatt tggcacttaa 20940aacatacagc agccgaagta catcttaaat ttcatttagt gaatatattg ctcacgagtc 21000cctctcttca tttcctgagg aagtagatcc ctattctctc ccatatggtg agaaaatctc 21060tttgctatcc agtttccctg tttacatttg ttcttccatc cccttctgat tcctcagcaa 21120agcagctgct ttgaaaagaa gctcaaagga ttaccttgaa atggttaccc actctccccc 21180ttttctgctc caagaagaaa gtttctttct gtattttccc agcattatta agagtcaaga 21240cttgaggcct taaggtacat ttcctatatt cttcttttgt tttaaacttg aaagcaaaat 21300aatcttccca agtctaccat gaggatgaat ttcagaatgc acaataactt acatttaccg 21360agaaaactgt taagtcagaa tcttctaaaa tagagataac aataacaagt gtgatgccag 21420tactcctatc ctgatacatt gtaaaaaatc atctccttcc ttactgtgtc ctacatcctt 21480ttttcttttc tttttttttg agatggagtt ttgctcttgt tgcccctgct ggagtgcaat 21540ggtgcaatct tggcttactg taacctctgc ctcccgggtt caagcaattc tcctgcctca 21600gcctccctag tagctgggat tacaggtgcc caccaccatg cccagctaat tttttgtatt 21660tttagtagag gtggggtttc cccatgttgg ccaggctggt ctcgaactcc tgaccttagg 21720tgatccgccc accttggtct cccaaattgc tgggattaca ggtgtgagcc actgcgcccg 21780gccacatctt cctttcttaa tgaatttaga gtctcctcct acatcaaaat ggtattagag 21840agaatgctaa ggataattcc atggatctca cgataaacaa ttggaataaa agagcttgcc 21900acttagtcat gattattagc atgaaaatgt gatgatgcag cctattttgg gggagggatg 21960aagtgtcatt gaatcagctc atgagtagtc tggcatgcga ctttctgttt cttgcaaaca 22020agttgcatga taggggaaca tcaacactat gccagtgtgt ccctgaagaa aaggtagctg 22080gttttgtttt ggtcaaagta ggaatgagtc tggaaaggga ttcagtttta tgaacattgt 22140tttgcctgct ttagaaaaga tgacagatag ataagagtca accaatgctt tgggaagaga 22200taagattagc atttaaaagc ttaacccatg tcatcacttc tgggtatgtc ttccctgagt 22260gaaatcttat tctttgcatg cattttgaat gctgaatcac catgaataat ttttatgaat 22320ggctacttta gaaatgttaa tggattgcaa ctcattgggg aaataaaaaa tttatggttg 22380cgtattacaa taaattctaa ataaagcagt atatggaatt gttgacttct cagagacttc 22440cagtgaactt actgcaggac agtgtctaca tttcagctct gtgtgatact ttcagtgttt 22500tcgctttttt aatatgcccc tttttgtcaa taaaggaaag aaactccact ttatacttac 22560gtagtttctt cagaaaggga gctctgggga ttttatagcc aggtcagccc ttcaattttg 22620ggccacatct accaggagaa gcatttttca atgattgttc attcattaaa attgtctatt 22680tacgttgtcc ctgcccttct tggagcttgt agtctactca tggttttata gcctagtttt 22740ccacttacat ggacctaggg aagtattaaa ggtacaaaga aaatggttaa aagttcaaac 22800tgagaagcat ttagctaatc gtcactattt gaatttagaa gttattgagc ctaagagtga 22860gaaggatgac aatggtttat acccaactga tgaacaaaac cctcctgttt atccttcact 22920cattcattct aaatatattt acgaggggct tcttcacacc tgccattgcc ttagtcagtc 22980tgacaaaaca gggagggact gaaagtccat atggcacctt atggtgctca cagtcaagtg 23040taagaaacag ataataagca aaactggttg gcgatgttgc aattctattg tgtaaatatg 23100gaaaccagct gctgaggatt gaattgtaca ttgtccagcc ttcatctctg ccccattata 23160tgtacctggg tttattctct agtaacacaa tttagatagt gccagtacct tggtctgtac 23220catataaaag agaattttat tgaaggagaa cttttatgag caattcagta gcagttagca 23280atacatacaa tccacaaata tggaggatta tgtactatga gaaaggacca aatcaaggtg 23340ctagaggtaa gaatgacgga taagaataat ataatcctca ttcttatacc atgtatattc 23400ttgttgggag gaacagatga taagagtaaa ctaattgggt aatcaaaata atttctgaga 23460atgaaaagtg cttaagggaa agcaaaactg tgatgtgatg tgatgtgata agaggtgctc 23520cttaagacag gttacctggg aaggcttctc tgagaaggtg atatttgaac tgagtcctga 23580atgatgagaa caacttgcca ttggcaaagc ccaaaaagga gaaaactggc agcagcaagt 23640acaaagggct tgaggcagga attaccttcc aggtatttca ggaacataaa ggaggccagt 23700gtggctagag catggtggga agataaaggt tactaaggcg taaacatgga gttggacaag 23760tctgtcttcc gaaggtctct gtaggccttg attaggagtt ggagtcttac tgtaaatgta 23820atgggaaggc attgggagat tttaagcagg tagtgatcat gttttatata ttcccagaag 23880aagatactat tgtgagtaat aagtctgaaa gcaaagaagg ctatttagag gatccctgta 23940atcttccagt aagagtgtgc taagagctta gcacaagcta aagttgtaca agatgaaatg 24000gagagaaatg agttttgaga tttattttgc tggtagaaaa tatgatcaga tgtggaaata 24060ggggactggg aagaatcaag gatagtgccc agatttttgg cttaaaccac cattcttcaa 24120gtttgggcat gatgttacta ggaatgcttg ttctagacag caatgacaat agccatagca 24180aacatttata aaggtgtatt atttgacagg tactatacta aatgcccact tacataacta 24240tatttaatcc tcagcaaacc tataaagtca gttctgttat tacacccatt tacagatgag 24300gaaactgagg cccagaatga taaaataact ttccaacggc cataaggatt gtgtcaaagt 24360agaaattaag atatgggcag tctgattcta aggttgatgt tttccaaccc cctcctctat 24420accacttctc tgtgcatgag taaacacact gttttctttc tttctttctt tttttttttt 24480tttttttttt ttttgagaca gagtctcgct ttgttaccca ggcttgagtg cagtggtgcg 24540atctcggctc accgcaacct ccgcctccca ggttcaagcg attctcctgc ctcagccccc 24600caggtagctg ggactacagg cgcatgccac catgcccggc taattttttt atttttagta 24660gagacggggt ttcactgtgt tggccaggct ggtctcgaac tcctgacctt gtgatccgcc 24720cgcctcagcc tcccaaagtg ctgggattac aggcgtgagc cacagcgccc agccaacaca 24780ctgttttcta aagctgtttt ggatatatta gtgaagctga agttgacttt tgaaactttg 24840ctatttaggt taacattcac ctgttgccca aacaattttt gcctcacttt tatactaggt 24900agaaaagagc ctgcctgctt ggaagagcgg atccaaatcc aagctcaggg ggcagtttca 24960caacctcttt atttcacttt ccagggctga tctttaatct atggggaggc agaggataaa 25020tgagattgtt tacaattatt atgtgcaatt aaaaagaaaa cacatttttt aaaaaatggg 25080ttgtattgtg aatgattgta tagtcattta ctgttagcaa agcttttaga atatttaagg 25140gatcaataaa gataaaatta taactgtagc atcaactgtg gtttaactgt agctaactga 25200actagctata gttaactatt aactgtaatt aacaaaataa taattagagc attaatagtc 25260atgtaatgtt ctcatagatc tgccaattgg cttgtgagtg ctttacataa cattgtctta 25320tgtaatcttt tcaaaactgc aaaaacatgt tatcatcaca agaaaactga gtctcagaga 25380ggttacataa tttgctctgg attacaacac gtgacagcac caggatttga atccatatct 25440tactttgata acgttctctg ttttaaatgt catcatcttc ataaacagtg agcctgcaat 25500tgacatatat tgttatcaga agaacaagtc ctgggcatgt cattgaacct gcagttgtag 25560agtattgtta ccagagaaat gtgtctcagg catgtcaatg aatctgcaat tgaagtttat 25620tgttaccaga ggaacttgtc ccaggcatgc caactgtaaa tgctttgctg tgtttcagct 25680ggtagaattg tattaaactc tcatggattg cctataaaaa attggtcttt tttaaaattt 25740aaaataatcc ttatttttca acttgaattg tgaaacacta atgggaaatt tctaaggaaa 25800attttgacca cccttaggag aaataaatgc ttaattactg ggctttttcc ccttgttttg 25860cttaggattt aattttggac aatatagtct actctgcctg attgaaacct gggatatact 25920tcaaagagta tattgaagtg catttgatgg cctgtggaat ttttggaagg gccgtaggag 25980cagactgcaa gctgagcttc cagaacactc ccagcactca gaacttaagt ccactacagt 26040ctgtaaatat gttgcggcca gtgcagaaag ctgttactgc tgctgactgg agggtcttgc 26100tgcccgcagt accaactaag ccaataaaat gcatgcccac agccagcctg tcttctcgag 26160tatttccttc ccatatcaga tcttgcagtc acgcctaatt ggcagaaccg aagtcccatc 26220tacaaccctg gataacttaa agggcgtctg ataaatggag tttttagctt tccagccatg 26280atagtttaga gggatatgct aggagaatat tggaatggca attgaaagag acaatcaaca 26340gtattctgtg cactcaatac acttaacttt tcttatcttt tttgttttta atttccatag 26400gttattgggg aacagggggt gtttggttac atgagtaagt tctttagtgg tgatttgtga 26460gattttggtg cacccatcac ccaagtagta tacactgtac ccaatttgtc atcttttatc 26520cctcaccccc ctcaagttac tctaaatcaa gagcacctag tatgtagccc ttcaatcaaa 26580tacaacgttg tatatctttt aagacatttt tgttttgttt gttttcttta acaatataac 26640actcatgaac tatctaaaat attatatctc agactttata gagtacctaa taatcacata 26700gagggcttac taaaagatgt gctttcctga gatgcaccct cagaaatttg gattccctct 26760gacatgcagg aatctgtatt ttaattaaac accccagatg attacacttt catctaaaca 26820taacacagct gactcccttg gtttcagttg gaacaccttc agtgaaagaa ggaacatttt 26880aacagtttat tccatcccac taaaaaggca gttggtatgt ctcacacaca ttgcataatt 26940cgcaaagcca gtttgtgacc tttgccaatc aagcttgtca tctgatgtga cagagaaaac 27000gatcaagatt gtgaacgttc tttgagctca tgtctctagc tccactcatt ttgatgatat 27060ttggaagcag ttaaagtaga gtataaagat ttagtttatt ctcatgtagt aaaagatcca 27120ggttggtggg tggcgggtcg tgaggggtgg ggggaacttt gttgaaaatt ttctgagctg 27180cttcctcaga tttttattta tttatttatt tatttaaaag gtcttgtgag actaaacttg 27240tcagattttc agtagcagat gtgaaaatac atatgagcac attaattaca tccatagtgg 27300gaaaaaaaca caaaaaggtt ctaggaaatt caggcattca gtctcagatt gggcacttgg 27360gatcagatgg caactgaagc ctatgctcat ttcacatcac caaatggttt ggagttcaaa 27420atattttaga gattttaaat aattacataa tttttgattt acatgagatc tcactttcag 27480aagcaagtca cgtatttgaa agagtataaa ttatgattta tgtttctgta gctgttttta 27540gagagctaca gccctcagat tcttgtacat cagggtgatg tacaaccctc agattcttgt 27600acatcagggt agttcaattt taagagcagc tagttggcat aagagtaaac ggtgcccgtg 27660tccatggagt gctgaagatg cagatttcag aactttcatg aatttttaaa attatgtttc 27720catactttgg ccggtgttgg gggaggtctg ctcgtcttat ttctttgaaa atacacactt 27780attgcagaga gttggcggta gatatataaa cctagctaac ttaaagggca tctgagaaat 27840ggcgttttta gttttccaga cgtgatagtt gagaattaaa ctagtaccca tcagttttct 27900agactagctc agtctgttac acatcattat cctcacagaa attagaatta gtcattagct 27960cgcatctcct taaaattaag tggattatac tgtgtagggg atttttttcc cctctgcttt 28020gtataactgc taaagagctt tatgaatagc taatgatcta ctttttaaac cattgttact 28080aattagatag tctatactgc tgtcacaaat gcttactcta ttatgtacat ctttattaaa 28140attagtaaca gtaaaaacct gaacactcag gccctatatt cttgcctgag gtaccagtct 28200tttatcaggg atttttacac agggtcaaaa cttctgagct cacctggctc tgttaacccc 28260aggtgagttg ttccctattc ctaggccatt taaaaaagag aaatgctgag tttattattt 28320tagtcccact ctttcagggc tgtgtgttcc cttctaccaa gaccgaagtg aacttttcta 28380gccctttcca tgatcactca ctctcttctt tcttttttgg caaactgtag taatataccc 28440ttgaaattcc ctcatactca ttcactcctg tagtccatcc actgtgattg acttttagtt 28500ccatcaactc aatggcaact gccctcatta aggtcagctc cattaagtac tctttagtaa 28560acatcatatt gacatctttt agtgcttgtt ttattcattg ccctctccct ttggtatttt 28620ttttccattg gctgctgtac cccacaatca actgatttcc ctctcatgta tttggcttca 28680cattctctca tctcctacac agatagttca tgatagaatg gtatctatct atggtatggt 28740agtattcact aaggtttttg gtgataaact cccagactcc tgctgcctct tctcttccta 28800catatctgga tcagcagata gcaccacttt ttacacagtc acccaaactg gaacctggaa 28860gtcatttgtg cctcttcccc ttttgaccct accctctttt cattagtcac caagatcttc 28920cgttctactt ccatatcctg ttgaacgtgt tcatttctct ccattctcac tgccactgcc 28980ctagttaagg cccttatcat ctcagctttg gattacttca gtgacctcct aatgggcctc 29040atgcccccag gtttgttgcc ctctctcccc cagtccatcc ttcattctga tgcctgataa 29100accttttaac aataaatcag atcttgacac tcccttgctt aaaatcattc caggtctccc 29160cagtgctttt aaaatgaaat ctaaatcaat tagcaaggag ggcctctgca tatctctcta 29220gccttgcctt tttttttttt tttgcttatt ctttgaaact tattttccaa tggagtaaaa 29280aaatcacaga attctaagaa catgccttgt ttccacactt ctgtgggcac ctgctacttc 29340ctctctcagg ggtcttctcc ctcactaact ttgagttggc tgcccccgat cacctttgga 29400gaagcttctc cccccagaaa ctatcagccc ctccctcaaa tgcccctgtc ccccaaatgc 29460ccccctcccc aagtcaggtt tagctgtact tggggcaggg ggcagttgag ggaggggctg 29520ggccttttct ctctgactcc caccactgtg ctgtgttctc tagtagcttg agcttcctga 29580ggccagtgat catagtcttt ctgtctttat cttatgtatt tattagatac tccatacatg 29640tctgttaaat gaagaagtgg cccctgggat aatgttacta gaagtctgaa agcacccaat 29700tttgttttta taagaagaaa caacttatcg ctttattttt tgggtttctc ccctccctct 29760agctatgtaa atttttgttt aagctatagc taagtgtaac ttgatttacc tgctcgactc 29820atttattcaa cctggtaata acttgacatg taatttcaat ttgaaactga ttgagttaaa 29880gagacctata tgccttctgt aacatgaccc tttaaaataa aatttagtag ttcattaaat 29940ataaatgaag tcagttttat tatctaactg gcctagtatt tttctttcca gcatagatag 30000tgtaggtttt ggtttatatc caatccagca atttcctagc aaagtggcca taaggaagtt 30060acctaatctc tcccgaattg tgttttgctc acctgcagag tagggtaata atatctacct 30120ggttgatggg aggattacag gaaataatat ctgcaaactg cttgtcctat gcacagcccc 30180ttagaagtca atatattttg agttatttcc actttaagta gtatttacat gtagaacgag 30240gactgattat taaagggtga atttccccaa atttttctca tgcattgaat accttgaaat 30300ctttaaattc ccgttacatt aaattccctt attatattac attgtctgcc ttttcatatt 30360aagcaacttc tttctagttt ttttttgtta taagagcata ttcattacat attttaatga 30420tcacccatat tttaaaacat ttttagctta cttcttcaca aaagggaagt tagaagagta 30480tctttacaag attttggcac ccaccaattt ggactccaag ttggtctgag tttcaattag 30540ttactctggt aacatctgtt ttattgatag tagttaagtt tcaaaaacta acaaatctac 30600taaatgtttc agttgtccct ttaagattac ttaacagtgc ttagagaaga attttagtct 30660gtgtaagatg attaggtatt ttccttataa tgtttaatgg taaattttta agcaattaaa 30720aagatctgaa aaaagaagtc atatagaact gaatggttat tcaccaaatg aggatcctgt 30780tgttggttta catagattat ttttggaaga tatggttcat atgagttaag aagacctgaa 30840ctcaggacca attaccatac ctgtatcttg gtaacttgct

ttcctgtctt tgggtagttg 30900agccagttgg ctctcttcca aaatacattt gggctacaag ggaactgcct gcttcaatag 30960ctggtatgca ggattaaaga gaagatgaat gttgtttttg gcagcagctg ctgttttgtt 31020cagggcagct ctaataccat ttcgtaggtc tgtgcataaa gaaaatgagc tacacaaaca 31080cttaactctc tttttgactt tgtgacgtct gtaatatgga ccctgacaat gcatatttta 31140aaatcacttc tcatattcat agagaaaaaa aggctttttt tctgtatttt atttttttgt 31200ttttccaagt ttagaggatt ggtatcttct ggtttcttga acagcatatg tccccctgat 31260attcttaggc tgcatgcatt ttaatttcta ctgaaggaaa ggaatctgca gtgtgactta 31320tgtgttgcaa cacacgttca gattaagtcc atggtttgtt ttgggtgttt gtagctcctt 31380ttcctggcat gtggcctttt actgtaagga aagctctgtg ttcaggaggg attggttatt 31440gagtgaatag tccactggag tctgctgatg tttgttctca acaaaaatga aaattaagca 31500agacctttgc ctttgaggtc ttcaacctca tcttcttgta aatccctccc accccagact 31560tctttaaagt acttttctag tactgtctgc tactgtctgc tcccagaaaa ggaccagcca 31620catctgtcac tgatgttaac ataaatttta gtcagtatta cttccctcta ccagaactgt 31680ttctactgca tgccttggtt gtaacagtgc acacagattt ctcattattt tttttcctaa 31740tcatgtaaat gatttgaagg caacatttag caatgaaagg attattggaa ttttgtgtgt 31800tgtgggggaa taaacggggg acacttttga tgcatgtttc tgaaaacaaa aatcgagaac 31860tacccaggac gatatgtgtc tatacaatga aagactatcc agaaatgagg ggttatcaca 31920caatgagctt gacctagaga gaaaagttgt acaatgataa aacaaagtag ccaaagaaaa 31980atgtcatata cagctgcaga gaatgttaac agaaaaaaaa aatagatgat tacccaaact 32040gaaaagatac attttagagc agaagtcaca aacacaggga cattttgctt cagtgtatct 32100ccattagggt gtgtgtgagc gtgtgtgcac acccacatat ttaggcaccc tattccctca 32160gagtttccag aaattggaag aagaggaaga gaaaacaaag ataaatattt tgttgatata 32220ttggaacatg tcatttaatt ggaagtcagt tgtctgttaa ctgaagctac tctaaacaca 32280gttcagaact aagaaataaa tgaaaaaggt gtgtgaactc aaagtcagtg tcaaaatatt 32340catgtgctaa aggtcatact ctttgtatat gggagatcta ctctataaag tcttttctca 32400gaaagtttta ctcaaacatt tttgaagtac ttgtcttctg gtttcctagc cgatgcaaag 32460cccataaacc acacaacaaa taattgcata acacccatag tttgaatagt ttctcatttg 32520attcttacta ctagttcatg agatgggtaa ggcaaaaatt atttttacta tcattttata 32580ggctgataaa tggatactca gggagattag ctgacttgtt tgaagtcaca tgggaaggga 32640ctgagaggtt ttctggatcc atgttctgcc catggtttcc aaaagcgttt ttttttttta 32700atttcttctt gcccttggaa acctttgttt aaacagaatc ttactcagag gcatgaacaa 32760ataaaacaac aaaaaaagct gagctgctct gcttaaaata aagggtgcgg gccaataatt 32820ctggcagagg caattcactt tcctcatttg gaatagcttg accatcacta cactacacct 32880ttcattcatt cattcattcc aaaagtattt caattgagct gtgttttatt tttctattgc 32940tgctgtaata aattaccaaa aatttagtga ctggccgggt gcagtggctc atacctgtaa 33000tctcagcatt ttgggaggct gaagtgggtg gattacctga gctcaggagt tcaagaccag 33060cctggccaac atggtgaaac tctgtctcta ccaaaaatac aaaaaaaaaa agaaaagaaa 33120aaggccagcc atggtggcag gcacctataa tcccagctac ttgggaggct gaggcaggag 33180aatcacttga atctgggagg cggaggttgc agtgagccga gattgcacca ctgcatccca 33240gcctgagcaa cagagagaga ctctgtcaaa aaaaaaaaaa aaaaattatc ataattttga 33300agatcaaatg ttcaaaataa ggctcactta catcaaagta tcagttggac tatgttgctt 33360tccaaaggct ctaggggaga atctatttct ttctcttttg cagcttctag aagctgccta 33420catttcttga ctcttggtcc cttctctcta ttttcaatgc cagcaaggac atgttgagtc 33480cttttcaggt cacatttccc cgacccttct tccatcatcg cacctctctc tgaacagagc 33540tggaaaatag tctccatttt taaggaccat ggggattaga ttggacccag ggaatctagg 33600atagtctctt catctctagt tcttaacgta atcatgtctg gaaagtccat tttgccatgt 33660gtggtgatgt attcacaagt tccaagaatt aggacatgga catctttggg gccattatac 33720ttcctgctgt aagctcccat atgggacaca tgttgtgttc atcatcaaga gtgcaggaga 33780ctagaaatag gcatgttagc agtaacaata tgttgcaagt aatgacctga caatctgggg 33840gaaacaggaa aactaaggaa caaacaaatg aaacctaaaa taaattttta aaagttatac 33900aaacaagtac agtaatttgg aaccacttgg agaagggaag ctacaactac ataactgaga 33960tactttgaaa gtcttccttt tatttgatga acctgaaccg tcacagaaaa gcaaagttta 34020atatttcaat tcactgttta agaaatcaaa gtgtgtgtac aatattagaa agacttctgt 34080gtagaaaagt taatactgaa aagtttcttt tttatataaa gaagtaagct ctgcatttca 34140gctctctcct gagtatactg atgaaggagg tgtcattact gcttggcaaa aatctcaata 34200tttctaactt gattatctcc ctgtcctaat gtttagctat gccagaatga attctttaat 34260tttatgaaag gtggatattt ttaatggtag caaagtatat gcaaaattaa attacaatat 34320gaatttattt ataatggctt taaaataaag ggctttaaga aatataggtt tttcagagat 34380atttgaatgt tcataagcta acatagattt aattttagca ttgacccagt ggatttttac 34440aaaatgaaca aaactgtgtc tttgtaagat atttgaattt aactgatgca aaaaaaattt 34500atgttgccct gatcaaatca ttcttcaatt attatagcag tctgtgttga tatattttaa 34560ggaactatca aaggacatat attaaaaatt atgatgactt gaataatata gactattcag 34620ctgtggcatg gaatatggaa cagacttgtc tccatatgct tgatatttgc aaaagtatta 34680aagttttaga gatctgctat cgcatcacag ttctcacaaa ccatctaaca gatttgcagg 34740agttctggga cagtgtagag gggctaaaag gacaggagtt agaattggac agacctgggc 34800ttactgattt caccactcag aagtcaccaa acctccctgt acctcaactc cttcatcttt 34860aaaattagga tatatgaaag ttattttttt ccttattaga tatccaagat gcttggtgta 34920tagtaaatgt tcagtaactt ttagctctat tcatggaatt gttcccaaaa cacaaccagc 34980atctgcattc acattccagc gctctgcctg atagcattga actacctata ctcctattta 35040aagccacact tcctacttga gaactgaatc tcaccccttc tttcctgctc aaggctgttg 35100cccaagcagt cccctctctt ctgacctata ttgccaattg cttcctccta ctggatcatt 35160cccataggca tacactcatt ctggtatttc aaccttctta caaataaaac tctcttcacc 35220cccaatttct tgccagctac ttccccatct tggttttatt tatttatgtg tttgtttatc 35280cagttgctct cctttgaagg aaaactcttc aaaacagctg ttcatactca ctgtttctgg 35340ttaatctcat ctcctgcttt ctaaaaccca ctgtagactg gctttttccc ataccacttc 35400aataaaactg ctcttgttaa tattaccagt gaccttaatg ttgccaaacc agtggtcagt 35460tatcagtcct tatcttactc ggcctataac tggcatttga catagtttga ccctattttc 35520cttaaagtac ttttttaaag tcttggcttc cttccatgat ctttttgctt tgctctcttg 35580gactcttttg tgggttctca ctcttctgtc agatccctta aggttagaac accatagcat 35640gcgcagttct ttgttctttt ttatactttt actcactccc ttggtgatct catccactca 35700tttgcttata tatggaaggt tatcatattt atatctgcac cccaagtttc cctcctaagt 35760ccagatttaa atatcttgac ctggcattat cactagaata tctcaaagac atcttaaact 35820taacatactc aaaatttaac tcctgatctg cctcccaaac cttctctacc cccagtcttc 35880cccatatcag ttgatggcaa caccatcttt ccataccaaa gttcttatag tcatctttga 35940ctctttctct caaatcgtac tctgtcagga aattctcatg gttcagcctt cagaagatgt 36000ccagaatcta accactttca gtcaatgtat tactataacc ctttcttgat tggattacag 36060cagcaacctc ccaacttgtc ggtttcattt tctacctgtt gtcaccctaa agtctatttt 36120tattatagca atcagactga tcttatttaa ttaatttatt tttattctga agtatatatc 36180cctcctctgc tcaaaatctt gcaatgaatc ctattttatt caagtaagag tcaatgtctg 36240ggcaataggc ttcaacgccc tacatgagct gtcctctcca ttgtctctct gacatcatca 36300tcttctactg ctccctaatt tactcttgct atgtcttaaa taacagatgc cttaaataga 36360ccttaggacc ttggcaccag cttttctctc tgtatgcagt actcttccca cagatgtctg 36420cgtggatcac tgccttagct ttgtccaatc tttggtgaaa ttttgtcttc ttaatgagac 36480acttatcaat attaccctgt taaaaatttc aagctgtttc cacccatact gcaatctctg 36540gaaagaaaag agtacagtta gttctatggc tcttctcgag actcagagat gcctggtgat 36600ttgaccctcc ctataagtct tctttcaaca aggactcaag aaacttttag gtgtacatct 36660atacttcctc caagggaatt cagctgaagt tactaggtag caggcactga tgtgtgcttt 36720attacaagca tatgtcatgc actgatatgt attaatattt tattgttgag actggtactg 36780gtgggggtag ctgctctaga aaatgaatag aagttcatgc catatcccta aaaagctgct 36840cagctagcaa ggtataagca gaaggaatga ttctacctac ccaacatcac ctatcattgc 36900cacacgtatc ttgagtgttc agtgactttc tgatattaga gatggcctgg gagtctgctc 36960tgcttgtgct gtatgtactc ttttgttatc taactttttt tttttttggt gatgtagtct 37020agcttttgtc gccaggctag agtgcagtgg cgcaatcttg gctcactgca acctccgcct 37080cacgggttca aacgattctc ctgcctcagc ctcccgagta cctgggatta caggcatgca 37140ctgccacacc cagctaattt ttgtattttt agtagagatg gggattcacc atgttggcca 37200ggctggtctc gatctcctga cctcgtgatc cgcccacctc ggcctcctaa agtgctggga 37260ttacaggcat gagccaccac gcccggccaa cctaaccttt cttaaaactc agcagtacac 37320ttggtgctgg gcaaagcctt ggtgctgggc aaagcagtac acatggtgac tcatgactgt 37380gattgtcact ctgaacccaa gatcaccagt aaggcagagt agacattatg atcccaatcc 37440cctttacctt gctctaatct tatttttccc atagcttttg acaccttttg acatactgta 37500taacctagtt acttatgttt gttgattaag ccttatgagg gcaggaattt tgttcattat 37560ataggcctag cacctaggac aatgcctggt atacaccaga tactcaataa atacttgtta 37620gatgaatgag aaaacaaacg tttctttagt atttattcaa tgccaggcgt aaaggattgc 37680gaagaccagt aacagaattt ctgccattac atttctttga gcaaaggata tgattatggt 37740catttctgag attggtccat tacagagatc cttagagaaa gttcccaaat cacatcagaa 37800agaggggttt agagaaatat atttccaggg aggttacgtc ttagcaatga caatataaaa 37860gaaagctgac tttaataaat tatatttaca catgtgaagg aagactgtta ttaagggaca 37920gtttgtatgt aggggttgtc tgtccaaaaa gaagaatcct caaacagaaa taatttatac 37980aacttagtct tcactgtggt ccaagaaaga tgatactatg aagagtgtga ccccacaatg 38040ctagaagtag ccttaaataa aattatgcag gattactgct aaaatagacc atggtggtaa 38100gggaataaaa ggatttcaat cttgaataca aaccactata tccatgtaat tagcattgta 38160gttttagaaa ttttagcaaa taataatcat tatttgttct agtataatat ctagttagca 38220tgcagatatt aattgaaaat aataatccag ttccttttta atatggctag ctatgtaaat 38280agagaagatt tctagccatg tgcatctgga ctcttgtgta attttaagat tttacagaag 38340aatggtatct cataaaagaa tgtgcacata ttttaagatg tcttcacatc tgcatggttt 38400tgtcatgctg tttatttttc ctggaacgtt catcttttct ttgtcagccc ttgcaagtcc 38460tccttgtcct cctaaatcca gccacagtca cattttcttt ctggcttttc ttggcactgt 38520cttcctgcaa ctggttctcc cccttaaggt taattgatta attgcttcct cttctgtgtt 38580ccgaaagtgt ttatcacaag atgtgagtgt tatttattta cctgcctctg agtttcaagg 38640gcatcttcaa catctttgtt accctagtga ctggcgttgt tcctgttgca taatatatat 38700tcaataaatg gttattgaat ttgagcatca ggatatattt ttaccttgac cactgattga 38760cccagcaaat atttattcag gactttttat atgcaagaca ctgtgttatg tgttgtgaag 38820atactacaaa gataaatcag ttgcacaagt tcttgaaact ctacagtgta ataaggaaaa 38880ataagtcatg cataaaagca actataatac ataatagaaa atgttatttt caagccgatg 38940tgtaggttat gtgtgttcga gagagagaga gagaagacag attactttct gctagggttc 39000aagaatgcct tcctgttggc taaggaaata ttttccttaa gtggctaaaa agctgtgttt 39060caaaatattc ttttgatgtc tcacaaattc agtggaattc tcttaggtct aaaaatatac 39120atctctctca ctttaacttg gtgtgctatt gtagattatt ggattaaagc actgctcagg 39180gattatgctg cttcttgcca agcagtctac atttaaagta gaaataagat gtttcttttg 39240gtgccataag gtatacattt tatgcattct ctagttttta gaagataccc taagggctaa 39300gtctttaaca tgctgctaca agtttattcc taattgccat tgggaaattg gctgaagaaa 39360gtttttaaca aaagttaaca atattgtcat tgagagaata attcaaaatg gattttaact 39420aaaagctttt aaaaactttg gtgagcatag cttgaatgcg taatatttaa ttgcatttaa 39480gccaataaca tatattagac tggtcttttt gtgcatcaag gcattagatg ttaaaagttt 39540gaatgattac agatcttaac tgatgatcac caagcaattt ttctgttttc atttagactt 39600ggattctcac ttgcatttat cttcagctgc tcctatttaa tcctctcgtc aaaactgaag 39660ggatctgcag gaatcgtgtg actaataatg taaaagacgt cactaaattg gtaagtaagg 39720aatgctttac cgtgctgtgt aaaaaagagc tgtggctctt tttcctgtgc ttgttgataa 39780aagatttaga tttttcttgc cccaaagtaa tgttttccta aagtggggaa agtaatcact 39840gggttacaat aaagggttta tagaaagcag gtagtgagat atttagggtc atggataatt 39900tgttggtaaa actggctagt tgcacaccac tgctgtgact gcttctttgc tggtcttctc 39960cccatccttc ataggcagtg aaggaccttg gagagttcgc tgtgtgctga tgggcttgcc 40020ccagcttgtt ccccataatc tctccagtgg gtttcccagc atgttctatt ccccttcaca 40080tgtcttccta ctcttcttta aaaagtctaa cgaaaggaaa tctgaaatgg ctattctccc 40140aattcaatca gcaggaagac cctgtcacat gtcagtgggt gtttgctcct tcagggaaca 40200tagagaggtg attcattgcc cacatgttga agggactcat ctccctggtt tgtcacattg 40260aactcttccc tcagcgaaag catttgcatt gcttccccaa gtcactgcca gctccatgct 40320atgcttctcc agatgctttc tctcaagatc aagccacaga gtgggcataa cacatctgga 40380atgcatcctt cccaaccgtc acgatctagg ggagatctta catgcttttt tgaaccatgt 40440atgatttcgg gatgaacaaa gcttgagaag caatttttaa attcctagtt atttaaatac 40500tgactagaga acggaggtga tgcgagaata ggtaagaaat gagggttgtg atatggagaa 40560aaggatgaaa agtttatttt ttgaggtact gggataagga gtagaagggg ctctatttat 40620cttccctcct cttgatatat attttctggt gtatatgtct tattttgggt ttataataga 40680acttcaaaac agaagtttaa ggccactgtt gaagtcttgc tagcttagcc tcaatataga 40740attagaaaca cacagacaca atttatacaa attccttagg acccagagag ttcaggttta 40800aaagcattct tccatgcagc agcagcagat gtatcagtat gtcttacttg agggaactgt 40860gagcaaagcc atgcaagagt tgtctgctat atacccagtt agctattgtg tatagggatg 40920ggggtgaggg gggagtactt tttttttttt aaaaacatac tgtttaaaaa tagatcatgt 40980tttttttcct tgagttgctt gttagtttgt gacccaagaa acaatactaa aaggacagta 41040tgtgtttaaa atctaacttt tgtgaatgac ctggattcta gtctagactt aacatgtgat 41100ggctgtgtga ccttagtcaa ggcacttgac cccttctagc ctcagtttct tcatctgtta 41160aatagagata atatgtcagc gggatctgag gataaattgt aataatgcag cctaagccca 41220tagtgagcac agtgcctagc acatagtaaa tatttgacac acagcacata gtaaacagtg 41280ttagacactg ttactatttt atgattatgt gaacttaggg aagaggaacc atactaagtg 41340cctcagttca gagctgaaat gagtgtcagg tggcattact ggaagtagga tggtagtaag 41400caccatgaga tcagtcacta tcttagctcc catctgacta cttacgccca tgcctaaatt 41460catttatcaa taaattgttc tcagattcta gaagctattg acaagaccaa gcaggatcat 41520acagaaaatg ttatagtaaa cttgtgcgag atgacctagt ctagcaaaca tgcacatgat 41580cttagctttc tataaggaag ataaagtttt tagagacaag atgaaggaaa aaaataagtc 41640tacaaagcaa gatagaccct tgaagatgta atggtgcaaa actacttttt aatgtatgtt 41700atcttaatac ccaatggtct aagtcagttg ttggctaaat ttttcatctg ggaattttaa 41760tggtgatatg agaagggcgg ggaaagcaaa gtagatgatg ttcattaaat gtttgctggg 41820tgagtgaaga agaggaaatg atgtgcttta ctttagtgag catcacaatc acctgtcatg 41880ctgatgaaaa ttatgggtac ctctattcac aatagcaaag acttggaacc aacccaaatg 41940tccatcaata atagaagact ggataaagaa aatttcacaa atatacacca tggaatacta 42000tgcagccata aaaaaagatg agttcatgtc ctttgaaggg acatggatga agctggaaac 42060aattattctc agcaaaatat cacaagaaca gaaaaccaaa caccgcatgt tctcactcat 42120aagtgggagt tgaacaatga gaacacaagg acacagggag gggaacatta cacactgggg 42180cctgttgggg ggtgggggcc tggaagaggg atagcgtttg gagaaatacc taatgtaaat 42240gacgagctga tgggtgcagc aaagcaacat gatacatgta tacctatgta acaaacctgc 42300acgttgtgca catgtaccct agaacttaaa gtataataat aataataaaa attacaggtg 42360cctgggacta caccaagaaa ctactttggt aagtctgaaa aagggatcca aacatctata 42420tttttaacaa aattctgaaa ggaggcttaa gagcaattgc tctagatggt ctcttctgtc 42480attgtttaga aaccaaaatg tcttgactct tatttaatct caattcctat tatgactagt 42540tatcatctgt gttgtataga cttataagga tgccagatta atacaaatct ccaaaatatg 42600tttaaagtag gtcactactt tagaaccagc agatcacagt ctctcaggga cttatttaag 42660gtacagctct aagggcatca ctataccgaa tctcaacttg tggagatggg gtgtgagaac 42720cctaatgttt ccaactagca tctacataaa agttatttat gtagaagtat caggactgct 42780taggccataa tgtttataga gtgtgtgtgt gtgtgtgcat gtgtgtgtgt gtattttaat 42840gtttgaaaag aagtggagtg ttcctatcct aacttactaa tttcatttat tcaaacagca 42900acatgagtga ctgttgagat ttatggatgt agataatcac agcctttctc tatacatggt 42960ggggcagagc atcttgaagg aagtcgggaa tactaggttt ttttaatacc agctccacta 43020ttacctggct gtgtggcctt ggacaaatca ctgaatcttt ctggcattca atttcctaat 43080ttttcaaatg aggaacatgg acatgatcaa tagttattaa tctagggctg acaaacttca 43140aatgcttaca catgaagctc agaattgtgc tgccttcata tgccttatgt tcatttctga 43200ggagagtgtg tagcttttat catttctcag agaaatctaa gaccccaaag aagataagaa 43260ctactaaatt agagaaactt ggaggccttt ttcagctttg aaattatttt attccataat 43320gatgtttttt tcaggttgca cctcttggaa taattcaata ataactcgac tgaaggaggt 43380tatataggat gagaggaatt tgctttgcag acataacctt gctacatttt atgtgaggga 43440aagaaagtgg cctgacagtc catttttgcc tgtgctcaca gttattcctt caatggaaat 43500cattcctttg cagcccctac cagtttcatc tttgcattag gtggcaacct gcactgtgcc 43560cacacacttg aagttttttt gcttgaagtg attctcatgt cacagaagcc catgatttat 43620gactttaatg tcaaattttt tttttaattt cttcttgaac actttgtact gcccccacgg 43680tccccgtggt ttttattcga aatgcttctg gcctgtttac taagcataat agttcatcta 43740tttacagaac attttgcttt gtttctaaaa aaatgttgca acatttgttt taaatggcaa 43800actcagactt acctaaagaa agggtcaagt atctgggaag aggatgaatc aagcttttat 43860aacttgtatc tatgggatag ttaggttgca gattttagta gttgtaattg ttcatatagg 43920aaggaaatcc agatattgaa aaagttcaaa ctagttttct ctgtgtattc tccctactct 43980tctctttgtt tgtgacgact ttgtttcttc atttgaggca atttaactca atatgatttt 44040gttgctaaaa agaaaaaatg ttttgtcttt aactttctat tgggtaatga cttgtcttcc 44100tcaaaatgtt tcttaaggtc ccatctccat ctcgtaaagg tgaaattact ctgtcttttc 44160aaggtcagtc tacaaattag taagatagca ttagaacaaa ggaatctatt gtcttcctgg 44220tcttgtgata ggcacttatt tcaagttggc tatatttata catgattcat ggagattatc 44280gtaatgagag attataaaaa tatcctcaga taacacattt tcacactttg aatagacagt 44340tttgattaat tctgattatt tttcttcttc aaatattatt gttctcaagt agaattacca 44400ctgcattaaa gtgaagaatt ggagtctctg aatgtcccca tttctggatg aattctaagc 44460ttttcttcag atgacctagg tatatcatta atagtgaaga tttcacataa ttttgtttga 44520ccttcgaaaa ataatagttt gaaagatagt tttcaaattt tattttatta ggttgcaata 44580tttagcattt taatttctgc tcctgtggtc agcagaaaat tctttctcta tttaaagcta 44640aatttaggac attttgtatt ttaaaaaaag gactattaga aataatattt attaaaactc 44700tagatagatt gctatattat caagcatatg agtcttattg ttaggaaaac attgttcttc 44760ctttatagaa taaactactt ggataatatg tcttctgctc attgaatttt tatttttatc 44820ttttgtagag atgtgggtcc cattatgttg cccaggctgg tctcaaactc ctggccccaa 44880gagatcctcc cacctctgcc tctcaaaatt ctgggattac aggtgtaagc caccatgtgc 44940agcctgctcc ttggtttttt agaacaatag ttatgagtac acagacacca tgcagaaaca 45000ctttgcatgg attgtctcct ttacttttca taaccttaca agataactat ttttagccca 45060gtatattggt tagaaagctg agatctagtg gtatttagta atctgattat catcacttgg 45120gaaatagttg agtagaggtt tgacccctta actacctatc attgaggcac agtaatgcag 45180ttcataattt ataatgcctt cccagtagtt taagatgtaa gtttcaacta caattaatga 45240attatcattg taacctactg aattttattt cctgaatagt agtcacctta tcttactgtg 45300attaggaagc tatcatattg tagaagctca gttagttact taattggaat catcatcata 45360aggtgtgtca tcatcataag gtatgtcatc atcatgaggt attatacact ccctaaatat 45420tgtagtttat aatctcttct atttgaaaga aaaaattgtt atctatcttt atttagatat 45480tctttactga ggaacgatga ccttgaaaag cctcctgaag ttgtcatctt aatttttttt 45540tgagacaggg tgtctatcac ccagtctggg gtacagtggt gcaatcatgg ctcactggag 45600cctcaaactc ctgggctcaa gtgatcctcc tgcctcagcc tctctagtag ctgggactcc 45660agagtagttg ccaccatacc tgactaattt tttaaatttt tttcttgaga tggactctca 45720ctttgttgcc aagtctggtc tcaaactcct ggctttaagc aatcctctca ccttggcttc 45780ttaaagcagt aggattacag gtatgaacca ctgtgccctg ccaatttatt tttaatcact 45840aattgagtta ccattttaaa aaagtccagt tacatttgaa aactataggt aatacacctg 45900gttggttaga tttttagtca atattagtgg tagataattt

tttaggagaa acaaagaagc 45960aaaaagcttt tttggggggt gttatttctg aaggttgaac aactaacata gtaagctact 46020gtacctgagt gaacgatgat agaagtgtag agcttttttc ctctacagcc atgattatta 46080aactttgcta ggtctgaaaa tcatagactc tttccccaga aaagttgaca taggcacagc 46140attttgtaaa agtgtcaagg gagtcataga taactaatga attttgattc ctgccttgga 46200cttgggagag acagcataac aagtaagggt cttgcaaaga tgtccaaggt ggccactgat 46260ctcatgagat ttagggacct cagtgaccct tgaaccatga gatccaaagg tttttagatc 46320agtgatctag agaggaagtt aagtgttctg actttgagtt atttttcttc tgagtccaaa 46380tacctgcagg gtgtggtatg cagccagtgc cagtttggct tgcaaagact tcttgttggc 46440caagtctcca gcatttccaa atgctgttct cccatggatg tttttccagt gtcactggac 46500atggtcaatc attaagacat ccaaagaaga cagtaaatgg ttcccaccta gcacagttag 46560ctcatcatcc agtaacttgt ttttgcagtt ttttatttta caatctggca tagtattgac 46620atccttgcac tccaaaccag agctcattag aatgagttgg cctttggtaa aattttccgt 46680caattaattt ttttttgcca tgaatttctt tagtagaaca gatttgtttt ttaaaatacg 46740tagccttggt tacttatcca cttaacccta ccaataggga atactccctg gactctagac 46800ctggttcctg cttttctagt ttccagtttg catgtaaaca tttaatgtat attcaaattc 46860ctagtggtct tctaatcttt tgtctgtctc tgttttctct ttgctccaaa ctgtcttccc 46920atactaatct tggtatttta ctaccaggtt ccttctctgc tgaaacacat tcagtgttgc 46980atcaccactt acgtaagtaa gtcaaaccag actgaattta tagaaattat ctagtttgcc 47040ctctttatct tacagacaag gagtagaact tccacagaga tgatgctatg tgtctaagca 47100gaagcaactt gctgtggagt tctgatggta gaactgttcc tactgtacct tactacctac 47160catttgacct ccctagctca acatttggag ggtctctctc aaaggcagtt ttcctggaat 47220atttcttatc tcttaactgg tctccctaac acattatgag ctttcccatt tctttgcctt 47280tgcataagac ccttcctcta tctagactgt cccactccct cgcatttagt attaatagaa 47340attcatactc attcacatac catgcctaac taagcactta gtattttata tgagttattt 47400tatccttata aatccatcag ctagattcta tatgattccc atgtgacaca tgtaaaaatt 47460taggctgaag tagatgaagt gacttgccca agacttctga gttagtaggt ttcagggcca 47520ggatccaacc tgagtggtct gtctccagaa cctgtgctat taatccatac attaaattca 47580ctgcctctaa ggagtatttt ccaagttgca ggccataggc tccctctact ctgtgaaact 47640ttccacagtg atatatatca gagaagcaga atttatttca tggttatctg tgctttctgg 47700aaggtgagtc aatatactac ggggttgaat atttcctacc caaaatgctt gggaccaaaa 47760gtgtttcaga tttcattttt ttttttcaga ttttagaata tttgtggttt tttcttttta 47820atatactttc aactattatt tttgattcag cagctacatt tgaaggttta ttacttgggt 47880atattgtatg atgctgaggt ttggggtatg attgatccca tcacccaggt agtgaacatg 47940gtacccaata ggtagttttt ttaaccctgt cttcctgctt cctatctagt tgtccctggt 48000atctattgtt gccatcttta tttccatgag tatgcagtgt ttagtcccca cttgtaagtg 48060agaacatgca atatttggtt ttctgttcct gtgttaattc actgacaatg atggcctcca 48120gctgcatcca tattcctgcg aaagacatga ttttattgtt tttttttgtg gctgcatagt 48180attccatggt gtatatgtat cacattgtct ttattcagtc cattgttgat aggcacatag 48240gtcgattcca tgtctttcta ttgcgaatag tactgtgatg aacgtacaaa tgcatgggtt 48300tatttggtag aaggacttat tttggatata cacccagcaa tgggattgct gggttgaatg 48360gtagttctga attctttgag aaatccccaa atttctttcc acagtggctg aacagtgtat 48420atgcattccc ttttctctgc agccacacca gcatctgttg ttttttgaca tttttttaaa 48480ttatacttta agttctgggg tacaagtgca gaacgtgcag tttgttacat aggtatacac 48540gtgccatggt ggtttgctgc agccatcaac ccctcaccta cattaggtat ttctcctagt 48600gctatccttc gcctagcccc ccaataccca acaggccctg gtgtgtgatg tttccctccc 48660tgtgtccatg tgttctcatt gttcaactcc cacttatgag tgagaacatg tggtgtttgg 48720ttttctgttg ttgtgttagt ttactgagaa tgattgtttc cagcttcatc catgtctctg 48780caaaggacat gaactcatcc tttttatggc tgcatattat tccatgctgt atatgtgcca 48840cattttcttt atccagtcta tcactgatgg acatttgggt tggttccaag tctttgctat 48900tgtgaatagt gccgcaataa acatacgtgt gaatgtgtct ttatagtaga atgatttata 48960atcctttgtg tatataccca gtaatgggat cgctgggtca aatggtattt ctggttctag 49020atccttgagg aatcgctaca ctatcttcca caatggttga actaatttac actcccacca 49080acatgacagc cattctaacc cctatgagat ggtgtcttat tgtggttttg atttgcattt 49140ctctgttgat tagtgaggtt aagcaatttt tcatatgttt cttggcccct tgcatatctt 49200cttttgagaa atgtctgtgt cttttgctta cttgttgttt atggggttat tttttgcttg 49260ttgaattgtt taagttcctt atagattctg gatattagat ctttgttgga tgcatagttt 49320acaaatattt tcccccattc tgtaggttgt ctatttactc tgttagtagt ttcttttgct 49380gtgtagtagc tctttagttt aattagctcc caattgtcat tttttgtttc tgttggaatt 49440gcttttgagg acttagtcat aaattattcc ccaaggccaa tgtccagaat ggtgtttact 49500aagttttctt ccagcatttt tacagtgtgg ggccttacat tttaatcttt aattcctttt 49560gagttaattt ttgtatgtgt tgaatagtag gggtctagtt ccattcttct gcatatagct 49620agccagttat cccagcacca tttattgaat agggagtcct ttccctattg tgtatgtttg 49680ttgactctgt caaagatgac atggcttgta tgcatgtagc tttattttag atttcagatt 49740agagatgttc aatctgtata aatgcttaga aagtaatatt aacacaataa tttattaagc 49800gaccacaaat atctacaact tgactttcta agttctcttc caaatgtaat tagaaggaaa 49860ccaatgaaaa ccaaggacgt aataatggga tactgcgcta taggtagatg tgctgagccc 49920cagcacacaa agccagggtt tgcatcctac ctgcaccagt tactcctaat gcgtgtacct 49980tggggatacc ggttaacctc tctgagtttc catgacctca tttgtaaaat aagcttaata 50040taatctctca tggggttgtg gtagagactg tatatgaaaa cttgttttaa attataaagc 50100acgtttcaaa aacagtatta tttttcagtc tcaccagaaa ttgaatatca gtaaatcagc 50160ttaacgaata aattacttta ttattataca ctgccctacc accagtggta gaatgtagat 50220ttcagtgttt ttatcaggga acttcctata agccttcttt gaaccaatat ttgtatataa 50280attattcatg aaatggaatt ggtcaccata ctctttacat tttctttgct ttttagacat 50340atactcagat cccacacttt ctaaatacta atctagttga ttgtattttt gtatttttaa 50400aataatataa attatgctta agaaagggct ggattgttaa gcagcttgtg ttcagaatac 50460ctgccacttc ttgctgcgtt tttctatctg gaatgccttt aatttacctg gtttattcaa 50520aatgagatag aatattgctg cttacttatc gctttcacat ctgcaacagg tctgctctgt 50580ctttgtcact agggtttaca taaaatgact agacgctttt tcctttcaaa gatgaggatg 50640aagattaaga agagaaaaga aattatggaa gcttggtgga aagtttgaga ggaagaaagt 50700taagtgtaaa gaataaggga gttaacgcca aaaataaatg aacaaaaatg ccgtcatgtc 50760tggcagctca ggtgtgtttt caataagcag caatggaatg gacacggtgg ctcatgactg 50820taatcccagc attttgggag gccaaggcag gcagatcact tgaggccagg agtttgagac 50880cagactggcc aacatagcaa aaccccgtct ctgctaaaaa tacaaaaatt agccaggtgt 50940agtggtgcgt gcctgtaatc ccagaggctg acacatgcga ggattcctta agcccaggag 51000gtggaggttg gagtgagcca ggattgtgct actgcactcc agcctgggtg acagagtgag 51060actctgtacc aaaaaaagaa aaaaaaagca gcaatgattc taatatttta tttagggtgg 51120tatgaagaag gttgatggtg aagattcaca agtggaaaat acccagaaca agtgcccaga 51180gggtgtagag tgtgaaaata tgtagtggag ctaagagaca agggaggctc ttctctccag 51240ggacctgagt tctagtctag acttaacatg tgatggctgt gtgaccttag tcaaggcact 51300tgaccccttc tagcctcagt ttcttcatct gttaaataga ggtaatatgt cagcgggatc 51360tgaggataaa ttgtaataat gcagcccaag cccatagtga gcacagtgcc tagcacatag 51420taaatatttg atacgtgtta gacactgtta ctattttatg attatgtgaa cttagggaag 51480aggaaccata ctaagtgcct cagttcagag ctgaaatgag tgtcaggtgg cattactgga 51540agtaggatgg tagtaagcac catgagatca gttactatct tagctcccat gtgactactt 51600atgcccatgc ctaaattcat ttatcaataa attgttctca gattctagaa gctattgaca 51660agaccaagca gaatcataca gaaaatgtta tggtaaactt gtgcgagatg acctagtcta 51720gcaaacatgc acatgatctt agctttctat aaggaagata aagtttttag agacaaggtg 51780aaggaaaaaa taagtctaca aagcaagata gacccttgaa gatgtaatgg tgcaaaacta 51840ctttttaata tatgttatct taatacccaa tggtctaagt cagttgttgg ctaaattatt 51900catctgggaa ttttaatggt gacataagag aaggttgcgg acagcaaagt tggtacagtg 51960tgtaggaagg aagagaacct agaactctgt aacttactag ggttatatgc ctccaaagtc 52020agttgccggt atagctctca gtaaagaaca tggattgagt gtttataatg tgtttggcac 52080attcatctgt cttaattcta aatgaccatt tgacataggt atcttgatct tcattgtata 52140aatgagagga accaacatac agagtttaga tttgtcccag attcctttgt cctaaaccta 52200ttgcttattc acttatgcca gaatgactga tgtaagtaag tcagacgtta tttaaagcct 52260tcaaaagaac ctaggcaaat atttgagact tctcaaatgt tcagccgaag gtaagagaat 52320taagttacat cacaaaatga atctttctaa attatttctt ataaagcatt tgcatctcat 52380aagtagcatt gtttttttga agtttcactg atcatttttt aggttcaaca taaaatagtt 52440ctcaaatttg ggtgcacatc ggaatcacct ggaggacttg ttaagtaaga ttcagtagct 52500ctggtatggc ctgagaatat gcatttctga cagattccct attgctggca catcactttg 52560aggtccacat tttgagaacc actgacataa aaagcatgta ttttatgatt aatcctatgg 52620aggaacaagt aggaaaatag tccttgaggg gaattccaag atcacaggtg gaagctgaaa 52680ttcagatcat gtttccaaaa ctcagtaggt tatacctagc caggcataac tgaatttgga 52740gtctaaaaga tctgtattat cactttttta ttttgaagga tgccttttga ttacagaggg 52800aaatcaagga ttaaaaatca atatacatgt aaatattgaa attcattggt aactttaaaa 52860agcacaacag ttttgtgtgc ttttctccaa agcactacaa atatgattaa ttgatgtata 52920agaattttct tatggaattt ttttttttgt ctctgtaggt ggcaaatctt ccaaaagact 52980acatgataac cctcaaatat gtccccggga tggatgtttt ggtatgtaaa ctacatttct 53040gagtttcatt ttagtagctc atagaagaaa tgggatcatt catattgaga tagtacacta 53100gctgctattt aggagcttgc ttattgtcag gatttgaaga atttatcttt ggaatttgac 53160ttgcaggctt ttttttcccc ctcttaagca actgttaaac tttatcagtg atcatttcag 53220cacatatatc ctagacctgt atagttgttc ttcactatcc ttaggaaatt ggttccagga 53280caccttccga ccctccatcc tccccactca ctccaccatc aggataccca aatcctagga 53340tgctcaagtc atttctatga atggcatagt atttgcatat aacctatgca cattcttctg 53400tatactttat ttttaaattt ttatttttca ccatggaagg cacactggaa ctcctgtgtg 53460ttccaggagt ttagttccag gaatttggaa ctaaattaaa ttatttctaa attaattcta 53520aattaattct aatactgaat ataatgccta tattacataa atcattgtta tgctctgctt 53580ttaaaatttg ctttatgttt tcttattgtt tgaatatttt ccatctttgg ttggttgaat 53640ccgtggatac agagggccat ctgtacttcc aaagtaaata cgattatctt ctttagacct 53700agccacagat tttctcattt tccttaagca tccccctaaa aaaggacttc agtgtaagtt 53760tatgtaaatt ctttttcata gctgacctac ctcacttgtt taatttgaaa gcacaatttg 53820atttactagc agtgcaatag attcttagaa ccttacaaca agtgtatgtg tttactatta 53880taagtactac tctgttgctg aaaccaagaa attttaatta tttttctcat tacacagtat 53940cagaattcat acattaagaa atatttcaag catagagaat aatgtaatat ctgtatacct 54000atcacagagc ttatcaaacc ttaccattgt gcaatatttg cttcaaatat atttttaata 54060atgaaacatt acaaatgtaa ctaatgccct atgtatgctc cttctctgtc ttcattaatc 54120cccagtttgt ctcttagtgg taacacttat tctgagctgg actttttcat tccaatgtgt 54180gctttcaggg cttgtaatgt ggacagcatt cactagtgtt atgtagagca ggaggagcat 54240tctcttatat gtttactgca gtgtatgcac ccatacataa gacatggaac tttttgaata 54300tttaataatt ttatgttaag cttctatact ttgaagttcg cttatatctt tcaacatgct 54360ttgacaagtt gttttatgtt gactcatttc aactactgta ctgtgtttct tgagtaaatt 54420taaaaccatt tgagttactc tcttctctac aactctcatc tttccataat ctaccaaaac 54480ttattaaaca tctactttgt gtaggttttg accaattttt gtggatcagt ccatcaaagt 54540tgttaccaat ttaaacacat cattatgtgc ttgtgcttaa aatgcatttc aaagactaca 54600aggctgtcat ttcaagagaa tgtcaaagct acatttcttt ttattagtaa gcattttagc 54660atgaactacc taaagttttt tttagaattc cttagaacat gtatcttcat gttgaaccat 54720ttatcctcaa gcctatgtca atgtctatcc tattgagtta gggggagttc tctggaaata 54780gaaatggaaa attattagca gcctgaaagt taattaattt tattatttgg ttactaagca 54840agtatttagt taggaaaaac taaccaggag ccagcacttt ggcagacact tgaagaaaca 54900aacacacacc aattaggcaa tgatctttag ttcaaaaagt ttagtaattg ggaaggcagg 54960caggggggat tatgtggcct ttttataaca cttttccatt tgcaaaatga tttcacatat 55020taattttctt ggctgattag aaaaactctg aggaaggtaa aatagctatt atgattccta 55080ttttacaaat gatatattga gctttacagc gcaaaagtat tttgcccaag gctatacaat 55140tagaacgtga cagagtctta aaaagaaacc cgtaggtcat cagactctta accctgggtt 55200acggaaagtg aggatctata taattacaac acttagggaa agtgtttgaa gggaagcctc 55260taggtaataa agacccaccc cagatccttc ctgactgcag cagcatttag gattgtgatt 55320gccaagcacc caaagataac ctgctgctct ttgaaaaaca aatgctaaac tgaaaacaac 55380taaatagaat tttttgtgct ggtataatca tagaatatgc taaatcatag atttcaatgt 55440cggcttgtga tagcaagagc caagcttgtg aaaataaagc tgattcactg ccacatattt 55500tgtgcttgtt tctgtaattg agaaaaatct actattgatt ctggcatgct atactggcac 55560aactggagaa acactgttct tcttggccac caccatgggc aagaaaaaca aggccacgtt 55620tagagcattg gacagaacca ggggtctcac cctttgtttt accttcattt tgacatgttc 55680gtttctgcga agaccagagg agtttcccag tggggaggcc tctgattatt tgcgagtggt 55740cagagtccca gctgccttat cttttgtctt taaacaatag agggtggagt gggaatgggt 55800gccaaaaaga ttctctggtg ccaaagcaat aaaattctgc ccatgggcac tgggcagaag 55860aaagctctag aatagaatac acttttattt ttatttttat tttgccttca ttttaaaaaa 55920cagctaaatt ctcctagaaa agggagttca cttttccaag gtccttctta gaaaattacc 55980tagcagattc tgtctcacaa atagcagggt gtgatcaacc acttagtatt taaatctaac 56040gataatcacc aatgctattt taaactcttt gaaactatta tattgccgga ctgttgaaat 56100ttcctttaac ttcctaacct taaagaatac ctggatattt ctggtctttt acatttactg 56160gtaatgttat tgtagaaaga ataaaaatgg cattgctgaa ttgttgccac acaccttcat 56220tcacttgaac ttccatagca tttacataaa ctgtgggttg cttgtaatat ttgagtttaa 56280aaatcttcat ttttaaaaaa atttttttat ttttagctat tatggataca taagagttgt 56340gtatatttat ggggtacacg tgatattttg atgcaaccat acagtgtgta atgatcaaat 56400cagagtaact ggaatattca tcagctcaac tttacaattt ctttgtgtta ggaacattcc 56460aattccactc ttttagttat tttgaaacag acatttttat taactgtgcc acctattttg 56520ctactgaaca ctggatctta ttcgtttttc aatttttact taaagtatat attcatcagt 56580tttcttttag ttttttattt gcaaataatt ttaaatttac aaaaacgttg caagaatatt 56640taaagaacac ccatgtgccg tttaatccag attcacctat ttcaagtatt tttaacattc 56700tgcctcattt gctttatcag ttgctttctt tctatgtatg catgtgtgtt ttattatgtt 56760tctgaacttc ttgaataagt tgtatactga tgcatacatg gccatttact cttaaatact 56820tcagtgtgta tttcctatgt gaaggatatt ttcttaccta aacacagtac atttgcaact 56880tcaacacatt ttacaataat tatagtactt taatctaggt ctgcatttca atttagcttt 56940tttgtttttg tttttgtttt ttgttttttg catttctttt ccttcagtat aagatcctat 57000ctaggattgg aattttcatt ttgataccat gtaaataatg actcccaaag atgccttgtt 57060ttgggaatat attacctgac atggcaaaaa ggactttgca gatgtaatta aattaaggat 57120ttatagatgg aaaagttatt ttggattatc caggtggtcc cactctgatt tacatgagtt 57180cttataaaag ggaggcagga gtgtcagagc cagagaggga gatttgaaaa tattacactg 57240ctggcttcaa agatggaaga gggattcaca agccaaggaa cgcacatggc ctctgaaagc 57300tggaaaaggc aaagaaaatg atttttcctt tacagcatcc aaaagaaaca cagccctaat 57360gaataccttg gtattagcct attcagacct attttaaact tctgacctcc agaactgtaa 57420aataacaaat ttgtgctgct ttaagccacc aagtttgtag tactttgtta tagcagcaat 57480aataagctac tacatcctgt ctctctagtc tcctttaaga tttcctcagg ctttctttgt 57540ctttatggca ctgacatttt tgaatgagac agtcctcccc tttaagaaaa gtgtttctct 57600ttgcaacatt tgattgatgt ttcttaatgc ttctgtggag cttatgcaat tcaagtgaca 57660atgctacgaa agtgatgatt tgtgctcagg taaccacatc cagaggcaca cagagtccat 57720cctctcctca tggtgctgtt tgttttgatc atctggttaa gatgttcaac tgtttatgta 57780tatttactca ttcacaattt ggggagagac actttaaggc catataagca atccttctcc 57840ttctcaaagt ttctcccaca gatttagcat ctattgatga tacttacctg aaccaatctt 57900tgctatcatt gtcagttttt cgttcagact ttttaaaggc acagctaaat atggaagtga 57960ttggcagcat gtgatgccac tctaacaata agccttccaa acttcttccc tcttcagtac 58020ccttcctcag gtatgcacaa tttcacttct tggctattct catctttttt tgcctggccc 58080tgttacttac ctctgggctc tttccatttt ctttgatacc tgaggattta ttatttgatg 58140ttttgattct tcctgaatag acagagtgat ttaaaaagaa atacatccaa tcatttcatt 58200cctgtttctt tatattcatt gaatggcttt ttgctgctct taggaaacag gaaaaggagc 58260tgcttcaaca aggcctccaa gactgtagtg atctagctgg gcacagtggc tcacgcctgt 58320aatctcagca ttttgggagg ctgagacagg tggatcatga ggtcaggaga tcgagaccat 58380cctggttaac acagtgaaac cccgtctata ctaaaaatac agaaaattag ctgggcgtgg 58440tggcgggcgc ctgtagtccc agctactcag gaggctgaga caggagaatg gcgtgaacct 58500gggaggtgga gttttcagtg agccaagatt gcaccactgc actccaggct gggtgacaga 58560gcgagactcc gtctcaaaaa aaaaaaaaaa aaaaaaaaga ctatactgat ctggcacctt 58620tccacatcct ttttcagtcc tttatcgtta tcgttaccag ttccttgctg ccacatagct 58680tcttgcctgg ctttctctat tcctggaatc ctgattggtt cttttgtcta gctaacacct 58740gattctttta gattttggtt ttgatttttg atgaaaccaa attgttctac tggaaactct 58800tctcatcgca ctcctcagag tagcagtttt ggatgtattt gtgggatgat ttaatagata 58860atcttccacc aacctgtaaa cttcacaaag gtaggaacta tattcttttt gctctgcatt 58920agattctcag tgcctcatgg atgctcatat gcatttaatg aatatttgct gaaagaaagt 58980gaagaaagga aggaggaaga aaatttccct accatataat catttgtaat ttccttgagg 59040cacagatttg aacttagttc acaaggttat aatgatgagg tgagtgccta taactcgtca 59100cacagtatct gcactgcaac gttattatct tcttttagat acgtgtgaat tttttgaaat 59160attttaggtg taagaattgt attatgcaat tttggggaat cacagcttct ttaagccttt 59220ggttctactg ctttaattga gttatttttg cattcaagaa cctgcttatt gttcccattt 59280actaaaggct gagagccaga ttcctgaacc taatgttcat gactctctgt aatccttccc 59340taccctaacc aactcatctg aatacccatt attccccaac acaaacttgc cacctcagcc 59400acatgcatca ctgcctccta aaataatgcg cctgttctca aaggatgccc tttcttctcc 59460catccactta tccaaattct acccatagaa tcatgcaata tcttggggct taacccatct 59520tagtggtcat ctcctccaag gtcagtttaa gatttctctt tcacacaagg acattttctg 59580ctctttcagc ccacattttc tctcccatgt atcatttgat caggtgctac tctgtgctga 59640gtgttttgct tgatgctgga gcttggtggt cagcaatact atcattgcac ctgctatgtg 59700gaccctatct tttagtttat tatcttgtga cttctatgga atttgtccag aagctattca 59760tcttgggtac atgatcatat attgcttaca tagagcttaa tattttaagt gttatatatt 59820tgtctcaaag tattatcgtg agaaataatt aaaatacaaa aaatatgagt tgtggtgtgc 59880atataataac atcagatacc atcattatta ttttgttgtt gtgttctctt tttactatat 59940tttgactatc tacagagcaa ggttgaaatc ttttactcct gtgtatctcc tcaccattct 60000ttagacaatc caatgcatat agtcgctcct caaaaaatag ttatgaataa acatttaact 60060cttggcaaaa atgctagatt tccaaatctt gaaggagccc atcttattag tttagattcc 60120tgtggttttc actaatagta aaagtcccat tattataagg acactaaaga atagcatgtc 60180acctaagggc atgggtgcaa caggccccat gaaaagactg gaaccaggat tggaaactat 60240cagtttttcc ttctccatct ctgaatctct gtttctccgc ctcattctat ttgtgcagat 60300attttctcct ttgttccagt cacttggtag aatgtaactg cccaagagct gccaggttta 60360catatttcaa tttcagccct attcagagat taattatcag tttctaatga tcaataccaa 60420ttttggggag gaacgttctg gttggcctat ccaagtcagg tgtctaccat catccaacca 60480actatggcta cagggaaagt tcaccgtttg taagcatgga ttctgaggct ccactcctat 60540gatgcggcat tctctctata aaaatagacg tcagccaaat agacacttgt agtcaactat 60600ttatctgcct cgatctccca tgtaataaga aagtatttat ttaaaaggaa taagcataaa 60660atgttagttg aatgaataaa atatgacatt aaatgctatt ttaagtagtt cttttgtata 60720ctgtactaca gcacaggggc ccgtcttcct aataagtgat tatattaaga tgctgttcag 60780attatctagg ccattattct ctgatagaaa tacaatggaa gccacgttca taatcttaat 60840tgtctggtag caacactaaa aaaaaaataa ataaaaacga gcagctgaaa ttaagtataa 60900tgaaatttat ttagccccat gtatccaaaa tattgtcatt tcaccttaaa atcaatataa 60960aaattattaa tggaatgttt tatatgtttc atattattca

aaatccacag tgtgttttac 61020atctgcaaca tattacagtt catacactaa attttcatta gaaatagtta ttccatattt 61080agatttttta ataaaattta ccattgagaa gatagtcaca tttctgtgtt ttccaaacat 61140acttaaaagt tccagcctct aaatagggtg atattattga cttttcaatt taaagaaatt 61200ataattaagt aaaatgaaaa ttttagctct gcagtcacat tcaatgcatt ttgagtgaga 61260ggagctgcag atggatagtg ggattcccat attggccagc atcgattaga atttatttta 61320tagatggaga aactgaagcc ctgaatagat taaatgactt gcctgaaatc acaaagtgaa 61380ttgacagagt tgggactaaa aactgggcct ctagttcatc gttgctgccc tcactcccca 61440aacccctgtg ctgttccttc actgttctat tttagggcgg gcctaggggc catttatagg 61500tctgccctgg gaaataacac atatttagag tgctttggtt gattaccatc ttaatcagat 61560tgagatttct acggcacttt aattcaaaca agaaatgtaa agttagatgc ctaaagtgaa 61620cactgaagac ccaaattaat tgtttaggga gaggactaaa ataaatcatt acaaaaccca 61680gctaatccaa agagaagtta agatttcgga gactacactg gcaacatgaa catgcaatat 61740tttgcaaagt gcttttatag gaaactttta aatttgcctg cccatggaat gtttgagagt 61800agaatgtcag ctttctagcc acaacgaaaa aaaccaaaag aacagtaaat aaagtccaca 61860aatggcaatt gtcacaattc ctcagagaaa tttaccttca ttggtatgac tctggattaa 61920ctgaaatgaa acagaaagaa ccatttctat tccaaagaga tttcttaatt gagccgaaga 61980atcgtgtctg gtccagacgt gagaccacat gggtcatagt gaattcagca gaaaaaaaca 62040cttcagtttt atgtgctttc actattctca tggctactag aacaaaatga gtgatcgaga 62100gtcctggttt ctcgcgtttc tgtgtaaatt tgttatgtaa cctggaacga gacacttcac 62160tactctaagc cttgattttc ccgcttatac aattggtgag actggtcctc agtattcctt 62220ctaggtctgg aattctggtg attctgatga catggctcta gattttgatt gcacatccca 62280gaggtagatg ctgaaagcac aaacagaaag gtacacttcg tgccctaggc atgcatacgt 62340ctaattccag caagactgaa gtttgacacg aattaaagaa agtaaaacaa aaacaaactg 62400ggtggtaatg ctggtattta gattaatcag attagaaggc tgggtgaata ctccttatga 62460atattcttag aaacctcata gtttatgagc caggcttttc actctgagaa tctctaattg 62520gtatttcacc aactcctgtg aaaagaacgt tctcactggg tagatactag ctttaggccc 62580tataagaaag aacatagata tctctctgtg aatagctacc aaattgaaag taatgtaaag 62640tgcttcagtg aagaagtgtt acagtattat tttgtctact ttcagcttct taatccgtaa 62700atcttaccgt gactgctctt aaagactgtc tgtggggtgg gcgcggtggc tcactcctgt 62760aatcccagca ctttgggaaa ccgagatggg tggatcacct ggagtcagga cttcaagatt 62820agcctagcca acatggtgaa atcccatctc tactaaaaaa acaaaagtta gctgggcgtg 62880gtggcagatg cccgtaatcc caactactca ggaggctgaa gcaggagaat cgcttgaacc 62940caggaggcag aggttgcagt gagctgagac cacgccattg cactccagcg tgggcgagaa 63000gagcaagact gtctcaaaaa aaaaaaaaaa aaaaagactg tgcatctaat accaatgtgg 63060aataatgtgt caaattattt cacctaataa accagcacag tttcttgaat gaggcaggag 63120atagagagga gaaagcatgc cagaattagc aagtctccag gatgatttta aaacacatgg 63180tttatttcag ccggtttctc aaatttactg gttctggaac tacagagatt ctgtgtcaca 63240tattctgtgg aattaatata ttacctttta aaaatttatg cttagtctta gaatgtgcat 63300atatttctgt tgttggtctt tggaaacagt catattccta gttatgttta gaatactttt 63360aattttattc attttaaatt atatatttta tttaatcatt tagcgagatc atgatttggt 63420gttcatggta ctttaaaatt cctaaggaag ttcacagttt ttgactgaat tgtcagtggt 63480agtgagatgc tagcagtcca atatcattga gtcaatatca ttgagtcagt ttggattcta 63540gaataataac atgttataag tacataggga tagaaagcac atgtttttag tagtcctagc 63600tagagctgct ataattaacc aagaaaacat aagacagaag ctatcaagaa agaaggtaag 63660aaattgagag catggagtca agaaaaaagt ttttacactc aaagcagaat ctatatgttt 63720cttcttttgg aaacaataca tacaggcgaa taaagcatca aattacatgc acatcaggaa 63780tttaaaagca atcatttgtg tcacctttga catcctgtat gactgtcaat tttcttaatc 63840ctttatattt tctagtgaaa gagagaacat tatatcttac tttcttctaa tgaatatcat 63900tgagtaaata tttatataaa actatttaag tttaaaatat ctctgaaaaa taagaaacag 63960tgataatatc atttattaag gcaaacaaaa agtacgtttg taaatttgac aatctttaat 64020catttatttc ataatcataa tcgtggagga attgaaaaac agaaagacct gctatcacct 64080cacaaatgaa atacttagtt cattgtgttt tattctcctt gcaaatcatt gacgatattt 64140aattaaaaat tacaaaactg ctagacattc cagtcaaaac tagtttcctc tcttacaagc 64200aggcagatca aaactgactg acatttaata ttatttgttt accttcattt tatctttaat 64260aaactcaaga atcttaaaaa gccaatatat acatggttaa acaaattact gaaaaataga 64320actttgaatt tatgattcag ccttttccaa agaaacaata gcacaagcaa aaataaaaaa 64380tgagtgaaac atgattacaa gtgttttaga aatacacgat ttttcagctg tgtaggcacg 64440tagtttatat caactatcaa caaattgttc atgactttag aagggcaaac attagagtag 64500gattagattg tggaatttcc atggtttaac tctaagaaag cacaggattg gggttggaaa 64560ttttactgga aagagttgat tgtttagact gttgatttag ttatgtgaaa atgggaggag 64620gtagggtaat agctaaatag gaactagaat ataaaatgta aacatagagt tcggttgaaa 64680atagtgatca acagaagtgg cagtaagtac cccagaattc ctcagcttaa aaataagaaa 64740caaatcaaca aaacattttc tactttaacc acaaaaaaaa agcaaaatga acaaagtacc 64800cattcatata agacaactac cacagaaact aaaggaatgt cttaattcaa aacaaggaat 64860ttgattttgg ttcttacttg ccagagatcc taggcaagca tactcataaa gtgaggaaat 64920agagtcctaa gagattaata cctggaattt cattgttaga aaaatttgca caatttcaac 64980ggaatgaatt ggtgaaagct actctatcac ataaatacta ttcaaatagc aatttacagt 65040gtatgctgtg ttccatatcc taggaaatct gttcattgtt gatagggctg acggtctata 65100aggaattatc actccctaaa aggaaagaaa cttgcagagc aattcagcat ccaaggaaag 65160actgacagct ttgaaagaga cctgataatg atgcaagtag gaacttgcat gtgcttgaag 65220taagcactga tttaacaaaa caaagactct gttctgcaca tactttggaa acacatattc 65280tttgagaact tggttatgac gagttgaaat ggccacttga actaaattcc catgttgaga 65340tttgtttatc cttcttatct gaagccagag ccaacaatcc taaaagtttt gagtgacatt 65400taatggttcc aggaattggg gcttacctat ctgcagataa acctgcttcc tttttggctg 65460tattatatta agtaaaacag ttctgttaaa tgaatataaa gtggtccctt ctatacttta 65520gaataaataa atgacttgag ccagatgcag tgactcacgc ctctaatccc agcattttag 65580aatgctgagg agggaggatt gcttgaggcc aggagttcaa gagcagcctg agcagcaaag 65640caagatccca tttatataaa aaaattaaaa attagccaca cttggtagca tgcgccttgt 65700agtgccagct acttgggagg ctgaagtggg aagatcactt gggcccagga attcaacatt 65760gcagtgaacc atgattgcac ccctgcacta cagcccagga ctcagagaga gagcctgtct 65820ctgtaaaaaa ttaaagcata aataaataaa taaataaata aaaggcttga cttttcccac 65880agagatattc tgctggtaaa tagagaaata aagcttagta gtttgattta ttttccatga 65940catcagttac tctgagtaca atatttttgt tacaagttgt attaaattat ttgtgctttt 66000aatgagccaa ttcaatgtta ttcttaataa caatgttatt tttgtcactg cagcatttca 66060gcagtgttca aaatacatca gagtttccac ttaaaagatc tttataagag aaaagataac 66120acattaaaat catataaagc atgcaaactt acaacctcct cctcctgtta caagagtccc 66180tcctcctatt acaatagtcc ctcctcctcc tgtcacacta gtcccttctc ttcctgttac 66240aataacccct gtcctcctat tacaacattt taagtaatgt aatattaatt ttaaaaatct 66300ggccaggcac ggtggttcat gcttgtaatc ccagcacatt gggaagctga gacgggtgga 66360tcatttgagg tcaggaagtt tgagacagcc tggccaacat ggtgaaactt cctctctact 66420aaaaataaaa aagtagccag gcatggtggc aggcacttgt aatctgagct actcgagagg 66480ctgaggcagg agaatcactt gagtaactaa aacgatagct ttgaagagta ctccgagttt 66540tatggcactt acttattaaa atagctgttt tgtctctttt ttcatatctt gcagccaagt 66600cattgttgga taagcgagat ggtagtacaa ttgtcagaca gcttgactga tcttctggac 66660aagttttcaa atatttctga aggcttgagt aattattcca tcatagacaa acttgtgaat 66720atagtggatg accttgtgga gtgcgtgaaa gaaaactcat ctaaggtaac tttgtgttca 66780ttgggattat ttttcattac gcttctctaa aaacccatgc ttcttggtgc tgttggggaa 66840aatgaggcac ctttatttat gatattttga ttgtataaac ttcaaattta aaaatcttgt 66900tcagatgagc aaagaaaaca agtatttgca gttatactgc aatactgaag tgcacattca 66960aacttaaatg ttgtcatcta atacaaaaat aaaacttatt tacattacaa tgcaaaatca 67020gtcctgcctt tatctttatt cttcataagt gaagtccttg aaacctttct attaagttac 67080aagtgttcat ttaaatggaa aattgtctgt aataaacaca tatcctttta tgcataatgt 67140aatcacccca taggtcacag caaaggaata cattttggct cctcaaagtc attggataac 67200tgatcatcac caacttcagg atatgggcat gacatatgga ataggcaatg tactccagaa 67260aataagagtg agatattgga gtttaaattc tggctctgtc gctaatgagt tctgtgagcg 67320taacttcttt gtacctcact ttctttatct ttaaaatggg ggcaatgcca gaattcatct 67380aacagggtcg acgttaggtt atttgccatg agtactttaa caccctacct agcaaaataa 67440atattaataa aatttgatgt tttttcatca cactgtgtct atagcaaaat catgcttagg 67500agacctgatt tggtatacag ttcaaatggc caaagattag cttaggagac ttatgtaagt 67560ttaaattcaa agttttaagc aggattctgt accttaatac ctaaatgctt tttcctgtgt 67620cccatctaaa ttccagctat tctgtaagac ccaacttcaa tcccatttgt actatcctat 67680cttataattc ttaccatact aataatccaa tcactacctt taatctttgt attattttct 67740aatttccaaa gtcagaatgt ggtgttaacc actctttcat attactactg catatagtgt 67800gccaatggca ttcaaaagaa gtagagatca caaaaagtag agtgcatcac acaaggtgtc 67860acaaaggaaa taacatgtgc attagagcct taaagatgag ttaggtaaat gagttaggtc 67920aatggtgatg ttgggaagaa cactgcaaac aacaaagaat gtgcatttac taatcatggc 67980actttggtgg gaacccagga gattaccaat tatatttctc agttgttttg gagattgagt 68040tactgtatat aaagcactta gaatagtcca tagtaagtaa cagatagttg tttattttgg 68100ttacctactt ttctttataa tccattatgt atccctacaa catagctttg atttgctttg 68160gggaatttta tattaatttt atatcaatta atcataccat ttttattatt ttgggacttg 68220cctctttcac tcagctttag gtttagaaga tgtatccact ttaaagcata tagctgtagt 68280taaccttctc ttgaattgaa ctccactaat tcaatatatc actagttatt aagccactct 68340gtctggattg acataagtga tacagatatt gctgcaaaaa aactttttta tatgtctgat 68400ggtatatatg tgcaatagtt tcagtaggtt atttacctga gaatggaatt tctgagtcac 68460aggatggtca catgtctaac tttactggat aatatcaaat gtaccaactt acattatctc 68520agcagtatgt aagagttccc ttgccaatat ttagtatcat ctgactttct aatttttgga 68580tattagagca tgcattatgg tgtggttttt tttttttttt tctaaagact ggaagttctt 68640gtaatggagg ggtcttttaa aaataaaccc tcatgttttt aatgagcatc tctaaagaaa 68700aatatcccca gcttgttttt aaggttattt attttatttt attttttgcc atgacattgt 68760gttcactgcc ccagattcaa cttgtgatcc cactgggatc actaccctgc attaccaatc 68820tgaattacat acgttaaaac agccatctaa aagtgctagt tgtaagagtc taaatacttg 68880aatctttgag agacatattt atagtccatt atcttcacct cagttaagtc tgaagactat 68940ttgaaaaatg taatcctatt ttttcttcta ggatctaaaa aaatcattca agagcccaga 69000acccaggctc tttactcctg aagaattctt tagaattttt aatagatcca ttgatgcctt 69060caaggacttt gtagtggcat ctgaaactag tgattgtgtg gtttcttcaa cattaagtcc 69120tgagaaaggt aagacatgta agcatttcca gttcaaatgt aaacaacaaa cttaaatctt 69180ccctatgtag taagaatcta cctctgtgtt aagctgtagc aagatacatg catgtacgtc 69240taataaaaaa gcagatatca atagcacaga agaaactaat gattgtagat ttgtgggttt 69300ctaacccaaa gcaatattca tcagtttcaa attagtgagc tgtttgtgag taaacaatat 69360atgagatggc cactcatacc ttttctcatc taatatttgc cagtaattac aattatatta 69420ttctgtacct ccagtctact taatggtcct tctcaaattt tttcttaagg atctaactct 69480aagtcacaaa ttctgatcct ccccagttac tctgtattaa gactcttttt tcccttcact 69540gggtccctat aatacaaacc tggattcatt gctttctttt tttaatgaga aattaccatc 69600aatgtgtcat tatttgtatt tattaacata tttattcaat aagtatttgt tctacaaggt 69660gataatggtg gatttcttag taaaatagaa tagcaggacc cagtatgaaa taaataggat 69720ctgaatactc tagatttggg aagcagacct ggcaagtaat ctggtattgc acttcacctt 69780tcagggactc tataactcat acaaatcacc atataacact gacacattat tgctttctat 69840ttagattcca gagtcagtgt cacaaaacca tttatgttac cccctgttgc agccagctcc 69900cttaggaatg acagcagtag cagtaatagt aagtacatat atctgattta atgcatgcat 69960ggctccaatt agcacctata ggagtattgc atgggctttc aaggaaactt ctacatttat 70020tattattgat actgttctgt tactgttatt ccttttatgg tcttcttgag acttaagttt 70080gtagaattaa atttccctag agctggagat aatgtttaga gaattaggcc aataaatttt 70140ctgctgaggt tattttaaat aagacataaa attaatttta gaaatatgat ttatgccttt 70200tgttgaatca ttaacatata ttctgtgtgg aatgtgtctg acaaaaacgg aagtttaaaa 70260ttaagtgtat aaatgctggt tgttaacata tattctgtgt ggaatgtgtc tgacaaaaac 70320ggaagtttaa ttgtataaat gctggttgta tagaattcca aagatatttt cagatttgtg 70380tatatcagta tttactattg cgcatttgga aatttgatga ttttgttatt ttaccacaga 70440attatatgct tgatgagaat aggcttattt ttctgaaatt tctgtcctct tttgccaaga 70500acccaatttt ctttgaaaat ccagacaaag gtcattataa agtttattgt atttactaaa 70560tactctctag cctcatggaa ggtgcaggaa gaataaaaac catcttgtga tctaggctaa 70620aaaagtcatc ccttaggact tataaatggg gcactgtcat ttgttcaggg acttagaaga 70680cctatctgtg aaaagaagta atcaatttat tactaattca taatagcttg ttacaatgca 70740gaacatttaa aaaaactcta aaatacaatt tacttatttt ggaatttttt gccaccaaaa 70800cctgacctga agtgatttga ggctatttat agtctattta tctttctaaa agtgaatata 70860tattttacta atgaaatatt aatgtatttg attacagaat gctacccagg attcactggg 70920gatagtatat gttatacatt atgtgcagga cagaatcttc tgaaacctaa aattctaaat 70980tacgtagcat atcagttcta aaatgtgtca aaaagggact agagccctgt ttgtgtgtta 71040gtcaattaag ggttttctta ccaactatgg attttattaa cattttcaaa ggagcaactt 71100ttgtcttctt aaattctctg cttatatttc attgatttct aatctttatt atttccttcc 71160atttgctttg tgtttcattt actccacttt ttctagcttt tcgaagtgga aaaattagat 71220cattgatttt aaactatttt tgctaattac aagcacttaa aggtataaat tttcttaaat 71280acactgcttt agctgcacct cacacatttt tgtgttgtga ttttattatt tttctaaata 71340tttgtcaact gtgatttctt ctttgattca tggataatat aaaagtgcgt tgtttgatgt 71400caaaaagttt gagattttcc tatatatctt attgttacta gtctatgaat tttgttgtca 71460gaaaacatac tctataattt ttttgaaatg tactgaaact attttatgta tgttatttaa 71520ctgttgaatg cagcattcaa atttggccaa aatggttgat agtgttgttt atatcttccc 71580ttccttatta agctttttgt ctaattttta ttaattattg agaaaaggga attgaaaata 71640ttaacttttg attgtggatt tgtctgattt tccatttggt tttgggtttt ttgcttcata 71700tattttaaag ctttattatt atgggcttat acattgatta ttgttagatc ttccctgttt 71760aattgacctt ttaatcttac tgaaatatcc ctcattatct ctgatagtgc tttttgtctt 71820gaagtttatt ttgtctgata ttagagcaaa tctttcttat gcttactgtt ttcatggtat 71880atgtttggct ttcctttatt ttcagtctgt tgtctgtatt tttaaagtac atctctagta 71940gacaacatac acttgggctt cactatttta tttagtgtat ctctgattgt taattagagt 72000gcttcatcca tttctatgta ataaatcatt ggtatagttg gattttaatc caccatttcc 72060ttacttgttt tttatctgtc ccatttgtgt attgaccttc tgttccttat ttctagccct 72120ttttttttgg ttaattattt tttttctagt gtttaattag tgttggattt tcaacagcac 72180ccttttgcat tattttgtgg ttattttgga aattaaagta tacactttaa cttgtaatat 72240tctacttaga gttactattg tagcacttga gaaaaaaatg caagaacctt gaaacagggt 72300agttttattt acccataccc atcttttgtg tcatcattga catatattaa catatctaca 72360tcagttataa atttcacaat acagtctaca atacacaatg cagttcaatt atttctttga 72420actgtcatac gtattttaaa gtaagagaag aagaaatata ttgtctttta tattaacctg 72480catatttatt atttcctgta cccttcattc cctcttgtca gtccaggttt ccatttggta 72540ttaatttcct tcaggctgaa aaacattcta tagcatctct tataatacag atctgctggc 72600aacaaattct tttttctttt taattggaaa atatctttgt tttgtcttcc atgtagcagg 72660tactttttac tagatattga agtctgggta cacaggattt tttttctttt agtcttttaa 72720aaatatcact atagtattcc tgcagttatt gtttctgaaa aaaataaaaa aggtagccac 72780catttgtctt atttttaccc tgtatgtaca gggtcatttc ttcctctgcg ttctttcaaa 72840gtgttctcct ttttaacatt tgagctagaa tatgcctatg agtggttttc tttgtattta 72900tctttcttga gatttgtcaa gcttttggat ctatatattt gttttccctc aaatttgggg 72960aaaaaatagt cataaatttt tcaaattttt taatccattt tatctatcct ctcattctga 73020gatactactt gcatatatgt tagcagattg ctagtgtgtc ataggtcgca gaagttttgt 73080tcatttattt atcctctttt ctctgtgttc ttctgagtaa tttctacttg tctgctttca 73140agttcattga ccttttgttc tgccatctct gatctgctga atttttcatc cagtgaatta 73200ttctatttat gaatttatat tttctgttct agaatgtcca gttaattgtt taatgtattt 73260catatttctc tgctggaatt ttccatctgc atactcatta tgaaaaaaaa ttcttttttg 73320tcacttaaat ataatagctg ctctaaagtt cttgtctcca aatttcaaca cataggtcat 73380ctcagagttg atgtctattg acttcttcct ctctgattag gggtcacatt tccttttaaa 73440tggttagtga ttgtttaata tatgctggat attatgaatt ctatgctgtt gaatatcttt 73500cagattctat agcgtttctt tagaggctat tgaattgtgt tttggtagga ggttcgaata 73560gtagcagatg aggtttatct tttctcccag ccttgttttt aaactttctt gggtacaagg 73620gaaagtagcc ttattcctaa ggcatggaat atctgagata tctgtaaagt gctcagaaaa 73680tctctccaat attgcttgtc ttaataattt tgtccccatt gttctgtgtc tgctagggcc 73740tctactaaga ttatagcttc ccatgagttt ggtgtttggt tatttttttt tattataatt 73800ttattttttg gtcaggtttt gtgacttctc tactggtgca tatacagctt attattggac 73860cagtaactta aggagattca tctagggatt tctgatgctc cttctttgca cagctccctt 73920cttccaaata acctagatct agaaactcca gctgcatcgg cagccctgaa cttcaatctc 73980tgactcctta gcttagtgag gccattggtc cttgtttgaa ctctgtcttc cacacctcaa 74040tctgaaagtg caccccagac aggagagctg aaagccagaa aaccagggca aacatgagcc 74100tcacttagtg tggatgccct gtctcgtgaa ttacacattt gtactgctgt tcactaattt 74160ctgaaaagag ttgcctcata atttgtctag aattataatt acttaggttg gaaaggctag 74220tctgctacca tcgtatttgg aaacaaaagt ctttcctact aattttaaaa gacaaactaa 74280atataaaata cctattcaat cccgtgcctc tgaatctgag attacttttt aaattgcaag 74340atgaatggac agccttcttt tttgttcact gtttcttttt tcttcctttt taaaaaattt 74400tgagtcttgc attgtcagca acttttagct aggtgatgtc actcatgaat atgccactta 74460gatacctctg cacaggacaa gattttgtgg tgctaacatt cagagagaaa cagacgtaaa 74520taaaatactc atagtacagt atacaatatg tatcataata gaattatgca aatgttttta 74580ggtgtagcac aaagggggta agactttttt gggagaaggg ttagggaaat ttgaagaaag 74640catgctagtt ttcagggata actcaaaagc tggccatgtc agagtaccag ggcatatgaa 74700aacttattac agagatgtga gtgtgtaagg aaaagtcatg ggagaatgga ataacataca 74760aacctattag ttatccaaag ggaattatgt ttatagcata acagtaatat cctttatgat 74820aagcaaagag aatgtttaaa agatgtttta tgtttgagat tcttgcagac agtacctaaa 74880tgatagagaa agaaatccta actttgacat tatcttatac attttctaga aatagtcttg 74940acccaacctt cctggactca tataccagtc aagttaaact cttttgactc ttcggtcaaa 75000tggaagatca taatacctcc ttcacagagt ttctgtgagt tttaagtgag ttagtacaca 75060tgtgcttgga gacataactg atgtcttata agagttcatc attaagtact agaacaatga 75120gatgcaataa gactttggat cagtaattat gttgaacctt tagtgagctc tggatctgaa 75180tcaaggaaga atactcattt gcttgtttca taagcaccaa gataaccatg tcacaggcat 75240ggtgctccta aaaatgctca ctctaaaaag aagttcagca aaattatagc acaattcttt 75300gtgttctgta tttctacaga tgttggttat tattagtcaa atgaaacaag tcagttccca 75360agatttctgg acagtgaata gttttggtaa taaggagata taattaatat ggtaataacc 75420caggaatttg ggattactca aatccggccc agatgagctt cctctccaca taaattctct 75480gcaggctatc tttgcataaa atcaaaactt tgttgtcagc ttaagttttg tccccaaata 75540cccttcattt gggccttgag gcatgcaatt caaatctcat tgattcctac tcgatagctg 75600cagtggaaag tattgaaagt cattccctgg aggtataagc aaatctcttc tgataaagtt 75660ctttttgtct gcctgcaaga gggacagtag tatccctgat ggaccaaagg cctggttctt 75720aagccccacg catgaacaag tgaaataaaa aaagtgaggt aacattgacc tctatgggag 75780gtgaagctca gtaacctatg ggcgcttccc ccaaccctcc tctccactgc cgccccgcca 75840tccagtcttt gctgtttctc tccctccctg tctcctgcag gaagattgct gctgtgtctt 75900agtcagactc caggattgtg tattggctca cagtcttagg gatggaactg tcaacatggt 75960agtcaccgca ctctcagaga gcaagtctga gctttcttta gtaaggcctc ccacacagct 76020ttgttggaag ccccttcctt agatgctatc cttctgcctg

agaaaggttt taggtagcaa 76080attccagaat atctcactct ttagtaagct gctttttgct tttgagtctg gtctttcctt 76140tggtctgagt ggtgaatttc aagacttagg tttggtcttt gtatagaaca agaagaacat 76200ggtgtttggc taccttgatt acagggcatc tggctgaccc aacgtagata atggagttct 76260ccatagcatc ttcctagctt gttagagttt tccattggat tttaaggaca aaatcttaat 76320taatttattt gttctctctg tgtacattga ttatggcctc ttaaggaggt tacctagtaa 76380atctattgta attacatacc agcaatccca atagataatg ggggccacag cagattggcc 76440ttcaaggaga accaaaacag ccaaatgatt tatttacaaa cagtattatt aaaattcctc 76500aaatggtgcg aattgtttgt aaccctgaat ccatttcata tagagagcac ttttatagat 76560agtagttaaa tcccaaacga gtccacgtaa tcttacataa tccttaaaat agctctgtta 76620ttgtaatgat agtactttaa ttataccaaa tcccatttta ttactcagat ttattttggt 76680gagaaaattc acattgggct ccaacttgag atggtaaaac atatctccca tggggctata 76740ggatgtgggg tgggccagga ggaggaatcc taaggaggat attcatggtt atgtgaatgg 76800aagaccctat ttctgacaac tccctacgtg gggatcacca gcaccttatt gttcatccct 76860cactgtctcc tcccccaaca cacacacaca cacacacaca cacacgtgag cacacatcca 76920tacatcccat cattagagtc tccttggaaa cataccattt ataggatgag ggaatttcct 76980gctctcagtt cacatatggt agaggaatgg gctgttcctg tcctcaccat agtagataaa 77040atctatttct gacttatcaa gttaaatcta tgaattaatt tttccctgta agatgttgat 77100acacacatag tagatctatt attttatatt atgtactcta ggtactttgt atgttaaata 77160ggtataggta tatagaaaaa tctgtcaaaa taagtttcat gttggaagag acccttgtgg 77220gagatgtgtt ttgtttattt aattcgtgaa tcattctgct cctttttttt ttttaaagac 77280agagtcttgc tctgttgccc aggctggagt gcagtgacat gatctcggct cattacgact 77340tccgcctccc gggttcatgc gattctcctg cctcagcctc cgtagtagcc gggattacag 77400gcatgcgcca ccacacccag ctaatttttg tatttttagt agagacgggg ttttaccgtg 77460ttggctgggc tggtcttgaa ctctggatct caggtgatct gcttgactcg gcctcccaaa 77520gtgctgggat tacaggggtg agccaccaca tccaacctaa attcatgaat cattttgctt 77580ctcatggaag agccaggcag gatcaagaaa ggtgtcccaa accaattcca tatatgttct 77640ctctcattga gtttcttatt tagacgtagt atgggacaac actagtgcct tcaaccattc 77700cccttagtgt tagtcttaag ctttccaaca ttaaaagtcc aaactcttag tccgttgcac 77760agtggcatcc gacaaatgtg tgttgaagat atgaacgtgg aaacagccat gattctaagc 77820atgagatacc taaaaatact ataaccacag tgcacataca attatttaat attttcaaca 77880gtccaaacat agctgaatgc atacagatct tgatccaggt tgtaaaatgt catgtttaaa 77940atttctgagc aagctgtcaa gagtgagaaa gcaaaaagat tttagaagat aaaggcatgg 78000gagatagatt tgcttgatag tataagtcat tttcccccca gaaatatatt aagtctgtgt 78060tccatatgtg tcagaattac ccaagccaga tctctttctg tctctgtgat ttctttcttt 78120tttaatgcat tcatcttttg gagactacat ggatttatat tgccttgcaa aagcaatatt 78180cagcagaaaa atcaatcttt tgaatacttt aaacagatat aggtccagaa tgtgttacag 78240aaacagttaa aaacaaccac agcataagag caaaacttct agaatggata tgctgtattc 78300atcagtgtgt tctttaaatt atagggaagg ccaaaaatcc ccctggagac tccagcctac 78360actgggcagc catggcattg ccagcattgt tttctcttat aattggcttt gcttttggag 78420ccttatactg gaaggtaagt ggtaccattc ctttttttaa aaatatgcta tgtttacata 78480aattatcatc tttttttcct caagaaatga tcctttaaga aaacagtgaa ttctacctta 78540gcttataatc taaacaaaat ttaaatttta taaagtttcc tgtttctcat tatgtctgga 78600gacaatccct ctagctgata attcacgctt aagaattagg aactggtgtt aatagcctga 78660actaaatatg aaaaggaaga tctgacgaaa atcatatgat atacttaaaa gacaaataga 78720aatgtcattt gtattatctg ctattttgga tttctactct tttgttctct tcttattgaa 78780gcctgtctaa aattttaaaa tgtgcaaaaa tcattgtgac tcagcttctt gaaatgtctt 78840ttgaatttga ttaagtgtca gcttctttat tactctttag cccccaacat cctaaaaatg 78900ttgagttgat agtaccatgc agctttcata attttttatt acaaagatat tttgcatgct 78960acatgctgaa aaaactgtta ttggagttat tgccataaaa gataaaagtg gagtccactt 79020acctcttaaa tattagacca ttcattgatt attttacagt atatgtcttt cttctttttc 79080cagaagagac agccaagtct tacaagggca gttgaaaata tacaaattaa tgaagaggat 79140aatgagataa ggtattttgt tttgctaaat gtgtgcccaa tcaagcatga cattgccatt 79200tcacacactg tgtacctgcc cataatgtct ttaagaagtc cttcactcat gacagtagct 79260cctaaccagt gagtcccaac tctatccatg tttctgatgt ctcactctct cttcagtgct 79320gcatctccat ttacctacag aataccttca cataattgtc ttcatgactc ttcttagggt 79380ttgtaaaggg gttacctctg ctttcttatt taagtaaggc agcagaatct tctcagactc 79440ctggttgtgg aactctggag gcacatttgc tcatgttctg tcttttccac acttttatca 79500aaacatactg attcagctct tggagcgtct tcctaagctt agttctttca gtaggaagca 79560ttatcatcac tgacatcttt atgatctcat gagttgctgc cactgatctt ttctttagcc 79620cttctaaact caacctttag tcacactctt ctgtaaaccc tggttcatag tgtgacttct 79680tattcaaaat acgttcagtg attttctctt gtatatttta tatatctttt attttactct 79740ggctttcaga gcccttaact attctcaaca tactcgttgt tttctctaaa gctgaataca 79800aaatttgctg taagatgact ttccattcac tgtagctggc tcttgtcatt ctttcacctt 79860accctatact gcccagcact catatgttcc cttacctttg attataattt tcatttggtg 79920tgttgccttc tctcatgtca tgcctgtatg tatacacaga cacatatgaa atgcatatag 79980gcatgcttgg tatgtgtata tgcatataca gagaaagaaa tgttttaact acttggaaag 80040actaccttaa gacaaatgaa gtcttccctc ttccctatag taataagaag gtaggctccc 80100cattcaattt tgcaatcttc tgctactata tttacagaaa agctgccttt tacaatgccg 80160agatcatggt gtacctcaga atctctgacc aagagcaaat aagcattttt tcttattgtt 80220tttcagtatg ttgcaagaga aagagagaga gtttcaagaa gtgtaattgt ggcttgtatc 80280aacactgtta ctttcgtaca ttggtaagtt tttttcttct ttcctttttt tttctttttt 80340ttattatact ttaagttcta gggtacatgt gcacaatgtg caggtttgtt acgtatgttt 80400acatgtgcca tgttggtgtg ctgcacccat taactcgtca tttacattag gtatatctcc 80460taatgctatc ccttccccct ccccccaccc catgacaggc tccattgtgt ggtgttccct 80520accctgtgtc caagtgttct cattgttcaa ttcccaccta tgagtgagaa cacgcagtgt 80580ttggttttct gtccttgcga tagtttgctc agaatgatgg tttccagcct cagtcataca 80640tgtgcatgtg tctttagagc agcatgattt ataattttat aatcctttgg gtatataccc 80700agtaatggga tgcctgggtc aaatggtgtt tctagttcta gatccctgag gaatcaccac 80760actgacttcc acaatggttg aactacttta cagtcccacc aacagtgtaa aagtgttcct 80820atttctccac atcctctgca gcacctgttg tttcctgact ttttaatgat caccattcta 80880actggtgtga gatggttatc tcattgtggt tttgatttgc atttctctga tggccagtga 80940tgatgagcat tttttcacgt gtctgttggc tgcataaata tcttcttttg agaagtgtct 81000gttcatatcc ttcgcccact ttttgatggg gttgtttgat tttttcttgt aaatttgttt 81060aagttctttg cagattctgg atattagtcc tttgtcagat gggtagattg taaaaatttt 81120aattcaaact gaaatattta gcaagaacta tacagcatat gagatgccaa agtttagaaa 81180caaacttcat tagtaagtct tctatcaagc agatgtcagt atgttggctg aagctgttac 81240ataattgaaa tgtgcatatc taattcattg tgtattctcc agttttgaaa ggtaagcagt 81300gtttgtcctg actagtggat tcatctagtg tgggatatgc acaaaaatta acagttgtat 81360gtttacttag actgtttttg agaccagaaa attaattaga caagggaaat atagcaaatt 81420aggactaaaa ttaagtatta ctaatagaat aaaaatatat ggaatcaatt ttatttaggt 81480gtgaccacta taacatgact gtatcccatg tgttagatgc tgaaacaaga gaaaaccaga 81540atgtttccct gccattttca ggaaaaagag aaaacatcca aatatcttag agtcaggaga 81600gtaagatttt ccaggtcaat tgccaccttc cacatgacta gtaataatta tgcattataa 81660tagttcagag cccaggatgc acggaacata aacgcatggc agacaggtgc tgtgactgtg 81720tgggtctgcg gagggattgt gggagaaagg agagtaagat gtggctagaa ggtactggta 81780tcattatctc ctgtcccgtc tgctttctct gctcccttgc tgcagatact gttgcgggaa 81840gacccacagc tgcattaact taatcagtat tgattcatgc tgtccctcac ttttgcttct 81900gctaattggc tgttattttt gttactggca tggaatcctg agagcattta ccacagttgc 81960atgcgaatat tctctcctct ttcaaattcc tcatttcttc tgtgcattcc ctcattgtca 82020aaggattatt tactctcctc tttattacct agatagtctg tggtggattc cttctttacc 82080ctgcaagtca ctgcccttcc agctttaact cctgtcccag gttcagagat ttgtctctag 82140acttaatctt cttcagtgga actgagtcat aaattatctt gcctcctgta tgagcttact 82200tttatggtta tacagcaaca ttttttaaca actcaagata acatgatata attatgttgc 82260acatttctct aacactgtaa tactttattc tacacactct gtcactttta ccttcaatca 82320tactcttgat taacatgtcc gtagcatgaa tagcaccctc tattttctgc tgcccattgc 82380cacttctttc taggctcatt cttcctgtca aataatgtaa atggatagac cgtttgcttg 82440acacttttta ttttccgaag caatatttcc caagctgaaa ccatgctttt tcctagaaat 82500tcattttttt gcctttgaac cataagatga gattcactgt tttctttttt ttttccttcc 82560ttttcttctt cttgacagcc atttcagttc tctcaagtgg cattcacaaa ccacacatta 82620aatggttaat tgagtcttaa atgctcaaat gctcaagtaa tgtcctctgt tgtttggttt 82680gggattgcat tgctgggatc ctgttttggc acacaggtct ttaatcatgt ggttttaatt 82740acgacattcg tcccatgaca tttgggagtg gattaggata cggaaaccca ggccatttgt 82800ttaggaggaa aaaagatggt attttaaagc aaccatagtc aaacaactta tgaatgtgtt 82860gaatagtcag ttagcattcc aagaaaagag ttaggtcttt ctgtttcttg ctgtaccctc 82920aacacctaac ttgctgcccg tgcattgttg gctctctata aatgtttgtt gaatgagaga 82980ctgagtgagt gactcagccc taaatgaaag tttaagcaaa tggcaaagaa gattgttctt 83040ctgaagtagg gtttgttttt atcttttgtg tgaatgtgtg tatttaatgt attgggttgg 83100attaatgaga cagcaaggta tgcatatcaa gggggaatat aatcacatga gaaattaact 83160aataaattat ttttcttaag gtatgaatta ttactcatgt ataaaatgaa tcacattagc 83220caggttaaag aagccagaat tatccagaaa aaaacccagc tcacttattc agtagtcaat 83280gggaaataga tggcagaatg aatgaaaaca tgactcattc tccttcacaa agtgccttac 83340tcgtcatctc tgcaaagctc caccaaggct caggtcacat actgccacct acaaaaacct 83400gacttcccaa atgaaaataa tctctatttc tggccctcta aatagatttc atatgacact 83460tggtaggaca tcacagtcac ctaaatgaac attttgtgcc cttttcaaaa ccataaactc 83520ttagggggca ggattttttt ttcttaattt agttttggag ccccactaaa caatgtcact 83580tatactgtgc acatataaat gatgaataac tatttgttga ataaattaat ggaaagatga 83640atgaatatct cagaattaca ataagatgag aataaaatat atgaaaataa aaacagaaat 83700aaacttgtat gtgtctatga gtacataaga attcaaatgc tgagggaaaa gtaggaaaag 83760aggtagaaaa gggccagttc gtcctcagaa tctgaaaacc ccaaagtaaa aatgtgtgta 83820tatcatttgt aatcatctat tgactaccca ttatatgaag taaaccctta catttggcct 83880tctaccagca gagatggaaa accctccaag gaactcttaa gtctagttcc tagataggtg 83940tgtagtagag ggaaggtctg ggagaacaaa ctgtgatgtg gttggagaga aaatcttcta 84000aaaaataaga ataaccgtag taaaaaatgc caagacagaa ataaaatgtg gagttgcaat 84060tatacattta aaaaatagat ataatttgga ttataatcat tttcttggtt aacaaggaag 84120gggaaaaata caaggaatga ctcaggttta tggcaaggat ttctccctga aatggaagtt 84180tcaccaacta aaacaaatta aatcaagaca aatattggaa cactagaata gagaacatat 84240gggctagttc tcttaaggaa ataacctata attggaaaat gaaaagaagg agggtttaat 84300atcttaacta caaggaacaa acgtggaagg aatgtattga actagaaaca acaacaaaaa 84360aagttgcacc taggaataaa taggccagga gtggtagctc attcctgtaa tcccagtacc 84420ttggggggct gaggtgggag gatcacttga gcccaggagt tcgagaccag cctgggaaac 84480ataggaagac cccatctcta caaataattt aaaaattggg tgggcatagt ggtatgcacc 84540tatggtctca gctactcagg aggctgaggc aggaggattg cctgagcccg agaggttgag 84600gctgtaataa gccatgatca aatcacagca ttccaacctg ggtgatagag caaggcctgg 84660tctcaaaata taaaaataaa aaattaactg aagtgtgttt atttgacacc atgcgtgaga 84720acagctatgt gaaaagttaa aaatttactt ggtcacatag cttatgccaa atatccatag 84780tagcatccta caaaaacctt tttcatattg ggttgtggat agcaaagatt aagatagata 84840aataaaacag atggattaca ttaaaaactt tatgttaata cgttacagga aacttttgct 84900ctttactctt cccacaaatg tgattccaac aaactctagc aggtgcattg atggatgcaa 84960agcttaatat aactttgttg tgtcctcaaa ttgtttatag cctacaaaag gaaatcagac 85020atatacataa taaccaattt ttaaaagtga actaaaagtt tacagagtta ctttgatcaa 85080agtccaggcc tctgcatatg ttcatacaag ttgtgaattc aacacagttc tagatactgg 85140aatgaatgac atattagaat tgagcacacc caaatacagt tacctatttt tgaaaggaga 85200gagaaatcac taacaatagg agaaaagctt cactttgtaa gtggtggagg tagcatctgt 85260actgagcaca tggcgtaaac taagtcagta tagtcccaaa gtagaaactg tgttcaaggg 85320tcagtttgga cagttggatt ccagtggaag tcacatataa agagttaaat gctgagaaga 85380aagggtgagc atcggtttga gaaagggttt gaattccagg aatggggctt aaatgccttg 85440tgtcacataa ggaggtattg ggagaaaatg gtgtaggaaa gatgtgggtg acaatcaaag 85500ctgaggtgtg agaaaactaa tcgggtgttg gtatggcaag atgtacggga gtggttaact 85560agagaaattg agtccactta ccaggccagt ctttaaataa gatgtgagag agaatgacaa 85620ctgaattagc atcagggcca ggaaataaga aagcatagac catttgaaga cattgcagta 85680aaagaatcaa caaatcctag ttactaatgg gatgtgagac ttggcatcac aggcagaact 85740caagaatgat ttggagacta tgggacttac cagagtgcgg tggcattaac aggaaatgga 85800aaatcaggag aagaagctgt tggtaaatat tattgaaact aaccagtggc caaatatgtt 85860agactggaac ttcagataac agtcaggcct ggaaagaatg atttgcaatt tgagagttat 85920ctgcaaagaa tcttatttga gaccttgaaa gagaaattgc ttagtcacag aaatggcccc 85980gtgtggttga tagtaaatag catgggtttg attgtggaga ggttcactaa agaaatacaa 86040acaaaatttt ggtatatctg agctaagggg gaagaaaata aatcttagaa ttggaataaa 86100ggacaagctg aagacaagct tggaaattta ggcctcaagg cttggcaatg gttggttacc 86160aaatagcaag acctgtgatt ctttgctaga tgtttcctaa gactttcaaa gctttgggag 86220tttggtttct gtagatacat tgtaaagcag atgattagaa aggattaggc ttggattcaa 86280gccttgactc tttggtgtag aatactagat gaatttatat cagttatcag ttcatgtctt 86340ttaaaccctg gtttttcatg tgtaatgagg gttagtgata aatgtactct tcttatttca 86400caagggttct tgcaagaacc caggttatca ttataaaaat tgtagttttg ttctatggga 86460aaacaggtgc cagtctttac gtttattttc tttgctaagt gacccgggga caaaaatcaa 86520aaatagattt ccaatcttct ctacatttaa ttctctctct ctctctttct ttcttgagag 86580tctttctttc ttaagagtct tgctctgtag cccaggctgg agtgcagtgg cgtgatctca 86640gctcactgca acctccgcct cctgggttga agggggtttt gtgcctcacc cccagagtag 86700ctgggactac agatgtgcac taacacaccc agctaatttt tgtattttta gtagagacag 86760ggtttcacca tgttggccag gctggtctca aactcctgac ttcagatgat ccgcctacct 86820cagcctctca aagtgctggg attacaggtg tgagccacca tgcctgtcct atatttaatt 86880ctctatatta agctttgtgt ttccaattat ttatacattt aaaaaataca tccttttgct 86940ttttgtgatt cccttttgct ttctaaccat gcttttaaaa tatcttctat atattttgga 87000tttctgaagt tttatcttag tatctcttaa tttttgttct agatttaatt ctgatttcct 87060aaatacaggt tgtctagctt ctagagttct ctgcttggta aattgttgta taagacagtt 87120tttaatatat tacatttaga gagactggaa aaatattgca attcttatta cactatggaa 87180ttttatggct aatctgtgta atgatctcaa tactttaaaa acagtgatct gtgatgctgg 87240gttgataaac aaggacaaaa gtagtgaaag agattgagca gagaagacga gaaatgcaaa 87300acacatttat ttagagactg ggattgaaga tgggatgata ccatctatat gtattttttt 87360taagtagaga aggaaataca aatactttac tggaaagttg aaacatctga aagctagtag 87420attaacacaa gtactgaata actaaaacaa aatatggctt tcatgcttca acaagtacaa 87480taagctatgc tgaggccaaa cgaactttcc ccagaacttt gttgggatgc ggtgccaaga 87540ttacagtctt tctttttttg gttggaagta ttgaacgaaa tctttctgct gctatttcct 87600cagctggaag aaatgattgc tagaacctaa ctagatagtc aagatactag agcggctcag 87660tggccatgtg aggctattac ttgttggtgg ccattgacca gcttctccac ctgtcttagg 87720gatctctttc atgctactcc atctatctcc atttctctaa tctccactct ccagacactt 87780tgttcgtcaa tccttctccc ttcacccctg tttactacct tttataatct tgctagagaa 87840attcaaagct aactaagttg ataattatat tatcacaaac tgagatcatt ggtagaatgt 87900caggaggtat gaaagtcaca gggtgaaata tttattgagt gtctattaag tgccagacat 87960tattataaga gtggaggtac agcagtaaaa gagatagaga agctccctgc cctcatggag 88020cttacatttt aatgggagga catagataat aaacaaaaag taagattgtt tcagataatg 88080tgttaccaag gagatcatag aaaatgactg atcactgcat ctaagaaaat atttatgaag 88140ttcttttctg ctatataagt gaagcataca ttgcaaattt gttactcatt ctgtggtcat 88200atatgatttc ggctattgag atttttaaaa aggcagaagg cagatacacc ataatctctc 88260ttttcttcct tcctttctag gctggtaaca gttcatgttt gcttcataaa tgaagcagct 88320ttaaacaaat tcatattctg tctggagtga cagaccacat ctttatctgt tcttgctacc 88380catgacttta tatggatgat tcagaaattg gaacagaatg ttttactgtg aaactggcac 88440tgaattaatc atctataaag aagaacttgc atggagcagg actctatttt aaggactgcg 88500ggacttgggt ctcatttaga acttgcagct gatgttggaa gagaaagcac gtgtctcaga 88560ctgcatgtac catttgcatg gctccagaaa tgtctaaatg ctgaaaaaac acctagcttt 88620attcttcaga tacaaactgc agcctgtagt tatcctggtc tctgcaagta gatttcagct 88680tggatagtga gggtaacaat ttttctcaaa gggatctgga aaaaatgttt aaaactcagt 88740agtgtcagcc actgtacagt gtagaaagca gtgggaactg tgattggatt tggcaacatg 88800tcagctttat agttgccgat tagtgatatg ggtctgattt cgatctcttc ctgatgtaaa 88860ccatgctcac ccatatccca ctatacaaat gcaaatggtt gcctggttcc atttatgcaa 88920gggagccagt actgaattat gccttggcag aggggagact ccaaaagagt catcgcagga 88980agaagttaag aacactgaac atcagaacag tctgccaaga aggacattgg catcctggga 89040aagtccgcct tttcccttga ccactatagg gtgtataaat cgtgtttgca aaatgtgtta 89100tgatgtgttt atattctaaa actattacag agctatgtaa agggacttag gagaaaatgc 89160tgaatgtaag atggtcccat ttcaatttcc accatgggag agcctaaaaa taaattatga 89220catttagtat ctaaggttag aaaaccacgc ccacatgcta atatgggtgt tgaaaactag 89280gttacttata atgcaaggaa tcaggaaact ttagttattt atagtataat caccattatc 89340tgtttaaagg atccatttag ttaaaatcgg gcactctata ttcattaagg tttatgaatt 89400aaaaagaaag ctttatgtag ttatgcatgt cagtttgcta tttaaaatgt gtgacagtgt 89460ttgtcatatt aagagtgaat ttggcaggaa ttcccaagat ggacattgtg cttttaaact 89520agaacttgta agacattatg tgaatatccc ttgccaattt tttttataat aagaaaacat 89580ctgactaaag tcaaagaatg atttcttatg gtttattttg atgaaagttc ttttaacatg 89640tcttgaatgt acacataaag gaatccaaag ctttccattc taacttaatc tttgtgataa 89700cattattgcc atgttctaca accgtaagat gacagttttc aatgtagtga cacaaaaggg 89760catgaaaaac taactgctag ctttcctttc atttcaaaag tccaagaatt tctagtatat 89820ttggatttta gcttctgttc aaagcaaatc cagatgcaac tccagtaagt ggcctttgct 89880cttttttgta ccaaagagcc cagatgattc ctacagtccc tttcttctct aacatgctgt 89940ggttccttaa atatgagtaa tttctctaag atataaccca ggtgctttga gaagctgcat 90000taaggtgttc aggccctcag atatcacatg gtacacttga ttagtaataa aaccagagat 90060caatttaaat tgctgatagg tcctgtctca gtgtgtggca ttgactgttt tcaggaaaat 90120agatacagat taatatgagt tatgcgtgta ggttgtgtat agattgagaa gatagatact 90180tctcaatcta gtagtttgat ttatttaacc aatggtttca gtttgcttga gcatatgaaa 90240atcctgctta atgtgcttaa gagtataata aatgtgtact tttgtcctca aacctagtag 90300ctgggtttta acactcatgg acatggtctt aatcaatgga gttaaataaa caaattcagc 90360aagttattaa atctgacatg gtaggagagg ggagatgtgt cctgcttatt aaatgtgttg 90420gtccattgaa agttacatgg attgccaatt tttaaaacac taaagttgaa taaaatgcat 90480gaacaataga aaaatgctga acattatttt ggatgctagc tgcttggaca ttaactgtgt 90540tatttctgct ttgagatgaa aatatatatt tatctttgct tattttatcc cagatgtgtt 90600ctgaatatcc ttcttcataa atcatggaaa actcactgct gagatagtaa accatgaaat 90660cgccttttca gttggtgcca tgtatctgac agttccatct tggaaggttt caaaattacc 90720ttttaaaatg atctcagaag tctgtagatt ctcaatgata ctgaaagctt tgcacctctt 90780tggtagaaac caggtctatt tagaaaatgg ctttatgata aatgttgcct cctgagtgat 90840aatgaagtgt tcctggatat tgtattgtaa tttaatgtgc ttaccacact gccacatttt 90900aatgagtcag agaaaaatta atttttcttc aatacaataa tagaacaagt agcctattct 90960cttaaaaagt atgtgaaaag aaaattatga aaaaatatgc atacctaatg aagtattggt 91020tttagtaaga attaaataca tttcattgag ctttaaagta ctttggagaa actttggggc 91080acgttttcct actctaattc aactaaagtt ataaataaag

agaaaaactc attcagaaat 91140catggatttt aaaaatattt tactgcagcc aagttttcat ttcaaaatgt aatttcagtt 91200tggagctttt aggcattatg tatatttaaa aaatatattc ttcaaaaatg cattttggca 91260tggtgggatg gatgttgcaa aagatatccg gagcctccag tctgtcatta actgatatgg 91320taaatcacct ctcttctttg ggtctcaatt ttttatttat ctatatggta aactcagaga 91380tcactcctta ggggtgagtc ctattgcaat atgaccgaca aagaagacaa aatagcattg 91440aaactaaccc atacaaaata tccaactctg gattctgtga ataagtatct tgaccataaa 91500aagtcattgc tgttcttgtt tctaatgtaa atagtgtcca ttagtaaaag tgaaattcag 91560tcttaagtag ggtgaattgg atcaccattt acacaagaga tggctttttc ctttgcttga 91620ataaacattt tggatcacct ccaaagaatg aaaaccagta gtacgtttta gtcatattag 91680tcaggatgag aaactataag atgtgtgtaa catttggaaa tgcaccaaag tgagcgttta 91740aatcttctca ttttattgaa aactaagagc agaaaatgta aaatgctcat gaaggttttg 91800aatgccaaaa gatattttag aatcaattta taaaggggta attcattaat tacactttaa 91860aattggaaag tgggataaga aatctaaagt aaaccagctt atctttgaaa caatattatt 91920ttgaaattgg ctttaaaata aaaccattca gattgaaatt ctaattagct catttgtgga 91980gtttgatcac acaattcata atgttgctgc tttccattaa ctagtcttga aatgcctttg 92040tttgtaaaaa taaaataatg gtactttcat tttataacaa ggtgtttttt tcaagaaata 92100atccatgcta aaatggatat ttgtgatcct gaaatgttta ctaagcattg taaatttatt 92160tataactgcc atctccaact acatccttat gatgttttta acaataaaat taaaacaact 92220gttaaactaa aaaccacacc gttttccagt acttgatctc tgagctacaa tactcactaa 92280atataatttt ccaatcaaaa tattctattc tatattctaa gggttaatat gtgattatag 92340tgtccacttg ccaccatttt tttaaatcaa tggacttgaa aagtattaat ttagatggat 92400gcgcagatat accctcagtt cagtcataga ttggagtttg catataataa tgtaaatgta 92460tgtcgacact attctaaata gttctattat gactgaaatt taattaaata aaaaaggttg 92520taaaatgtga tgtgtatgtg tatatactgt atgtgtactt tttaaaatag gtgtatgtcc 92580caaccctttt ttatacaggt ttgaatttaa aattacatga tatatacata tactttattg 92640ttctaaataa agaattttat gcactctcat aaatttaccc tgtcatttta tatcttaagg 92700ttatcacata tgtctacaac atggttgaac atttattaag ctacagtcaa aattttctgt 92760ggaaactttt gtcattaatc tagattttat taagctacag tcaaaatttt ctatggaaac 92820ttttgtcatt aatctagatt ttgttgaaat atggacacaa ggggataaga aattatttaa 92880attcaataaa tatttattga ttactaaaat aagtaatcaa ggaaaataaa aaatgaatta 92940acacgacgtt tgctttcaag agatttataa cttgtctgga gaaaagaaat gcaatagata 93000tatagaaaag cactataata gatatagata cagaaaatag atacagatat agaaaagcag 93060tataatatgt atagtccctg aagtaaatac ctacatgcaa aacagaggtc cagttaatgt 93120gcttagggca tgcaggaagg agaaatggat acagaataag ggagactgtg gaggaattga 93180tgtttgggct aatacttaga aggatgagac aaatttttat agtgagaatt atgcccagtc 93240agggcagcat tttccataga aagaagttat aaaatttcta agcttatttt accaactcaa 93300acaggtatgt ccttagcata caccaaataa aaagacacac ggagagataa gaccagaccc 93360ttcaatgtaa cagtgagtgc agacactatt cttagggtcc ctgagaagcc tagactcctt 93420tgcttgtttc tgagaagagg aatgacgaga tcaaaacgca ctacggatcc tgtgatttag 93480gcagtagata atcatcctgc ttaaccactg gttgctattt gactctcaag cacataccct 93540tgctatatga caccacactt attcacccac catttctttt cagaatttgc tgattctctc 93600tcctctgttt accttttata ttgccaactt cgaccaatat atgactagat aactagaata 93660gcctcctaac tgatcacatg attctacttt cacccttttt caatcagttc atcacattat 93720agtcaataat agggagacat ctggtgtgtg tgtgtgtgtg tatgtgtgtg tgtgtgtgtg 93780tgtgtgtgtg tgtgtttctg gctaaaacac ttggatggct tctcattatt ttttagctaa 93840agcccaaact cttcaacaca gcctacaagg ctctacttga gccatctcat cttttttgtt 93900ttgttttgtt ttggattgct gattgttttt gcctttaatc cataaacaac atgaaggttg 93960ggcccataaa tactaatata actgcagtcc taaaatttag tgcaaattat tagtggaatt 94020gaaacaaaat gtgatctaaa tagctatcac ctgttgtact gagaagttat tttttaagat 94080ttttattatg gacaatttga aatatatgca aaagtcgata agatagtgta ataaattcat 94140ataaccataa ccctacttca accttactca actcctgaat aatcttgttt tatctaaact 94200ccctatccac tccccatttt gtatcatttt caggaaagta cttaaaagta tttgatctgg 94260aaacatttct atgtatatct ctgttaactt tcagaaagcg taaccctgat acccttgtta 94320tacctaagaa aagttattaa caatttctta ataccaagga gccaacaagt atttacattt 94380taactctttt acactgtaca ctgcagtatt gcactagagt agggtataaa atacagttaa 94440aagtccccct tgaacattaa agttagtttc cctggaggta aacactgcta attattcctc 94500attaacttct agaatttttt ggcatgtaca aactatatat acatgtatat acaatactag 94560tctctaactt tatctctttc atttattagg agaacgcaga acactacatc agcctagata 94620aatctttacc cactgcacca taataaccag gaaattatag ataggcatta cga 94673811PRTHomo sapiens 8Glu Gly Ile Cys Arg Asn Arg Val Thr Asn Asn1 5 10

Claims

1. A method of treating a fibrotic or tissue remodeling disease comprising administering a therapeutically effective amount of a stem cell factor inhibitor to a subject with or at risk for a fibrotic or tissue remodeling disease.

2. The method of claim 1, wherein the inhibitor is an isolated antibody or antigen-binding fragment thereof.

3. The method of claim 2, wherein the antibody is a monoclonal antibody or antigen-binding fragment thereof.

4. The method of claim 2, wherein the antibody or antigen-binding fragment thereof specifically binds to stem cell factor.

5. The method of claim 2, wherein the antibody or antigen-binding fragment thereof specifically binds to a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 1 and SEQ ID NO: 8.

6. The method of claim 1, wherein the inhibitor is a small interfering RNA.

7. The method of claim 1, wherein the subject has an abnormal activity of stem cell factor or abnormal collagen production.

8. The method of claim 1, wherein the disease is fibrosis, a remodeling disease, or a pulmonary disease.

9. The method of claim 1, wherein the disease is idiopathic pulmonary fibrosis, chronic obstructive pulmonary disease, acute respiratory distress syndrome, cystic fibrosis, peribronchial fibrosis, hypersensitivity pneumonitis, asthma, sclerodoma, inflammation, liver cirrhosis, renal fibrosis, parenchymal fibrosis, endomyocardial fibrosis, mediastinal fibrosis, nodular subepidermal fibrosis, fibrous histiocytoma, fibrothorax, hepatic fibrosis, fibromyalgia, gingival fibrosis, or radiation-induced fibrosis.

10. The method of claim 2, wherein the antibody or antigen-binding fragment thereof is delivered into an airway of the subject by intranasal administration.

11. The method of claim 1, wherein said administering the inhibitor reduces an activity of a receptor and/or reduces an interaction of stem cell factor with a receptor.

12. The method of claim 11, wherein the receptor is a receptor tyrosine kinase.

13. The method of claim 11, wherein the receptor is c-Kit.

14. The method of claim 1, wherein administering the inhibitor to a subject results in a direct inhibition of fibroblast activation.

15. A composition comprising an isolated antibody or antigen-binding fragment thereof that specifically binds to stem cell factor.

16. The composition of claim 15 wherein the isolated antibody or antigen-binding fragment thereof specifically binds to a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 1 and SEQ ID NO: 8.

17. The composition of claim 15 wherein the antibody is a monoclonal antibody or antigen-binding fragment thereof or the antibody is a humanized antibody or antigen-binding fragment thereof.

18. A method comprising the steps of providing an inhibitor of stem cell factor and administering the inhibitor to a cell or tissue.