COMPOSITIONS AND METHODS FOR REGULATING PRODUCTION OF A PRECURSOR PROTEIN

Abstract

The present disclosure relates to one or more agents, therapies, treatments, and methods of use of the agents and/or therapies and/or treatments for increasing production of a TLR3 precursor protein. Embodiments of the present disclosure can be used as a therapy or a treatment for a subject that has a condition whereby the subject's immune system is, or is likely to become, dysregulated and where the production of the TLR3 precursor protein may result in an increased production of a functional and bioavailable TLR3 protein product, which may be of therapeutic benefit.

Description

TECHNICAL FIELD

[0001] The present disclosure generally relates to compositions and methods for regulating production of a precursor protein. In particular, the present disclosure relates to compositions and methods for regulating production of a precursor protein that can be post-translationally modified to produce a protein of the toll-like receptor family.

BACKGROUND

[0002] The mammalian immune system can differentiate between self and foreign matter. A number of cascades of signaling molecules and immune cells are characterized by their ability to recognize foreign matter and to call upon the production and stimulation of effector cells of the immune system to kill, break down, consume, or sheath the foreign matter in order to protect a host.

[0003] One mechanism by which the immune system can differentiate between self and foreign matter relates to a class of pattern recognition receptors that can detect molecular patterns that foreign microbes present. These pattern recognition receptors can be present on the phospholipid bilayer of various cell types and these cells can become activated upon a binding event between the pattern recognition receptors and the foreign molecular patterns. Activation of these cells reflects triggering of the innate immune system and the acquired immune system.

[0004] It is known that under various conditions the immune system can become dysregulated. A dysregulated immune system can cause further damage to the host, thereby preventing healing. It may also result in a loss of homeostatic controls and/or a chronically stimulated immune system.

SUMMARY

[0005] Some embodiments of the present disclosure relate to compositions and methods that upregulate the production of a precursor protein. The precursor protein may be subjected to one or more post-translational modification processes to produce a protein that is a member of the toll-like receptor (TLR) family of proteins. In some embodiments of the present disclosure, the precursor protein is a precursor of TLR3. When the precursor protein is subjected to one or more post-translational modification processes, the TLR3 protein product is bioavailable and functionally equivalent to a TLR3 protein that is produced within a subject without the benefit of the embodiments of this disclosure.

[0006] In some embodiments of the present disclosure, the compositions described herein comprise a vector of plasmid deoxyribonucleic acid (DNA) that includes an insert sequence of nucleic acids. The insert sequence encodes for the production of the precursor protein and the insert sequence may also include a backbone sequence of nucleic acids that facilitate introduction of the insert sequence into one or more of a subject's cells. Within the subject's cells, the insert sequence is expressed and/or replicated. Expression of the insert sequence by one or more cells of the subject results in an increased production of the precursor protein. In some embodiments of the present disclosure, the methods that upregulate the production of precursor protein relate to methods of manufacturing and administering the composition.

[0007] Some embodiments of the present disclosure relate to compositions and methods that can be used as a therapy or a treatment for a subject that has a condition whereby the subject's immune system is, or is likely to become, dysregulated.

[0008] Some embodiments of the present disclosure relate to a recombinant virus vector (RVV). The RVV comprises a nucleotide sequence encoding production of the precursor protein and an inverted terminal repeat.

[0009] Some embodiments of the present disclosure relate to a composition that comprises a nucleotide sequence according to the present disclosure (SEQ ID No. 3) of which at least a portion can be expressed in a target cell.

[0010] Some embodiments of the present disclosure relate to an insert for use with an RVV, wherein the insert has a nucleotide sequence according to SEQ ID 1.

[0011] Some embodiments of the present disclosure relate to a method of making an agent/target cell complex, the method comprising a step of administering a therapeutically effective amount of the agent to a subject, wherein the agent/target cell complex increases the subject's production of the precursor protein.

[0012] Some embodiments of the present disclosure relate to a pharmaceutical agent that comprises an agent, a pharmaceutically acceptable carrier and/or an excipient. Administering the pharmaceutical agent to a subject may increase the subject's production of the precursor protein.

[0013] Some embodiments of the present disclosure relate to a method of treating a condition. The method comprises a step of administering to a subject a therapeutically effective amount of an agent that upregulates the subject's production of the precursor protein.

[0014] Some embodiments of the present disclosure relate to a use of an agent for treating a condition, wherein the agent upregulates the subject's production of the precursor protein.

[0015] Embodiments of the present disclosure relate to at least one approach for inducing endogenous production of the precursor protein. A first approach utilizes one or more gene vectors containing nucleotide sequences for increasing the endogenous production of the precursor protein. The one or more vectors can be administered to a subject to increase the subject's production of the precursor protein.

[0016] Without being bound by any particular theory, the one or more vectors may be expressed in cells of the subject that already have the capability to produce, and do or may have produced, the precursor protein. These cells are referred to herein as capable cells. Therefore, the administration of the one or more vectors may increase the production of the precursor protein within those capable cells. The one or more vectors may also be expressed in other cells of the subject that do not produce the precursor protein when the subject is in a homeostatic state, an infection state or a disease state. These cells are not capable of producing the precursor protein without the one or more vectors of the present disclosure being expressed therein and, therefore, these cells may be referred to herein as incapable cells. The administration of the one or more vectors may induce the incapable cells to start producing the precursor protein.

[0017] The precursor protein can be subjected to one or more post-translational modification processes, which results in subject cells that are producing the precursor protein to produce a final protein-product that is bioavailable and functional. In some embodiments of the present disclosure, the protein product may be a receptor protein that is capable of participating in a binding event with a ligand. When a binding event occurs, the receptor protein may initiate one or more changes in the internal cell-signaling and metabolism of the subject cells. In some embodiments of the present disclosure, the protein product is a membrane-bound receptor protein. In the case of capable cells, the one or more vectors of the present disclosure may cause the capable cells to be more sensitive to ligands by increasing the number of membrane-bound receptors that can participate in binding events. In the case of incapable cells, the one or more vectors may cause incapable cells to become able to participating in binding events with ligands. In other words, the embodiments of the present disclosure may cause incapable cells to become responsive to a ligand that binds to the membrane-bound receptor product.

[0018] In some embodiments of the present disclosure, the membrane-bound receptor protein is a member of the toll-like receptor (TLR) family. Without being bound to any particular theory, the embodiments of the present disclosure may cause capable cells to increase their participation in the innate immune system, the acquired immune system; binding events with other TLR ligands or combinations thereof. Furthermore, incapable cells that produce the precursor protein due to the one or more vectors of the present disclosure, may now be able to participate in binding events with TLR ligands whereas such binding events would not otherwise occur.

[0019] Without being bound by any particular theory, TLR3 is a known component of the innate and acquired immune systems. TLR3 participates in activating immune cells upon a binding event with foreign matter due to the ability of TLR3 to recognize patterns that foreign matter present. Furthermore, pharmaceutical agents may also participate in binding events with TLR3 and these pharmaceutical TLR3 agonists may increase the efficacy of other chemotherapies for cancer, such as inhibitors of checkpoint proteins. For example, increased expression and/or functionality of TLR3 may increase the activity of polyIC, a synthetic double stranded RNA. Without being bound by ant particular theory, the embodiments of the present disclosure may ultimately increase the amount of TLR3 expressed and functioning in a subject and that subject may then further benefit from treatment with other oncolytic viruses, as compared to treatment with oncolytic viruses alone.

DETAILED DESCRIPTION

[0020] Unless defined otherwise, all technical and scientific terms used herein have the meanings that would be commonly understood by one of skill in the art in the context of the present description. Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present disclosure, the preferred methods and materials are now described. All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the publications are cited.

[0021] As used herein, the singular forms "a", "an", and "the" include plural references unless the context clearly dictates otherwise. For example, reference to "an agent" includes one or more agents and reference to "a subject" or "the subject" includes one or more subjects.

[0022] As used herein, the terms "about" or "approximately" refer to within about 25%, preferably within about 20%, preferably within about 15%, preferably within about 10%, preferably within about 5% of a given value or range. It is understood that such a variation is always included in any given value provided herein, whether or not it is specifically referred to.

[0023] As used herein, the term "activity" is used interchangeably with the term "functionality" and both terms refer to the physiologic action of biomolecule.

[0024] As used herein, the term "agent" refers to a substance that, when administered to a subject, causes one or more chemical reactions and/or one or more physical reactions and/or or one or more physiological reactions and/or one or more immunological reactions in the subject. In some embodiments of the present disclosure, the agent is a plasmid vector.

[0025] As used herein, the term "ameliorate" refers to improve and/or to make better and/or to make more satisfactory.

[0026] As used herein, the term "biomolecule" refers to a carbohydrate, a protein, an amino acid sequence, a nucleic acid, a lipid, a primary metabolite, a secondary metabolite or another metabolite that is found within a subject. A biomolecule may be endogenous or exogenous to a subject.

[0027] As used herein, the term "cell" refers to a single cell as well as a plurality of cells or a population of the same cell type or different cell types. Administering an agent to a cell includes in vivo, in vitro and ex vivo administrations and/or combinations thereof.

[0028] As used herein, the term "complex" refers to an association, either direct or indirect, between one or more particles of an agent and one or more target cells. This association results in a change in the metabolism of the target cell. As used herein, the phrase "change in metabolism" refers to an increase or a decrease in the one or more target cells' production of deoxyribonucleic acid (DNA), ribonucleic acid (RNA), one or more proteins, and/or any post-translational modifications of one or more proteins.

[0029] As used herein, the terms "dysregulation" and "dysregulated" refer to situations or conditions wherein homeostatic control systems have been disturbed and/or compromised so that one or more metabolic, physiologic and/or biochemical systems within a subject operate partially or entirely without said homeostatic control systems.

[0030] As used herein, the term "effector molecule" refers to a molecule within a subject that can directly or indirectly regulate the metabolic activity of a target cell by increasing or decreasing the production of DNA, RNA and/or amino-acid sequences and/or by increasing or decreasing any post-translational modifications of one or more proteins.

[0031] As used herein, the term "endogenous" refers to the production and/or modification of a molecule that originates within a cell of a subject.

[0032] As used herein, the term "excipient" refers to any substance, not itself an agent, which may be used as a component within a pharmaceutical composition or a medicament for administration of a therapeutically effective amount of the agent to a subject. Additionally, or alternatively, an excipient may, either alone or in combination with further chemical components, improve the handling and/or storage properties and/or permit or facilitate formation of a dose unit of the agent. Excipients include, but are not limited to, one or more of: a binder, a disintegrant, a diluent, a buffer, a taste enhancer, a solvent, a thickening agent, a gelling agent, a penetration enhancer, a solubilizing agent, a wetting agent, an antioxidant, a preservative, a surface active agent, a lubricant, an emollient, a substance that is added to mask or counteract a disagreeable odor, fragrance or taste, a substance added to improve appearance or texture of the composition and/or a substance that is used to form the pharmaceutical compositions or medicaments. Any such excipients can be used in any dosage forms according to the present disclosure. The foregoing classes of excipients are not meant to be exhaustive but are provided merely to be illustrative of what a person of skill in the art would know and would also recognize that additional types and combinations of excipients may be used to achieve delivery of a therapeutically effective amount of the agent to a subject through one or more routes of administration.

[0033] As used herein, the term "exogenous" refers to a molecule that is within a subject but that did not originate within the subject.

[0034] As used herein, the terms "inhibit", "inhibiting", and "inhibition" refer to a decrease in activity, response, or other biological parameter of a biologic process, disease, disorder or symptom thereof. This can include but is not limited to the complete ablation of the activity, response, condition, or disease. This may also include, for example, a 10% reduction in the activity, response, condition, or disease as compared to the native or control level. Thus, the reduction can be a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or any amount of reduction in between the specifically recited percentages, as compared to native or control levels.

[0035] As used herein, the term "medicament" refers to a medicine and/or pharmaceutical composition that comprises the agent and that can promote recovery from a disease, disorder or symptom thereof and/or that can prevent a disease, disorder or symptom thereof and/or that can inhibit the progression of a disease, disorder, or symptom thereof.

[0036] As used herein, the term "patient" refers to a subject that is afflicted with a disease or disorder. The term "patient" includes human and veterinary subjects.

[0037] As used herein, the term "pharmaceutical composition" means any composition comprising, but not necessarily limited to, an agent to be administered a subject in need of therapy or treatment of a disease, disorder or symptom thereof. Pharmaceutical compositions may include additives such as pharmaceutically acceptable carriers, pharmaceutically accepted salts, excipients and the like. Pharmaceutical compositions may also additionally include one or more further active ingredients such as antimicrobial agents, anti-inflammatory agents, anesthetics, analgesics, and the like.

[0038] As used herein, the term "pharmaceutically acceptable carrier" refers to an essentially chemically inert and nontoxic component within a pharmaceutical composition or medicament that does not inhibit the effectiveness and/or safety of the agent. Some examples of pharmaceutically acceptable carriers and their formulations are described in Remington (1995, The Science and Practice of Pharmacy (19th ed.) ed. A. R. Gennaro, Mack Publishing Company, Easton, Pa.), the disclosure of which is incorporated herein by reference. Typically, an appropriate amount of a pharmaceutically acceptable carrier is used in the formulation to render said formulation isotonic. Examples of suitable pharmaceutically acceptable carriers include, but are not limited to: saline solutions, glycerol solutions, ethanol, N-(1(2, 3-dioleyloxy)propyl)-N,N,N-trimethylammonium chloride (DOTMA), dioleolphosphotidylethanolamine (DOPE), and liposomes. Such pharmaceutical compositions contain a therapeutically effective amount of the agent, together with a suitable amount of one or more pharmaceutically acceptable carriers and/or excipients so as to provide a form suitable for proper administration to the subject. The formulation should suit the route of administration. For example, oral administration may require enteric coatings to protect the agent from degrading within portions of the subject's gastrointestinal tract. In another example, injectable routes of administration may be administered in a liposomal formulation to facilitate transport throughout a subject's vascular system and to facilitate delivery across cell membranes of targeted intracellular sites.

[0039] As used herein, the phrases "prevention of" and "preventing" refer to avoiding the onset or progression of a disease, disorder, or a symptom thereof.

[0040] As used herein, the terms "production", "producing" and "produce" refer to the synthesis and/or replication of DNA, the transcription of one or more sequences of RNA, the translation of one or more amino acid sequences, the post-translational modifications of an amino-acid sequence, and/or the production of one or more regulatory molecules that can influence the production and/or functionality of an effector molecule or an effector cell. For clarity, "production" is also be used herein to refer to the functionality of a regulatory molecule, unless the context reasonably indicates otherwise.

[0041] As used herein, the terms "promote", "promotion", and "promoting" refer to an increase in an activity, response, condition, disease process, or other biological parameter. This can include, but is not limited to, the initiation of the activity, response, condition, or disease process. This may also include, for example, a 10% increase in the activity, response, condition, or disease as compared to the native or control level. Thus, the increase in an activity, response, condition, disease, or other biological parameter can be 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, or more, including any amount of increase in between the specifically recited percentages, as compared to native or control levels.

[0042] As used herein, the term "prophylactic administration" refers to the administration of any composition to a subject, in the absence of any symptom or indication of a disease or disorder, to prevent the occurrence and/or progression of the disease or disorder within the subject.

[0043] As used herein, the terms "signal molecule", "signalling molecule" and "regulatory molecule" can be used interchangeably and refer to a molecule that can directly or indirectly affect the production and/or functionality of an effector molecule or effector cell. Signal molecules can be enzymes or other types of biomolecules that can act as a direct ligand on a target cell or they may influence the levels or functionality of a downstream ligand or a receptor for a ligand.

[0044] As used herein, the term "subject" refers to any therapeutic target that receives the agent. The subject can be a vertebrate, for example, a mammal including a human. The term "subject" does not denote a particular age or sex. The term "subject" also refers to one or more cells of an organism, an in vitro culture of one or more tissue types, an in vitro culture of one or more cell types, ex vivo preparations, and/or a sample of biological materials such as tissue and/or biological fluids.

[0045] As used herein, the term "target cell" refers to one or more cells and/or cell types that are deleteriously affected, either directly or indirectly, by a dysregulated immune system and/or a disease process. The term "target cell" also refers to cells that are not deleteriously affected but that are cells in which it is desired that the agent interacts.

[0046] As used herein, the term "therapeutically effective amount" refers to the amount of the agent used that is of sufficient quantity to ameliorate, treat and/or inhibit one or more of a disease, disorder or a symptom thereof. The "therapeutically effective amount" will vary depending on the agent used, the route of administration of the agent and the severity of the disease, disorder or symptom thereof. The subject's age, weight and genetic make-up may also influence the amount of the agent that will be a therapeutically effective amount.

[0047] As used herein, the terms "treat", "treatment" and "treating" refer to obtaining a desired pharmacologic and/or physiologic effect. The effect may be prophylactic in terms of completely or partially preventing an occurrence of a disease, disorder or symptom thereof and/or the effect may be therapeutic in providing a partial or complete amelioration or inhibition of a disease, disorder, or symptom thereof. Additionally, the term "treatment" refers to any treatment of a disease, disorder, or symptom thereof in a subject and includes: (a) preventing the disease from occurring in a subject which may be predisposed to the disease but has not yet been diagnosed as having it; (b) inhibiting the disease, i.e., arresting its development; and, (c) ameliorating the disease.

[0048] As used herein, the terms "unit dosage form" and "unit dose" refer to a physically discrete unit that is suitable as a unitary dose for patients. Each unit contains a predetermined quantity of the agent and optionally, one or more suitable pharmaceutically acceptable carriers, one or more excipients, one or more additional active ingredients, or combinations thereof. The amount of agent within each unit is a therapeutically effective amount.

[0049] In embodiments of the present disclosure, the pharmaceutical compositions disclosed herein comprise an agent as described above in a total amount by weight of the composition of about 0.1% to about 95%. For example, the amount of the agent by weight of the pharmaceutical composition may be about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 1.1%, about 1.2%, about 1.3%, about 1.4%, about 1.5%, about 1.6%, about 1.7%, about 1.8%, about 1.9%, about 2%, about 2.1%, about 2.2%, about 2.3%, about 2.4%, about 2.5%, about 2.6%, about 2.7%, about 2.8%, about 2.9%, about 3%, about 3.1%, about 3.2%, about 3.3%, about 3.4%, about 3.5%, about 3.6%, about 3.7%, about 3.8%, about 3.9%, about 4%, about 4.1%, about 4.2%, about 4.3%, about 4.4%, about 4.5%, about 4.6%, about 4.7%, about 4.8%. about 4.9%, about 5%, about 5.1%, about 5.2%, about 5.3%, about 5.4%, about 5.5%, about 5.6%, about 5.7%, about 5.8%, about 5.9%, about 6%, about 6.1%, about 6.2%, about 6.3%, about 6.4%, about 6.5%, about 6.6%, about 6.7%, about 6.8%, about 6.9%, about 7%, about 7.1%, about 7.2%, about 7.3%, about 7.4%, about 7.5%, about 7.6%, about 7.7%, about 7.8%, about 7.9%, about 8%, about 8.1%, about 8.2%, about 8.3%, about 8.4%, about 8.5%, about 8.6%, about 8.7%, about 8.8%, about 8.9%, about 9%, about 9.1%, about 9.2%, about 9.3%, about 9.4%, about 9.5%, about 9.6%, about 9.7%, about 9.8%, about 9.9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90% or about 95%.

[0050] Where a range of values is provided herein, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the disclosure. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges, and are also, encompassed within the disclosure, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure.

[0051] In some embodiments of the present disclosure, an agent is a plasmid vector for introducing into a target cell for reproduction or transcription of an insert that comprises one or more nucleotide sequences that are carried within the plasmid vector. In some embodiments of the present disclosure, the plasmid vector is a viral vector. In some embodiments of the present disclosure, the vector is an adeno-associated virus vector.

[0052] In some embodiments of the present disclosure, the insert comprises one or more nucleotide sequences that encode for production of a precursor protein. The precursor protein is substantially similar, or substantially the same, as a translational product of a gene whose expression results in increased production of a protein that can participate in the innate immune system and/or the acquired immune system. In some embodiments of the present disclosure, the precursor protein is susceptible to one or more post-translational modification processes to create a protein product that acts as a membrane-bound receptor protein. The protein product can bind with extra-cellular ligands and initiate one or more intracellular signal pathways that change the metabolism of the associated cell.

[0053] In some embodiments of the present disclosure, the precursor protein is substantially similar, or substantially the same, as the translational product of the TLR3 gene. Following one or more post-translational modification processes, the precursor protein is converted into a protein product that has substantially similar, or substantially the same, function as the toll-like receptor 3 (TLR3) protein.

[0054] In some embodiments of the present disclosure, the precursor protein is a translational product of a nucleotide sequence that is substantially similar or substantially the same as SEQ ID No. 1, provided herein below.

[0055] In some embodiments of the present disclosure, the precursor protein has an amino acid sequence that is substantially similar or substantially the same as SEQ ID No. 2, provided herein below.

[0056] Without being bound by any particular theory, the precursor protein can be subjected to one or more post-translational modification processes within a subject's cell so that it is converted into a TLR3 protein product that has the same functionality and bioavailability as a TLR3 protein that a subject would produce without the embodiments of the present disclosure.

[0057] The present disclosure relates to one or more agents, therapies, treatments, and methods of use of the agents and/or therapies and/or treatments for initiating or upregulating production of the TLR3 precursor protein. Some embodiments of the present disclosure relate to methods for making a complex between at least one particle of an agent and at least one target cell of a subject for initiating or increasing production of the TLR3 precursor protein. The at least one target cell can be a capable cell or an incapable cell. For clarity, a capable cell is a cell that already has the capability to produce, and do or may have produced, the precursor protein. Therefore, the administration of the one or more vectors may increase the production of the precursor protein within those capable cells. The one or more vectors may also be expressed in an incapable cell. For clarity, an incapable cell is a cell of the subject that does not produce the precursor protein when the subject is in a homeostatic state, an infection state or a disease state. Incapable cells are not capable of producing the precursor protein without the one or more vectors of the present disclosure being expressed therein. The administration of the one or more vectors may induce the incapable cells to start producing the precursor protein. The embodiments of the present disclosure may cause at least a portion of a subject's capable cells and a portion of incapable cells to produce, in aggregate, a greater amount of the precursor protein which may then be subjected to one or more post-translational modification processes to produce a functional and bioavailable TLR3 protein product. As such, the embodiments of the present disclosure can be used as a therapy or a treatment for a subject that has a condition whereby the subject's immune system is, or is likely to become, dysregulated. For example, it is known that administering TLR3 agonists in animal models increases the efficacy of certain checkpoint protein inhibitors. Furthermore, it is known that the tumor cells of certain types of cancers may have dysfunctional TLR3 proteins. As such, the embodiments of the present disclosure may enhance the efficacy of therapies that include the administration of one or more checkpoint protein inhibitors, such as therapies that are directed at treating various types of cancer. Furthermore, the embodiments of the present disclosure may also increase the population of functional TLR3 protein products that are present on the surface of the subject's cells, including tumor cells. This may increase the efficacy of chemotherapy agents that acts as ligands for TLR3, such as those chemotherapy agents that may mimic or form segments of double stranded RNA.

[0058] In some embodiments of the present disclosure, the agent can be administered to the subject by an intravenous route, an intramuscular route, an intraperitoneal route, an intrathecal route, an intravesical route, a topical route, an intranasal route, a transmucosal route, a pulmonary route, and combinations thereof.

[0059] In some embodiments of the present disclosure, the agent can be administered to the subject by pipetting a dose of the agent into an in vitro cell culture, perfusing or immersing an ex vivo cell or tissue preparation with a solution that comprises the agent, mixing a biological fluid sample with a solution or substrate that comprises the agent, or combinations thereof.

[0060] Some embodiments of the present disclosure relate to an agent that can be administered to a subject with a condition that could benefit from an increased production of a TLR3 precursor protein that can be modified into a functional TLR3 protein. When a therapeutically effective amount of the agent is administered to the subject, the capable and incapable cells of the subject may increase the translational production of the TLR3 precursor protein, which then may be modified into functional and bioavailable TLR3 membrane-associated receptor protein.

[0061] In some embodiments of the present disclosure, administering a therapeutic amount of the agent to a subject upregulates the production, functionality or both of a TLR3 protein product by increasing the translational production of the precursor protein.

[0062] In some embodiments of the present disclosure, the agent is a vector used for gene therapy. The gene therapy is useful for increasing the subject's endogenous production of the TLR3 precursor protein. For example, the vector can contain one or more nucleotide sequences that that cause increased production of the precursor protein in both capable cells and incapable cells in which the vector is expressed.

[0063] In some embodiments of the present disclosure, the vector used for gene therapy is a virus that can be enveloped or not, replication effective or not, or combinations thereof. In some embodiments of the present disclosure, the vector is a virus that is not enveloped and not replication effective. In some embodiments of the present disclosure, the vector is a virus of the Paroviridae family. In some embodiments of the present disclosure, the vector is a virus of the genus Dependoparvaovirus. In some embodiments of the present disclosure, the vector is an adeno-associated virus (AAV). In some embodiments of the present disclosure, the vector is a recombinant AAV.

[0064] The embodiments of the present disclosure also relate to administering a therapeutically effective amount of the agent. In some embodiments of the present disclosure, the therapeutically effective amount of the agent that is administered to a patient is between about 10 and about 1.times.10.sup.16 TCID.sub.50/kg (50% tissue culture infective dose per kilogram of the patient's body weight). In some embodiments of the present disclosure, the therapeutically effective amount of the agent that is administered to the patient is about 1.times.10.sup.13 TCID.sub.50/kg. In some embodiments of the present disclosure, the therapeutically effective amount of the agent that is administered to a patient is measured in TPC/kg (total particle count of the agent per kilogram of the patient's body weight). In some embodiments the therapeutically effective amount of the agent is between about 10 and about 1.times.10.sup.16 TCP/kg.

[0065] Some embodiments of the present disclosure relate to a method for making a complex within a subject. The method comprises a step of administering a therapeutically effective amount of the agent to the subject. The complex comprises at least one particle of agent and one or more target cells. When the complex is formed, it affects a change in metabolism of the one or more target cells, which results in the target cells upregulating the production of the TLR3 precursor protein. Examples of a target cell include both capable cells and incapable cells, such as but not limited to: an innate immune cell, an acquired immune cell, an adrenal gland cell; a bile duct cell; a chondrocyte; a cochlear cell; a corneal cell; an endocardium cell; an endometrial cell; an endothelial cell; an epithelial cell; a fibroblast; a hair follicle cell; a hepatocyte; a lymph node cell; a mucosal cell; a myocyte; a neuron; a glomeruli cell; an optic nerve cell; an osteoblast; an ovarian tissue cell; a pancreatic islet beta cell; a pericardium cell; a platelet; a red blood cell (RBC); a retinal cell; a scleral cell; a Schwann cell; a T cell; a testicular tissue cell; a thyroid gland cell; a uveal cell; a tumor cell, or combinations thereof.

[0066] Some embodiments of the present disclosure relate to a therapy, or method of treating a condition, that can be administered to a subject with the condition. The therapy comprises a step of administering to the subject a therapeutically effective amount of an agent that will upregulate the subject's production of the TLR3 precursor protein. The increased production of the TLR3 precursor protein may result in increased levels of functional and bioavailable TLR3 protein product, which may reduce deleterious effects of the condition upon the subject.

[0067] Below are examples of nucleotide sequences of each may be present in the insert. As will be appreciated by those skilled in the art, minor modifications, substitutions or replacements of a select few nucleotides or amino acids in the sequences provided below will not substantially impact the physiologic or biologic effect of such modified sequences, as compared to the sequences provided herein below. Any such modified sequences are also contemplated by the present disclosure.

TABLE-US-00001 (nucleotide sequence for transcription of precursor protein) SEQ ID No. 1 atgcgccaga ccctgccgtg catttatttt tggggcggcc tgctgccgtt tggcatgctg 60 tgcgcgagca gcaccaccaa atgcaccgtg agccatgaag tggcggattg cagccatctg 120 aaactgaccc aggtgccgga tgatctgccg accaacatta ccgtgctgaa cctgacccat 180 aaccagctgc gccgcctgcc ggcggcgaac tttacccgct atagccagct gaccagcctg 240 gatgtgggct ttaacaccat tagcaaactg gaaccggaac tgtgccagaa actgccgatg 300 ctgaaagtgc tgaacctgca gcataacgaa ctgagccagc tgagcgataa aacctttgcg 360 ttttgcacca acctgaccga actgcatctg atgagcaaca gcattcagaa aattaaaaac 420 aacccgtttg tgaaacagaa aaacctgatt accctggatc tgagccataa cggcctgagc 480 agcaccaaac tgggcaccca ggtgcagctg gaaaacctgc aggaactgct gctgagcaac 540 aacaaaattc aggcgctgaa aagcgaagaa ctggatattt ttgcgaacag cagcctgaaa 600 aaactggaac tgagcagcaa ccagattaaa gaatttagcc cgggctgctt tcatgcgatt 660 ggccgcctgt ttggcctgtt tctgaacaac gtgcagctgg gcccgagcct gaccgaaaaa 720 ctgtgcctgg aactggcgaa caccagcatt cgcaacctga gcctgagcaa cagccagctg 780 agcaccacca gcaacaccac ctttctgggc ctgaaatgga ccaacctgac catgctggat 840 ctgagctata acaacctgaa cgtggtgggc aacgatagct ttgcgtggct gccgcagctg 900 gaatattttt ttctggaata taacaacatt cagcatctgt ttagccatag cctgcatggc 960 ctgtttaacg tgcgctatct gaacctgaaa cgcagcttta ccaaacagag cattagcctg 1020 gcgagcctgc cgaaaattga tgattttagc tttcagtggc tgaaatgcct ggaacatctg 1080 aacatggaag ataacgatat tccgggcatt aaaagcaaca tgtttaccgg cctgattaac 1140 ctgaaatatc tgagcctgag caacagcttt accagcctgc gcaccctgac caacgaaacc 1200 tttgtgagcc tggcgcatag cccgctgcat attctgaacc tgaccaaaaa caaaattagc 1260 aaaattgaaa gcgatgcgtt tagctggctg ggccatctgg aagtgctgga tctgggcctg 1320 aacgaaattg gccaggaact gaccggccag gaatggcgcg gcctggaaaa catttttgaa 1380 atttatctga gctataacaa atatctgcag ctgacccgca acagctttgc gctggtgccg 1440 agcctgcagc gcctgatgct gcgccgcgtg gcgctgaaaa acgtggatag cagcccgagc 1500 ccgtttcagc cgctgcgcaa cctgaccatt ctggatctga gcaacaacaa cattgcgaac 1560 attaacgatg atatgctgga aggcctggaa aaactggaaa ttctggatct gcagcataac 1620 aacctggcgc gcctgtggaa acatgcgaac ccgggcggcc cgatttattt tctgaaaggc 1680 ctgagccatc tgcatattct gaacctggaa agcaacggct ttgatgaaat tccggtggaa 1740 gtgtttaaag atctgtttga actgaaaatt attgatctgg gcctgaacaa cctgaacacc 1800 ctgccggcga gcgtgtttaa caaccaggtg agcctgaaaa gcctgaacct gcagaaaaac 1860 ctgattacca gcgtggaaaa aaaagtgttt ggcccggcgt ttcgcaacct gaccgaactg 1920 gatatgcgct ttaacccgtt tgattgcacc tgcgaaagca ttgcgtggtt tgtgaactgg 1980 attaacgaaa cccataccaa cattccggaa ctgagcagcc attatctgtg caacaccccg 2040 ccgcattatc atggctttcc ggtgcgcctg tttgatacca gcagctgcaa agatagcgcg 2100 ccgtttgaac tgttttttat gattaacacc agcattctgc tgatttttat ttttattgtg 2160 ctgctgattc attttgaagg ctggcgcatt agcttttatt ggaacgtgag cgtgcatcgc 2220 gtgctgggct ttaaagaaat tgatcgccag accgaacagt ttgaatatgc ggcgtatatt 2280 attcatgcgt ataaagataa agattgggtg tgggaacatt ttagcagcat ggaaaaagaa 2340 gatcagagcc tgaaattttg cctggaagaa cgcgattttg aagcgggcgt gtttgaactg 2400 gaagcgattg tgaacagcat taaacgcagc cgcaaaatta tttttgtgat tacccatcat 2460 ctgctgaaag atccgctgtg caaacgcttt aaagtgcatc atgcggtgca gcaggcgatt 2520 gaacagaacc tggatagcat tattctggtg tttctggaag aaattccgga ttataaactg 2580 aaccatgcgc tgtgcctgcg ccgcggcatg tttaaaagcc attgcattct gaactggccg 2640 gtgcagaaag aacgcattgg cgcgtttcgc cataaactgc aggtggcgct gggcagcaaa 2700 aacagcgtgc attaa 2715 (amino acid sequence for TRL3 precursor protein) SEQ ID No. 2 MRQTLPCIYFWGGLLPFGMLCASSTTKCTVSHEVADCSHLKLTQVPDDLPTNITVLNLTH NQLRRLPAANFTRYSQLTSLDVGFNTISKLEPELCQKLPMLKVLNLQHNELSQLSDKTFA FCTNLTELHLMSNSIQKIKNNPFVKQKNLITLDLSHNGLSSTKLGTQVQLENLQELLLSN NKIQALKSEELDIFANSSLKKLELSSNQIKEFSPGCFHAIGRLFGLFLNNVQLGPSLTEK LCLELANTSIRNLSLSNSQLSTTSNTTFLGLKWTNLTMLDLSYNNLNVVGNDSFAWLPQL EYFFLEYNNIQHLFSHSLHGLFNVRYLNLKRSFTKQSISLASLPKIDDFSFQWLKCLEHL NMEDNDIPGIKSNMFTGLINLKYLSLSNSFTSLRTLTNETFVSLAHSPLHILNLTKNKIS KIESDAFSWLGHLEVLDLGLNEIGQELTGQEWRGLENIFEIYLSYNKYLQLTRNSFALVP SLQRLMLRRVALKNVDSSPSPFQPLRNLTILDLSNNNIANINDDMLEGLEKLEILDLQHN NLARLWKHANPGGPIYFLKGLSHLHILNLESNGFDEIPVEVFKDLFELKIIDLGLNNLNT LPASVFNNQVSLKSLNLQKNLITSVEKKVFGPAFRNLIELDMRFNPFDCTCESIAWFVNW INETHTNIPELSSHYLCNTPPHYHGFPVRLFDTSSCKDSAPFELFFMINTSILLIFIFIV LLIHFEGWRISFYWNVSVHRVLGFKEIDRQIhQFEYAAYIIHAYKDKDWVWEHFSSMEKE DQSLKFCLEERDFEAGVFELEAIVNSIKRSRKIIFVITHHLLKDPLCKRFKVHHAVQQAI EQNLDSIILVFLEEIPDYKLNHALCLRRGMFKSHCILNWPVQKERIGAFRHKLQVALGSK NSVH (nucleotide sequence for plasmid vector) SEQ ID No. 3 cagcagctgc gcgctcgctc gctcactgag gccgcccggg caaagcccgg gcgtcgggcg 60 acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc 120 atcactaggg gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca 180 tgctctagga cattgattat tgactagtgg agttccgcgt tacataactt acggtaaatg 240 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc 300 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa 360 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca 420 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta 480 cttggcagta catctacgta ttagtcatcg ctattaccat ggtcgaggtg agccccacgt 540 tctgcttcac tctccccatc tcccccccct ccccaccccc aattttgtat ttatttattt 600 tttaattatt ttgtgcagcg atgggggcgg gggggggggg gggcgcgcgc caggcggggc 660 ggggcggggc gaggggcggg gcggggcgag gcggagaggt gcggcggcag ccaatcagag 720 cggcgcgctc cgaaagtttc cttttatggc gaggcggcgg cggcggcggc cctataaaaa 780 gcgaagcgcg cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc 840 cgccgcctcg cgccgcccgc cccggctctg actgaccgcg ttactaaaac aggtaagtcc 900 ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc cctcctcacg gcgagcgctg 960 ccacgtcaga cgaagggcgc agcgagcgtc ctgatccttc cgcccggacg ctcaggacag 1020 cggcccgctg ctcataagac tcggccttag aaccccagta tcagcagaag gacattttag 1080 gacgggactt gggtgactct agggcactgg ttttctttcc agagagcgga acaggcgagg 1140 aaaagtagtc ccttctcggc gattctgcgg agggatctcc gtggggcggt gaacgccgat 1200 gatgcctcta ctaaccatgt tcatgttttc tttttttttc tacaggtcct gggtgacgaa 1260 cagggtaccg ccaccatggt gtccaaggga gaggagctgt tcaccggagt ggtgcccatc 1320 ctggtggagc tggacggcga tgtgaatggc cacaagttta gcgtgtccgg agagggagag 1380 ggcgacgcaa cctacggcaa gctgacactg aagttcatct gcaccacagg caagctgccc 1440 gtgccttggc caaccctggt gaccacactg acatacggcg tgcagtgttt ttctcggtat 1500 ccagaccaca tgaagcagca cgatttcttt aagagcgcca tgcccgaggg ctacgtgcag 1560 gagaggacaa tcttctttaa ggacgatggc aactataaga ccagagccga ggtgaagttc 1620 gagggcgaca cactggtgaa ccggatcgag ctgaagggca tcgactttaa ggaggatggc 1680 aatatcctgg gccacaagct ggagtacaac tataattccc acaacgtgta catcatggcc 1740 gataagcaga agaacggcat caaggtgaac ttcaagatcc gccacaatat cgaggacggc 1800 tctgtgcagc tggccgatca ctaccagcag aacaccccta tcggcgacgg acccgtgctg 1860 ctgcctgata atcactatct gtctacacag agcgccctgt ccaaggaccc aaacgagaag 1920 agggatcaca tggtgctgct ggagttcgtg accgcagcag gcatcacact gggcatggat 1980 gagctgtata agcgaaaaag aagatcaggt tcgggtgcgc cagtaaagca gacattaaac 2040 tttgatttgc tgaaacttgc aggtgatgta gagtcaaatc caggtccagg atccatgcgc 2100 cagaccctgc cgtgcattta tttttggggc ggcctgctgc cgtttggcat gctgtgcgcg 2160 agcagcacca ccaaatgcac cgtgagccat gaagtggcgg attgcagcca tctgaaactg 2220 acccaggtgc cggatgatct gccgaccaac attaccgtgc tgaacctgac ccataaccag 2280 ctgcgccgcc tgccggcggc gaactttacc cgctatagcc agctgaccag cctggatgtg 2340 ggctttaaca ccattagcaa actggaaccg gaactgtgcc agaaactgcc gatgctgaaa 2400 gtgctgaacc tgcagcataa cgaactgagc cagctgagcg ataaaacctt tgcgttttgc 2460 accaacctga ccgaactgca tctgatgagc aacagcattc agaaaattaa aaacaacccg 2520 tttgtgaaac agaaaaacct gattaccctg gatctgagcc ataacggcct gagcagcacc 2580 aaactgggca cccaggtgca gctggaaaac ctgcaggaac tgctgctgag caacaacaaa 2640 attcaggcgc tgaaaagcga agaactggat atttttgcga acagcagcct gaaaaaactg 2700 gaactgagca gcaaccagat taaagaattt agcccgggct gctttcatgc gattggccgc 2760 ctgtttggcc tgtttctgaa caacgtgcag ctgggcccga gcctgaccga aaaactgtgc 2820 ctggaactgg cgaacaccag cattcgcaac ctgagcctga gcaacagcca gctgagcacc 2880 accagcaaca ccacctttct gggcctgaaa tggaccaacc tgaccatgct ggatctgagc 2940 tataacaacc tgaacgtggt gggcaacgat agctttgcgt ggctgccgca gctggaatat 3000 ttttttctgg aatataacaa cattcagcat ctgtttagcc atagcctgca tggcctgttt 3060 aacgtgcgct atctgaacct gaaacgcagc tttaccaaac agagcattag cctggcgagc 3120 ctgccgaaaa ttgatgattt tagctttcag tggctgaaat gcctggaaca tctgaacatg 3180 gaagataacg atattccggg cattaaaagc aacatgttta ccggcctgat taacctgaaa 3240 tatctgagcc tgagcaacag ctttaccagc ctgcgcaccc tgaccaacga aacctttgtg 3300 agcctggcgc atagcccgct gcatattctg aacctgacca aaaacaaaat tagcaaaatt 3360 gaaagcgatg cgtttagctg gctgggccat ctggaagtgc tggatctggg cctgaacgaa 3420 attggccagg aactgaccgg ccaggaatgg cgcggcctgg aaaacatttt tgaaatttat 3480 ctgagctata acaaatatct gcagctgacc cgcaacagct ttgcgctggt gccgagcctg 3540 cagcgcctga tgctgcgccg cgtggcgctg aaaaacgtgg atagcagccc gagcccgttt 3600 cagccgctgc gcaacctgac cattctggat ctgagcaaca acaacattgc gaacattaac 3660 gatgatatgc tggaaggcct ggaaaaactg gaaattctgg atctgcagca taacaacctg 3720 gcgcgcctgt ggaaacatgc gaacccgggc ggcccgattt attttctgaa aggcctgagc 3780 catctgcata ttctgaacct ggaaagcaac ggctttgaga aattccggtg gaagtgttta 3840 aagatctgtt tgaactgaaa attattgatc tgggcctgaa caacctgaac accctgccgg 3900 gagcgtgttt aacaaccagg tgagcctgaa aagcctgaac ctgcagaaaa acctgattac 3960 cagcgtggaa aaaaaagtgt ttggcccggc gtttcgcaac ctgaccgaac tggatatgcg 4020 ctttaacccg tttgattgca cctgcgaaag cattgcgtgg tttgtgaact ggattaacga 4080

aacccatacc aacattccgg aactgagcag ccattatctg tgcaacaccc cgccgcatta 4140 tcatggcttt ccggtgcgcc tgtttgatac cagcagctgc aaagatagcg cgccgtttga 4200 actgtttttt atgattaaca ccagcattct gctgattttt atttttattg tgctgctgat 4260 tcattttgaa ggctggcgca ttagctttta ttggaacgtg agcgtgcatc gcgtgctggg 4320 ctttaaagaa attgatcgcc agaccgaaca gtttgaatat gcggcgtata ttattcatgc 4380 gtataaagat aaagattggg tgtgggaaca ttttagcagc atggaaaaag aagatcagag 4440 cctgaaattt tgcctggaag aacgcgattt tgaagcgggc gtgtttgaac tggaagcgat 4500 tgtgaacagc attaaacgca gccgcaaaat tatttttgtg attacccatc atctgctgaa 4560 agatcgctgt gcaaacgctt taaagtgcat catgcggtgc agcaggcgat tgaacagaac 4620 ctggatagca ttattctggt gtttctggaa gaaattccgg attataaact gaaccatgcg 4680 ctgtgcctgc gccgcggcat gtttaaaagc cattgcattc tgaactggcc ggtgcagaaa 4740 gaacgcattg gcgcgtttcg ccataaactg caggtggcgc tgggcagcaa aaacagcgtg 4800 cattaatacc cctatgacgt gcctgattat gcctgactct agaataatca acctctggat 4860 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt 4920 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc 4980 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg 5040 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc 5100 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa 5160 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat 5220 tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc 5280 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt 5340 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag 5400 acgagtcgga tctccctttg ggccgcctcc ccgcctaagc ttatcgatac cgtcgagatc 5460 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac 5520 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc 5580 ttatcatgtc tggatctcga cctcgactag agcatggcta cgtagataag tagcatggcg 5640 ggttaatcat taactacaag gaacccctag tgatggagtt ggccactccc tctctgcgcg 5700 ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg 5760 cggcctcagt gagcgagcga gcgcgccagc tggcgtaata gcgaagaggc ccgcaccgat 5820 cgcccttccc aacagttgcg cagcctgaat ggcgaatgga attccagacg attgagcgtc 5880 aaaatgtagg tatttccatg agcgtttttc ctgttgcaat ggctggcggt aatattgttc 5940 tggatattac cagcaaggcc gatagtttga gttcttctac tcaggcaagt gatgttatta 6000 ctaatcaaag aagtattgcg acaacggtta atttgcgtga tggacagact cttttactcg 6060 gtggcctcac tgattataaa aacacttctc aggattctgg cgtaccgttc ctgtctaaaa 6120 tccctttaat cggcctcctg tttagctccc gctctgattc taacgaggaa agcacgttat 6180 acgtgctcgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt 6240 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc 6300 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg 6360 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat 6420 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg 6480 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct 6540 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa 6600 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt 6660 taaatatttg cttatacaat cttcctgttt ttggggcttt tctgattatc aaccggggta 6720 catatgattg acatgctagt tttacgatta ccgttcatcg attctcttgt ttgctccaga 6780 ctctcaggca atgacctgat agcctttgta gagacctctc aaaaatagct accctctccg 6840 gcatgaattt atcagctaga acggttgaat atcatattga tggtgatttg actgtctccg 6900 gcctttctca cccgtttgaa tctttaccta cacattactc aggcattgca tttaaaatat 6960 atgagggttc taaaaatttt tatccttgcg ttgaaataaa ggcttctccc gcaaaagtat 7020 tacagggtca taatgttttt ggtacaaccg atttagcttt atgctctgag gctttattgc 7080 ttaattttgc taattctttg ccttgcctgt atgatttatt ggatgttgga attcctgatg 7140 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatggtg cactctcagt 7200 acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac 7260 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc 7320 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc 7380 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca 7440 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat 7500 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa 7560 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt 7620 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag 7680 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt 7740 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg 7800 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag 7860 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta 7920 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg 7980 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta 8040 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac 8100 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt 8160 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca 8220 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag 8280 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta 8340 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag 8400 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt 8460 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat 8520 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta 8580 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa 8640 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt 8700 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag 8760 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta 8820 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca 8880 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag 8940 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa 9000 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga 9060 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc 9120 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc 9180 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt 9240 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt 9300 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag 9360 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa 9420 tg 9422 (nucleotide sequence for inverted terminal repeat 1) SEQ ID 4 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt 60 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact 120 aggggttcct 130 (nucleotide sequence for inverted terminal repeat 2) SEQ ID 5 aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg 60 ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc 120 gagcgcgc 128 (nucleotide sequence for eGFP mouse-human optimized) SEQ ID 6 cagcagctgc gcgctcgctc gctcactgag gccgcccggg caaagcccgg gcgtcgggcg 60 acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc 120 atcactaggg gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca 180 tgctctagga cattgattat tgactagtgg agttccgcgt tacataactt acggtaaatg 240 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc 300 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa 360 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca 420 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta 480 cttggcagta catctacgta ttagtcatcg ctattaccat ggtcgaggtg agccccacgt 540 tctgcttcac tctccccatc tcccccccct ccccaccccc aattttgtat ttatttattt 600 tttaattatt ttgtgcagcg atgggggcgg gggggggggg gggcgcgcgc caggcggggc 660 ggggcggggc gaggggcggg gcggggcgag gcggagaggt gcggcggcag ccaatcagag 720 cggcgcgctc cgaaagtttc cttttatggc gaggcggcgg cggcggcggc cctataaaaa 780 gcgaagcgcg cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc 840 cgccgcctcg cgccgcccgc cccggctctg actgaccgcg ttactaaaac aggtaagtcc 900 ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc cctcctcacg gcgagcgctg 960 ccacgtcaga cgaagggcgc agcgagcgtc ctgatccttc cgcccggacg ctcaggacag 1020 cggcccgctg ctcataagac tcggccttag aaccccagta tcagcagaag gacattttag 1080 gacgggactt gggtgactct agggcactgg ttttctttcc agagagcgga acaggcgagg 1140 aaaagtagtc ccttctcggc gattctgcgg agggatctcc gtggggcggt gaacgccgat 1200 gatgcctcta ctaaccatgt tcatgttttc tttttttttc tacaggtcct gggtgacgaa 1260 cagggtaccg ccaccatggt gtccaaggga gaggagctgt tcaccggagt ggtgcccatc 1320 ctggtggagc tggacggcga tgtgaatggc cacaagttta gcgtgtccgg agagggagag 1380 ggcgacgcaa cctacggcaa gctgacactg aagttcatct gcaccacagg caagctgccc 1440 gtgccttggc caaccctggt gaccacactg acatacggcg tgcagtgttt ttctcggtat 1500 ccagaccaca tgaagcagca cgatttcttt aagagcgcca tgcccgaggg ctacgtgcag 1560

gagaggacaa tcttctttaa ggacgatggc aactataaga ccagagccga ggtgaagttc 1620 gagggcgaca cactggtgaa ccggatcgag ctgaagggca tcgactttaa ggaggatggc 1680 aatatcctgg gccacaagct ggagtacaac tataattccc acaacgtgta catcatggcc 1740 gataagcaga agaacggcat caaggtgaac ttcaagatcc gccacaatat cgaggacggc 1800 tctgtgcagc tggccgatca ctaccagcag aacaccccta tcggcgacgg acccgtgctg 1860 ctgcctgata atcactatct gtctacacag agcgccctgt ccaaggaccc aaacgagaag 1920 agggatcaca tggtgctgct ggagttcgtg accgcagcag gcatcacact gggcatggat 1980 gagctgtata ag 1992 (nucleic acid sequence for HA tag) SEQ ID 7 tacccctatg acgtgcctga ttatgcc 27

Example 1--Expression Cassette

[0068] Expression cassettes for expressing a precursor protein were synthesized by Genscript. Each cassette contained a signal peptide, the precursor protein that may be followed by a self-cleaving 2A peptide sequence, a signal peptide and the human lambda constant domain. The synthesized precursor protein expression cassettes were cloned into the pAVA-00200 plasmid backbone containing the CASI promoter1, multiple cloning site (MCS), Woodchuck Hepatitis Virus post-transcriptional regulatory element (WPRE), Simian virus 40 (SV40) polyadenylation (polyA) sequence all flanked by the AAV2 inverted terminal repeats (ITR). pAVA-00200 was cut with the restriction enzymes KpnI and XbaI in the MCS and separated on a 1% agarose gel. The band of interest was excised and purified using a gel extraction kit. Each precursor protein expression cassette was amplified by PCR using Taq polymerase and the PCR products were gel purified and the bands on interest were also excised and purified using a gel extraction kit. These PCR products contained the precursor protein expression cassettes in addition to 15 base pair 5' and 3' overhangs that align with the ends of the linearized pAVA-00200 backbone. Using in-fusion cloning2, the amplified precursor protein or expression cassettes are integrated with the pAVA-00200 backbone via homologous recombination. The resulting plasmid vectors contained the following 5' ITR, CASI promoter the precursor protein expression cassette, WPRE, SV40 polyA and ITR 3', per SEQ ID No. 3.

Example 2--Experimental Data

[0069] Table 1 below summarizes data obtained from HEK293 cells that were administered an AAV vector of SEQ ID No. 3 and TLR3 protein levels were compared against control HEK293 cells that did not receive the AAV vector of SEQ ID No. 3 (control). Speicifically, about 2.5.times.10.sup.6 HEK293 confluent cells were administered about 1.times.10.sup.8 AAV vectors of SEQ ID No. 3. About 48 hours later, expression levels of human TLR3 mRNA were determined by qRT-PCR, and converted to expressed TLR3 protein levels.

TABLE-US-00002 TABLE 1 TLR3 Protein Levels Molecule Control (ag) AAV recipient (ag) P-value TLR3 1 23 0.0076 n.b. ag = attogram 1 .times. 10.sup.-18 grams

[0070] As shown in Table 1, cells that received the AAV of SEQ ID No. 3 that include SEQ ID No. 1 that encoded for increased production of a TLR3 precursor protein demonstrated statistically significant higher amounts of TLR3 than the control cells that did not receive the AAV. Without being bound by any particular theory, the cells that were administered the AAV had higher levels of TLR3 than cells that did not receive the AAV.

Sequence CWU 1

1

712715DNAArtificial SequenceSynthetic Sequence 1atgcgccaga ccctgccgtg catttatttt tggggcggcc tgctgccgtt tggcatgctg 60tgcgcgagca gcaccaccaa atgcaccgtg agccatgaag tggcggattg cagccatctg 120aaactgaccc aggtgccgga tgatctgccg accaacatta ccgtgctgaa cctgacccat 180aaccagctgc gccgcctgcc ggcggcgaac tttacccgct atagccagct gaccagcctg 240gatgtgggct ttaacaccat tagcaaactg gaaccggaac tgtgccagaa actgccgatg 300ctgaaagtgc tgaacctgca gcataacgaa ctgagccagc tgagcgataa aacctttgcg 360ttttgcacca acctgaccga actgcatctg atgagcaaca gcattcagaa aattaaaaac 420aacccgtttg tgaaacagaa aaacctgatt accctggatc tgagccataa cggcctgagc 480agcaccaaac tgggcaccca ggtgcagctg gaaaacctgc aggaactgct gctgagcaac 540aacaaaattc aggcgctgaa aagcgaagaa ctggatattt ttgcgaacag cagcctgaaa 600aaactggaac tgagcagcaa ccagattaaa gaatttagcc cgggctgctt tcatgcgatt 660ggccgcctgt ttggcctgtt tctgaacaac gtgcagctgg gcccgagcct gaccgaaaaa 720ctgtgcctgg aactggcgaa caccagcatt cgcaacctga gcctgagcaa cagccagctg 780agcaccacca gcaacaccac ctttctgggc ctgaaatgga ccaacctgac catgctggat 840ctgagctata acaacctgaa cgtggtgggc aacgatagct ttgcgtggct gccgcagctg 900gaatattttt ttctggaata taacaacatt cagcatctgt ttagccatag cctgcatggc 960ctgtttaacg tgcgctatct gaacctgaaa cgcagcttta ccaaacagag cattagcctg 1020gcgagcctgc cgaaaattga tgattttagc tttcagtggc tgaaatgcct ggaacatctg 1080aacatggaag ataacgatat tccgggcatt aaaagcaaca tgtttaccgg cctgattaac 1140ctgaaatatc tgagcctgag caacagcttt accagcctgc gcaccctgac caacgaaacc 1200tttgtgagcc tggcgcatag cccgctgcat attctgaacc tgaccaaaaa caaaattagc 1260aaaattgaaa gcgatgcgtt tagctggctg ggccatctgg aagtgctgga tctgggcctg 1320aacgaaattg gccaggaact gaccggccag gaatggcgcg gcctggaaaa catttttgaa 1380atttatctga gctataacaa atatctgcag ctgacccgca acagctttgc gctggtgccg 1440agcctgcagc gcctgatgct gcgccgcgtg gcgctgaaaa acgtggatag cagcccgagc 1500ccgtttcagc cgctgcgcaa cctgaccatt ctggatctga gcaacaacaa cattgcgaac 1560attaacgatg atatgctgga aggcctggaa aaactggaaa ttctggatct gcagcataac 1620aacctggcgc gcctgtggaa acatgcgaac ccgggcggcc cgatttattt tctgaaaggc 1680ctgagccatc tgcatattct gaacctggaa agcaacggct ttgatgaaat tccggtggaa 1740gtgtttaaag atctgtttga actgaaaatt attgatctgg gcctgaacaa cctgaacacc 1800ctgccggcga gcgtgtttaa caaccaggtg agcctgaaaa gcctgaacct gcagaaaaac 1860ctgattacca gcgtggaaaa aaaagtgttt ggcccggcgt ttcgcaacct gaccgaactg 1920gatatgcgct ttaacccgtt tgattgcacc tgcgaaagca ttgcgtggtt tgtgaactgg 1980attaacgaaa cccataccaa cattccggaa ctgagcagcc attatctgtg caacaccccg 2040ccgcattatc atggctttcc ggtgcgcctg tttgatacca gcagctgcaa agatagcgcg 2100ccgtttgaac tgttttttat gattaacacc agcattctgc tgatttttat ttttattgtg 2160ctgctgattc attttgaagg ctggcgcatt agcttttatt ggaacgtgag cgtgcatcgc 2220gtgctgggct ttaaagaaat tgatcgccag accgaacagt ttgaatatgc ggcgtatatt 2280attcatgcgt ataaagataa agattgggtg tgggaacatt ttagcagcat ggaaaaagaa 2340gatcagagcc tgaaattttg cctggaagaa cgcgattttg aagcgggcgt gtttgaactg 2400gaagcgattg tgaacagcat taaacgcagc cgcaaaatta tttttgtgat tacccatcat 2460ctgctgaaag atccgctgtg caaacgcttt aaagtgcatc atgcggtgca gcaggcgatt 2520gaacagaacc tggatagcat tattctggtg tttctggaag aaattccgga ttataaactg 2580aaccatgcgc tgtgcctgcg ccgcggcatg tttaaaagcc attgcattct gaactggccg 2640gtgcagaaag aacgcattgg cgcgtttcgc cataaactgc aggtggcgct gggcagcaaa 2700aacagcgtgc attaa 27152904PRTArtificial SequenceSynthetic Sequence 2Met Arg Gln Thr Leu Pro Cys Ile Tyr Phe Trp Gly Gly Leu Leu Pro1 5 10 15Phe Gly Met Leu Cys Ala Ser Ser Thr Thr Lys Cys Thr Val Ser His 20 25 30Glu Val Ala Asp Cys Ser His Leu Lys Leu Thr Gln Val Pro Asp Asp 35 40 45Leu Pro Thr Asn Ile Thr Val Leu Asn Leu Thr His Asn Gln Leu Arg 50 55 60Arg Leu Pro Ala Ala Asn Phe Thr Arg Tyr Ser Gln Leu Thr Ser Leu65 70 75 80Asp Val Gly Phe Asn Thr Ile Ser Lys Leu Glu Pro Glu Leu Cys Gln 85 90 95Lys Leu Pro Met Leu Lys Val Leu Asn Leu Gln His Asn Glu Leu Ser 100 105 110Gln Leu Ser Asp Lys Thr Phe Ala Phe Cys Thr Asn Leu Thr Glu Leu 115 120 125His Leu Met Ser Asn Ser Ile Gln Lys Ile Lys Asn Asn Pro Phe Val 130 135 140Lys Gln Lys Asn Leu Ile Thr Leu Asp Leu Ser His Asn Gly Leu Ser145 150 155 160Ser Thr Lys Leu Gly Thr Gln Val Gln Leu Glu Asn Leu Gln Glu Leu 165 170 175Leu Leu Ser Asn Asn Lys Ile Gln Ala Leu Lys Ser Glu Glu Leu Asp 180 185 190Ile Phe Ala Asn Ser Ser Leu Lys Lys Leu Glu Leu Ser Ser Asn Gln 195 200 205Ile Lys Glu Phe Ser Pro Gly Cys Phe His Ala Ile Gly Arg Leu Phe 210 215 220Gly Leu Phe Leu Asn Asn Val Gln Leu Gly Pro Ser Leu Thr Glu Lys225 230 235 240Leu Cys Leu Glu Leu Ala Asn Thr Ser Ile Arg Asn Leu Ser Leu Ser 245 250 255Asn Ser Gln Leu Ser Thr Thr Ser Asn Thr Thr Phe Leu Gly Leu Lys 260 265 270Trp Thr Asn Leu Thr Met Leu Asp Leu Ser Tyr Asn Asn Leu Asn Val 275 280 285Val Gly Asn Asp Ser Phe Ala Trp Leu Pro Gln Leu Glu Tyr Phe Phe 290 295 300Leu Glu Tyr Asn Asn Ile Gln His Leu Phe Ser His Ser Leu His Gly305 310 315 320Leu Phe Asn Val Arg Tyr Leu Asn Leu Lys Arg Ser Phe Thr Lys Gln 325 330 335Ser Ile Ser Leu Ala Ser Leu Pro Lys Ile Asp Asp Phe Ser Phe Gln 340 345 350Trp Leu Lys Cys Leu Glu His Leu Asn Met Glu Asp Asn Asp Ile Pro 355 360 365Gly Ile Lys Ser Asn Met Phe Thr Gly Leu Ile Asn Leu Lys Tyr Leu 370 375 380Ser Leu Ser Asn Ser Phe Thr Ser Leu Arg Thr Leu Thr Asn Glu Thr385 390 395 400Phe Val Ser Leu Ala His Ser Pro Leu His Ile Leu Asn Leu Thr Lys 405 410 415Asn Lys Ile Ser Lys Ile Glu Ser Asp Ala Phe Ser Trp Leu Gly His 420 425 430Leu Glu Val Leu Asp Leu Gly Leu Asn Glu Ile Gly Gln Glu Leu Thr 435 440 445Gly Gln Glu Trp Arg Gly Leu Glu Asn Ile Phe Glu Ile Tyr Leu Ser 450 455 460Tyr Asn Lys Tyr Leu Gln Leu Thr Arg Asn Ser Phe Ala Leu Val Pro465 470 475 480Ser Leu Gln Arg Leu Met Leu Arg Arg Val Ala Leu Lys Asn Val Asp 485 490 495Ser Ser Pro Ser Pro Phe Gln Pro Leu Arg Asn Leu Thr Ile Leu Asp 500 505 510Leu Ser Asn Asn Asn Ile Ala Asn Ile Asn Asp Asp Met Leu Glu Gly 515 520 525Leu Glu Lys Leu Glu Ile Leu Asp Leu Gln His Asn Asn Leu Ala Arg 530 535 540Leu Trp Lys His Ala Asn Pro Gly Gly Pro Ile Tyr Phe Leu Lys Gly545 550 555 560Leu Ser His Leu His Ile Leu Asn Leu Glu Ser Asn Gly Phe Asp Glu 565 570 575Ile Pro Val Glu Val Phe Lys Asp Leu Phe Glu Leu Lys Ile Ile Asp 580 585 590Leu Gly Leu Asn Asn Leu Asn Thr Leu Pro Ala Ser Val Phe Asn Asn 595 600 605Gln Val Ser Leu Lys Ser Leu Asn Leu Gln Lys Asn Leu Ile Thr Ser 610 615 620Val Glu Lys Lys Val Phe Gly Pro Ala Phe Arg Asn Leu Thr Glu Leu625 630 635 640Asp Met Arg Phe Asn Pro Phe Asp Cys Thr Cys Glu Ser Ile Ala Trp 645 650 655Phe Val Asn Trp Ile Asn Glu Thr His Thr Asn Ile Pro Glu Leu Ser 660 665 670Ser His Tyr Leu Cys Asn Thr Pro Pro His Tyr His Gly Phe Pro Val 675 680 685Arg Leu Phe Asp Thr Ser Ser Cys Lys Asp Ser Ala Pro Phe Glu Leu 690 695 700Phe Phe Met Ile Asn Thr Ser Ile Leu Leu Ile Phe Ile Phe Ile Val705 710 715 720Leu Leu Ile His Phe Glu Gly Trp Arg Ile Ser Phe Tyr Trp Asn Val 725 730 735Ser Val His Arg Val Leu Gly Phe Lys Glu Ile Asp Arg Gln Thr Glu 740 745 750Gln Phe Glu Tyr Ala Ala Tyr Ile Ile His Ala Tyr Lys Asp Lys Asp 755 760 765Trp Val Trp Glu His Phe Ser Ser Met Glu Lys Glu Asp Gln Ser Leu 770 775 780Lys Phe Cys Leu Glu Glu Arg Asp Phe Glu Ala Gly Val Phe Glu Leu785 790 795 800Glu Ala Ile Val Asn Ser Ile Lys Arg Ser Arg Lys Ile Ile Phe Val 805 810 815Ile Thr His His Leu Leu Lys Asp Pro Leu Cys Lys Arg Phe Lys Val 820 825 830His His Ala Val Gln Gln Ala Ile Glu Gln Asn Leu Asp Ser Ile Ile 835 840 845Leu Val Phe Leu Glu Glu Ile Pro Asp Tyr Lys Leu Asn His Ala Leu 850 855 860Cys Leu Arg Arg Gly Met Phe Lys Ser His Cys Ile Leu Asn Trp Pro865 870 875 880Val Gln Lys Glu Arg Ile Gly Ala Phe Arg His Lys Leu Gln Val Ala 885 890 895Leu Gly Ser Lys Asn Ser Val His 90039422DNAArtificial SequenceSynthetic Sequence 3cagcagctgc gcgctcgctc gctcactgag gccgcccggg caaagcccgg gcgtcgggcg 60acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc 120atcactaggg gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca 180tgctctagga cattgattat tgactagtgg agttccgcgt tacataactt acggtaaatg 240gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc 300ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa 360ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca 420atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta 480cttggcagta catctacgta ttagtcatcg ctattaccat ggtcgaggtg agccccacgt 540tctgcttcac tctccccatc tcccccccct ccccaccccc aattttgtat ttatttattt 600tttaattatt ttgtgcagcg atgggggcgg gggggggggg gggcgcgcgc caggcggggc 660ggggcggggc gaggggcggg gcggggcgag gcggagaggt gcggcggcag ccaatcagag 720cggcgcgctc cgaaagtttc cttttatggc gaggcggcgg cggcggcggc cctataaaaa 780gcgaagcgcg cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc 840cgccgcctcg cgccgcccgc cccggctctg actgaccgcg ttactaaaac aggtaagtcc 900ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc cctcctcacg gcgagcgctg 960ccacgtcaga cgaagggcgc agcgagcgtc ctgatccttc cgcccggacg ctcaggacag 1020cggcccgctg ctcataagac tcggccttag aaccccagta tcagcagaag gacattttag 1080gacgggactt gggtgactct agggcactgg ttttctttcc agagagcgga acaggcgagg 1140aaaagtagtc ccttctcggc gattctgcgg agggatctcc gtggggcggt gaacgccgat 1200gatgcctcta ctaaccatgt tcatgttttc tttttttttc tacaggtcct gggtgacgaa 1260cagggtaccg ccaccatggt gtccaaggga gaggagctgt tcaccggagt ggtgcccatc 1320ctggtggagc tggacggcga tgtgaatggc cacaagttta gcgtgtccgg agagggagag 1380ggcgacgcaa cctacggcaa gctgacactg aagttcatct gcaccacagg caagctgccc 1440gtgccttggc caaccctggt gaccacactg acatacggcg tgcagtgttt ttctcggtat 1500ccagaccaca tgaagcagca cgatttcttt aagagcgcca tgcccgaggg ctacgtgcag 1560gagaggacaa tcttctttaa ggacgatggc aactataaga ccagagccga ggtgaagttc 1620gagggcgaca cactggtgaa ccggatcgag ctgaagggca tcgactttaa ggaggatggc 1680aatatcctgg gccacaagct ggagtacaac tataattccc acaacgtgta catcatggcc 1740gataagcaga agaacggcat caaggtgaac ttcaagatcc gccacaatat cgaggacggc 1800tctgtgcagc tggccgatca ctaccagcag aacaccccta tcggcgacgg acccgtgctg 1860ctgcctgata atcactatct gtctacacag agcgccctgt ccaaggaccc aaacgagaag 1920agggatcaca tggtgctgct ggagttcgtg accgcagcag gcatcacact gggcatggat 1980gagctgtata agcgaaaaag aagatcaggt tcgggtgcgc cagtaaagca gacattaaac 2040tttgatttgc tgaaacttgc aggtgatgta gagtcaaatc caggtccagg atccatgcgc 2100cagaccctgc cgtgcattta tttttggggc ggcctgctgc cgtttggcat gctgtgcgcg 2160agcagcacca ccaaatgcac cgtgagccat gaagtggcgg attgcagcca tctgaaactg 2220acccaggtgc cggatgatct gccgaccaac attaccgtgc tgaacctgac ccataaccag 2280ctgcgccgcc tgccggcggc gaactttacc cgctatagcc agctgaccag cctggatgtg 2340ggctttaaca ccattagcaa actggaaccg gaactgtgcc agaaactgcc gatgctgaaa 2400gtgctgaacc tgcagcataa cgaactgagc cagctgagcg ataaaacctt tgcgttttgc 2460accaacctga ccgaactgca tctgatgagc aacagcattc agaaaattaa aaacaacccg 2520tttgtgaaac agaaaaacct gattaccctg gatctgagcc ataacggcct gagcagcacc 2580aaactgggca cccaggtgca gctggaaaac ctgcaggaac tgctgctgag caacaacaaa 2640attcaggcgc tgaaaagcga agaactggat atttttgcga acagcagcct gaaaaaactg 2700gaactgagca gcaaccagat taaagaattt agcccgggct gctttcatgc gattggccgc 2760ctgtttggcc tgtttctgaa caacgtgcag ctgggcccga gcctgaccga aaaactgtgc 2820ctggaactgg cgaacaccag cattcgcaac ctgagcctga gcaacagcca gctgagcacc 2880accagcaaca ccacctttct gggcctgaaa tggaccaacc tgaccatgct ggatctgagc 2940tataacaacc tgaacgtggt gggcaacgat agctttgcgt ggctgccgca gctggaatat 3000ttttttctgg aatataacaa cattcagcat ctgtttagcc atagcctgca tggcctgttt 3060aacgtgcgct atctgaacct gaaacgcagc tttaccaaac agagcattag cctggcgagc 3120ctgccgaaaa ttgatgattt tagctttcag tggctgaaat gcctggaaca tctgaacatg 3180gaagataacg atattccggg cattaaaagc aacatgttta ccggcctgat taacctgaaa 3240tatctgagcc tgagcaacag ctttaccagc ctgcgcaccc tgaccaacga aacctttgtg 3300agcctggcgc atagcccgct gcatattctg aacctgacca aaaacaaaat tagcaaaatt 3360gaaagcgatg cgtttagctg gctgggccat ctggaagtgc tggatctggg cctgaacgaa 3420attggccagg aactgaccgg ccaggaatgg cgcggcctgg aaaacatttt tgaaatttat 3480ctgagctata acaaatatct gcagctgacc cgcaacagct ttgcgctggt gccgagcctg 3540cagcgcctga tgctgcgccg cgtggcgctg aaaaacgtgg atagcagccc gagcccgttt 3600cagccgctgc gcaacctgac cattctggat ctgagcaaca acaacattgc gaacattaac 3660gatgatatgc tggaaggcct ggaaaaactg gaaattctgg atctgcagca taacaacctg 3720gcgcgcctgt ggaaacatgc gaacccgggc ggcccgattt attttctgaa aggcctgagc 3780catctgcata ttctgaacct ggaaagcaac ggctttgaga aattccggtg gaagtgttta 3840aagatctgtt tgaactgaaa attattgatc tgggcctgaa caacctgaac accctgccgg 3900gagcgtgttt aacaaccagg tgagcctgaa aagcctgaac ctgcagaaaa acctgattac 3960cagcgtggaa aaaaaagtgt ttggcccggc gtttcgcaac ctgaccgaac tggatatgcg 4020ctttaacccg tttgattgca cctgcgaaag cattgcgtgg tttgtgaact ggattaacga 4080aacccatacc aacattccgg aactgagcag ccattatctg tgcaacaccc cgccgcatta 4140tcatggcttt ccggtgcgcc tgtttgatac cagcagctgc aaagatagcg cgccgtttga 4200actgtttttt atgattaaca ccagcattct gctgattttt atttttattg tgctgctgat 4260tcattttgaa ggctggcgca ttagctttta ttggaacgtg agcgtgcatc gcgtgctggg 4320ctttaaagaa attgatcgcc agaccgaaca gtttgaatat gcggcgtata ttattcatgc 4380gtataaagat aaagattggg tgtgggaaca ttttagcagc atggaaaaag aagatcagag 4440cctgaaattt tgcctggaag aacgcgattt tgaagcgggc gtgtttgaac tggaagcgat 4500tgtgaacagc attaaacgca gccgcaaaat tatttttgtg attacccatc atctgctgaa 4560agatcgctgt gcaaacgctt taaagtgcat catgcggtgc agcaggcgat tgaacagaac 4620ctggatagca ttattctggt gtttctggaa gaaattccgg attataaact gaaccatgcg 4680ctgtgcctgc gccgcggcat gtttaaaagc cattgcattc tgaactggcc ggtgcagaaa 4740gaacgcattg gcgcgtttcg ccataaactg caggtggcgc tgggcagcaa aaacagcgtg 4800cattaatacc cctatgacgt gcctgattat gcctgactct agaataatca acctctggat 4860tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt 4920ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc 4980tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg 5040caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc 5100accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa 5160ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat 5220tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc 5280tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt 5340ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag 5400acgagtcgga tctccctttg ggccgcctcc ccgcctaagc ttatcgatac cgtcgagatc 5460taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac 5520aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc 5580ttatcatgtc tggatctcga cctcgactag agcatggcta cgtagataag tagcatggcg 5640ggttaatcat taactacaag gaacccctag tgatggagtt ggccactccc tctctgcgcg 5700ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg 5760cggcctcagt gagcgagcga gcgcgccagc tggcgtaata gcgaagaggc ccgcaccgat 5820cgcccttccc aacagttgcg cagcctgaat ggcgaatgga attccagacg attgagcgtc 5880aaaatgtagg tatttccatg agcgtttttc ctgttgcaat ggctggcggt aatattgttc 5940tggatattac cagcaaggcc gatagtttga gttcttctac tcaggcaagt gatgttatta 6000ctaatcaaag aagtattgcg acaacggtta atttgcgtga tggacagact cttttactcg 6060gtggcctcac tgattataaa aacacttctc aggattctgg cgtaccgttc ctgtctaaaa 6120tccctttaat cggcctcctg tttagctccc gctctgattc taacgaggaa agcacgttat 6180acgtgctcgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt 6240gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc 6300gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg 6360gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat 6420tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg 6480ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct 6540atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa 6600aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt 6660taaatatttg cttatacaat

cttcctgttt ttggggcttt tctgattatc aaccggggta 6720catatgattg acatgctagt tttacgatta ccgttcatcg attctcttgt ttgctccaga 6780ctctcaggca atgacctgat agcctttgta gagacctctc aaaaatagct accctctccg 6840gcatgaattt atcagctaga acggttgaat atcatattga tggtgatttg actgtctccg 6900gcctttctca cccgtttgaa tctttaccta cacattactc aggcattgca tttaaaatat 6960atgagggttc taaaaatttt tatccttgcg ttgaaataaa ggcttctccc gcaaaagtat 7020tacagggtca taatgttttt ggtacaaccg atttagcttt atgctctgag gctttattgc 7080ttaattttgc taattctttg ccttgcctgt atgatttatt ggatgttgga attcctgatg 7140cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatggtg cactctcagt 7200acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac 7260gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc 7320gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc 7380ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca 7440ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat 7500tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa 7560aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt 7620tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag 7680ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt 7740tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg 7800gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag 7860aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta 7920agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg 7980acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta 8040actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac 8100accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt 8160actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca 8220cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag 8280cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta 8340gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag 8400ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt 8460tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat 8520aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta 8580gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa 8640acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt 8700tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag 8760ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta 8820atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca 8880agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag 8940cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa 9000agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga 9060acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc 9120gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc 9180ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt 9240gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt 9300gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag 9360gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa 9420tg 94224130DNAArtificial SequenceSynthetic Sequence 4ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt 60ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact 120aggggttcct 1305128DNAArtificial SequenceSynthetic Sequence 5aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg 60ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc 120gagcgcgc 12861992DNAArtificial SequenceSynthetic Sequence 6cagcagctgc gcgctcgctc gctcactgag gccgcccggg caaagcccgg gcgtcgggcg 60acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc 120atcactaggg gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca 180tgctctagga cattgattat tgactagtgg agttccgcgt tacataactt acggtaaatg 240gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc 300ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa 360ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca 420atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta 480cttggcagta catctacgta ttagtcatcg ctattaccat ggtcgaggtg agccccacgt 540tctgcttcac tctccccatc tcccccccct ccccaccccc aattttgtat ttatttattt 600tttaattatt ttgtgcagcg atgggggcgg gggggggggg gggcgcgcgc caggcggggc 660ggggcggggc gaggggcggg gcggggcgag gcggagaggt gcggcggcag ccaatcagag 720cggcgcgctc cgaaagtttc cttttatggc gaggcggcgg cggcggcggc cctataaaaa 780gcgaagcgcg cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc 840cgccgcctcg cgccgcccgc cccggctctg actgaccgcg ttactaaaac aggtaagtcc 900ggcctccgcg ccgggttttg gcgcctcccg cgggcgcccc cctcctcacg gcgagcgctg 960ccacgtcaga cgaagggcgc agcgagcgtc ctgatccttc cgcccggacg ctcaggacag 1020cggcccgctg ctcataagac tcggccttag aaccccagta tcagcagaag gacattttag 1080gacgggactt gggtgactct agggcactgg ttttctttcc agagagcgga acaggcgagg 1140aaaagtagtc ccttctcggc gattctgcgg agggatctcc gtggggcggt gaacgccgat 1200gatgcctcta ctaaccatgt tcatgttttc tttttttttc tacaggtcct gggtgacgaa 1260cagggtaccg ccaccatggt gtccaaggga gaggagctgt tcaccggagt ggtgcccatc 1320ctggtggagc tggacggcga tgtgaatggc cacaagttta gcgtgtccgg agagggagag 1380ggcgacgcaa cctacggcaa gctgacactg aagttcatct gcaccacagg caagctgccc 1440gtgccttggc caaccctggt gaccacactg acatacggcg tgcagtgttt ttctcggtat 1500ccagaccaca tgaagcagca cgatttcttt aagagcgcca tgcccgaggg ctacgtgcag 1560gagaggacaa tcttctttaa ggacgatggc aactataaga ccagagccga ggtgaagttc 1620gagggcgaca cactggtgaa ccggatcgag ctgaagggca tcgactttaa ggaggatggc 1680aatatcctgg gccacaagct ggagtacaac tataattccc acaacgtgta catcatggcc 1740gataagcaga agaacggcat caaggtgaac ttcaagatcc gccacaatat cgaggacggc 1800tctgtgcagc tggccgatca ctaccagcag aacaccccta tcggcgacgg acccgtgctg 1860ctgcctgata atcactatct gtctacacag agcgccctgt ccaaggaccc aaacgagaag 1920agggatcaca tggtgctgct ggagttcgtg accgcagcag gcatcacact gggcatggat 1980gagctgtata ag 1992727DNAArtificial SequenceSynthetic Sequence 7tacccctatg acgtgcctga ttatgcc 27

Claims

1. A recombinant virus vector (RVV), the RVV comprising: a. a nucleotide sequence encoding a toll-like receptor 3 (TLR3) precursor protein; and b. an inverted terminal repeat.

2. The RVV of claim 1, wherein the nucleotide sequence encoding the TLR3 precursor protein is SEQ ID No. 1.

3. The RVV of claim 1, wherein the precursor protein has an amino acid sequence of SEQ ID No. 2.

4. The RVV of claim 1, wherein the inverted terminal repeat is SEQ ID No. 4 or SEQ ID No. 5.

5. The RVV of claim 1, wherein the inverted terminal repeat is a first inverted terminal repeat of SEQ ID No. 4 and a second inverted terminal repeat of SEQ ID No. 5 and, wherein the nucleotide sequence encoding a toll-like receptor 3 (TLR3) precursor protein is positioned between the first inverted terminal repeat and the second inverted terminal repeat.

6. The RVV of claim 1, wherein the RVV is an adeno-associated virus.

7. A composition that comprises a nucleotide sequence according to SEQ ID No. 1 that can be expressed in a target cell.

8. A pharmaceutical composition comprising the composition of claim 7 and one or more pharmaceutically acceptable carriers and/or one or more excipients.

9. An insert for use with a recombinant virus vector (RVV), wherein the insert has a nucleotide sequence according to SEQ ID No. 1.

10. A method of making an agent/target cell complex, the method comprising a step of administering a recombinant virus vector (RVV) to a target cell for forming the agent/target cell complex, wherein the agent/target cell complex causes the target cell to increase production of a TLR3 precursor protein.

11. The method of claim 10, wherein the RVV comprises a nucleotide sequence according to SEQ ID No. 3 for increasing the target cell's production of the TLR3 precursor protein.

12. The method of claim 10, wherein the RVV comprises a nucleotide sequence according to SEQ ID No. 1 for increasing the target cell's production of the TLR3 precursor protein.

13. The method of claim 10, wherein the TLR3 precursor protein has an amino acid sequence of SEQ ID No. 2.

14. The method of claim 10, wherein the TLR3 precursor protein is subjected to one or more post-translation modification processes within the target cell to produce a TLR3 protein product.

15. The method of claim 10, wherein the target cell is one or more of an innate immune cell, an acquired immune cell, an adrenal gland cell; a bile duct cell; a chondrocyte; a cochlear cell; a corneal cell; an endocardium cell; an endometrial cell; an endothelial cell; an epithelial cell; a fibroblast; a hair follicle cell; a hepatocyte; a lymph node cell; a mucosal cell; a myocyte; a neuron; a glomeruli cell; an optic nerve cell; an osteoblast; an ovarian tissue cell; a pancreatic islet beta cell; a pericardium cell; a platelet; a red blood cell (RBC); a retinal cell; a scleral cell; a Schwann cell; a T cell; a testicular tissue cell; a thyroid gland cell; a uveal cell; a tumor cell, and combinations thereof.