Novel chemokine-like protein fragments

Abstract

The present invention relates to a TAFA protein, a fragment thereof, or a variant thereof, which has the ability to increase the length or branching point of neurites. In addition, the present invention relates to a therapeutic use of a pharmaceutical composition containing: a polypeptide comprising the amino acid sequence of the TAFA protein, a fragment thereof, or a variant thereof; a nucleic acid molecule encoding the polypeptide; a vector carrying the nucleic acid molecule; a recombinant viral particle comprising the vector and a capsid protein; a cell comprising the vector; a cell transformed with the vector; or a combination thereof. The composition of the present invention promotes recovery from damaged retinas (for example, cone cells of photoreceptors, etc.) or increases the length of neurites in neurons, and thus can be advantageously used for preventing or treating retinal neurological diseases and / or neuropathic pain.

Description

Novel chemokine-like protein fragments

[0001] The present invention provides a TAFA protein, a TAFA protein fragment, or a variant thereof, which has the ability to increase neurite length or branch point.

[0002]

[0003] The TAFA family of proteins is abundantly expressed throughout the brain and consists of five proteins, TAFA1 to TAFA5. These proteins contain structurally conserved cysteine ​​residues, and the C, CC, and CXC motifs between the cysteines are associated with chemokines. TAFA1 to TAFA4 share a high degree of similarity in the number of cysteines and the spacing between them, while TAFA5 has fewer cysteines.

[0004] These proteins are evolutionarily highly conserved in vertebrates and have been shown to be important for normal central nervous system function. TAFA1 knockout mice have been reported to exhibit reduced body weight, decreased anxiety behavior, and impaired fear memory (Lei X, Liu L, Terrillion CE, et al. FASEB J. 2019;33(12):14734-14747. and Yong HJ, Ha N, Cho EB, et al. Sci Rep. 2020;10(1):3969.). Conversely, knockout of TAFA2 and TAFA3 has been reported to increase anxiety behavior (Choi JH, Jeong YM, Kim S, et al. Proc Natl Acad Sci U S A. 2018;115(5):E1041-E1050. and Kim S, Lee B, Choi JH, Kim JH, Kim CH, Shin HS. Sci Rep. 2017;7(1):16503.). TAFA4-null mice have been reported to exhibit allodynia and hyperalgesia (Delfini MC, Mantilleri A, Gaillard S, et al. Cell Rep. 2013;5(2):378-388.). Behavioral changes such as increased depressive behavior and loss of spatial memory ability were reported in TAFA5 knockout mice (Huang S, Zheng C, Xie G, et al. FAM19A5 / TAFA5, a novel neurokine, plays a crucial role in depressive-like and spatial memory-related behaviors in mice. Mol Psychiatry. 2021;26(6):2363-2379.)

[0005] Despite the important roles these proteins play in the central nervous system, very little is known about the functions of each protein and their role in pathophysiology or therapeutics, and limited information is available about their roles outside the central nervous system.

[0006]

[0007] The matters described as background technology above are only intended to enhance understanding of the background of the present invention, and should not be taken as an admission that they correspond to prior art already known to those skilled in the art.

[0008]

[0009] An object of the present invention is to provide a polypeptide having the ability to increase neurite length and / or branch point.

[0010] The above polypeptide can reverse or restore the loss or dysfunction of cone cells, which are one of the photoreceptors.

[0011]

[0012] The above polypeptide may comprise an amino acid sequence of a TAFA (TAFA Chemokine Like Family Member) protein, a fragment thereof, or a variant thereof.

[0013] The above polypeptide may comprise the amino acid sequence (from N-terminus to C-terminus) of the following general formula 7:

[0014] General Formula 7

[0015] X1-X2-X3-GTCEV-X4-A-X5-H-X6- CCN-X7-N-X8-IEE- -X20-X21-X22-WWC-X23-M-X24-PC-X25-X26-GE-X27-CK-X28-LPD-X29-X30-GW-X31-C-X32-X33-G-X34-K-X35-KTT-X36-X37-X38-X39

[0016] In the above general formula 1,

[0017] X1 is either non-existent, V, I, or L,

[0018] X2 is K, E, R or Q,

[0019] X3 is G, T, Q, P or A,

[0020] X4 is V or I,

[0021] X5 is A, L, V or I,

[0022] X6 is R or L,

[0023] X7 is K, R or Q,

[0024] X8 is R or K,

[0025] X9 is R or L,

[0026] X10 is V or G,

[0027] X11 is K or N,

[0028] X12 is F or L,

[0029] X13 is P or S,

[0030] X14 is Q or K,

[0031] X15 is R, H or Q,

[0032] X16 is A, N, S or T,

[0033] X17 is A, Q, R, K or T,

[0034] X18 is D or E,

[0035] X19 is S or A,

[0036] X20 is I, E, L, A or V,

[0037] X21 is Q, G or E,

[0038] X22 is K or R,

[0039] X23 is H, Q or E,

[0040] X24 is E, Q, N, D, S or H,

[0041] X25 is L, V or M,

[0042] X26 is E, D, P, L or A,

[0043] X27 is E or D,

[0044] X28 is V, T, A or I,

[0045] X29 is L, N, R, Y, S or Q,

[0046] X30 is S, K or T,

[0047] X31 is S or M,

[0048] X32 is S, A or Y,

[0049] X33 is S, T or R,

[0050] X34 is N or H,

[0051] X35 is V or I,

[0052] X36 is R or K,

[0053] X37 is either non-existent or is V, A, G, M or N,

[0054] X38 is either non-existent or is T, I, N, F or S, and

[0055] X39 is either absent or is R, H, V, K, I, or Q.

[0056] The above polypeptide may comprise one or more amino acid sequences selected from the group consisting of amino acid sequences set forth in SEQ ID NOs: 87 to 141.

[0057] The above polypeptide may be composed of a sequence of 8 to 61 amino acids.

[0058] The above polypeptide may comprise a sequence having at least 50% sequence identity with the amino acid sequence of SEQ ID NO: 1, SEQ ID NO: 15 or SEQ ID NO: 87.

[0059] The amino acid sequence of the above polypeptide may include the amino acid sequence of the following general formula 1 (from the N-terminus to the C-terminus):

[0060] <General Formula 1>

[0061] X1-GE-X2-CK-X3-L

[0062] In the above general formula 1

[0063] X1 is E, D, P, L or A,

[0064] X2 is D or E, and

[0065] X3 is T, V, I or A.

[0066] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 1 may include the amino acid sequence of SEQ ID NO: 142.

[0067] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 1 may include an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 14.

[0068] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 1 may be composed of 8 to 43 amino acid sequences.

[0069] The amino acid sequence of the above polypeptide may include the amino acid sequence of the following general formula 2 (from the N-terminus to the C-terminus):

[0070] <General Formula 2>

[0071] IV-X4-X5-X6-WWC-X7-M-X8-PC-X9-X1-GE-X2-CK-X3-L

[0072] In the above general formula 2

[0073] X1 is E, D, P, L or A,

[0074] X2 is D or E,

[0075] X3 is T, V, I or A,

[0076] X4 is I, E, A, L or V,

[0077] X5 is Q, E or G,

[0078] X6 is K or R,

[0079] X7 is E, H or Q,

[0080] X8 is E, Q, N, D, S or H, and

[0081] X9 is L, M or V.

[0082] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 2 may include the amino acid sequence of SEQ ID NO: 143.

[0083] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 2 may include an amino acid sequence selected from the group consisting of SEQ ID NOs: 28 to 51.

[0084] The amino acid sequence of the above polypeptide may include the amino acid sequence of the following general formula 3 (from the N-terminus to the C-terminus):

[0085] <General Formula 3>

[0086] X1-GE-X2-CK-X3-LPD-X4-X5-GWSCS-X6-GNK-X7-KTTKVTR

[0087] In the above general formula 3

[0088] X1 is E, D, P, L or A,

[0089] X2 is D or E,

[0090] X3 is T, V, I or A,

[0091] X4 is Y, S or L,

[0092] X5 is S or T,

[0093] X6 is S or T, and

[0094] X7 is V or I.

[0095] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 3 may include the amino acid sequence of SEQ ID NO: 144.

[0096] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 3 may include an amino acid sequence selected from the group consisting of SEQ ID NOs: 52 to 58.

[0097] The amino acid sequence of the above polypeptide may include an amino acid sequence selected from the group consisting of SEQ ID NOs: 152 to 171.

[0098] The amino acid sequence of the above polypeptide may include the amino acid sequence of the following general formula 4 (from the N-terminus to the C-terminus):

[0099] General Formula 4

[0100] G-X1-VAGTT-X2-X3-X4-PSCV-X5-X6-X7

[0101] In the above general formula 4

[0102] X1 is Q or K,

[0103] X2 is R, H or Q,

[0104] X3 is A, N, S or T,

[0105] X4 is R, A, Q, K, or T,

[0106] X5 is D or E,

[0107] X6 is A or not present, and

[0108] X7 is either S, A or non-existent.

[0109] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 4 may include the amino acid sequence of SEQ ID NO: 145.

[0110] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 4 may include an amino acid sequence selected from the group consisting of SEQ ID NOs: 15 to 27.

[0111] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 4 may be composed of a sequence of 15 to 46 amino acids.

[0112] The amino acid sequence of the above polypeptide may include the amino acid sequence of the following general formula 5 (from the N-terminus to the C-terminus):

[0113] <General Formula 5>

[0114] X8-IEE-X9-SQT-X10-X11-CSC-X12-X13-G-X1-VAGTT-X2-X3-X4-PSCV-X5-X6-X7

[0115] In the above general formula 5

[0116] X1 is Q or K,

[0117] X2 is R, H or Q,

[0118] X3 is A, N, S or T,

[0119] X4 is R, A, Q, K, or T,

[0120] X5 is D or E,

[0121] X6 is A or does not exist,

[0122] X7 is S, A or not present,

[0123] X8 is R or K,

[0124] X9 is R or L,

[0125] X10 is V or G,

[0126] X11 is K or N,

[0127] X12 is F or L, and

[0128] X13 is P or S.

[0129] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 5 may include the amino acid sequence of SEQ ID NO: 146.

[0130] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 5 may include an amino acid sequence selected from the group consisting of SEQ ID NOs: 59 to 74.

[0131] The amino acid sequence of the above polypeptide may include the amino acid sequence of the following general formula 6 (from the N-terminus to the C-terminus):

[0132] <General Formula 6>

[0133] G-X1-VAGTT-X2-X3-X4-PSCV-X5-X6-X7-IV-X8-X9-KWWC-X10-M-X11-PC-X12

[0134] In the above general formula 6

[0135] X1 is Q or K,

[0136] X2 is R, H or Q,

[0137] X3 is A, N, S or T,

[0138] X4 is R, A, Q, K or T,

[0139] X5 is D or E,

[0140] X6 is A or does not exist,

[0141] X7 is S, A or not present,

[0142] X8 is I, A, V or L,

[0143] X9 is Q or E,

[0144] X10 is H or Q,

[0145] X11 is N, D, S or H, and

[0146] X12 is L or M.

[0147] The amino acid sequence of a polypeptide comprising the amino acid sequence of the above general formula 6 may include the amino acid sequence of SEQ ID NO: 147.

[0148] The amino acid sequence of the polypeptide comprising the amino acid sequence of the above general formula 6 may include an amino acid sequence selected from the group consisting of SEQ ID NOs: 75 to 85.

[0149] The amino acid sequence of the above polypeptide may include an amino acid sequence selected from the group consisting of SEQ ID NOs: 172 to 184.

[0150] Another object of the present invention is to provide a nucleic acid molecule encoding the polypeptide.

[0151] Another object of the present invention is to provide a vector comprising the nucleic acid molecule.

[0152] Another object of the present invention is to provide a recombinant viral particle comprising the vector and capsid protein.

[0153] The above virus may be AAV.

[0154] Another object of the present invention is to provide a cell comprising the above vector.

[0155] Another object of the present invention is to provide a cell transformed with the above vector.

[0156] Another object of the present invention is to provide a composition comprising the polypeptide, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

[0157] The above composition may be a pharmaceutical composition.

[0158] The above pharmaceutical composition may be a composition for preventing or treating retinal neuropathies.

[0159] The above retinal neurological disease may include retinopathy, choroidal neovascularization, macular disease, macular degeneration, retinal degeneration, macular edema, retinal edema, macular swelling, retinal swelling, color vision abnormality, retinal cell degeneration, retinal vascular occlusion, retinal detachment, hereditary retinal disease, or a combination thereof.

[0160] The above pharmaceutical composition may be a composition for preventing or treating neuropathic pain.

[0161] The above neuropathic pain may be allodynia, hyperalgesia, hyperesthesia or dysphagia.

[0162] The above neuropathic pain may be central neuropathic pain or peripheral neuropathic pain.

[0163] The neuropathic pain may be neuralgia, deafferentation pain syndrome, complex regional pain syndrome, or neuropathy (central or peripheral).

[0164] Another object of the present invention is to provide a method for producing a composition, comprising a step of producing a composition comprising the polypeptide, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

[0165] Another object of the present invention is to provide a therapeutic use for the manufacture of a medicament comprising the polypeptide, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

[0166] Another object of the present invention is to provide a method for preventing or treating a disease or disorder in a subject in need thereof, comprising administering to the subject a composition comprising the polypeptide, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

[0167]

[0168] Other objects and advantages of the present invention will become more apparent from the detailed description, claims and drawings below.

[0169]

[0170] I. Definition

[0171] Throughout this disclosure, the term "at least" preceding a number or series of numbers is understood to include the number following the term "at least" and all subsequent numbers or integers that would logically and obviously be included in the context. For example, the number of nucleotides in a nucleic acid molecule must be an integer. For example, "at least 18 nucleotides of a 21 nucleotide nucleic acid molecule" means that 18, 19, 20, or 21 nucleotides have the given characteristic. When "at least" precedes a series of numbers or a range of numbers, it is understood that "at least" can modify each of the numbers in the series or range. "At least" is also not limited to integers (e.g., "at least 5%" includes 5.0%, 5.1%, and 5.18%, regardless of the number of significant digits).

[0172] When aspects are described herein as "comprising," it is understood that other similar aspects described in terms of "consisting of" and / or "consisting essentially of" are also provided.

[0173] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure relates. For example, Concise Dictionary of Biomedicine and Molecular Biology, Juo, Pei-Show, 2nd ed., 2002, CRC Press; Dictionary of Cell and Molecular Biology, 3rd ed., 1999, Academic Press; and Oxford Dictionary Of Biochemistry and Molecular Biology, Revised, 2000, Oxford University Press provide one of ordinary skill in the art with general dictionaries of many of the terms used in this disclosure.

[0174] Units, prefixes, and symbols are indicated in their Systeme International de Unites (SI) formats. Numerical ranges include the numbers that define them. Unless otherwise indicated, amino acid sequences are written from left to right in the amino to carboxyl direction. The headings provided herein are not intended to limit the various aspects of the disclosure, which may be incorporated by reference into the specification as a whole. Accordingly, the terms defined immediately below are more fully defined by reference to the specification as a whole.

[0175] The term "about" is used herein to mean approximately, approximately, or within the range. When used with a numerical range, the term "about" modifies that range by extending the boundaries above and below the stated value. Typically, the term "about" modifies the stated value by, for example, a 10% change above or below the stated value.

[0176] The term "family with sequence similarity 19" or "FAM19", or "TAFA", as used herein, refers to the proteins belonging to the TAFA family of five proteins (also known as the FAM19 family) and expressed primarily in the brain and spinal cord, also referred to as TAFA proteins or TAFA polypeptides. FAM19A1 is also known as TAFA1, FAM19A2 is also known as TAFA2, FAM19A3 is also known as TAFA3, FAM19A4 is also known as TAFA4, and FAM19A5 is also known as TAFA5.

[0177] The human TAFA1 gene encodes a 133-amino acid sequence, and although there is disagreement about the length of the signal peptide and the mature protein, it is predicted that the human TAFA1 protein consists of a 35-amino acid signal peptide and a 98-amino acid mature protein. TAFA1 is highly expressed in the frontal, temporal, occipital, and parietal cortices, and is lowly expressed in the basal ganglia, thalamus, and cerebellum. Cell experiments have shown that TAFA1 affects the differentiation fate of neural stem cells, inhibiting astrocyte differentiation and promoting neuronal differentiation of neural stem cells. TAFA1 knock-out (KO) mouse experiments have shown that TAFA1 can regulate motor activity, anxiety-responsive behavior, learning and memory, and somatosensory function.

[0178] The human TAFA2 gene encodes a 131-amino acid sequence. Although there is disagreement about the length of the signal peptide and the mature protein, it is predicted that the human TAFA2 protein consists of a 30-amino acid signal sequence and a 101-amino acid mature protein. TAFA2 is abundantly expressed in the occipital and frontal cortices, and medulla oblongata within the central nervous system. Injection of TAFA2 recombinant protein into the third ventricle of mice increased food intake and meal frequency, and increased energy expenditure and respiratory exchange ratio. This suggests that TAFA2 may play a role in the regulation of food intake and energy metabolism. Furthermore, inhibition of TAFA2 in zebrafish and mice is known to increase anxiety-related behaviors.

[0179] The human TAFA3 gene encodes a 133-amino acid sequence. Although there is disagreement about the length of the signal peptide and the mature protein, it is predicted that the human TAFA3 protein consists of a 30-amino acid signal peptide and a 103-amino acid mature protein. It is known that TAFA3 expression is increased in microglia in a mouse model of transient focal cerebral ischemia, and that TAFA3-treated microglia polarize into anti-inflammatory microglia. Furthermore, knocking out TAFA3 in a mouse model results in three major behavioral deficits observed in autism spectrum disorders: decreased response to social novelty, impaired social communication, and increased repetitive behaviors, suggesting that TAFA3 is involved in the normal function of social relationship formation.

[0180] The human TAFA4 gene encodes a 140-amino acid sequence, and although there is disagreement about the length of the signal peptide and the mature protein, it is expected that the human TAFA4 protein consists of a 45-amino acid signal peptide and a 95-amino acid mature protein. TAFA4 is mainly expressed in sensory neurons in the peripheral nervous system. TAFA4 protein is specifically expressed in low-threshold mechanoreceptors (C-low-threshold mechanoreceptors) and appears to reduce pain by regulating the activity of interneurons, particularly GABAergic neurons.

[0181] The human TAFA5 gene encodes a 132-amino acid sequence, and although there is disagreement about the length of the signal peptide and the mature protein, it is predicted that the human TAFA5 protein is composed of a 43-amino acid signal sequence and an 89-amino acid mature protein. TAFA5 is highly expressed in the basal ganglia region and cerebellum. It is known that TAFA5 expression increases in the hypothalamus of mice by inflammatory stimuli such as TNF-α, and that knocking out TAFA5 partially restores the decreased food intake, body weight loss, and increased inflammatory cytokines induced by TNF-α.

[0182] For the above TAFA1 to TAFA4 mature proteins, high sequence identity is shown (e.g., human TAFA4 has 73.7% sequence identity with TAFA1, 85.3% sequence identity with TAFA2, and 81.1% sequence identity with TAFA3). On the other hand, TAFA5 has low sequence identity with TAFA1 to 4 (e.g., human TAFA5 has 48.9% sequence identity with TAFA1, 51.1% sequence identity with TAFA2, 47.7% sequence identity with TAFA3, and 50.0% sequence identity with TAFA4).

[0183] Human TAFA4 (TAFA Chemokine Like Family Member 4) protein (SEQ ID NO: 299) is a member of the TAFA protein family (TAFA1-5) that is abundantly expressed throughout brain regions. TAFA4, consisting of 140 amino acids, is evolutionarily highly conserved among vertebrates, with 95 or 93 amino acids remaining, excluding the 45 or 47 amino acids containing the signal sequence at the front. Furthermore, among the TAFA protein family, TAFA1 to 4 have been confirmed to have highly conserved interspecies sequence identity for the 93 amino acid sequences. i) The human mature TAFA4 amino acid sequence exhibits at least 90% sequence identity compared to TAFA4 of mammals and amphibians, at least 95% sequence identity compared to TAFA4 of birds, and at least 85% sequence identity compared to TAFA4 of reptiles and fish; ii) The human TAFA1 amino acid sequence exhibits at least 90% sequence identity compared to TAFA1 of mammals and amphibians, at least 95% sequence identity compared to TAFA1 of birds, and at least 85% sequence identity compared to TAFA1 of reptiles and fish; iii) The human TAFA2 amino acid sequence exhibits at least 90% sequence identity compared to TAFA2 of mammals and amphibians, at least 95% sequence identity compared to TAFA2 of birds, and at least 80% sequence identity compared to TAFA2 of reptiles and fish; iv) The human TAFA3 amino acid sequence exhibits at least 75% sequence identity compared to TAFA3 of mammals and amphibians, and at least 80% sequence identity compared to TAFA3 of birds. Compared to TAFA3 of reptiles and fish, it shows more than 75% sequence identity, confirming that it shows very high sequence identity regardless of species.

[0184] The term "TAFA protein fragment" as used herein refers to a fragment of TAFA1 to TAFA4 proteins that has the ability to increase the length and / or branching point of neurites. The above fragments are 8 (e.g. TAFA fragment 3.7), 9, 10, 11, 12, 13, 14, 15, 16, 17 (e.g. TAFA fragment 2.5), 18, 19, 20, 21, 22 (e.g. TAFA fragment 7), 23, 24, 25 (e.g. TAFA fragment 2.5+3.7), 26, 27, 28, 29 (e.g. TAFA fragment 3), 30, 31 (e.g. TAFA fragment 2), 32 (e.g. TAFA fragment 5), 33, 34, 35, 36, 37, 38, 39 (e.g. TAFA fragment 2.5+7; e.g. TAFA fragment 2+3.7), 40, 41, 42, It may be composed of, but is not limited to, an amino acid sequence of 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54 (e.g., TAFA fragment 5+7), 55, 56, 57, 58, 59, 60 (e.g., TAFA fragment 2+3) or 61 (e.g., TAFA fragment 5+3).

[0185] The term "variant of a TAFA protein fragment" as used herein refers to a polypeptide in which some of the amino acids of a polypeptide constituting a TAFA1 to TAFA4 protein fragment are replaced with other amino acids. The variant is preferably a functional variant. The term "functional" refers to a variant derived from a fragment of a TAFA1 to TAFA4 protein that has the ability to increase the length and / or branching point of a neurite, like the "TAFA protein" or the "TAFA protein fragment." The variant is preferably a cross-species variant (e.g., a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142 to 147). A person skilled in the art can easily produce a polypeptide by utilizing the differences in non-conserved sequences excluding conserved sequences among the amino acid sequences of TAFA1 to TAFA4 fragments of each known species (for example, by replacing the fourth sequence D with E in EGEECKVL (SEQ ID NO: 6), which is a fragment 3.7 sequence of rabbit TAFA2, with reference to EGEDCKVL (SEQ ID NO: 1), which is a fragment 3.7 sequence of human TAFA4, or by replacing the first sequence E with P and the fourth sequence D with E in PGEECKVL (SEQ ID NO: 9), which is a fragment 3.7 sequence of pig TAFA3).

[0186] The amino acid substitution may be a substitution of 1 to 22 amino acids. For example, when the TAFA protein fragment simultaneously includes fragments 5 and 7, 1 to 22 amino acid substitutions are possible, when the TAFA protein fragment simultaneously includes fragments 2 and 3, 1 to 19 amino acid substitutions are possible, when the TAFA protein fragment only includes fragment 2, 1 to 12 amino acid substitutions are possible, when the TAFA protein fragment only includes fragment 5, 1 to 13 amino acid substitutions are possible, when the TAFA protein fragment only includes fragment 3, 1 to 7 amino acid substitutions are possible, when the TAFA protein fragment only includes fragment 7, 1 to 9 amino acid substitutions are possible, when the TAFA protein fragment only includes fragment 2.5, 1 to 7 amino acid substitutions are possible, and when the TAFA protein fragment only includes fragment 3.7, 1 to 3 amino acid substitutions are possible.

[0187] Preferably, the number of amino acid substitutions may be 1 to 20, 1 to 19, 1 to 18, 1 to 17, 1 to 16, 1 to 15, 1 to 14, 1 to 13, 1 to 12, 1 to 11, 1 to 10, 1 to 9, 1 to 8, 1 to 7, 1 to 6, 1 to 5, 1 to 4, or 1 to 3.

[0188] The term "adeno-associated virus" (AAV) as used herein refers to a single-stranded DNA virus, a helper-dependent human parvovirus. The genome size is approximately 4.6 kbp, and the N-terminal portion of the genome encodes the rep gene, which is involved in viral replication and viral gene expression, and the C-terminal portion encodes the cap gene, which encodes the viral capsid protein, and is composed of repeat regions (ITRs) with approximately 145 base pairs inserted at both ends. The function of the 145 bp inverted terminal repeats (ITRs) with a T-shaped structure is to function as an origin of replication during viral genome replication and act as the primary packaging signal. ITR is the only cis-acting base sequence required to create a recombinant AAV construct, and it has enhancer activity in the presence of Rep protein, but very weak activity in the absence of Rep protein. Therefore, when cloning a foreign gene into a recombinant AAV construct, this is taken into consideration and the enhancer, promoter, pA, etc. are appropriately configured to create an expression construct (RJ Samulski and N Muzyczka, Annu. Rev. Virolo. 2014. 1:427-451). Four proteins are translated from the rep gene, which are classified as rep78, rep68, rep52, and rep40 according to their molecular weights, and perform important functions in AAV DNA replication. Four proteins are translated from the cap gene, among which VP1, VP2, and VP3 proteins are structural proteins that constitute AAV particles, and assembly-activating protein (AAP) promotes the formation (assembly) of AAV particles by the structural proteins.For efficient replication of the above adeno-associated viruses, some proteins and RNAs derived from helper viruses such as adenovirus or herpes simplex virus are required (Muzyczka N. Curr Top Microbiol Immunol 158, 97-129, 1992).

[0189] AAV includes, but is not limited to, AAV type 1, AAV type 2, AAV type 3 (including types 3A and 3B), AAV type 4, AAV type 5, AAV type 6, AAV type 7, AAV type 8, AAV type 9, AAV type 10, AAV type 11, AAV type 12, AAV type 13, AAVrh.74, snake AAV, avian AAV, bovine AAV, canine AAV, equine AAV, ovine AAV, goat AAV, shrimp AAV, the AAV serotypes and phylogenies disclosed by Gao et al. (J. Virol. 78:6381 (2004)) and Moris et al. (Virol. 33:375 (2004)), and other AAV now known or later discovered. For example, FIELDS et al. See VIROLOGY, volume 2, chapter 69 (4th ed., Lippincott-Raven Publishers). In some embodiments, "AAV" includes derivatives of known AAV. In some embodiments, "AAV" includes modified or artificial AAV. In some embodiments, "AAV" includes AAV with a modified capsid.

[0190] As used herein, the terms "administering," "administering," and grammatical variations thereof refer to introducing a composition (e.g., a polypeptide comprising the amino acid sequence of a TAFA protein, fragment thereof, or variant thereof described herein) into a subject via a pharmaceutically acceptable route. The composition is introduced into the subject by any suitable route, including intratumoral, oral, intrapulmonary, intranasal, parenteral (intravenous, intraarterial, intramuscular, intraperitoneal, or subcutaneous), rectal, intralymphatic, intrathecal, periocular, intraocular, or topical administration (e.g., eye drops, intranasal, etc.). Administration includes self-administration and administration by another. The composition or formulation exerts its intended function through a suitable route of administration. For example, if the suitable route is intravenous, the composition is administered by introducing the composition or formulation into a vein of the subject. Pharmaceutical compositions may be formulated for topical or topical application, such as topical application to the eyes, skin, and mucous membranes, in the form of eye drops, gels, creams, and lotions, or for intracisternal or intrathecal application. Topical administration includes topical administration, transdermal delivery, administration to mucous membranes (e.g., nose, mouth, rectum, etc.), or inhalation therapy. The compositions may be administered alone or in combination with other pharmaceutically acceptable excipients.

[0191] As used herein, the term "intraocular" refers to within or beneath the ocular tissue. The term "intraocular administration" as used herein refers to any administration that can deliver a composition to sub-Tenon, subconjunctival, suprachoroidal, subretinal, intravitreal, or similar locations within the eye. In some aspects, intraocular administration includes suprachoroidal, subretinal, and intravitreal administration.

[0192] The term "conserved," as used herein, refers to nucleotides or amino acid residues in a polynucleotide sequence or polypeptide sequence, respectively, that appear unchanged at the same position in two or more sequences being compared. Relatively conserved nucleotides or amino acids are those that are more conserved among related sequences than are nucleotides or amino acids that appear at different positions in the sequences.

[0193] The term "amino acid" as used herein includes the 20 standard amino acids (arginine (R), lysine (K), histidine (H), glutamic acid (E), aspartic acid (D), glutamine (Q), asparagine (N), leucine (L), isoleucine (I), valine (V), methionine (M), phenylalanine (F), tryptophan (W), tyrosine (Y), glycine (G), alanine (A), serine (S), threonine (T), proline (P), cysteine ​​(C), etc.) that are naturally incorporated into peptides, as well as D-isomers and modified amino acids. In addition, the peptides may include non-standard amino acids that have undergone post-translational modification. Post-translational modifications may include, but are not limited to, phosphorylation, glycosylation, acylation (e.g., acetylation, myristoylation, and palmitoylation), alkylation, carboxylation, hydroxylation, glycation, biotinylation, ubiquitinylation, changes in chemical properties (e.g., beta-elimination deimidation, deamidation), and structural changes (e.g., formation of disulfide bridges). The peptide may be a wild-type peptide identified and isolated from a natural source. Alternatively, the peptide may be an artificial mutant comprising an amino acid sequence in which one or more amino acids are substituted, deleted, and / or inserted. Amino acid changes in the wild-type polypeptide as well as in the artificial variants include conservative amino acid substitutions that do not significantly affect the folding and / or activity of the protein.For example, the conservative substitutions may include basic amino acids (arginine (R), lysine (K), and histidine (H)), acidic amino acids (glutamic acid (E) and aspartic acid (D)), polar amino acids (glutamine (Q) and asparagine (N)), hydrophobic amino acids (leucine (L), isoleucine (I), valine (V), and methionine (M)), aromatic amino acids (phenylalanine (F), tryptophan (W), and tyrosine (Y)), and small amino acids (glycine (G), alanine (A), serine (S), and threonine (T)). Amino acid substitutions that generally do not alter specific activity are known in the art. The most common exchanges may include Ala / Ser, Val / Ile, Asp / Glu, Thr / Ser, Ala / Gly, Ala / Thr, Ser / Asn, Ala / Val, Ser / Gly, Tyr / Phe, Ala / Pro, Lys / Arg, Asp / Asn, Leu / Ile, Leu / Val, Ala / Glu, and Asp / Gly.

[0194] In some embodiments, two or more sequences are said to be "completely conserved" or "identical" if they are 100% identical to each other. In some embodiments, two or more sequences are said to be "highly conserved" if they are at least 70% identical, at least 80% identical, at least 90% identical, or at least 95% identical to each other. In some embodiments, two or more sequences are said to be "highly conserved" if they are about 70% identical, about 80% identical, about 90% identical, about 95%, about 98%, or about 99% identical to each other. In some embodiments, two or more sequences are said to be "conserved" if they are at least 30% identical, at least 40% identical, at least 50% identical, at least 60% identical, at least 70% identical, at least 80% identical, or at least 90% identical, or at least 95% identical to each other. In some embodiments, two or more sequences are said to be "conserved" if they are about 30% identical, about 40% identical, about 50% identical, about 60% identical, about 70% identical, about 80% identical, about 90% identical, about 95% identical, about 98% identical, or about 99% identical to each other. Conservation of sequence can apply to the entire length of a polynucleotide or polypeptide, or to portions, regions, or features thereof.

[0195] The terms "complementary" and "complementarity" refer to two or more oligomers (i.e., each comprising a base sequence) or between an oligomer and a target gene that are associated with each other by the Watson-Crick base-pairing rules. For example, the base sequence "TGA(5'→3') is complementary to the base sequence "ACT(3'→5'). Complementarity may be "partial" if fewer than all of the bases of a given base sequence match another base sequence according to the base-pairing rules. For example, in some embodiments, the complementarity between a given base sequence and another base sequence may be about 70%, about 75%, about 80%, about 85%, about 90%, or about 95%. Thus, in certain embodiments, the term "complementary" refers to an identity or complementarity of at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% with a target nucleic acid sequence. Or, to continue the example, there can be "complete" or "perfect" (100%) complementarity between a given base sequence and another base sequence. In some embodiments, the degree of complementarity between base sequences significantly affects the efficiency and strength of hybridization between the sequences.

[0196] The term "downstream" refers to a nucleotide sequence located 3' to a reference nucleotide sequence. In certain embodiments, the downstream nucleotide sequence relates to the sequence following the start of transcription. For example, the translation initiation codon of a gene is located downstream of the transcription start site.

[0197] The term "enhancer," as used herein, refers to a portion of DNA that contains sequences capable of providing enhanced transcription and, in some cases, can act independently of its orientation relative to another regulatory sequence. An enhancer may function in concert with or in addition to a promoter and / or other enhancer elements.

[0198] The terms "excipient" and "carrier" are used interchangeably and refer to an inert substance added to a pharmaceutical composition to further facilitate administration of a compound, for example, a polynucleotide comprising a foreign gene and a non-translated nucleic acid sequence as described herein.

[0199] The term "exon" refers to a nucleic acid sequence that appears in the mature form of an RNA molecule after a portion of a nucleic acid encoding a protein or a portion of a preprocessed (or precursor) RNA has been removed by splicing. The mature RNA molecule may be messenger RNA (mRNA) or a functional form of a non-coding RNA such as rRNA or tRNA.

[0200] As used herein, the term "expression" refers to the process by which a polynucleotide produces a gene product, such as RNA or a polypeptide. This includes, but is not limited to, transcribing a polynucleotide into messenger RNA (mRNA) and translating the mRNA into a polypeptide. Expression produces a "gene product." As used herein, a gene product may be a nucleic acid, such as RNA, produced by transcription of a gene. As used herein, a gene product may be a nucleic acid or a polypeptide translated from a transcript. The gene products described herein further include nucleic acids subjected to post-transcriptional modifications, such as polyadenylation or splicing, or polypeptides subjected to post-translational modifications, such as phosphorylation, methylation, glycosylation, lipid addition, association with other protein subunits, or proteolytic cleavage.

[0201] The term "identity," as used herein, refers to overall monomer conservation between polymer molecules, e.g., polynucleotide molecules. The term "identical," without any further modifiers, e.g., polynucleotide A is identical to polynucleotide B, means that the polynucleotide sequences are 100% identical (100% sequence identity). Describing two sequences as being, for example, "70% identical" is equivalent to describing them as having, for example, "70% sequence identity."

[0202] For example, calculating the percent identity of two polypeptide or polynucleotide sequences can be performed by aligning the two sequences for optimal comparison purposes (e.g., gaps can be introduced in one or both of the first and second polypeptide or polynucleotide sequences for optimal alignment, and non-identical sequences can be ignored for comparison purposes). In certain embodiments, the length of the aligned sequences for comparison purposes is at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or 100% of the length of the reference sequence. Thereafter, the amino acids at corresponding amino acid positions, or in the case of polynucleotides, the bases, are compared.

[0203] If a particular position in the first sequence is occupied by the same amino acid or nucleotide as the corresponding position in the second sequence, the molecules are identical at that position. The percent identity between two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps that need to be introduced for optimal alignment of the two sequences and the length of each gap. Sequence comparison and determination of the percent identity between two sequences can be performed using a mathematical algorithm.

[0204] Suitable software programs for aligning different sequences (e.g., polynucleotide sequences) are available from several sources. One suitable program for determining percent sequence identity is bl2seq, part of the BLAST suite of programs available from the U.S. government's National Center for Biotechnology Information's BLAST website (blast.ncbi.nlm.nih.gov). bl2seq performs comparisons between two sequences using the BLASTN or BLASTP algorithm. BLASTN is used to compare nucleic acid sequences, while BLASTP is used to compare amino acid sequences. Other suitable programs include Needle, Stretcher, Water, or Matcher, which are part of the EMBOSS suite of bioinformatics programs and are also available from the European Bioinformatics Institute (EBI) at www.ebi.ac.uk / Tools / psa.

[0205] Sequence alignment can be performed using methods known in the art, such as MAFFT, Clustal (ClustalW, Clustal X, or Clustal Omega), MUSCLE, etc.

[0206] Different regions within a single polynucleotide or polypeptide target sequence aligned with a polynucleotide or polypeptide reference sequence may each have their own percent sequence identity. Note that percent sequence identity values ​​are rounded to the nearest decimal place. For example, 80.11, 80.12, 80.13, and 80.14 are rounded down to 80.1, and 80.15, 80.16, 80.17, 80.18, and 80.19 are rounded up to 80.2. Also note that length values ​​will always be integers.

[0207] In certain embodiments, the percent identity (%ID) of a first amino acid sequence (or nucleic acid sequence) to a second amino acid sequence (or nucleic acid sequence) is calculated as %ID = 100 Х (Y / Z), where Y is the number of amino acid residues (or nucleobases) that score as identical matches in an alignment of the first and second sequences (either by visual inspection or by a particular sequence alignment program) and Z is the total number of residues in the second sequence. If the first sequence is longer than the second sequence, the percent identity of the first sequence to the second sequence will be higher than the percent identity of the second sequence to the first sequence.

[0208] Those skilled in the art will appreciate that the generation of sequence alignments for calculating percent sequence identity is not limited to binary sequence-sequence comparisons processed entirely by primary sequence data. It will also be appreciated that sequence alignments can be achieved by integrating sequence data with data from heterogeneous sources, such as structural data (e.g., crystallographic protein structures), functional data (e.g., mutation locations), or phylogenetic data. A suitable program for integrating heterogeneous data to create multiple sequence alignments is T-Coffee, available at www.tcoffee.org or alternatively, for example, from EBI. It will also be appreciated that the final alignment used to calculate percent sequence identity can be structured automatically or manually.

[0209] As used herein, the term "intron" refers to a segment of DNA (intervening sequence) within a gene that is spliced ​​out of the mRNA transcribed from the gene before export from the cell nucleus, without coding for part of the protein produced by the gene. "Intron sequence" refers to the nucleic acid sequence of an intron. Such sequences are also referred to herein as "untranslated nucleic acid sequences." Accordingly, introns are regions of DNA sequence that are transcribed along with the coding sequence (exon) but are removed during the formation of mature mRNA.

[0210] As used herein, the term "intron fragment" refers to a fragment derived from a full-length intron sequence (e.g., a full-length EF-1α intron A sequence). Such fragment is meant to exclude the full-length intron. In some embodiments, the "intron fragment" comprises the minimum number of nucleotides or configurations necessary to achieve an expression level that exceeds the expression level achieved with a corresponding construct lacking all nucleotides of EF-1α intron A. Therefore, the intron fragment of the present disclosure (also referred to herein as a "untranslated nucleic acid sequence") is not particularly limited as long as it comprises a fragment of an EF-1α intron and can increase the expression of a foreign gene. As demonstrated herein, in some embodiments, an intron fragment (i.e., an untranslated nucleic acid sequence) can increase the transcription of a foreign gene, thereby increasing the expression of the foreign gene. Therefore, in some embodiments, the intron fragments described herein may be untranslated regulatory elements.

[0211] As used herein, the terms "isolated," "purified," "extracted," and grammatical variations thereof are used interchangeably and refer to a preparation of a desired composition of the present disclosure, e.g., a polynucleotide comprising a polypeptide or a nucleic acid sequence encoding the same, comprising a TAFA protein, or fragment or variant thereof, that has undergone one or more purification steps. In some embodiments, isolation or purification as used herein is a process of removing or partially removing (e.g., fractionating) a composition of the present disclosure, e.g., a polypeptide or polynucleotide described herein, from a sample containing contaminants.

[0212] In some embodiments, the isolated composition has no detectable undesirable activity, or alternatively, the level or amount of undesirable activity is below an acceptable level or amount. In other embodiments, the isolated composition has an amount and / or concentration of the desired composition of the present disclosure that is greater than or equal to an acceptable amount and / or concentration and / or activity. In other embodiments, the isolated composition is concentrated relative to the starting material from which the composition is obtained. Such enrichment may be at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, at least about 99.9%, at least about 99.99%, at least about 99.9999%, at least about 99.9999%, or greater than 99.9999% relative to the starting material.

[0213] In some embodiments, the isolated preparation is substantially free of residual biological products. In some embodiments, the isolated preparation is 100%, at least about 99%, at least about 98%, at least about 97%, at least about 96%, at least about 95%, at least about 95%, at least about 94%, at least about 93%, at least about 92%, at least about 91%, or at least about 90% free of any biological contaminant. Residual biological products may include abiotic materials (including chemicals) or unwanted nucleic acids, proteins, lipids, or metabolites.

[0214] The term "linked" as used herein refers to a first amino acid sequence or polynucleotide sequence that is covalently or non-covalently joined to a second amino acid sequence or polynucleotide sequence, respectively. The first amino acid or polynucleotide sequence may be directly joined or juxtaposed to the second amino acid or polynucleotide sequence, or alternatively, an intervening sequence may covalently join the first sequence to the second sequence. For example, since a TAFA protein is a polypeptide comprising fragments 1, 2, and 3 joined together, the fragments of the TAFA protein may be in the form of fragments 2 and 3 joined together, or in the form of fragments 5 and 7 joined together. The term "linked" refers not only to the fusion of a first polynucleotide sequence to a second polynucleotide sequence at the 5'-end or 3'-end, but also includes inserting the entire first polynucleotide sequence (or the second polynucleotide sequence) into any two nucleotides within the second polynucleotide sequence (or the first polynucleotide sequence). The first polynucleotide sequence may be linked to the second polynucleotide sequence by a phosphodiester bond or a linker. The linker may be, for example, a polynucleotide.

[0215] The term "retinal neuropathic disease" as used herein refers to a disease, illness, or condition that affects or is related to a part or region of the nerves within the retina or macula of the eye. The retinal neuropathic disease may be a disease, illness, or condition caused by damage to the nerves of all or part of the retina or macula. The retinal neuropathic disease may be caused by dysfunction or damage to the nerve cells of the retina or macula.

[0216] NaIO3 (sodium iodate) has been reported to induce damage and / or degeneration of the retina throughout part or all of the outer and inner parts of the retina, and damage to the retinal pigment epithelium (RPE) and photoreceptors (e.g., rod and cone cell layers) (AE-H. Koh, et al. Journal of Photochemistry & Photobiology, B: Biology 196 (2019) 111514). Therefore, using the NaIO3 model, it is possible to confirm the preventive or therapeutic effect of diseases caused by damage to the retina or macular nerve, or damage to the retinal pigment epithelium (RPE) and photoreceptors (e.g., rod and cone cell layers). Examples of the above diseases include retinopathy, choroidal neovascularization, macular disease, macular degeneration, retinal degeneration, macular edema, retinal edema, macular swelling, retinal swelling, color vision abnormalities, retinal cell degeneration, retinal vascular occlusion, retinal detachment, hereditary retinal diseases, or a combination thereof.

[0217] As used herein, the term "retinopathy" refers to a disease or damage to the retina (i.e., the tissue lining the inner surface at the back of the eye that captures images passing through the cornea and lens) or the cells of the retina.

[0218] As used herein, the term "diabetic retinopathy" (DR) refers to retinopathy caused by complications related to diabetes. Depending on the severity of the disease, DR may be asymptomatic, cause mild vision problems, or lead to blindness. DR is the result of microvascular retinal changes. Hyperglycemia-induced intramural pericyte death and basement membrane thickening can lead to vessel wall dysfunction. This damage alters the formation of the blood-retinal barrier and makes retinal blood vessels more permeable. Perivascular cell death can be induced when hyperglycemia sustainably activates protein kinase C-δ (PKC-δ) and p38 mitogen-activated protein kinase (MAPK), which are encoded by protein kinase C-δ, thereby increasing the expression of Src homology-2 domain-containing phosphatase-1 (SHP-1), a previously unknown protein tyrosine phosphatase that acts as a target of PKC-δ signaling. This signaling cascade can lead to PDGF receptor dephosphorylation and decreased downstream signaling from this receptor, resulting in "perivascular cell death." Small blood vessels, such as those in the eye, are particularly vulnerable to dysglycemic control. Excessive glucose and / or fructose accumulation can damage small blood vessels in the retina.

[0219] DR can be divided into two distinct stages (Wu L., el al., World J Diabetes 4(6): 290-294 (2013)). The first stage, called nonproliferative diabetic retinopathy (NPDR), is associated with early diabetic retinopathy. NPDR usually has no obvious symptoms and is associated with mild vision distortion caused by blood vessels leaking fluid into surrounding tissues. The only way to detect NPDR is with fundus imaging, which can show microaneurysms (tiny, blood-filled bulges in the artery walls). If left untreated, patients with DR can progress to a second, more advanced stage, called proliferative diabetic retinopathy (PDR). PDR is characterized by abnormal new blood vessel formation (neovascularization), which can rupture and bleed, blurring vision. Other symptoms of PDR include floating spots or dark lines in the field of vision ("floaters"), vision changes, impaired color vision, dark or blank areas in the field of vision, pain, patchy vision, and complete loss of vision.

[0220] The term “diabetic retinopathy” includes all types of diabetic retinopathy, including but not limited to nonproliferative diabetic retinopathy (NPDR), proliferative diabetic retinopathy (PDR), diabetic macular edema, and diabetic macular edema.

[0221] In some respects, PDR occurs after the onset of NPDR (e.g., NPDR is initially diagnosed and the disease progresses to PDR). In other respects, PDR occurs independently of NPDR. The term "diabetic retinopathy" as used herein also includes all types of diabetic retinopathy, regardless of cause, and any and all symptoms of diabetic retinopathy. Non-limiting examples of risk factors for diabetic retinopathy include duration of diabetes, genetics, excessive alcohol consumption, smoking, hypertension, obesity, dyslipidemia, high cholesterol, kidney disease, pregnancy, and kidney damage.

[0222] As used herein, the term "maculopathy" refers to any pathological condition of the macula, the central region of the retina responsible for highly sensitive and accurate vision. In some aspects, the terms "maculopathy" and "retinopathy" may be used interchangeably (i.e., only the macula is affected). In some aspects, the maculopathy is diabetic maculopathy.

[0223] Diabetic maculopathy occurs when the macula becomes affected by retinal changes caused by diabetes. The term encompasses two distinct eye conditions: diabetic macular edema and diabetic ischemic maculopathy. These two types of maculopathy are often comorbid, meaning that people with macular edema often also have ischemic maculopathy. Ischemic maculopathy occurs alongside macular edema and can occur even when macular edema is mild. In some aspects, retinal changes associated with diabetic maculopathy include decreased retinal potential, loss of pericytes, acellular capillary formation, vascular congestion, vascular dysfunction, vascular leakage, vascular occlusion, tissue swelling (edema), tissue ischemia, or any combination thereof within the subject's retina.

[0224] As used herein, the term "acellular capillary" means a capillary-sized vessel tube that does not have a nucleus anywhere along its length.

[0225] The term "vascular congestion," as used herein, refers to a type of vascular damage that is a significant factor in the pathogenesis of various ocular diseases disclosed herein (e.g., diabetic macular edema). Vascular congestion is associated with the accumulation of fluid (e.g., intravascular blood) within vascular tissue. In some aspects, vascular congestion may be caused by hyperglycemia (i.e., high blood sugar).

[0226] As used herein, the term "macular degeneration" refers to any number of disorders and conditions in which the central portion of the retina (i.e., the macula) degenerates or loses functional activity. Such degeneration or loss of functional activity may occur, for example, as a result of cell death, decreased cell proliferation, loss of normal biological function, or a combination thereof. Macular degeneration may result in and / or be manifested by changes in the structural integrity of the cells and / or extracellular matrix of the macula, changes in the composition of normal cells and / or extracellular matrix, and / or loss of function of macular cells. Such cells may be any cell type normally present in or near the macula, including RPE cells, photoreceptors (e.g., rod and cone cell layers), and / or capillary endothelial cells. Age-related macular degeneration is the most common form of macular degeneration, but the term "macular degeneration" does not necessarily exclude macular degeneration in patients other than elderly individuals. Non-limiting examples of macular degeneration include: age-related macular degeneration (wet or dry); Best's macular dystrophy, Sorsby's fundus dystrophy, Malattia Leventinese, Doyne's honeycomb retinal dystrophy, Stargardt disease (also called Stargardt's macular dystrophy, juvenile macular degeneration, or fundus flavimaculatus), and pigment epithelial detachment-associated macular degeneration.

[0227] As used herein, the term "age-related macular degeneration" (AMD) refers to a retinopathy that typically affects older adults and is associated with loss of central vision due to damage to the central portion of the retina (i.e., the macula). AMD is typically characterized by the progressive accumulation or aggregation of yellowish, insoluble extracellular deposits called drusen (accumulations of extracellular proteins and lipids, such as amyloid beta), within the macula (primarily between the retinal pigment epithelium (RPE) and the underlying choroid). The accumulation or aggregation of these deposits within the macula can cause gradual deterioration of the macula, resulting in impaired central vision. As used herein, the term "macula" refers to the central portion of the retina responsible for high-resolution color vision.

[0228] The etiology of age-related macular degeneration (AMD) remains poorly understood, although several theories have been proposed, including oxidative stress, mitochondrial dysfunction, and inflammatory processes. An imbalance between the production and breakdown of damaged cellular components leads to the accumulation of harmful byproducts, such as intracellular lipofuscin and extracellular drusen. Early atrophy is characterized by areas of retinal pigment epithelium (RPE) thinning or depigmentation, which precedes geographic atrophy in the early stages of AMD. In advanced stages of AMD, atrophy of the RPE (geographic atrophy) and / or the development of new blood vessels (neovascularization) leads to photoreceptor death and loss of central vision. In dry (nonexudative) AMD, cellular debris called drusen accumulate between the retina and choroid, causing retinal atrophy and scarring. In more severe wet (exudative) AMD, blood vessels grow in the choroid behind the retina (neovascularization), leaking exudates and fluids, which can lead to bleeding.

[0229] Depending on the extent of drusen present, AMD can be classified into three main stages: (i) early, (ii) intermediate, and (iii) advanced or late. Early AMD is characterized by the presence of multiple small drusen (e.g., less than about 63 microns in diameter) or several medium-sized drusen (e.g., between about 63 and 124 microns in diameter). During the early stage, patients have no obvious symptoms and no vision loss. Intermediate AMD is characterized by the presence of multiple medium-sized drusen or one or more large drusen (e.g., greater than about 125 microns in diameter). During this stage, some patients may begin to experience blurred spots in the center of their vision. Advanced or late AMD is characterized by damage to large areas of retinal tissue, leading to central blind spots and eventual loss of central vision. Based on the type of damage (e.g., presence or absence of neovascularization), advanced or late AMD can be further divided into two subtypes: (i) geographic atrophy (also called atrophic AMD) and (ii) wet AMD (also called neovascular or exudative AMD).

[0230] There are two main forms of AMD: (i) dry AMD and (ii) wet AMD. Unless otherwise specified, the term "age-related macular degeneration" includes both dry AMD and wet AMD. As used herein, the term "age-related macular degeneration" also includes all types of age-related macular degeneration, regardless of cause, and any and all symptoms of age-related macular degeneration. Non-limiting examples of symptoms associated with macular degeneration (e.g., age-related macular degeneration) include loss of central vision, distortions, decreased contrast sensitivity, blurred vision, difficulty adapting to low light, sudden onset and rapid worsening of symptoms, and decreased color vision. In some instances, macular degeneration (e.g., age-related macular degeneration) may cause macular edema (i.e., swelling of the macular area due to the collection of fluid and protein deposits on or beneath the macula).

[0231] As used herein, the term "dry AMD" (also known as atrophic age-related macular degeneration or non-exudative AMD) refers to all forms of AMD other than wet (neovascular) AMD. This includes early and intermediate forms of dry AMD, as well as an advanced form known as geographic atrophy. Patients with dry AMD tend to have minimal symptoms at an earlier stage; visual loss occurs more frequently as symptoms progress to geographic atrophy.

[0232] As used herein, the term "wet AMD" (also known as neovascular age-related macular degeneration or exudative AMD) refers to a retinal condition characterized by the presence of retinal neovascularization, the most advanced form of AMD. In wet AMD, blood vessels grow from the choroidal capillaries and, in some cases, the underlying retinal pigment epithelium (choroidal neovascularization or angiogenesis) through defects in Bruch's membrane. The organization of serous or hemorrhagic exudates escaping from these vessels can cause fibrovascular scarring of the macula, with concomitant degeneration of the neural retina, detachment and rupture of the retinal pigment epithelium, vitreous hemorrhage, and permanent loss of central vision.

[0233] As used herein, the term "angiogenesis" refers to the growth of new, abnormal blood vessels in different parts of the eye, which can cause bleeding and vision loss. As used herein, the term "choroidal neovascularization" refers to the abnormal growth of new blood vessels in the choroid (i.e., the vascular layer of the eye, which contains connective tissue and lies between the retina and the sclera). In wet AMD, new blood vessels can grow into the retina through the retinal pigment epithelium (RPE) and choroid, and can impair visual function due to blood and lipid leakage. As used herein, the term "retinal neovascularization" refers to the abnormal development, proliferation, and / or growth of blood vessels overlying or within the retina, for example, on the retinal surface. Retinal neovascularization can occur in many retinopathies associated with retinal ischemia, such as diabetic retinopathy, sickle cell retinopathy, Eales disease, ocular ischemia syndrome, carotid cavernous fistula, familial exudative vitreoretinopathy, hyperviscosity syndrome, radiation retinopathy, retinal vein occlusion, retinal artery occlusion, retinal embolism, birdshot retinochoroidopathy, choroidal melanoma, chronic retinal detachment, anterior ischemic optic neuropathy (AION), non-arteritic anterior ischemic optic neuropathy (NAION), and incontinentia pigmenti. Methods for detecting neovascularization are known in the art and include, but are not limited to, measuring the expression of CD31 (also known as platelet endothelial cell adhesion molecule, PECAM-1) and vascular endothelial growth factor (VEGF) in tissues. For example: For reference, see Schluter A., ​​et al., BMC Cancerl 8(1): 272 (2018).

[0234] The term "Dychromatopsia (Color vision deficiency)" used herein refers to a symptom in which colors cannot be distinguished well due to a problem in the function of cone cells, which are one of the photoreceptors in the retina. Cone cells are one of the photoreceptors, and color vision deficiency occurs due to functional defect or loss of the cone cells (Bennett J. Gene therapy for color blindness. N Engl J Med. 2009;361(25):2483-2484). It has also been reported that in the case of AMD patients, most cases show color vision deficiency due to accompanying functional decline or loss of the cone cells (O'Neill-Biba M et al., Loss of chromatic sensitivity in AMD and diabetes: a comparative study. Ophthalmic Physiol Opt. 2010;30(5):705-716). NaIO3 (sodium iodate) can be used as a model to confirm the above color vision abnormality, and NaIO3 causes color vision abnormality by inducing the death of cone cells by giving oxidative stress to photoreceptors (Wang J et al., Direct effect of sodium iodate on neurosensory retina. Invest Ophthalmol Vis Sci. 2014;55(3):1941-1953; Takeda A et al., New Insights Into Immunological Therapy for Retinal Disorders. Front Immunol. 2020;11:1431).In one embodiment, a polypeptide comprising the amino acid sequence of TAFA, a fragment thereof, or a variant thereof of the present disclosure protects cone cells and photoreceptors comprising the same from oxidative stress caused by NaIO3, and this mechanism indicates that the polypeptide and the polynucleotide encoding the same can function as an agent for preventing or treating color vision disorders.

[0235] The above color vision deficiency can be divided into color blindness and color weakness. Color blindness is the inability to completely perceive a specific color among the three primary colors, while color weakness is a condition in which the color can be perceived but appears differently due to a problem with the receptors. Color vision deficiency is a common symptom, occurring in approximately 5-8% of the entire male population. Among color vision deficiencies, deuteranomaly is the most common, accounting for 25-45% of all color vision deficiencies. This is followed by deuteranopia, protanopia, and protanomaly. However, the frequency of these three is similar, each accounting for approximately 1% of the entire male population. Trichromatic color deficiency or complete color blindness is very rare, with a frequency of approximately 0.005%.

[0236] The term "hereditary retinal disease" as used herein refers to a retinal disease in which the structure and function of retinal cells are abnormal due to a defect or abnormality in a gene. The time of occurrence and symptoms of the hereditary retinal disease vary depending on the causative gene. Non-limiting examples of hereditary retinal diseases include retinitis pigmentosa (RP), Leber congenital amaurosis, Stargardt's disease, Coats retinopathy, cone dystrophy, choroideremia, Usher syndrome, Best's Disease, X-linked retinoschisis, hereditary color vision deficiency, and unspecified hereditary retinal dystrophy.

[0237] The term "retinitis pigmentosa" (RP), as used herein, refers to a retinal disease in which the optic cells and retinal pigment epithelium cells of the retina are damaged (degenerated). Damage to the optic cells initially causes night blindness, followed by a progressive narrowing of the field of vision and eventual blindness. The primary cause is a defect in a gene involved in the mechanism by which light is converted into electrical signals within the optic cells. Genetic abnormalities have also been found in some retinal pigment epithelium cells, and these genetic abnormalities can cause extensive retinal damage.

[0238] The first noticeable symptom of retinitis pigmentosa is the progressive destruction of rod and cone cells, with rods being damaged before cones. If the destruction of photoreceptor cells progresses to include cones, central vision is lost, leading to complete blindness.

[0239] The term "Leber congenital amaurosis" as used herein refers to an inherited retinal disease that can cause congenital blindness at or shortly after birth. Children with Leber congenital amaurosis do not have functioning rods and cones in the retina, and both rod and cone responses are lost in an electroretinogram. Leber congenital amaurosis is a genetic disease, and mutations in the genes are found in 40 to 50% of patients. Of the 12 known genetic abnormalities, 11 mutations are inherited in an autosomal recessive pattern (GUCY2D, RPE65, SPATA7, AIPL1, LCA5, RPGRIP1, CRB1, CEP290, IMPDH1, RD3, RDH12), and in rare cases, an autosomal dominant pattern (CRX) is also present.

[0240] The term "Stargardt's disease" as used herein refers to a type of retinal dystrophy that is inherited in an autosomal recessive pattern. Stargardt's disease presents between the ages of 8 and 15, with gradual loss of central vision due to macular degeneration in both eyes. It is currently thought that a mutation in a gene called ABCA4 causes Stargardt's disease. The ABCA4 gene mutation causes the accumulation of a lipofuscin-like substance in the retinal pigment epithelium (RPE), leading to RPE cell death and loss of photoreceptors (e.g., rod and cone cell layers). Mutations in ABCA4 are associated with dysplasia of cones and rods, as well as severe retinal dystrophy.

[0241] The term "Coats retinopathy" used herein refers to a retinal vascular disease that causes dilatation and vascularization of retinal capillaries, leading to exudative retinal detachment by accumulation of exudate in the intraretinal and subretinal spaces. It is known to be caused by a somatic mutation in the NDP gene on the X chromosome, resulting in a deficiency of the norrin protein, which is necessary for retinal development.

[0242] The term "cone dystrophy" used herein refers to a disease in which the cone cells in the retina, which are responsible for color vision and central vision, degenerate due to a genetic abnormality, resulting in loss of central vision. Cone dystrophy is broadly divided into pure cone dystrophy and cone-rod dystrophy. The inheritance pattern varies, including autosomal dominant, recessive, and sex-linked, and it can also occur due to mutations in the individual without showing a genetic form.

[0243] The term "Choroideremia" as used herein refers to a rare X-linked progressive degeneration of the choroid, retinal pigment epithelium, and photoreceptors. In patients, a mutation in the CHM gene causes a deficiency of the Rab escort protein 1 (REP1) protein, which causes the photoreceptor cells in the retina to lose function and gradually die. The disease usually presents in males, and typically begins with night blindness in childhood. As the disease progresses, peripheral vision declines, with central vision initially preserved to some extent. Male patients in their 40s typically have good vision, but their visual fields become very narrow, and vision loss occurs around the age of 50 to 70. Some patients also experience a decline in color perception.

[0244] The term "Usher syndrome" as used herein refers to a genetic disorder characterized by progressive visual impairment accompanied by hearing loss. The hearing impairment in Usher syndrome is due to an abnormality in the inner ear, and the visual impairment is associated with retinitis pigmentosa (RP).

[0245] The term "Best Disease" as used herein refers to an inherited retinal disease caused by mutations in the BEST1 (VMD2) gene. It progresses slowly and can cause central vision loss. Mutations in the BEST1 (VMD2) gene can cause dysfunction of Bestrophin, a calcium-chloride channel protein in the basal plasma membrane of the retinal pigment epithelium, which can impair water movement through the retinal pigment epithelium, resulting in serous retinal detachment or retinal pigment epithelial detachment.

[0246] The term "X-linked Retinoschisis" used herein refers to a disease in which the inner retina is separated from the rest of the retina due to mutations in the retinoschisis gene (RS 1). Retinoschisis causes visual impairment due to abnormal separation of the nerve fiber layer, one of the ten layers that make up the retina. Juvenile retinoschisis is a rare X-linked recessive genetic disease with a worldwide prevalence of 1:120,000.

[0247] The term "neuropathic pain" refers to pain resulting from injury, damage, and / or dysfunction affecting any level of the central nervous system (CNS) and / or peripheral nervous system. The term "neuropathic pain" encompasses any and all types of neuropathic pain, regardless of its cause or any symptoms.

[0248] Neuropathic pain includes central neuropathic pain and peripheral neuropathic pain. As used herein, the term "central neuropathic pain" refers to pain resulting from a disorder, congenital defect, or damage to the central nervous system (i.e., the brain or spinal cord). As used herein, the term "peripheral neuropathic pain" refers to pain resulting from damage or infection of peripheral sensory nerves.

[0249] Symptoms of neuropathic pain may include persistent / chronic pain, spontaneous pain and allodynia (e.g., painful responses to normally nonpainful stimuli), hyperalgesia (e.g., typically only mild discomfort, such as a pinprick), hyperesthesia (e.g., excessive physical sensitivity to stimuli, especially skin stimuli), or dysalgesia (e.g., where a brief period of discomfort becomes prolonged and severe pain). In some embodiments, symptoms may be long-lasting and may persist even after the primary cause, if present, is resolved. Merck Manual, Neuropathic Pain, merckmanuals.com / professional / neurologic-disorders / pain / neuropathic-pain; Campbell JN and Meyer RANeuron52 (1): 77-92 (2006).

[0250] In some embodiments, the types of neuropathic pain may include (1) neuralgia, (2) deafferentation pain syndrome, (3) complex regional pain syndrome (CRPS), and (4) neuropathy (central or peripheral).

[0251] In some embodiments, neuropathic pain is neuralgia, which generally refers to pain radiating along the course of one or more specific nerves (e.g., cranial nerves) without obvious pathological changes in the nerve structure. Neuralgia includes, but is not limited to, trigeminal neuralgia (TN), atypical trigeminal neuralgia (ATN), occipital neuralgia, glossopharyngeal neuralgia, postherpetic neuralgia (due to shingles or herpes), peripheral nerve injury pain, sciatica, low back pain, and atypical facial pain. Chemical irritants, chronic kidney disease, diabetes, inflammation, trauma (including surgery), compression of a nerve by a nearby structure (e.g., a tumor), certain medications (e.g., cisplatin, paclitaxel, or vincristine), porphyria (a blood disorder), and infections (e.g., herpes zoster (shingles), HIV / AIDS, Lyme disease, or syphilis) can all lead to neuralgia.

[0252] In some embodiments, the neuropathic pain is a deafferentation pain syndrome, which may result from a loss of sensory input from a part of the body (e.g., caused by interruption of peripheral sensory fibers or nerves from the central nervous system). Deafferentation pain syndromes include, but are not limited to, pain resulting from brain or spinal cord injury, post-stroke pain, phantom pain, paraplegia, brachial plexus avulsion injury, and lumbar radiculopathies.

[0253] In some embodiments, the neuropathic pain is "Complex Regional Pain Syndrome" (CRPS), which is a chronic pain condition that most commonly affects the arms or legs. In some embodiments, CRPS occurs after injury, surgery, stroke, or heart attack. In certain embodiments, the CRPS is CRPS Type I (CRPS-I) (also known as reflex sympathetic dystrophy syndrome). Individuals without identified nerve damage are often classified as having CRPS-I. In other embodiments, the CRPS is CRPS Type II (CRPS-II) (also known as causalgia) associated with identified nerve damage.

[0254] In some embodiments, neuropathic pain is a neuropathy, which refers to pain resulting from functional or pathological changes (e.g., disease or injury) in a nerve. Neuropathy can often be clinically characterized by abnormalities in sensory or motor neurons. In certain embodiments, the neuropathy is a central neuropathy (e.g., functional or pathological changes in the central nervous system). In other embodiments, the neuropathy is a peripheral neuropathy (e.g., functional or pathological changes in one or more peripheral nerves, including motor, sensory, autonomic, or a combination thereof). In some embodiments, the peripheral neuropathy involves functional or pathological changes in a single nerve or group of nerves (i.e., mononeuropathy). In some embodiments, the peripheral neuropathy involves functional or pathological changes affecting multiple nerves (locally or systemically) (i.e., polyneuropathy). In some embodiments, the peripheral neuropathy affects both sides of the body approximately equally (i.e., symmetrical polyneuropathy). In some embodiments, peripheral neuropathy affects disparate areas of the body (e.g., mononeuropathy multiplex, multifocal mononeuropathy, or multiple mononeuropathy).

[0255] "Mononeuropathy," as described herein, is a peripheral neuropathy involving loss of movement or sensation in an area caused by damage or destruction of a single peripheral nerve or group of nerves. Mononeuropathy is most often caused by localized injury or trauma, such as persistent pressure or compression on a single nerve. However, certain systemic disorders (e.g., mononeuritis multiplex) can also cause mononeuropathy. In some embodiments, localized injury or trauma results in the destruction of the myelin sheath (covering) of a nerve or part of a nerve cell (the axon), which can slow or prevent the conduction of impulses along the nerve. In some embodiments, mononeuropathy can affect any part of the body. Examples of mononeuropathy include sciatic nerve dysfunction, general pelvic nerve dysfunction, radial nerve dysfunction, ulnar nerve dysfunction, cranial mononeuropathy VI, cranial mononeuropathy VII, cranial mononeuropathy III (compressive type), cranial neuropathy III (diabetic type), axillary nerve dysfunction, carpal tunnel syndrome, femoral nerve dysfunction, tibial nerve dysfunction, Bell's palsy, thoracic outlet syndrome, carpal tunnel syndrome, and sixth (abducens) nerve palsy. Finnerup NB et al., Pain 157(8): 1599-1606 (2016); National Institute of Neurological Disorders and Stroke, Peripheral Neuropathy Fact Sheet, available at ninds.nih.gov / disorders / peripheralneuropathy / detailj3eripheralneuropathy.htm. In some embodiments, the mononeuropathy pain is sciatica.

[0256] As used herein, "polyneuropathy" is a peripheral neuropathy involving loss of movement or sensation in an area caused by damage or destruction of multiple peripheral nerves. Polyneuropathy pain includes, but is not limited to, post-polio syndrome, post-mastectomy syndrome, diabetic neuropathy, alcoholic neuropathy, amyloid, toxins, AIDS, hypothyroidism, uremia, vitamin deficiencies, chemotherapy-induced pain, 2',3'-didexoicitidine (ddC) treatment, Guillain-Barré syndrome, or Fabry's disease. Finnerup NB et al., Pain 157(8): 1599-1606 (2016); National Institute of Neurological Disorders and Stroke, Peripheral Neuropathy Fact Sheet, available at ninds.nih.gov / disorders / peripheralneuropathy / detail 3peripheralneuropathy.htm. In some embodiments, the polyneuropathy is diabetic peripheral neuropathy. In certain embodiments, diabetic peripheral neuropathy is caused by high blood glucose levels (blood sugar) and / or high levels of fats (e.g., triglycerides) in the blood of a diabetic patient, which cause damage to the subject's peripheral nerves.

[0257] In some embodiments, the peripheral neuropathy disclosed herein can be classified based on the part of the nerve cell (e.g., the axon, the myelin sheath, or the cell body) that is damaged or affected. In some embodiments, the peripheral neuropathy is a distal axonopathy, which is due to metabolic or toxic alterations of the axon. In some embodiments, the metabolic disorder includes a deficiency syndrome such as diabetes, renal failure, malnutrition, and alcoholism. In some embodiments, the metabolic disorder is diabetes, and the distal axonopathy is diabetic neuropathy.

[0258] In some embodiments, peripheral neuropathy is a myelinopathy, resulting from a primary attack on the myelin sheath or myelinating Schwann cells, causing an acute failure of impulse conduction. The most common cause is acute inflammatory demyelinating polyneuropathy (AIDP; also known as Guillain-Barré syndrome), but other causes include chronic inflammatory demyelinating syndrome (CIDP), genetic metabolic disorders (e.g., leukodystrophies), or toxins.

[0259] In some embodiments, peripheral neuropathy is a neuropathic condition resulting from the destruction of peripheral nervous system (PNS) neurons. In some embodiments, the neuropathy is caused by a motor neuron disease, a sensory neuropathy (e.g., shingles), a toxin, or an autonomic dysfunction. In some embodiments, the neuropathy is caused by a neurotoxin, such as the chemotherapeutic agent vincristine.

[0260] Neuropathic pain can result from or be associated with a variety of etiologies (e.g., physical injury (e.g., trauma or repetitive stress), disease or disorder, exposure to a toxic substance, or a combination thereof). In some embodiments, neuropathic pain results from or is associated with a traumatic injury or injury, such as a nerve compression injury (e.g., nerve crush, nerve stretching, nerve entrapment, or incomplete nerve transection); a spinal cord injury (e.g., a hemisection of the spinal cord); damage or injury to a peripheral nerve (e.g., a motor nerve, a sensory nerve, or an autonomic nerve, or a combination thereof), limb amputation; a contusion; inflammation (e.g., inflammation of the spinal cord); or a surgical procedure. In some embodiments, neuropathic pain results from or is associated with repetitive stress, such as repetitive, slow, and / or forceful activities requiring movement of any group of joints for a prolonged period of time. Without being bound by any theory, the resulting irritation can cause ligaments, tendons, and muscles to become inflamed and swollen, constricting the narrow passages through which the nerves pass (e.g., ulnar neuropathy and carpal tunnel syndrome, which are neuropathies of a trapped or compressed nerve at the elbow or wrist).In some embodiments, neuropathic pain is caused by or associated with a disease or disorder, including, for example: an ischemic event (e.g., a stroke or heart attack), multiple sclerosis, a metabolic and / or endocrine disease or disorder (e.g., diabetes, metabolic disease, and hypertrophy, a condition caused by overproduction of growth hormone and characterized by abnormal enlargement of skeletal parts, including joints, resulting in nerve compression and pain), a disease of the small blood vessels that reduces oxygen supply to peripheral nerves and causes damage to nerve tissue (e.g., vasculitis, i.e., inflammation of the blood vessels), an autoimmune disease (e.g., Sjogren's syndrome, lupus, rheumatoid arthritis, and acute inflammatory demyelinating neuropathy, also known as Guillain-Barré syndrome), kidney disease, cancer or tumors (e.g., neoplastic tumors, neuromas, paraneoplastic syndromes, and toxicity from chemotherapy agents and radiation in cancer treatment), an infection (e.g., herpes zoster, varicella, shingles, Epstein-Barr virus, West Nile virus) (e.g., infections caused by viruses, cytomegalovirus and herpes simplex virus, acquired immunodeficiency syndrome (AIDS) or Lyme disease, bacteria such as diphtheria, and leprosy), inflammatory disorders, peripheral neuropathy (e.g., neuroma), genetic or de novo inherited disorders (e.g., Charcot-Marie-Tooth disorder, which causes muscle weakness and wasting of the legs and feet, gait abnormalities, loss of tendon reflexes, paraplegia, etc.), mononeuropathy or polyneuropathy. In some embodiments, the neuropathic pain arises from or is associated with an infectious agent (e.g., tick-borne infection, herpes varicella-zoster, Epstein-Barr virus, West Nile virus, cytomegalovirus, herpes simplex virus, AIDS). In some embodiments, neuropathic pain results from or is associated with exposure to toxic agents, including, for example, drugs, alcohol, heavy metals (e.g., lead, arsenic, mercury), industrial agents (e.g., fumes from solvents, adhesives), or nitrous oxide.

[0261] The term “neuropathic pain associated with a disease or disorder” refers to neuropathic pain that accompanies, is caused by, or results from a disease or disorder (e.g., as disclosed herein).

[0262] The terms "nucleic acid," "nucleic acid molecule," "nucleotide sequence," "polynucleotide," and grammatical variations thereof are used interchangeably and refer to a sequence of nucleotides linked by phosphodiester bonds. Polynucleotides are represented herein in the 5' to 3' direction. The polynucleotides of the present disclosure may be deoxyribonucleic acid (DNA) molecules or ribonucleic acid (RNA) molecules. Nucleotide bases are represented herein by single-letter codes such as adenine (A), guanine (G), thymine (T), cytosine (C), inosine (I), and uracil (U).

[0263] As used herein, the terms "operably linked" or "operably linked" mean that the DNA sequences to be linked are positioned adjacent to each other so that they perform the desired function. For example, if a particular promoter helps initiate transcription of a coding sequence (e.g., a foreign gene), such a promoter may be operably linked to the coding region. The promoter and coding region do not necessarily need to be adjacent, as long as this functional relationship is maintained.

[0264] The terms "pharmaceutically acceptable carrier", "pharmaceutically acceptable excipient" and grammatical variations thereof include all formulations approved by a regulatory agency of the United States Federal Government or listed in the United States Pharmacopeia for use in animals, including humans, as well as any carrier or diluent that does not produce undesirable physiological effects that would render the composition unsuitable for administration to a subject and does not abolish the biological activity and properties of the administered complex. Excipients and carriers that are useful in preparing pharmaceutical compositions and are generally safe, non-toxic and desirable are included.

[0265] The term "pharmaceutical composition" as used herein refers to one or more of the compositions described herein (e.g., polypeptides, polynucleotides, vectors, cells and / or recombinant viruses) mixed or mixed with or suspended in one or more other chemical components, such as pharmaceutically acceptable carriers and excipients.

[0266] As used herein, the terms "promoter" and "promoter sequence" are used interchangeably and refer to a DNA sequence capable of controlling the expression of a coding sequence or functional RNA. Typically, the coding sequence is located 3' to the promoter sequence. A promoter may be derived entirely from a native gene, composed of different elements derived from different promoters found in nature, or may even include synthetic DNA segments. Those skilled in the art will appreciate that different promoters can direct the expression of a gene in different tissues or cell types, at different developmental stages, or in response to different environmental or physiological conditions. A promoter that causes a gene to be predominantly expressed in most host cell types is commonly referred to as a "constitutive promoter." A promoter that causes a gene to be expressed in a specific cell type is commonly referred to as a "cell-specific promoter" or a "tissue-specific promoter." Promoters that direct gene expression during specific stages of development or cell differentiation are commonly referred to as "development-specific promoters" or "cell differentiation-specific promoters." Promoters that are induced and gene expression occurs after exposure or treatment of cells with inducing agents, biological molecules, chemicals, ligands, light, etc. are commonly referred to as "inducible promoters" or "regulatable promoters." Furthermore, because the precise boundaries of regulatory sequences are often not fully defined, it is generally accepted that DNA fragments of different lengths can have identical promoter activity.

[0267] The promoter sequence typically borders the transcription initiation site at its 3' end and extends upstream (5') to include the minimum number of bases or elements necessary to initiate transcription at a detectable level above background. Within the promoter sequence, a protein binding domain (consensus sequence) responsible for binding RNA polymerase, as well as a transcription initiation site (e.g., conveniently defined by mapping to nuclease S1), will be found. In some embodiments, the promoters usable in the present disclosure include tissue-specific promoters.

[0268] As used herein, the term "gene regulatory region" or "regulatory region" refers to a nucleotide sequence located upstream (5' noncoding sequence), within, or downstream (3' noncoding sequence) of a coding region that influences the transcription, RNA processing, stability, or translation of the associated coding region. A regulatory region may include a promoter, a translation leader sequence, an intron, a polyadenylation recognition sequence, an RNA processing site, an effector binding site, or a stem-loop structure. If the coding region is intended for expression in a eukaryotic cell, a polyadenylation signal and a transcription termination sequence will typically be located 3' to the coding sequence.

[0269] In some embodiments, the polynucleotides described herein (including, for example, foreign genes and non-translated nucleic acid sequences encoding polypeptides comprising the amino acid sequence of a TAFA protein, fragments thereof, or variants thereof) may include a promoter and / or other expression (e.g., transcriptional) regulatory elements operably associated with one or more coding regions. In operably associated, a coding region for a gene product is associated with one or more regulatory regions in such a way that expression of the gene product is under the influence or control of the regulatory region(s). For example, a coding region and a promoter are "operably associated" if induction of promoter function results in transcription of an mRNA encoding the gene product encoded by the coding region, and the nature of the linkage between the promoter and the coding region does not interfere with the ability of the promoter to direct expression of the gene product or with the ability of the DNA template to be transcribed. Expression control elements other than promoters, such as enhancers, operators, repressors, and transcription termination signals, may also be operably associated with the coding region to direct gene product expression.

[0270] As used herein, the terms "subject," "patient," "individual," and "host" and variations thereof are used interchangeably and refer to any mammalian subject to which any of the compositions described herein (e.g., polypeptides, polynucleotides, recombinant expression constructs, vectors, cells, pharmaceutical compositions, or recombinant viruses) are administered. Non-limiting examples include humans, livestock (e.g., dogs, cats, etc.), farm animals (e.g., cows, sheep, pigs, horses, etc.), and laboratory animals (e.g., monkeys, rats, mice, rabbits, guinea pigs, etc.), particularly humans, in need of diagnosis, treatment, or therapy. The methods described herein are applicable to both human prophylactic or therapeutic and veterinary uses.

[0271] As used herein, the phrase "subject in need thereof" includes subjects such as mammalian subjects that would benefit from administration of the compositions described herein.

[0272] The term "therapeutically effective amount," as used herein, refers to an amount of a reagent or pharmaceutical complex comprising a composition of the present disclosure (e.g., a polypeptide comprising a TAFA protein fragment or variant thereof, or a polynucleotide encoding the same) sufficient to produce a desired therapeutic, pharmacological, and / or physiological effect in a subject in need thereof. A therapeutically effective amount may be a "prophylactic amount," as prevention can be considered a treatment.

[0273] The term "foreign gene" as used herein refers to at least one polynucleotide (e.g., a polynucleotide encoding a polypeptide comprising a TAFA protein, or a fragment or variant thereof) or a polynucleotide region or an expression product of said polynucleotide or polynucleotide region, a polynucleotide encoding a polypeptide or multi-polypeptide, or a promoting or regulatory nucleic acid, encoded in a recombinant expression construct. In some embodiments, the foreign gene may be heterologous to the cell into which it is inserted (or transduced) (i.e., not naturally expressed in the cell).

[0274] The terms "treat," "treatment," or "treating," as used herein, refer to, for example, reducing the severity of a disease or condition, reducing the duration of the disease, improving or eliminating one or more symptoms associated with the disease or condition, or providing a beneficial effect to a subject suffering from the disease or condition without necessarily curing the disease or condition. The terms also include preventing or preventing the disease or condition or its symptoms.

[0275] The term "vector" or "construct" as used herein refers to any vehicle into which a nucleic acid or gene can be inserted, for example, a delivery vehicle into which a nucleic acid sequence can be inserted and replicated in a cell. The nucleic acid sequence that can be inserted into the vector can be exogenous or heterologous. The nucleic acid sequence can be a foreign gene. Examples of constructs include, but are not limited to, plasmids, cosmids, and viruses (e.g., AAV). Those skilled in the art can produce such vectors or constructs by standard recombinant techniques (see, e.g., Maniatis, et al., Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Press, Cold Spring Harbor, NY, 1988; and Ausubel et al., In: Current Protocols in Molecular Biology, John, Wiley & Sons, Inc, NY, 1994). As used herein, the term "expression vector" or "expression construct" refers to a vector or construct comprising a nucleotide sequence encoding at least a portion of a gene product to be transcribed. In some cases, the RNA molecule is then translated into a protein, polypeptide, or peptide. Expression constructs may include various regulatory elements. In addition to regulatory sequences that regulate transcription and translation, vectors and expression vectors may also include nucleotide sequences that provide other functions. Viruses usable in the present invention include, but are not limited to, retroviruses, herpes simplex viruses, lentiviruses, poxviruses, vaccinia viruses, rhabdoviruses, adenoviruses, helper-dependent adenoviruses, adeno-associated viruses (AAV), and the like.

[0276] The vector may be engineered to encode a selectable marker or reporter that provides for the selection or identification of cells incorporating the vector. Expression of the selectable marker or reporter allows for the identification and / or selection of host cells that integrate and express other coding regions included in the vector. Examples of selectable marker genes known and used in the art include genes conferring resistance to ampicillin, streptomycin, gentamicin, kanamycin, hygromycin, bialaphos herbicides, sulfonamides, and the like; and genes used as phenotypic markers, such as anthocyanin regulatory genes and isopentanyl transferase genes. Examples of reporters known and used in the art include luciferase (Luc), green fluorescent protein (GFP), chloramphenicol acetyltransferase (CAT), β-galactosidase (LacZ), β-glucuronidase (Gus), and the like. Selectable markers may also be considered reporters.

[0277] Typically, recombinant adeno-associated viruses (AAVs) are produced by triple transfection of host cells (e.g., HEK293 cells). These consist of 1) an AAV construction plasmid containing a gene expression cassette flanked by inverted terminal repeats (ITRs); 2) a "Rep-Cap plasmid" that provides the Rep proteins necessary for replication of the adeno-associated virus genome and the capsid proteins that constitute the viral particle; and 3) a "helper plasmid" that provides adenoviral proteins (E2a, E4) and RNA (VA RNA) that aid in the AAV life cycle. AAVs are produced when these three plasmids are transfected into cells such as HEK293 cells, which provide the adenoviral E1 and E3 genes.

[0278] As used herein, the term "dual helper plasmid" refers to a plasmid capable of providing two or more of the requirements for producing AAV in a cell. As will be apparent from the present disclosure, in some aspects, the dual helper plasmids described herein provide the requirements 2) a "Rep-Cap plasmid" and 3) a "helper plasmid" described above. For example, in some aspects, the dual helper plasmid comprises a rep gene, a cap gene, an E2a gene, an E4 gene, and a VA RNA gene.

[0279] The dual helper plasmids described herein not only include the genes described above, but the genes are also arranged in a specific configuration within the plasmid. For example, in some aspects, the E2a gene, the E4 gene, and the VA RNA gene are sequentially linked within the dual helper plasmid, and the rep gene and the cap gene (collectively referred to herein as the "rep-cap genes") are sequentially linked in a clockwise direction (5' to 3') between the 5' end of the E2a gene and the 3' end of the VA RNA gene. More specifically, in some aspects, the 5' end of the rep-cap gene is linked to the 5' end of the E2a gene, and wherein the 3' end of the rep-cap gene is linked to the 3' end of the VA RNA gene. In some aspects, the E2a gene, the E4 gene, and the VA RNA gene are sequentially linked, and the rep-cap gene is located counterclockwise (3' to 5') between the 5'-end of the E2a gene and the 3'-end of the VA RNA gene. More specifically, in some aspects, the 3'-end of the rep-cap gene is linked to the 5'-end of the E2a gene, and wherein the 5'-end of the rep-cap gene is linked to the 3'-end of the VA RNA gene.

[0280] The term "cell" as used herein includes eukaryotic and prokaryotic cells, and refers to any transformable cell capable of replicating the vector or expressing a gene encoded by the vector. The cell may be transfected, transduced, or transformed by the vector, which refers to a process in which an exogenous polynucleotide (nucleic acid molecule) is transferred or introduced into a host cell. The term "transformation" as used herein is used to include the terms transfected and transduced.

[0281] The (host) cell of the present invention is not limited, but is preferably an insect cell or a mammalian cell, more preferably Sf9 in the case of an insect cell, and HEK293 cells, HeLa cells, ARPE-19 cells, RPE-1 cells, HepG2 cells, Hep3B cells, Huh-7 cells, C8D1a cells, Neuro2A cells, CHO cells, MES13 cells, BHK-21 cells, COS7 cells, COP5 cells, A549 cells, MCF-7 cells, HC70 cells, HCC1428 cells, BT-549 cells, PC3 cells, LNCaP cells, Capan-1 cells, Panc-1 cells, MIA PaCa-2 cells, SW480 cells, HCT166 cells, LoVo cells, A172 cells, MKN-45 cells, MKN-74 cells, Kato-III cells, NCI-N87 cells, HT-144 cells, SK-MEL-2 cells, SH-SY5Y cells, C6 cells, HT-22 cells, PC-12 cells, NIH3T3 cells, etc. can be used. In some embodiments, the host cell is an isolated host cell.

[0282]

[0283] II. Polypeptides

[0284] According to one aspect of the present invention, the present invention provides a polypeptide having the ability to increase neurite length and / or branch point.

[0285] According to a preferred embodiment of the present invention, the polypeptide comprises an amino acid sequence of a TAFA (TAFA Chemokine Like Family Member) protein, a fragment thereof, or a variant thereof. According to a preferred embodiment of the present invention, the polypeptide comprises an amino acid sequence of the following general formula 7 (from the N-terminus to the C-terminus):

[0286] General Formula 7

[0287] X1-X2-X3-GTCEV-X4-A-X5-H-X6- CCN-X7-N-X8-IEE- -X20-X21-X22-WWC-X23-M-X24-PC-X25-X26-GE-X27-CK-X28-LPD-X29-X30-GW-X31-C-X32-X33-G-X34-K-X35-KTT-X36-X37-X38-X39

[0288] In the above general formula 1,

[0289] X1 is either non-existent, V, I, or L,

[0290] X2 is K, E, R or Q,

[0291] X3 is G, T, Q, P or A,

[0292] X4 is V or I,

[0293] X5 is A, L, V or I,

[0294] X6 is R or L,

[0295] X7 is K, R or Q,

[0296] X8 is R or K,

[0297] X9 is R or L,

[0298] X10 is V or G,

[0299] X11 is K or N,

[0300] X12 is F or L,

[0301] X13 is P or S,

[0302] X14 is Q or K,

[0303] X15 is R, H or Q,

[0304] X16 is A, N, S or T,

[0305] X17 is A, Q, R, K or T,

[0306] X18 is D or E,

[0307] X19 is S or A,

[0308] X20 is I, E, L, A or V,

[0309] X21 is Q, G or E,

[0310] X22 is K or R,

[0311] X23 is H, Q or E,

[0312] X24 is E, Q, N, D, S or H,

[0313] X25 is L, V or M,

[0314] X26 is E, D, P, L or A,

[0315] X27 is E or D,

[0316] X28 is V, T, A or I,

[0317] X29 is L, N, R, Y, S or Q,

[0318] X30 is S, K or T,

[0319] X31 is S or M,

[0320] X32 is S, A or Y,

[0321] X33 is S, T or R,

[0322] X34 is N or H,

[0323] X35 is V or I,

[0324] X36 is R or K,

[0325] X37 is either non-existent or is V, A, G, M or N,

[0326] X38 is either non-existent or is T, I, N, F or S, and

[0327] X39 is either absent or is R, H, V, K, I, or Q.

[0328] According to a preferred embodiment of the present invention, the polypeptide comprises at least one amino acid sequence selected from the group consisting of amino acid sequences set forth in SEQ ID NOs: 87 to 141.

[0329] According to a preferred embodiment of the present invention, the polypeptide may be composed of a sequence of 8 to 61 amino acids.

[0330] In some embodiments, the polypeptide may consist of a sequence of 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60 or 61 amino acids.

[0331] According to a preferred embodiment of the present invention, the polypeptide comprises a sequence having at least 50% sequence identity with the amino acid sequence of SEQ ID NO: 1, SEQ ID NO: 15 or SEQ ID NO: 87.

[0332] In some embodiments, the polypeptide has an amino acid sequence of SEQ ID NO: 1, SEQ ID NO: 15 or SEQ ID NO: 87 that is at least 50%, at least 51%, at least 52%, at least 53%, at least 54%, at least 55%, at least 56%, at least 57%, at least 58%, at least 59%, at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least Has at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, or at least 95% sequence identity.

[0333] According to a preferred embodiment of the present invention, the amino acid sequence of the polypeptide comprises the amino acid sequence of the following general formula 1 (from N-terminus to C-terminus):

[0334] <General Formula 1>

[0335] X1-GE-X2-CK-X3-L

[0336] In the above general formula 1

[0337] X1 is E, D, P, L or A,

[0338] X2 is D or E, and

[0339] X3 is T, V, I or A.

[0340] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of Formula 1 comprises the amino acid sequence of SEQ ID NO: 142.

[0341] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of Formula 1 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 14.

[0342] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of the general formula 1 may consist of a sequence of 8 to 43 amino acids.

[0343] In some embodiments, the polypeptide comprising the amino acid sequence of Formula 1 may be comprised of a sequence of 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42 or 43 amino acids.

[0344] According to a preferred embodiment of the present invention, the amino acid sequence of the polypeptide comprises the amino acid sequence of the following general formula 2 (from N-terminus to C-terminus):

[0345] <General Formula 2>

[0346] IV-X4-X5-X6-WWC-X7-M-X8-PC-X9-X1-GE-X2-CK-X3-L

[0347] In the above general formula 2

[0348] X1 is E, D, P, L or A,

[0349] X2 is D or E,

[0350] X3 is T, V, I or A,

[0351] X4 is I, E, A, L or V,

[0352] X5 is Q, E or G,

[0353] X6 is K or R,

[0354] X7 is E, H or Q,

[0355] X8 is E, Q, N, D, S or H, and

[0356] X9 is L, M or V.

[0357] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of the general formula 2 may comprise the amino acid sequence of SEQ ID NO: 143.

[0358] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of the general formula 2 may comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 28 to 51.

[0359] According to a preferred embodiment of the present invention, the amino acid sequence of the polypeptide comprises the amino acid sequence of the following general formula 3 (from N-terminus to C-terminus):

[0360] <General Formula 3>

[0361] X1-GE-X2-CK-X3-LPD-X4-X5-GWSCS-X6-GNK-X7-KTTKVTR

[0362] In the above general formula 3

[0363] X1 is E, D, P, L or A,

[0364] X2 is D or E,

[0365] X3 is T, V, I or A,

[0366] X4 is Y, S or L,

[0367] X5 is S or T,

[0368] X6 is S or T, and

[0369] X7 is V or I.

[0370] In some embodiments, the amino acid sequence of a polypeptide comprising the amino acid sequence of the general formula 3 may comprise the amino acid sequence of SEQ ID NO: 144.

[0371] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of the general formula 3 may comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 52 to 58.

[0372] In some embodiments, the amino acid sequence of the polypeptide may comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 152 to 171.

[0373] According to a preferred embodiment of the present invention, the amino acid sequence of the polypeptide may include the amino acid sequence of the following general formula 4 (from the N-terminus to the C-terminus):

[0374] <General Formula 4>

[0375] G-X1-VAGTT-X2-X3-X4-PSCV-X5-X6-X7

[0376] In the above general formula 4

[0377] X1 is Q or K,

[0378] X2 is R, H or Q,

[0379] X3 is A, N, S or T,

[0380] X4 is R, A, Q, K, or T,

[0381] X5 is D or E,

[0382] X6 is A or not present, and

[0383] X7 is either S, A or non-existent.

[0384] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of the general formula 4 may comprise the amino acid sequence of SEQ ID NO: 145.

[0385] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of the general formula 4 may comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 15 to 27.

[0386] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of the general formula 4 may consist of a sequence of 15 to 46 amino acids.

[0387] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of Formula 4 may be comprised of a sequence of 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45 or 46 amino acids.

[0388] According to a preferred embodiment of the present invention, the amino acid sequence of the polypeptide comprises the amino acid sequence of the following general formula 5 (from N-terminus to C-terminus):

[0389] <General Formula 5>

[0390] X8-IEE-X9-SQT-X10-X11-CSC-X12-X13-G-X1-VAGTT-X2-X3-X4-PSCV-X5-X6-X7

[0391] In the above general formula 5

[0392] X1 is Q or K,

[0393] X2 is R, H or Q,

[0394] X3 is A, N, S or T,

[0395] X4 is R, A, Q, K, or T,

[0396] X5 is D or E,

[0397] X6 is A or does not exist,

[0398] X7 is S, A or not present,

[0399] X8 is R or K,

[0400] X9 is R or L,

[0401] X10 is V or G,

[0402] X11 is K or N,

[0403] X12 is F or L, and

[0404] X13 is P or S.

[0405] In some embodiments, the amino acid sequence of a polypeptide comprising the amino acid sequence of Formula 5 may comprise the amino acid sequence of SEQ ID NO: 146.

[0406] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of the general formula 5 may comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 59 to 74.

[0407] According to a preferred embodiment of the present invention, the amino acid sequence of the polypeptide comprises the amino acid sequence of the following general formula 6 (from N-terminus to C-terminus):

[0408] <General Formula 6>

[0409] G-X1-VAGTT-X2-X3-X4-PSCV-X5-X6-X7-IV-X8-X9-KWWC-X10-M-X11-PC-X12

[0410] In the above general formula 6

[0411] X1 is Q or K,

[0412] X2 is R, H or Q,

[0413] X3 is A, N, S or T,

[0414] X4 is R, A, Q, K or T,

[0415] X5 is D or E,

[0416] X6 is A or does not exist,

[0417] X7 is S, A or not present,

[0418] X8 is I, A, V or L,

[0419] X9 is Q or E,

[0420] X10 is H or Q,

[0421] X11 is N, D, S or H, and

[0422] X12 is L or M.

[0423] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of the general formula 6 may comprise the amino acid sequence of SEQ ID NO: 147.

[0424] In some embodiments, the amino acid sequence of the polypeptide comprising the amino acid sequence of the general formula 6 may comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 75 to 85.

[0425] In some embodiments, the amino acid sequence of the polypeptide may comprise an amino acid sequence selected from the group consisting of SEQ ID NOs: 172 to 184.

[0426]

[0427] III. Nucleic Acids

[0428] According to another aspect of the present invention, the present invention provides a nucleic acid molecule encoding the polypeptide.

[0429]

[0430] The above polypeptide, TAFA protein, or fragment or variant thereof is as described in II.

[0431] Nucleic acid molecules useful in the present disclosure are not particularly limited, as long as the nucleic acid molecule can be translated into a polypeptide upon transfection into a cell. In some embodiments, the nucleic acid molecule encodes a polypeptide (e.g., a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142 to 147) or a fusion protein.

[0432] In some embodiments, the nucleic acid encodes a protein useful for the prevention or treatment of a disease or disorder, such as those described herein. In some embodiments, the nucleic acid encodes a peptide for the prevention or treatment of a specific disease, the peptide being intended for sustained expression within the body of the subject or patient.

[0433] In some embodiments, the nucleic acid molecule has at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity to a nucleic acid molecule encoding the TAFA protein or a fragment thereof, or a variant thereof.

[0434] In some embodiments, the nucleic acid encoding the polypeptide further comprises a sequence encoding a signal sequence.

[0435] In some embodiments, the polynucleotide described herein further comprises a regulatory element. Thus, in some embodiments, the polynucleotide comprises (1) a regulatory element, (2) an untranslated nucleic acid sequence described herein (e.g., an EF-1α intron or a fragment thereof), and (3) a foreign gene (e.g., a nucleic acid molecule encoding a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142-147).

[0436] The term "regulatory element" as used herein refers to a nucleic acid sequence that regulates (e.g., increases or decreases) the expression of an operably linked nucleic acid. Regulatory elements useful in the present disclosure include an enhancer (e.g., a CMV enhancer), a promoter (e.g., a CMV promoter, an EF-1α promoter, or a β-actin promoter), an exon (e.g., exon 1 or exon 2), a splicing donor sequence, an acceptor sequence, or a combination thereof. In some embodiments, the regulatory element may include a sequence for transcription termination (e.g., a poly A sequence), a sequence for stably expressing a foreign gene (e.g., a WPRE sequence), a sequence for reducing the occurrence of foreign gene-specific immunity (e.g., an miRNA target sequence), or a combination thereof.

[0437]

[0438] IV. A vector containing a nucleic acid encoding a polypeptide

[0439] According to another aspect of the present invention, the present invention provides a vector comprising the nucleic acid molecule.

[0440]

[0441] The nucleic acid molecule is as described in III.

[0442] As described herein, such vectors are useful for recombinant expression in host cells and cells targeting therapeutic intervention. In some embodiments, vectors useful for delivering a polynucleotide described herein (e.g., a nucleic acid molecule encoding a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142-147) include viral vectors. Examples of viruses that can be used as vectors in the present disclosure include, but are not limited to, retroviruses, herpes simplex viruses, lentiviruses, poxviruses, vaccinia viruses, rhabdoviruses, adenoviruses, helper-dependent adenoviruses, adeno-associated viruses (AAV), baculoviruses, and combinations thereof. In some embodiments, vectors that can be used in the present disclosure include non-viral vectors. Non-limiting examples of such vectors include plasmids, cosmids, yeast artificial chromosomes (YACs), bacteriophages, and combinations thereof.

[0443] In some embodiments, the vector further comprises one or more sequences selected from the group consisting of a promoter sequence, an enhancer sequence, an exon sequence, an intron sequence, a signal sequence coding sequence, a splicing donor sequence, and one or more adeno-associated virus inverted terminal repeat (ITR) sequences.

[0444]

[0445] According to another aspect of the present invention, the present invention provides a recombinant viral particle comprising the vector and capsid protein.

[0446]

[0447] According to a preferred embodiment of the present invention, the virus may be AAV.

[0448]

[0449] V. AAV

[0450] In some embodiments, the polynucleotide described herein (e.g., a nucleic acid molecule encoding a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOS: 142 to 147) is delivered into, for example, a cell using AAV. Adeno-associated virus (AAV), a single-stranded DNA virus, is a helper-dependent human parvovirus. The AAV genome has a size of about 4.7 kbp and is composed of an N-terminus encoding a rep gene involved in viral replication and expression of viral genes, a C-terminus encoding a cap gene encoding a capsid protein of the virus, and an inverted terminal repeat (ITR) having about 145 base pairs inserted at each terminus. The 145 bp ITR has a T-shaped structure, functions as an origin of replication during viral genome replication, and acts as a primary packaging signal. The ITR is the only cis-acting base sequence required to produce recombinant AAV (rAAV) constructs. The ITR has enhancer activity in the presence of Rep protein, but very weak activity in the absence of Rep protein. When cloning a foreign gene into a recombinant AAV construct, these characteristics are taken into account to appropriately configure enhancers, promoters, pA, etc. to produce an expression construct (RJ Samulski and N Muzyczka, Annu. Rev. Virolo. 2014. 1:427-451). Four proteins are translated from the rep gene. These proteins are classified into rep78, rep68, rep52, and rep40 according to their molecular weights, and perform important functions in AAV DNA replication. Four proteins are translated from the cap gene. Among these, VP1, VP2, and VP3 proteins are structural proteins that constitute AAV particles, and assembly-activating protein (AAP) promotes the formation (assembly) of AAV particles by the structural proteins.Efficient replication of adeno-associated viruses requires certain proteins and RNAs derived from helper viruses such as adenovirus or herpes simplex virus (Muzyczka N. Curr Top Microbiol Immunol 158, 97-129, 1992).

[0451] AAV possesses unique characteristics that make it an attractive vector for delivering foreign genes (transgenes) into cells. AAV infection of cells in culture is generally non-cytopathic, and natural infections in humans and other animals are asymptomatic and asymptomatic. Furthermore, AAV can infect many different mammalian cell types, potentially targeting numerous different tissues in vivo. AAV also possesses additional advantages that make it a particularly attractive viral system for gene transfer, including the ability to elicit a mild immune response compared to other forms of gene transfer and the ability to sustain expression in both dividing and quiescent cells based on non-integrating, episomal vector DNA. Furthermore, AAV can withstand the conditions used to inactivate adenoviruses (56°C to 65°C for several hours), making cryopreservation of rAAV-based vaccines less critical.

[0452] The type of 아데노-연관 virus that can be used in this introduction is AAVrh.10(AAVrh10), AAV-DJ(AAVDJ), AAV-DJ8(AAVDJ8), AAV1, AAV2, AAV2G9, AAV3, AAV3a, AAV3b, AAV3-3, AAV4, AAV4-4, AAV5, AAV6, AAV6.1, AAV6.2, AAV6.1.2, AAV7, AAV7.2, AAV8, AAV9, AAV9.11, AAV9.13, AAV9.16, AAV9.24, AAV9.45, AAV9.47, AAV9.61, AAV9.68, AAV9.84, AAV9.9, AAV10, AAV11, AAV12, AAV16.3, AAV24.1, AAV27.3, AAV42.12, AAV42-1b, AAV42-2, AAV42-3a, AAV42-3b, AAV42-4, AAV42-5a, AAV42-5b, AAV42-6b, AAV42-8, AAV42-10, AAV42-11, AAV42-12, AAV42-13, AAV42-15, AAV42-aa, AAV43-1, AAV43-12, AAV43-20, AAV43-21, AAV43-23, AAV43-25, AAV43-5, AAV44.1, AAV44.2, AAV44.5, AAV223.1, AAV223.2, AAV223.4, AAV223.5, AAV223.6, AAV223.7, AAV1-7 / rh.48, AAV1-8 / rh.49, AAV2-15 / rh.62, AAV2-3 / rh.61, AAV2-4 / rh.50, AAV2-5 / rh.51, AAV3.1 / hu.6, AAV3.1 / rh.9, AAV3-9 / rh.52, AAV3-11 / rh.53, AAV4-8 / r11.64, AAV4-9 / rh.54, AAV4-19 / rh.55, AAV5-3 / rh.57, AAV5-22 / rh.58, AAV7.3 / hu.7, AAV16.8 / hu.10, AAV16.12 / hu.11, AAV29.3 / bb.1, AAV29.5 / bb.2, AAV106.1 / hu.37, AAV114.3 / hu.40, AAV127.2 / hu.41, AAV127.5 / hu.42, AAV128.3 / hu.44, AAV130.4 / hu.48, AAV145.1 / hu.53, AAV145.5 / hu.54, AAV145.6 / hu.55, AAV161.10 / hu.60, AAV161.6 / hu.61, AAV33.12 / hu.17, AAV33.4 / hu.15, AAV33.8 / hu.16, AAV52 / hu.19, AAV52.1 / hu.20, AAV58.2 / hu.25, AAVA3.3, AAVA3.4, AAVA3.5, AAVA3.7, AAVC1, AAVC2, AAVC5, AAVF3, AAVF5, AAVH2, AAVrh.72, AAVhu.8, AAVrh.68, AAVrh.70, AAVpi.1, AAVpi.3, AAVpi.2, AAVrh.60, AAVrh.44, AAVrh.65, AAVrh.55, AAVrh.47, AAVrh.69, AAVrh.45, AAVrh.59, AAVhu.12, AAVH6, AAVH03, AAVH-1 / hu.1, AAVH-5 / hu.3, AAVLG-10 / rh.40, AAVLG-4 / rh.38, AAVLG-9 / hu.39, AAVN721-8 / rh.43, AAVCh.5, AAVCh.5R1, AAVcy.2, AAVcy.3, AAVcy.4, AAVcy.5, AAVCy.5R1, AAVCy.5R2, AAVcy.5R3, AAVCy.5R4, AAVcy.6, AAVhu.1, AAVhu.2, AAVhu.3, AAVhu.4, AAVhu.5, AAVhu.6, AAVhu.7, AAVhu.9, AAVhu.10, AAVhu.11, AAVhu.13, AAVhu.15, AAVhu.16, AAVhu.17, AAVhu.18, AAVhu.20, AAVhu.21, AAVhu.22, AAVhu.23.2, AAVhu.24, AAVhu.25, AAVhu.27, AAVhu.28, AAVhu.29, AAVhu.29R, AAVhu.31, AAVhu.32, AAVhu.34, AAVhu.35, AAVhu.37, AAVhu.39, AAVhu.40, AAVhu.41, AAVhu.42, AAVhu.43, AAVhu.44, AAVhu.44R1, AAVhu.44R2, AAVhu.44R3, AAVhu.45, AAVhu.46, AAVhu.47, AAVhu.48, AAVhu.48R1, AAVhu.48R2, AAVhu.48R3, AAVhu.49, AAVhu.51, AAVhu.52, AAVhu.54, AAVhu.55, AAVhu.56, AAVhu.57, AAVhu.58, AAVhu.60, AAVhu.61, AAVhu.63, AAVhu.64, AAVhu.66, AAVhu.67, AAVhu.14 / 9, AAVhu.t19, AAVrh.2, AAVrh.2R. AAVrh.8, AAVrh.8R, AAVrh.12, AAVrh.13, AAVrh.13R, AAVrh.14, AAVrh.17, AAVrh.18, AAVrh.19, AAVrh.20, AAVrh.21, AAVrh.22, AAVrh.23, AAVrh.24, AAVrh.25, AAVrh.31, AAVrh.32, AAVrh.33, AAVrh.34, AAVrh.35, AAVrh.36, AAVrh.37, AAVrh.37R2, AAVrh.38, AAVrh.39, AAVrh.40, AAVrh.46, AAVrh.48, AAVrh.48.1, AAVrh.48.1.2, AAVrh.48.2, AAVrh.49, AAVrh.51, AAVrh.52, AAVrh.53, AAVrh.54, AAVrh.56, AAVrh.57, AAVrh.58, AAVrh.61, AAVrh.64, AAVrh.64R1, AAVrh.64R2, AAVrh.67, AAVrh.73, AAVrh.74, AAVrh8R. AAVrh8R A586R 변이체, AAVrh8R R533A 변이체, AAAV, BAAV, 염소 AAV, 소 AAV, AAVhE1.1, AAVhEr1.5, AAVhEr1.14, AAVhEr1.14, AAVhEr1.8, AAVhEr1.16, AAVhEr1.18, AAVhEr1.35, AAVhEr1.7, AAVhEr1.36, AAVhEr2.29, AAVhEr2.4, AAVhEr2.16, AAVhEr2.16, AAVhEr2.30, AAVhEr2.31, AAVhEr2.31, AAVhEr2.36, AAVhER1.23, AAVhEr3.1, AAV2.5T, AAV-PAEC, AAV-LK01, AAV-LK02, AAV-LK03, AAV-LK04, AAV-LK05, AAV-LK06, AAV-LK07, AAV-LK08, AAV-LK09, AAV-LK10, AAV-LK11, AAV-LK12, AAV-LK13, AAV-LK14, AAV-LK15, AAV-LK16, AAV-LK17, AAV-LK18, AAV-LK19, AAV-PAEC2, AAV-PAEC4, AAV-PAEC6, AAV-PAEC7, AAV-PAEC8, AAV-PAEC11, AAV-PAEC12, AAV-2-pre-miRNA-101, AAV-8h, AAV-8b, AAV-h, AAV-b, AAV SM 10-2, AAV Shuffle 100-1, AAV Shuffle 100-3, AAV Shuffle 100-7, AAV Shuffle 10-2, AAV Shuffle 10-6, AAV Shuffle 10-8, AAV Shuffle 100-2, AAV SM 10-1, AAV SM 10-8, AAV SM 100-3, AAV SM 100-10, B P61 AAV, B P62 AAV, B P63 AAV, AAVrh.50, AAVrh.43, AAVrh.62, AAVrh.48, AAVhu.19, AAVhu.11, AAVhu.53, AAV4-8 / rh.64, AAVLG-9 / hu.39, AAV54.5 / hu.23, AAV54.2 / hu.22, AAV54.7 / hu.24, AAV54.1 / hu.21, AAV54.4R / hu.27, AAV46.2 / hu.28, AAV46.6 / hu.29, AAV128.1 / hu.43, true type AAV(ttAAV), UPENN AAV 10 and Japanese AAV 10 serotypes, but not limited thereto.

[0453] In some embodiments, the serotype of the adeno-associated virus is AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, or AAVrh10. In some embodiments, the serotype of the AAV is AAV2. In some embodiments, the serotype of the AAV is AAV5. In some embodiments, the serotype of the AAV is AAV8. In some embodiments, the serotype of the AAV is AAV9.

[0454] In some embodiments, for efficient expression or production of the adeno-associated virus, or for other purposes such as targeting, some amino acid sequences of the Rep protein and / or the Cap protein may be mutated, or new amino acid sequences may be added or deleted, and the sequences of the genes encoding them may also be mutated. The mutated AAV is also included in the AAV of the present invention as long as it functions as an AAV.

[0455]

[0456] VI. Cells

[0457] In some embodiments, the present disclosure provides a cell comprising any of the polynucleotides described herein (e.g., a nucleic acid molecule encoding a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142 to 147) or a cell comprising a vector comprising the polynucleotide.

[0458]

[0459] For the nucleic acid molecule, it is as described in III and for the vector, it is as described in IV.

[0460] For example, in some embodiments, the cell is transduced, transfected or transformed using a vector (e.g., an AAV vector) comprising a foreign gene described herein (e.g., a nucleic acid molecule encoding a polypeptide comprising a TAFA polypeptide or any one of the amino acid sequences of SEQ ID NOs: 142 to 147) and a non-translated nucleic acid sequence for expression of the foreign gene.

[0461] Without being bound by any theory, in some embodiments, a cell described herein (e.g., transduced with a polynucleotide comprising a non-translated nucleic acid sequence) is useful for producing a protein, such as one encoded by a foreign gene described herein (e.g., a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOS: 142-147). As described herein, in some embodiments, a non-translated nucleic acid sequence described herein (e.g., an EF-1α intron or fragment thereof) can enhance expression of a protein encoded by the foreign gene (the “encoded protein”) in a cell. Thus, in some embodiments, a cell described herein (e.g., transduced with a polynucleotide comprising a foreign gene and a non-translated nucleic acid sequence of the present disclosure) results in greater expression of the encoded protein compared to a reference cell. In some embodiments, the reference cell is transduced with the polynucleotide but lacks the non-translated nucleic acid sequence.

[0462] In some embodiments, the cells described herein can produce a protein encoded by the foreign gene in vitro. In certain embodiments, the cells described herein can produce the encoded protein in vivo (e.g., in a subject administered a polynucleotide described herein). In some embodiments, the cells described herein can produce the encoded protein both in vitro and in vivo.

[0463] In some embodiments, a cell that can be used to produce a protein encoded by a foreign gene (e.g., in vitro) comprises a host cell. As used herein, the term "host cell" is intended to include any cell of an organism that can be transduced with the expression construct or vector (e.g., an AAV vector) to replicate the expression construct or express the gene encoded by the expression construct. Such cells include eukaryotic cells and prokaryotic cells. As used herein, the term "transduction" is intended to encompass transfection and transformation. The host cell can be transduced, transfected, or transformed with the expression construct. This process refers to the transfer or introduction of an exogenous nucleic acid molecule into the host cell. In some embodiments, the host cell is an isolated host cell comprising the AAV vector. In some embodiments, the host cell is an isolated host cell transformed with the AAV vector.

[0464] In some embodiments, the host cell is a eukaryotic cell. In some embodiments, the host cell is selected from the group consisting of a mammalian cell, an insect cell, a yeast cell, a transgenic mammalian cell, and a plant cell. In some embodiments, the host cell is a prokaryotic cell. In some embodiments, the prokaryotic cell is a bacterial cell.

[0465] In some embodiments, the host cell is an insect cell. In some embodiments, the insect cell is Sf9. In some embodiments, the host cell is a mammalian cell. Non-limiting examples of mammalian cells that can be used in the present disclosure include HEK293, HeLa, ARPE-19, RPE-1, HepG2, Hep3B, Huh-7, C8D1a, Neuro2A, CHO, MES13, BHK-21, COS7, COP5, A549, MCF-7, HC70, HCC1428, BT-549, PC3, LNCaP, Capan-1, Panc-1, MIA PaCa-2, SW480, HCT166, LoVo, A172, MKN-45, MKN-74, Kato-III, NCI-N87, HT-144, SK-MEL-2, SH-SY5Y, C6, HT-22, PC-12, NIH3T3 cells, and combinations thereof.

[0466] In some embodiments, the cells that can be used to produce a protein encoded by a foreign gene described herein (e.g., in vivo) include human cells. In some embodiments, the human cells are cells of a subject to whom a nucleic acid molecule described herein has been administered. In certain embodiments, the human cells are derived from a donor (e.g., a healthy human subject).

[0467] In some aspects, the present disclosure provides a composition comprising an AAV vector or a host cell comprising or transformed with the AAV vector.

[0468]

[0469] VII. Composition

[0470] According to another aspect of the present invention, the present invention provides a composition comprising the polypeptide, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

[0471]

[0472] The polypeptides, nucleic acid molecules, vectors, recombinant viral particles and cells are as described in II to VI.

[0473] In some embodiments, the composition is a pharmaceutical composition.

[0474] In some embodiments, the present disclosure discloses a pharmaceutical composition comprising (a) a polypeptide described herein (e.g., a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142-147) or a nucleic acid molecule encoding the same, and (b) one or more pharmaceutically acceptable carriers. In some embodiments, the present disclosure discloses a pharmaceutical composition comprising (a) a vector (e.g., rAAV) or recombinant viral particle described herein, and (b) one or more pharmaceutically acceptable carriers. In some embodiments, the present disclosure discloses a pharmaceutical composition comprising (a) a cell described herein, and (b) one or more pharmaceutically acceptable carriers.

[0475] In some embodiments, the pharmaceutical compositions described herein comprise a polypeptide consisting of a sequence of 8 to 61 amino acids.

[0476] In some embodiments, the pharmaceutical compositions described herein comprise a polypeptide comprising a sequence having at least 50% sequence identity to the amino acid sequence of SEQ ID NO: 1, SEQ ID NO: 15, or SEQ ID NO: 87.

[0477] In some embodiments, the pharmaceutical composition is a pharmaceutical composition for preventing, improving, or treating a retinal neuropathic disease.

[0478] In some embodiments, the retinal neuropathic disorder is caused by damage to all or part of the retina or macula. In some embodiments, the retinal neuropathic disorder is a disorder caused by dysfunction or damage to cells of the retina or macula. In some embodiments, the retinal neuropathic disorder may be a disorder caused by dysfunction or damage to photoreceptor cells (e.g., rod cell and cone cell layers) and / or retinal pigment epithelial (RPE) cells of the retina or macula. In some embodiments, the retinal neuropathic disorder is retinopathy, choroidal neovascularization, macular degeneration, retinal degeneration, macular edema, retinal edema, macular swelling, retinal swelling, color vision defect, retinal cell degeneration, retinal vascular occlusion, retinal detachment, hereditary retinal disease, or a combination thereof. In some embodiments, the macular degeneration is age-related macular degeneration, Best macular dystrophy, Sorsby fundus dystrophy, Mallatia Leventinese, Doyne honeycomb retinal dystrophy, Stargardt disease (Stargardt macular dystrophy), myopic macular degeneration, or pigment epithelial detachment-associated macular degeneration. In some embodiments, the macular degeneration is age-related macular degeneration. In some embodiments, the age-related macular degeneration is wet or dry age-related macular degeneration. In some embodiments, the retinopathy is retinal dystrophy. In some embodiments, the retinopathy is diabetic retinopathy. In some embodiments, the diabetic retinopathy is nonproliferative diabetic retinopathy (NPDR), proliferative diabetic retinopathy (PDR), diabetic macular edema, or a combination thereof. In some embodiments, the retinal neuropathies are hereditary. It is a retinal disease. In some embodiments, the color vision deficiency is color blindness or color weakness. In some embodiments, the color vision deficiency is deuteranopia, protanopia, cyanopia, deuteranopia, red-weakness, or blue-weakness. In some embodiments, the color vision deficiency is panchromatic achromatopsia, trichromatic achromatopsia, or complete achromatopsia.According to one embodiment, the polypeptide of the present specification or the nucleic acid molecule encoding the same can be usefully utilized for the prevention or treatment of color vision deficiency by reversing or restoring destruction, loss or dysfunction of cone cells, which are one of the photoreceptors.

[0479] In some embodiments, the hereditary retinal disease is Retinitis pigmentosa (RP), Leber congenital amaurosis, Stargardt's disease, Coats retinopathy, Cone dystrophy, Choroideremia, Usher syndrome, Best's Disease, X-linked Retinoschisis, Hereditary color vision deficiency, or Unspecified hereditary retinal dystrophy.

[0480] In some embodiments, the pharmaceutical composition is a pharmaceutical composition for preventing, ameliorating, or treating neuropathic pain.

[0481] In some embodiments, the neuropathic pain is central neuropathic pain, i.e., pain resulting from an injury or damage that affects any level of the central nervous system (e.g., brain injury and spinal cord injury), including the central somatosensory nervous system, or pain caused by or associated with a disease or disorder such as seizures, multiple sclerosis, or lateral medullary infarction. In some embodiments, the central neuropathic pain may be spontaneous or induced by a stimulus. In some embodiments, the central neuropathic pain may include mechanical allodynia and cold allodynia. Symptoms of central neuropathic pain commonly include, for example, burning, aching, shooting pain, pressure, painful coldness, paresthesia, and dysesthesia (e.g., tingling, pins and needles, coldness, and pressure). The distribution of central neuropathic pain can range from small to large areas in the peripheral region, for example, or in spinal cord injury, areas that cover half the body, include one side of the face, or include the opposite side of the body or limbs. Central neuropathic pain due to spinal cord injury includes "at-level pain," which is perceived in a segmented pattern at the site of injury, and "below-level pain," which is felt below the site of injury. In some embodiments, the method reduces, reverses, alleviates, improves, suppresses, or dulls or prevents central neuropathic pain, symptoms associated with the pain, the underlying cause of the pain, or a combination thereof.

[0482] In some embodiments, neuropathic pain is peripheral neuropathic pain, pain caused by damage or injury to any part of the peripheral nervous system (e.g., damage to motor, sensory, autonomic, or a combination thereof), or pain caused by or associated with a disease or disorder. Damage or injury to motor nerves is associated with symptoms such as muscle weakness (e.g., weakness of the muscles of the back, legs, hips, or face), painful cramps and fasciculations (uncontrolled muscle spasms beneath the skin), muscle atrophy (severe shrinkage of muscle size), and decreased reflexes. Damage or injury to sensory nerves can cause a variety of symptoms, including hypersensitivity of pain and pain receptors in the skin, resulting in allodynia (e.g., severe pain to normally non-painful stimuli).

[0483] In some embodiments, the neuropathic pain that can be treated with the compositions of the present invention is neuralgia, including but not limited to: trigeminal neuralgia (TN) (e.g., pain within the facial or oral trigeminal nerve territory), atypical trigeminal neuralgia (ATN), occipital neuralgia, post-herpetic neuralgia (e.g., unilateral pain distributed over one or more spinal cutaneous or ophthalmic divisions of the trigeminal nerve), peripheral nerve injury pain (e.g., pain in the innervation territory of an injured nerve, typically distal to trauma, surgery, or compression), glossopharyngeal neuralgia (e.g., irritation of the ninth cranial nerve causing severe pain in the neck, tongue, and back of the ear), sciatica, low back pain, and atypical facial pain. In some embodiments, the neuralgia is caused by or associated with chemical irritation, inflammation, trauma (including surgery), compression of the nerve by a nearby structure (e.g., a tumor), or infection. In some embodiments, the neuropathic pain is afferent pain syndrome, including but not limited to: brain or spinal cord injury, pain after stroke, phantom pain, paraplegia, brachial plexus infarction injury, and lumbar radiculopathy. In some embodiments, the neuropathic pain is complex regional pain syndrome (CRPS), including CRPS1 and CRPS2. CRPS includes, but is not limited to. In some embodiments, symptoms associated with CRPS may include severe pain; changes in nails, bones, and skin; and increased sensation to touch in the affected limb. In some embodiments, the neuropathic pain is a neuropathy (e.g., central or peripheral). Non-limiting examples of neuropathic pain include, for example, single neuropathic pain (mononeuropathy) and polyneuropathic pain (polyneuropathy). In some embodiments, the neuropathic pain is diabetic peripheral neuropathy.

[0484] In some embodiments, the compositions of the present invention prevent, improve, or treat hyperalgesia. As used herein, the term "hyperalgesia" refers to an increased or heightened response to painful stimuli (e.g., a pinprick or a hot plate). In some embodiments, the hyperalgesia is directed to mechanical stimuli, such as a pinprick (mechanical hyperalgesia). In other embodiments, the hyperalgesia is directed to thermal stimuli, such as a hot plate (thermal hyperalgesia).

[0485] Pharmaceutically acceptable carriers that can be used in the present disclosure are those commonly used in formulations. Examples of such pharmaceutically acceptable carriers include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum acacia, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. The pharmaceutical compositions of the present disclosure may further comprise one or more additives selected from the group consisting of lubricants, wetting agents, sweetening agents, flavoring agents, emulsifying agents, suspending agents, and preservatives. Details of suitable pharmaceutically acceptable carriers and formulations can be found in Remington's Pharmaceutical Sciences (19th ed., 1995).

[0486] The pharmaceutical composition of the present disclosure is formulated to be compatible with its intended route of administration. Examples of suitable parenteral routes of administration include intravenous infusion, percutaneous administration, subcutaneous infusion, intramuscular infusion, intraocular (e.g., any administration capable of delivering to sub-Tenon, subconjunctival, suprachoroidal, suprachoroidal, subretinal, intravitreal, and similar locations), eye drops, intraventricular infusion, intrathecal infusion, intraamniotic infusion, intraarterial infusion, intraarterial infusion, intracardiac infusion, intracavernosal infusion, intracerebral infusion, cistern infusion, intracoronary infusion, intracranial infusion, intrathecal infusion, epidural infusion, intrahippocampal infusion, intranasal infusion, intraosseous infusion, intraperitoneal infusion, intrathoracic infusion, intraspinal infusion, intrathoracic infusion, intrathymic infusion, intrauterine infusion, intravaginal infusion, intraventricular infusion, intravesical infusion, subconjunctival infusion, intratumoral infusion, topical infusion, intraperitoneal infusion, and combinations thereof. In some aspects, intraocular administration includes suprachoroidal, subretinal, or intravitreal administration. In some aspects, topical injection includes ophthalmic, nasal, transdermal, oral, or rectal administration. The amino acid sequence of the polypeptide disclosed in the present invention (e.g., a TAFA protein fragment or variant thereof) may be comprised of 8 to 61 amino acids, preferably 8 to 30 amino acids, so that the desired preventive or therapeutic effect can be achieved through ophthalmic or nasal administration.

[0487] In some embodiments, the pharmaceutical composition is administered at a daily dosage of 0.0001 to 100 mg / kg.

[0488] The pharmaceutical compositions of the present disclosure may be formulated with one or more pharmaceutically acceptable carriers and / or excipients. The pharmaceutical compositions may be presented in unit dosage form or dispensed into multi-dose containers. The formulations may be in the form of solutions, suspensions, or emulsions in oily or aqueous media, or in the form of extracts, powders, granules, tablets, or capsules. The formulations may further include dispersing agents or stabilizers.

[0489]

[0490] VIII. Kit

[0491] The present disclosure also discloses kits comprising one or more polypeptides disclosed herein (e.g., a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142-147), one or more polynucleotides (e.g., including foreign genes and non-translated nucleic acid sequences), one or more vectors disclosed herein (e.g., AAV vectors), one or more cells disclosed herein (e.g., host cells comprising or transformed with said AAV vectors), any composition disclosed herein, or any combination thereof. In some embodiments, the kit comprises instructions for use.

[0492] The terms "kit" and "system" as used herein are intended to refer to at least one polynucleotide disclosed herein, one or more vectors disclosed herein (e.g., AAV vectors), one or more host cells disclosed herein, any pharmaceutical composition disclosed herein, or any combination thereof, in some embodiments in combination with one or more other types of elements or components (e.g., other types of biochemical reagents, containers, packaging such as commercial packaging, instructions for use, etc.).

[0493]

[0494] IX. Uses and Methods

[0495] IX.A. Production Method

[0496] The present disclosure also provides a method for producing a composition, comprising the step of producing a composition comprising the polypeptide (e.g., a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142 to 147), or a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

[0497] In some embodiments, the method comprises synthesizing a TAFA protein fragment or variant thereof described herein under suitable conditions.

[0498] In some embodiments, the methods comprise culturing a cell described herein (e.g., transduced with a polynucleotide comprising a foreign gene and a non-translated nucleic acid molecule) under suitable conditions and recovering the encoded protein. In certain embodiments, a method for producing a polypeptide encoded by a foreign gene comprises administering a polynucleotide of the present disclosure (e.g., comprising a foreign gene and a non-translated nucleic acid molecule) to a subject in need thereof, thereby producing the encoded polypeptide in the subject. Additional disclosures regarding such in vivo methods for producing polypeptides are provided elsewhere in this disclosure (see, e.g., Therapeutic Uses).

[0499] In some embodiments, the present disclosure provides methods for producing recombinant adeno-associated viral particles comprising a polynucleotide described herein (e.g., including a foreign gene and a non-translated nucleic acid sequence). In some embodiments, the method for producing such recombinant AAV comprises culturing a cell transfected with an AAV vector described herein under conditions conducive to the production of recombinant AAV. In some embodiments, the method further comprises isolating the produced recombinant viral particles.

[0500] In some aspects, the present disclosure provides recombinant viral particles produced by the method.

[0501] In some embodiments, the recombinant viral particle can be produced by transducing a cell with (i) an AAV vector comprising the foreign gene (e.g., a nucleic acid molecule encoding a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142 to 147) and (ii) a construct containing rep and cap genes. Additionally, (iii) a helper construct can be used to transduce the foreign gene into the host cell. In such embodiments, the helper construct can contain an E2A gene that promotes AAV genome replication and gene transcription, an E4 gene that allows AAV mRNA to export from the nucleus to the cytoplasm, and a VA region that generates two VA RNAs that serve to regulate translation.

[0502] In some embodiments, the three constructs described above may be replaced by two constructs for transduction of host cells. In such embodiments, the AAV construct comprises the foreign genes and non-translated nucleic acid sequences, and a separate construct comprises the rep and cap genes, the E2A gene, the E4 gene, and the VA region. Additional methods for producing AAV particles described herein are generally known in the art. See, for example, Clement et al., Mol Ther Methods Clin Dev 3: 16002 (March 2016); Clark, Kidney Int. 61: S9-15 (January 2002); and Xiao et al., J Virol 72(3): 2224-32 (March 1998), each of which is incorporated herein by reference in its entirety.

[0503] The present specification also discloses a recombinant viral particle comprising (a) a capsid protein and (b) the AAV vector.

[0504]

[0505] IX.B. Therapeutic Uses

[0506] The polypeptides described herein (e.g., a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOS: 142-147), nucleic acids (e.g., including foreign genes and non-translated nucleic acid sequences), vectors and recombinant viruses (e.g., rAAV) carrying such nucleic acids, cells comprising such nucleic acids or vectors, and the methods described herein have many in vitro and in vivo utilities. For example, the polypeptides, polynucleotides, vectors, e.g., AAV vectors, described herein can be administered to cells in culture, in vitro, or ex vivo, or to a human subject, e.g., in vivo, to prevent or treat disease. Thus, in some aspects, the present disclosure provides therapeutic uses of any of the polypeptides, polynucleotides (e.g., including foreign genes and non-translated nucleic acid sequences) described herein, a recombinant expression construct or vector described herein, a cell described herein, a pharmaceutical composition described herein, or a recombinant virus described herein. In some embodiments, the present disclosure discloses a method of expressing a foreign gene in a subject in need thereof, comprising administering to the subject a polynucleotide disclosed herein (e.g., comprising a foreign gene and a non-translated nucleic acid sequence), a vector disclosed herein, a recombinant virus disclosed herein (e.g., rAAV), a cell disclosed herein, or a pharmaceutical composition disclosed herein, wherein expression of the foreign gene in the subject is increased after said administration.

[0507] As described herein, the untranslated nucleic acid sequences of the present disclosure can increase expression of a foreign gene when the foreign gene is translated. Accordingly, in some embodiments, the present disclosure relates to a method of increasing expression of a foreign gene in a cell, comprising contacting the cell with any of the polynucleotides disclosed herein (e.g., a nucleic acid molecule encoding a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142-147), vectors, or recombinant viruses (e.g., rAAV). The contacting can occur ex vivo or in vivo. When the contacting occurs in vivo, the method can further comprise administering to the subject any of the polynucleotides, vectors, or recombinant viruses prior to the contacting.

[0508] In some embodiments, expression of the foreign gene (e.g., a nucleic acid molecule encoding a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142-147) after contacting is increased by at least about 1 fold, at least about 1.1 fold, at least about 1.2 fold, at least about 1.3 fold, at least about 1.4 fold, at least about 1.5 fold, at least about 1.6 fold, at least about 1.7 fold, at least about 1.8 fold, at least about 1.9 fold, at least about 2 fold, at least about 2.5 fold, at least about 3 fold, at least about 3.5 fold, at least about 4 fold, at least about 5 fold, at least about 6 fold, at least about 7 fold, at least about 8 fold, at least about 9 fold, or at least about 10 fold or more relative to baseline expression. In some embodiments, the baseline expression is expression of the foreign gene in the cell prior to contacting. In some embodiments, the reference expression is expression of a foreign gene within the cell that has not been in contact with a polypeptide, polynucleotide, vector, or recombinant virus described herein.

[0509] Another aspect of the present disclosure provides a method for preventing or treating a disease in a subject in need thereof, comprising administering to the subject an effective amount of any of the polypeptides (e.g., a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142-147), the polynucleotides (e.g., a nucleic acid molecule encoding a TAFA polypeptide or a polypeptide comprising any one of the amino acid sequences of SEQ ID NOs: 142-147), the vectors, the cells, the recombinant viruses, or the pharmaceutical compositions. As will be apparent from the present disclosure, the compositions (e.g., polypeptides, polynucleotides, recombinant expression constructs, cells, pharmaceutical compositions, or recombinant viruses) described herein can be used to prevent or treat any disease in question.

[0510] In some embodiments, the method may further comprise administering to the subject an additional therapeutic agent (e.g., an inhibitor of vascular endothelial growth factor ("VEGF") or an agent for neuropathic pain). In some embodiments, the additional therapeutic agent may be administered to the subject concurrently with, prior to, or subsequent to the administration of the polypeptide, polynucleotide, vector, cell, recombinant virus, or pharmaceutical composition.

[0511] The diseases that can be prevented, improved, or treated by the present disclosure are not limited and include all diseases that require a reduction in the number of drug administrations. Non-limiting examples of such diseases include retinal neuropathies. In some embodiments, the retinal neuropathies are selected from retinopathy, choroidal neovascularization, macular disease, macular degeneration, retinal degeneration, macular edema, retinal edema, macular swelling, retinal swelling, color vision abnormalities, retinal cell degeneration, retinal vascular occlusion, retinal detachment, hereditary retinal diseases, and combinations thereof.

[0512] In some embodiments, the retinal neuropathies preventable or treatable by the present disclosure include macular degeneration. In some embodiments, the macular degeneration includes age-related macular degeneration (AMD). Age-related macular degeneration can be divided into dry (atrophic) macular degeneration and wet (neovascular or exudative) macular degeneration. Age-related macular degeneration can also be divided into early AMD, intermediate AMD, and late or advanced AMD (geographic atrophy). In some embodiments, the retinal neuropathies preventable or treatable by the present disclosure include diabetic retinopathy. In some embodiments, the diabetic retinopathy is non-proliferative diabetic retinopathy (NPDR). In some embodiments, the diabetic retinopathy is proliferative diabetic retinopathy (PDR). In some embodiments, the diabetic retinopathy is diabetic macular edema. In some embodiments, diabetic retinopathy is any retinopathy associated with ischemic damage within the retina. Unless otherwise specified, the present disclosure may be used to prevent or treat all forms of AMD and / or diabetic retinopathy.

[0513] Additional, non-limiting examples of the above conditions include neuropathic pain.

[0514] In some embodiments, the neuropathic pain is central neuropathic pain, i.e., pain resulting from an injury or damage that affects any level of the central nervous system (e.g., brain injury and spinal cord injury), including the central somatosensory nervous system, or pain caused by or associated with a disease or disorder such as seizures, multiple sclerosis, or lateral medullary infarction. In some embodiments, the central neuropathic pain may be spontaneous or induced by a stimulus. In some embodiments, the central neuropathic pain may include mechanical allodynia and cold allodynia. Symptoms of central neuropathic pain commonly include, for example, burning, aching, shooting pain, pressure, painful coldness, paresthesia, and dysesthesia (e.g., tingling, pins and needles, coldness, and pressure). The distribution of central neuropathic pain can range from small to large areas in the peripheral region, for example, or in spinal cord injury, areas that cover half the body, include one side of the face, or include the opposite side of the body or limbs. Central neuropathic pain due to spinal cord injury includes "at-level pain," which is perceived in a segmented pattern at the site of injury, and "below-level pain," which is felt below the site of injury. In some embodiments, the method reduces, reverses, alleviates, improves, suppresses, or dulls or prevents central neuropathic pain, symptoms associated with the pain, the underlying cause of the pain, or a combination thereof.

[0515] In some embodiments, neuropathic pain is peripheral neuropathic pain, pain caused by damage or injury to any part of the peripheral nervous system (e.g., damage to motor, sensory, autonomic, or a combination thereof), or pain caused by or associated with a disease or disorder. Damage or injury to motor nerves is associated with symptoms such as muscle weakness (e.g., weakness of the muscles of the back, legs, hips, or face), painful cramps and fasciculations (uncontrolled muscle spasms beneath the skin), muscle atrophy (severe shrinkage of muscle size), and decreased reflexes. Damage or injury to sensory nerves can cause a variety of symptoms, including hypersensitivity of pain and pain receptors in the skin, resulting in allodynia (e.g., severe pain to normally non-painful stimuli).

[0516] In some embodiments, the method of the present invention treats one or more types of neuropathic pain, comprising administering to a subject in need thereof the polypeptide, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof. In some embodiments, the neuropathic pain that can be treated by the methods of the present invention is neuralgia, which includes, but is not limited to: trigeminal neuralgia (TN) (e.g., pain within the facial or oral trigeminal nerve territory), atypical trigeminal neuralgia (ATN), occipital neuralgia, post-herpetic neuralgia (e.g., unilateral pain distributed over one or more spinal cutaneous or ophthalmic divisions of the trigeminal nerve), peripheral nerve injury pain (e.g., pain in the innervation territory of an injured nerve, typically distal to trauma, surgery, or compression), glossopharyngeal neuralgia (e.g., irritation of the ninth cranial nerve causing severe pain in the neck, tongue, and back of the ear), sciatica, low back pain, and atypical facial pain. In some embodiments, the neuralgia is caused by or associated with chemical irritation, inflammation, trauma (including surgery), compression of the nerve by a nearby structure (e.g., a tumor), or infection. In some embodiments, the neuropathic pain is a central pain syndrome, including but not limited to: brain or spinal cord injury, post-stroke pain, phantom pain, paraplegia, brachial plexus infarction injury, and lumbar radiculopathy. In some embodiments, the neuropathic pain is complex regional pain syndrome (CRPS), including CRPS1 and CRPS2. CRPS includes, but is not limited to. In some embodiments, symptoms associated with CRPS may include severe pain, changes in nails, bones, and skin; and increased sensation to touch in the affected limb. In some embodiments, the neuropathic pain is a neuropathy (e.g., central or peripheral).Non-limiting examples of neuropathic pain include, for example, single nerve pathological pain (mononeuropathy) and polyneuropathy (polyneuropathy).

[0517] In some embodiments, the neuropathic pain results from or is associated with physical damage, including, for example, (1) traumatic injury or injury, including nerve compression (e.g., nerve crush, nerve stretching, nerve entrapment, or incomplete nerve transection); (2) spinal cord injury (e.g., hemisection of the spinal cord); (3) injury or damage to a peripheral nerve (e.g., motor, sensory, or autonomic nerve, or a combination thereof); (4) limb amputation; contusion; inflammation (e.g., inflammation of the spinal cord); or surgical procedure; and (5) repetitive stress, for example, repetitive, slow, and / or forceful activity requiring movement of a group of joints for a prolonged period of time (e.g., ulnar neuropathy and carpal tunnel syndrome). In some embodiments, the method treats neuropathic pain resulting from or associated with exposure to a toxic agent.

[0518] In some embodiments, the neuropathic pain arises from or is associated with one or more diseases or disorders, such as, for example: (1) an ischemic event (e.g., a stroke or heart attack), (2) multiple sclerosis, (3) a metabolic and / or endocrine disease or disorder (e.g., diabetes, metabolic disease, and hypertrophy, a condition caused by excessive production of growth hormone and characterized by abnormal enlargement of skeletal parts, including joints, resulting in nerve entrapment and pain), (4) a disease of the small blood vessels that reduces oxygen supply to peripheral nerves and causes damage to nerve tissue (e.g., vasculitis, i.e., inflammation of the blood vessels), (5) an autoimmune disease (e.g., Sjogren's syndrome, lupus, rheumatoid arthritis, and acute inflammatory demyelinating neuropathy, also known as Guillain-Barré syndrome), (6) a kidney disorder, (7) cancer or tumors (e.g., a neoplastic tumor, a neuroma, a paraneoplastic syndrome, and toxicity from chemotherapy agents and radiation for cancer treatment), (8) an infection (e.g., herpes). (9) inflammatory disorders, (10) peripheral nerve disorders (e.g., neuromas), (11) genetic or de novo genetic disorders (e.g., Charcot-Marie-Tooth disorder), (12) mononeuropathy, (13) polyneuropathy, or a combination thereof. In some embodiments, the neuropathic pain is caused by or associated with diabetes (type I or type II). In some embodiments, the neuropathic pain is diabetic peripheral neuropathy.

[0519] In some embodiments, the neuropathic pain results from or is associated with exposure to an infectious agent, including, for example, a tick infestation, herpes varicella-zoster, Epstein-Barr virus, West Nile virus, cytomegalovirus, herpes simplex virus, AIDS, or a toxic agent (e.g., drugs, alcohol, heavy metals (e.g., lead, arsenic, mercury), or industrial substances (e.g., fumes from solvents, adhesives), and nitrous oxide).

[0520] In some embodiments, the neuropathic pain is caused by or associated with physical injury, infection, diabetes, cancer therapy, alcoholism, amputation, multiple sclerosis, shingles, spinal surgery, sciatica (pain along the sciatic nerve), low back pain, neuralgia such as trigeminal neuralgia (e.g., pain in the facial or oral trigeminal region), neuropathic pain such as painful polyneuropathy (e.g., pain in the foot that may extend to include the lower extremities, thigh, and hand), or a combination thereof. In some embodiments, the neuropathic pain is trigeminal neuralgia. In some embodiments, the neuropathic pain is associated with muscle weakness in the back, legs, hips, or face. In some embodiments, the neuropathic pain is caused by compression of a nerve, such as a nerve in the leg, foot, hip, or facial nerve. In some embodiments, the neuropathic pain includes damage to the sciatic nerve. In some embodiments, the neuropathic pain is sciatica.

[0521] In some embodiments, the methods of the present invention can reverse, alleviate, improve, suppress, slow, or prevent one or more symptoms associated with neuropathic pain. Accordingly, in one aspect, the present invention provides a method for improving hyperalgesia, comprising administering to a subject in need thereof the polypeptide, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof. As used herein, the term "hyperalgesia" refers to an increased or heightened response to a painful stimulus (e.g., a pinprick or a hot plate). In some embodiments, the hyperalgesia relates to a pinprick (mechanical hyperalgesia). In other embodiments, the hyperalgesia relates to a thermal stimulus, such as a hot plate (thermal hyperalgesia). In some embodiments, the subject in need thereof has a chronic constriction injury (e.g., sciatica). In some embodiments, the subject in need thereof has diabetic peripheral neuropathy.

[0522] In some embodiments, administering to a subject in need thereof (e.g., a subject with neuropathic pain who has not been administered the polypeptide (e.g., a polypeptide comprising an imanoic acid sequence of a TAFA protein, a fragment of the TAFA protein, or a variant thereof), a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof, can result in the subject having a higher threshold to mechanical stimulation compared to a control group. The term "threshold to mechanical stimulation" herein refers to the amount of pressure (from a mechanical stimulus) before the subject responds to the stimulus (e.g., a pulling stimulus). Thus, a subject having a higher threshold can tolerate or resist a greater amount of mechanical stimulation compared to a subject having a lower threshold. In some embodiments, the methods of the present invention can increase a subject's threshold to mechanical stimulation by at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 150%, or at least 200% compared to a reference control (e.g., the subject's threshold prior to administration).

[0523] In some embodiments, administering to a subject in need thereof the polypeptide, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof increases the latency (i.e., the time interval between the stimulus and the response) to a thermal stimulus (e.g., a hot plate) of the subject compared to a reference control (e.g., a neuropathic pain subject who has not received the treatment). In some embodiments, the methods of the present invention can increase the latency to a thermal stimulus of the subject by at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 150%, or at least 200% compared to a reference control (e.g., a subject's threshold prior to administration).

[0524] In another aspect, the present invention provides a method for improving sensory nerve conduction velocity in a subject in need thereof. As used herein, the term "sensory nerve conduction velocity" (SNCV) refers to the speed at which electrical signals travel through a peripheral nerve. Healthy nerves transmit faster and stronger electrical signals than damaged nerves. See Chouhan S., J Clin Diagn Resl0(l): CC0l-3 (2016). Therefore, tests that aid in measuring SNCV (e.g., sensory nerve conduction velocity testing) may be useful in identifying potential nerve damage and / or dysfunction in a subject. In some embodiments, the methods of the present invention can increase the SNCV of a neuropathic pain subject by at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 150%, or at least 200% compared to a reference control (e.g., the subject's threshold prior to administration).

[0525] Animal models exist for studying neuropathic pain. Non-limiting examples of these animal models include: (1) the spinal nerve ligation (SNL) model, in which one or more spinal nerves leading to the foot are ligated and transected (see Kim SH and Chung JM., Pain 50:355-363 (1992)); (2) the partial sciatic ligation (PSL) model, in which a portion of the sciatic nerve is tightly ligated (see Seltzer et al., Pain 43:205-218 (1990)); (3) the chronic constriction injury (CCI) model, in which four loose chromic-gut ligatures are placed on the sciatic nerve, and an immune response to the sutures induces nerve swelling and nerve contraction; (4) the spared nerve injury (SNI) model, in which the common ganglion and tibial nerve are transected, sparing the sural nerve (see Deosterd I. and Woolf CJ, Pain 87: 149-158 (2000)); and (5) STZ-induced diabetic rats, in which STZ injection induces experimental diabetes in rats by inducing pancreatic edema and degeneration of islet beta cells (see, e.g., Akbarzadeh A. et al., Indian. J. Clin. Biochem. 22 (2): 60-64 (2007)). These models induce hyperalgesia in animals, which is manifested by an enhanced response to mechanical and / or thermal stimuli.

[0526] Mechanical hyperalgesia testing in animals includes the von Frey test, in which multiple von Frey monofilaments of varying bending forces are applied to the plantar surface. Paw withdrawal thresholds decrease dramatically after nerve injury. Li et al., Pain 85: 493-502 (2000); Therefore, in some embodiments, the methods of the present invention can increase paw withdrawal thresholds in animal models of neuropathic pain (e.g., a chronic constriction injury model).

[0527] Tests for thermal hyperalgesia can involve using a radiant heat source (e.g., a hot plate) focused on the plantar surface of the paw, and the reaction time to paw withdrawal is measured. After nerve injury, paw withdrawal is faster than before the injury. Kim SH and Chung JM. Pain 50: 355-363 (1992); In some embodiments, the methods of the present invention can increase paw withdrawal latency in animal models of neuropathic pain (e.g., diabetic peripheral neuropathy models).

[0528] Another aspect of the present disclosure provides a gene therapy or method for preventing or treating a disease that can achieve sustained foreign gene expression.

[0529] The viral delivery system described herein allows for administration of the compositions described herein at intervals of about 1 week, about 2 weeks, about 3 weeks, about 1 month, about 2 months, about 3 months, about 4 months, about 5 months, about 6 months, about 7 months, about 8 months, about 9 months, about 10 months, about 11 months, about 1 year, about 2 years, about 3 years, about 4 years, about 5 years, or about 10 years or more. In some embodiments, the interval is about 2 to about 3 months. In some embodiments, the interval is about 6 months. In some embodiments, the interval is about 1 year. In some embodiments, the interval is at least about 1 year. In some embodiments, the interval is at least about 2 years. In some embodiments, the interval is at least about 3 years. In some embodiments, the interval is at least about 4 years. In some embodiments, the interval is at least about 5 years. In some embodiments, the interval is at least about 10 years. That is, the use of the viral delivery system described herein can dramatically reduce the number of administrations of the composition, thereby avoiding the inconvenience caused by repeated administration of the composition to the physician, patient, or subject. Depending on the patient's symptoms or needs, the composition may be administered at least 2 to 3 times at 1 to 2 week intervals initially, and then once every 2 to 3 months, every 6 months, every year or more, or every 2 to 10 years or more.

[0530]

[0531] The features and advantages of the present invention are summarized as follows:

[0532] (i) The present invention provides a polypeptide (e.g., a polypeptide comprising an imanoic acid sequence of a TAFA protein, a TAFA protein fragment or a variant thereof) having the ability to increase the length or branching point of a neurite.

[0533] (ii) In addition, the present invention provides a therapeutic use of a pharmaceutical composition comprising the polypeptide, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

[0534] (iii) The composition of the present invention can be usefully used for the prevention or treatment of retinal nerve disease and neuropathic pain by restoring damaged nerves.

[0535]

[0536] Figure 1a shows the results of analyzing changes in neurite length by full-length TAFA4 protein in ganglion cells differentiated from the mouse dorsal root ganglion progenitor cell line MED17.11, and Figure 1b shows the results of analyzing changes in branch points by full-length TAFA4 protein in the ganglion cells. * indicates p value <0.05.

[0537] Figure 2 shows the results of fluorescence angiography (FA) analysis of the improvement in retinal damage when the control group (control AAV8) or experimental group (AAV8.mouse TAFA4) was administered subretinally to the NaIO3-induced retinal damage model. OD (oculus dexter) indicates the right eye, and OS (oculus sinister) indicates the left eye.

[0538] Figure 3 shows the results of measuring the change in the A-wave and B-wave amplitude of electroretinogram (ERG) between the control group (control AAV8) and the experimental group (AAV8.mouse TAFA4).

[0539] Figure 4 shows the results of comparing the interspecies sequence identity of full-length TAFA4 proteins.

[0540] Figure 5 shows the results of comparing the interspecies sequence identity of mature TAFA4 proteins.

[0541] Figure 6 shows the results of analyzing the improvement in retinal damage by subretinal administration of the control group (control AAV8) or experimental group (AAV8.human TAFA4, AAV8.gecko TAFA4, and AAV8.fish TAFA4) in the NaIO3-induced retinal damage model using fluorescence angiography (FA). OD (oculus dexter) indicates the right eye, and OS (oculus sinister) indicates the left eye.

[0542] Figure 7 shows the results of measuring the change in the A-wave and B-wave amplitude of electroretinogram (ERG) between the control group (control AAV8) and the experimental group (AAV8.human TAFA4, AAV8.gecko TAFA4, AAV8.fish TAFA4).

[0543] Figure 8 shows the results of analyzing the improvement in retinal damage by subretinal administration of the control group (control AAV8) or experimental group (AAV8.TAFA1, AAV8.TAFA2, AAV8.TAFA3, and AAV8.TAFA4) in the NaIO3-induced retinal damage model using fluorescence angiography (FA). OD (oculus dexter) indicates the right eye, and OS (oculus sinister) indicates the left eye.

[0544] Figure 9 shows the results of measuring the change in the A-wave and B-wave amplitude of electroretinogram (ERG) between the control group (control AAV8) and the experimental group (AAV8.TAFA1, AAV8.TAFA2, AAV8.TAFA3, and AAV8.TAFA4).

[0545] Figure 10 shows the results of comparing the interspecies amino acid sequence identity of full-length TAFA1, TAFA2, TAFA3, and TAFA4. Figure 10a shows a comparison of amino acid sequences 1 to 70, Figure 10b shows a comparison of amino acid sequences 71 to 140, Figure 10c shows a comparison of amino acid sequences 141 to 210, and Figure 10d shows a comparison of amino acid sequences 211 to 244.

[0546] Figure 11a shows the results of analyzing the change in neurite length by TAFA4 peptide fragments (TAFA4 fragments 1-3, F1-3) in ganglion cells differentiated from the MED17.11 cell line, and Figure 11b shows the results of analyzing the change in branch points by TAFA4 peptide fragments (F1-F3) in the above cells.

[0547] Figure 12 shows the sequence of a peptide of TAFA4 (TAFA protein and fragments thereof (F1-F8)) according to an embodiment of the present invention.

[0548] Figure 13a shows the results of analyzing the change in neurite length by TAFA4 peptide fragment (TAFA4 fragment 4-8, F4-F8) in ganglion cells differentiated from the MED17.11 cell line, and Figure 13b shows the results of analyzing the change in branch point by TAFA4 peptide fragment (F4-F8) in the above cells. ** indicates p value <0.01.

[0549] Figure 14 shows the results of analyzing the pain relief effects of TAFA4 and TAFA4 fragments 1, 2, 3, 5, 6, and 7 (F1, F2, F3, F5, F6, and F7) in a neuropathic pain model.

[0550]

[0551] Hereinafter, the present invention will be described in more detail through examples. These examples are intended solely to illustrate the present invention more specifically, and it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples, in accordance with the gist of the present invention.

[0552]

[0553] Example

[0554] [Example 1] Evaluation of the efficacy of TAFA4 on nerve cells

[0555] To evaluate the biological efficacy of TAFA4 on neurons, the mouse dorsal root ganglion progenitor cell line MED17.11 was differentiated into neurons, and changes in neurite length or branch point caused by recombinant human TAFA4 (hereinafter referred to as rhTAFA4) were evaluated.

[0556] MED17.11 cells were cultured in a 33°C incubator (5% CO2) using DMEM / F-12 culture medium (Gibco) supplemented with 10% fetal bovine serum (Gibco), 1% penicillin / streptomycin (Gibco), 5 ng / mL interferon-gamma (R&D systems), and 0.5% chicken embryonic extract (Sera lab) before differentiation. The method for evaluating the efficacy of TAFA4 is as follows. MED17.11 cells before differentiation were harvested using 0.25% trypsin-EDTA (Gibco). The harvested cells were centrifuged, the supernatant was removed, and the cell pellet was resuspended in differentiation induction medium. The differentiation induction medium used was DMEM / F-12 culture medium supplemented with 10% fetal bovine serum (Gibco), 1% penicillin / streptomycin (Gibco), 10 ng / mL Fibroblast growth factor 2 (R&D systems), 0.5 mM dibutyryl cAMP (Sigma), 25 μM Forskolin (Sigma), 5 μg / mL Y-27632 (Chemdea), 100 ng / mL beta Nerve growth factor (R&D systems), and 10 ng / mL Glial derived neurotrophic factor (R&D systems). MED17.11 cells resuspended in the differentiation induction medium were seeded at 1 X 10 per well in a 96-well cell culture plate (Thermofisher). 4Cells / mL were dispensed in 100 μL. The experimental group was treated with 1 μM rhTAFA4 (R&D systems, Table 1), and the control group (CTL) was treated with the same volume of phosphate-buffered saline (Gibco), and cultured in a 37°C incubator (5% CO2). During culture, cells were photographed using Incucyte (Satorius), and the neurite length and branch points in MED17.11 cells were analyzed using the Incucyte program. A total of three replicate experiments were performed, and statistical significance was confirmed using the Student's test.

[0557] Amino acid sequence of recombinant human TAFA4 peptidePeptide nameLength (aa)Amino acid sequence (Sequence, N'→C')Recombinanthuman TAFA4 (SEQ ID NO: 86)105SQHLRGHAGH HQIKQGTCEV VAVHRCCNKN RIEERSQTVK CSCFPGQVAG TTRAQPSCVE ASIVIQKWWC HMNPCLEGED CKVLPDYSGW SCSSGNKVKT TKVTR

[0558] At 4 days after treatment, it was confirmed that the neurite length and branch points of MED17.11 cells were statistically significantly increased in the rhTAFA4-treated group compared to the control group (Fig. 1a and Fig. 1b).

[0559]

[0560] [Example 2] Evaluation of the efficacy of TAFA in a retinal nerve damage model

[0561] 2-A. Production of AAV carrying TAFA1 to 4 and their interspecies variant genes

[0562] Recombinant AAV containing a transgene was prepared according to the method described in Korean Patent Application No. 10-2023-0068976, and genes encoding TAFA1 to 4 and their interspecies variants were inserted into the transgene.

[0563]

[0564] 2-B. Evaluation of the efficacy of TAFA4 in a sodium iodate (NaIO3)-induced retinal nerve damage model.

[0565] NaIO3 (sodium iodate) has been reported to induce direct damage to neurons in the retina and reduce the area of ​​neurites and retinal ganglion cells (Zui Tao et al., Molecular Neurobiology, 2013 Feb;47(1):241-60).

[0566] As confirmed in Example 1 above, TAFA4 statistically significantly increases the length and branching points of ganglion cell neurites compared to the control group. To evaluate whether TAFA4 improves retinal neuropathies in an actual in-vivo model, changes in fluorescence angiography (FA) and electroretinogram (ERG) following administration of AAV8.mouse TAFA4 were evaluated in a mouse model in which retinal damage was induced by administration of NaIO3.

[0567] Control AAV8 or AAV8.mouse TAFA4 (CAT311 promoter-mTAFA4) without transgene was administered subretinal injection (SRI) to C57BL / 6 mice (Orient Bio).

[0568] 5Х10 to ensure sufficient expression of Mouse TAFA4 8After subretinal injection (SRI) of AAV8 of vg into both eyes of mice, AMD model was induced by tail vein injection of 20 mg / kg NaIO3 for 56 days. Ten days after model induction (66 days after SRI administration), fluorescent contrast agent was injected via tail vein. After focusing the image on the fundus with a Micron-IV imaging camera (Phoenix), fluorescence angiography (FA) images were taken. Eleven days after model induction (67 days after SRI administration), scotopic ERG evaluation was performed. Mice were placed on the ERG stage, and ERG probes were contacted to the tail, head, and cornea, respectively, and the amplitudes of the A-wave and B-wave were measured. ERG analysis was performed using the program 'LabScribeERG (iWorx DataAcquisition Software)'.

[0569] On the FA images taken 10 days after model induction, we could confirm the leakage of fluorescent contrast agent due to damage to the outer retina in the NaIO3-administered group (control group, control-AAV8-administered) compared to the healthy (non-AMD-induced) animal group (naive, control AAV8-administered) group. In contrast, the group administered AAV8.mouse TAFA4 (experimental group) showed retinal findings that were almost normal, similar to the naive group (Fig. 2). The scotopic ERG results evaluated on the 11th day after model induction also showed retinal abnormalities due to NaIO3, similar to the FA, and both the A-wave and B-wave amplitudes of the potential evoked widths were significantly reduced. In contrast, the group administered AAV8.mouse TAFA4 (experimental group) showed recovery in both A-wave and B-wave amplitudes, and in particular, the B-wave amplitude recovered to a level close to normal (Fig. 3).

[0570]

[0571] 2-C. Evaluation of the efficacy of interspecies variants of TAFA4 in a sodium iodate (NaIO3)-induced retinal nerve damage model.

[0572] To determine whether the interspecies variants of TAFA4 also exhibit the same efficacy, the sequence identity of full-length TAFA4 proteins from mammals (Human, Monkey, Pig, Rabbit, Rat, Mouse), birds (Chicken), reptiles (Komodo dragon, Wall lizard, Fence lizard, Gecko), amphibians (Frog), and fish was compared, and the consensus sequence between each species was confirmed (Fig. 4). The amino acid sequence of the mature human TAFA4 protein (95 aa) showed a very high sequence identity regardless of species, with more than 90% sequence identity compared to mammals and amphibians, more than 95% sequence identity compared to birds, and more than 85% sequence identity compared to reptiles and fish (Fig. 5).

[0573] Since the protective effect of AAV8.mouse TAFA4 against retinal neural damage was confirmed in the sodium iodate (NaIO3)-induced retinal damage model, an experiment was conducted to determine whether TAFA4 of other species also has a protective effect against retinal damage. An experiment was conducted to determine the efficacy of human TAFA4 and reptile (gecko TAFA4; sequence identity 87.4%) and fish (fish TAFA4; sequence identity 86.3%) TAFA4, which have the lowest sequence identity with human TAFA4. Fluorescence angiography (FA) and electroretinogram (ERG) changes following administration of AAV8.human TAFA4, AAV8.gecko TAFA4, and AAV8.fish TAFA4 to a mouse model of AMD induced by NaIO3 administration were evaluated.

[0574] Control AAV8 or each strain of AAV8.TAFA4 was administered subretinal injection (SRI) to C57BL / 6 mice (Orient Bio).

[0575] 1Х10 to ensure sufficient expression of TAFA4 9 After subretinal injection (SRI) of AAV8 of vg into both eyes of mice, AMD was induced by intravenous administration of NaIO3 at a dose of 20 mg / kg for 42 days. Nine days after model induction (51 days after SRI administration), a fluorescent contrast agent was injected via the tail vein. After focusing the image on the fundus with a Micron-IV imaging camera (Phoenix), fluorescence angiography (FA) images were captured. Seven days after model induction (49 days after SRI administration), scotopic ERG evaluation was performed. Mice were placed on an ERG stage, and ERG probes were contacted to the tail, head, and cornea, respectively, and the amplitudes of the A-wave and B-wave were measured. ERG analysis was performed using the program 'LabScribeERG (iWorx DataAcquisition Software)'.

[0576] On the FA images taken on the 9th day after model induction, the leakage of fluorescent contrast agent due to damage to the outer retina was confirmed in the retina of the NaIO3-administered group (control group, control-AAV8 administration) compared to the healthy animal group (naive, control AAV8 administration). On the other hand, the groups (experimental group) administered AAV8.human TAFA4, AAV8.gecko TAFA4, and AAV8.fish TAFA4 showed retinal findings that were almost normal, similar to the naive group (Fig. 6). The scotopic ERG results evaluated on the 11th day after model induction also confirmed retinal abnormalities in which both the A-wave and B-wave amplitude potential evoked widths were significantly reduced by NaIO3. In contrast, the groups administered AAV8.human TAFA4, AAV8.gecko TAFA4, and AAV8.fish TAFA4 (experimental group) showed retinal abnormalities due to NaIO3. A-wave and B-wave amplitude potential-induced changes were restored (Fig. 7).

[0577]

[0578] 2-D. Protective effect of TAFA4 paralogs on retinal damage according to sequence identity in a sodium iodate (NaIO3)-induced retinal nerve damage model.

[0579] It is known that there is a high degree of sequence identity between TAFA4 and the TAFA protein family (TAFA1-3). Therefore, we investigated whether human TAFA1, TAFA2, and TAFA3 also exhibit retinal protective effects in a mouse model of NaIO3-induced retinal damage. Fluorescence angiography (FA) and electroretinogram (ERG) changes following administration of AAV8.TAFA1, AAV8.TAFA2, AAV8.TAFA3, and AAV8.TAFA4 were evaluated in a mouse model of AMD induced by NaIO3 administration.

[0580] Control AAV8 or AAV8.TAFA1 or AAV8.TAFA2 or AAV8.TAFA3 or AAV8.TAFA4 was administered subretinal injection (SRI) to C57BL / 6 mice (Orient Bio).

[0581] To confirm the retinal protective effect of TAFA1, TAFA3, and TAFA4, 1Х10 9 After subretinal injection (SRI) of AAV8 of vg into both eyes of mice, AMD model was induced by intravenous administration of NaIO3 at a dose of 20 mg / kg for 42 days. Nine days after model induction (51 days after SRI administration), fluorescent contrast agent was injected through the tail vein. After focusing the image on the fundus with a Micron-IV imaging camera (Phoenix), fluorescence angiography (FA) images were captured. Seven days after model induction (49 days after SRI administration), scotopic ERG evaluation was performed. Mice were placed on the ERG stage, and ERG probes were contacted to the tail, head, and cornea, respectively, and the amplitudes of the A-wave and B-wave were measured. ERG analysis was performed using the program 'LabScribeERG (iWorx DataAcquisition Software)'.

[0582] To confirm the retinal protective effect of TAFA2, 1Х10 9After subretinal injection (SRI) of AAV8 of vg into both eyes of mice, AMD model was induced by intravenous administration of NaIO3 at a dose of 20 mg / kg for 56 days. Nine days after model induction (65 days after SRI administration), fluorescent contrast agent was injected through the tail vein. After focusing the image on the fundus with a Micron-IV imaging camera (Phoenix), fluorescence angiography (FA) images were taken. Seven days after model induction (63 days after SRI administration), scotopic ERG evaluation was performed. Mice were placed on the ERG stage, and ERG probes were contacted to the tail, head, and cornea, respectively, and the amplitudes of the A-wave and B-wave were measured. ERG analysis was performed using the program 'LabScribeERG (iWorx DataAcquisition Software)'.

[0583] In FA images, the leakage of fluorescent contrast agent due to damage to the outer retina was confirmed in the NaIO3-administered group (control group, control-AAV8 administration) compared to the healthy animal group (naive, control AAV8 administration). On the other hand, the group administered AAV8.TAFA1, AAV8.TAFA2, AAV8.TAFA3, or AAV8.TAFA4 (experimental group) showed retinal findings that were almost normal, similar to the naive group (Fig. 8). In the scotopic ERG results, retinal abnormalities were confirmed in which both the A-wave and B-wave amplitude potential evoked widths were significantly reduced by NaIO3. In contrast, the group administered AAV8.TAFA1, AAV8.TAFA2, AAV8.TAFA3, or AAV8.TAFA4 (experimental group) showed retinal abnormalities due to NaIO3. A-wave and B-wave amplitude potential-induced changes were restored (Fig. 9).

[0584]

[0585] 2-E. Sequence identity comparison of interspecies variants of TAFA1-4

[0586] As confirmed in the above 2-C and 2-D, the retinal protective effect was confirmed in the interspecies mutants of TAFA4 and the paralogs of TAFA4, TAFA1 to 3, in the retinal damage model by NaIO3. Therefore, it was thought that the common sequence of various TAFA proteins would have a retinal protective effect, and to confirm the common sequence, the sequence identity of full-length TAFA1, full-length TAFA2, full-length TAFA3, and full-length TAFA4 proteins of mammals (Human, Monkey, Pig, Rabbit, Rat, Mouse), birds (Chicken), reptiles (Komodo dragon, Wall lizard, Fence lizard, Gecko), amphibians (Frog), and fish (Fish) was compared. The amino acid sequences between full-length TAFA1 to TAFA4 in vertebrates are shown in Figs. 10a to 10d.

[0587] The 93-nucleotide sequence of human TAFA4 (IKQGTCEVVAVHRCCNKNRIEERSQTVKCSCFPGQVAGTTRAQPSCVEASIVIQKWWCHMNPCLEGEDCKVLPDYSGWSCSSGNKVKTTKVTR, SEQ ID NO: 87) was confirmed to have very high sequence identity with the TAFA4 sequences of other species and the consensus sequences of human and fish TAFA1 to TAFA3. The sequence identity within the consensus sequence is shown in Table 2.

[0588] Sequence identity of the common sequence (93 aa) of each species of TAFA1, TAFA2, TAFA3, and TAFA4 and human TAFA4 TAFA Family Species Scientific name Length Sequence number Sequence identity TAFA4 Human Homo sapiens 9387100.0% TAFA1 Human Homo sapiens 918875.8% TAFA1 Monkey Macaca fascicularis 918975.8% TAFA1 Pig Sus scrofa 919075.8% TAFA1 Rabbit Oryctolagus cuniculus 919175.8% TAFA1 Rat Rattus norvegicus 919275.8% TAFA1 Mouse Mus musculus 919375.8% TAFA1 Chicken Gallus gallus 919475.8% TAFA1 Komodo dragon Varanus komodoensis919576.9%TAFA1Wall lizardPodarcis muralis919676.9%TAFA1Fence lizardSceloporus undulatus919776.9%TAFA1FrogXenopus tropicalis_isoform1929876.1%TAFA1FrogXenopus tropicalis_isoform2929976.1%TAFA1FishDanio rerio_isoform19310074.2%TAFA1FishDanio rerio_isoform29310175.3%TAFA2HumanHomo sapiens9310286.0%TAFA2MonkeyMacaca fascicularis9310386.0%TAFA2PigSus scrofa9310486.0%TAFA2RabbitOryctolagus cuniculus9310586.0%TAFA2RatRattus norvegicus_isoform19310682.8%TAFA2RatRattus norvegicus_isoform29310784.9%TAFA2MouseMus musculus9310884.9%TAFA2ChickenGallus gallus_isoform19310983.9%TAFA2ChickenGallus gallus_isoform29311087.1%TAFA2Komodo dragonVaranus komodoensis9311184.9%TAFA2Wall lizardPodarcis muralis_isoform19111285.7%TAFA2Wall lizardPodarcis muralis_isoform29311387.1%TAFA2Fence lizardSceloporus undulatus9311487.1%TAFA2GeckoGekko japonicus9211583.7%TAFA2FrogXenopus tropicalis9311684.9%TAFA2FishDanio rerio9311782.8%TAFA3HumanHomo sapiens9311882.8%TAFA3MonkeyMacaca fascicularis9311982.8%TAFA3PigSus scrofa9312084.9%TAFA3RabbitOryctolagus cuniculus9312182.8%TAFA3RatRattus norvegicus9312282.8%TAFA3MouseMus musculus9312383.9%TAFA3ChickenGallus gallus9312484.9%TAFA3Komodo dragonVaranus komodoensis9312586.0%TAFA3Wall lizardPodarcis muralis9312683.9%TAFA3Fencee lizardSceloporus undulatus9312786.0%TAFA3FrogXenopus tropicalis9312887.1%TAFA3FishDanio rerio9312981.7%TAFA4MonkeyMacaca fascicularis93130100.0%TAFA4PigSus scrofa93131100.0%TAFA4RabbitOryctolagus cuniculus9313296.8%TAFA4RatRattus norvegicus9313396.8%TAFA4MouseMus musculus9313495.7%TAFA4ChickenGallus gallus9313596.8%TAFA4Komodo dragonVaranus komodoensis9313694.6%TAFA4Wall lizardPodarcis muralis9313796.8%TAFA4Fence lizardSceloporus undulatus9313895.7%TAFA4GeckoGekko japonicus9313987.1%TAFA4FrogXenopus tropicalis9314092.5%TAFA4FishDanio rerio9314187.1%.

[0589] Sequence identity was confirmed using Clustal Omega.

[0590]

[0591] [Example 3] Evaluation of the efficacy of TAFA4 peptide fragments

[0592] Based on the results of Examples 1 and 2, the present inventors synthesized three TAFA4 peptide fragments (TAFA4 fragments 1-3, F1-F3) as shown in Table 3 to identify fragments exhibiting biological efficacy of TAFA4. The peptides were synthesized by request from AppClone.

[0593] Amino acid sequence of TAFA4 peptide fragments 1-3 Peptide name Length (aa) Amino acid sequence (Sequence, N'→C') TAFA4 fragment 1 (SEQ ID NO: 148) 35HQIKQGTCEV VAVHRCCNKN RIEERSQTVK CSCFP TAFA4 fragment 2 (SEQ ID NO: 75) 31GQVAGTTRAQ PSCVEASIVI QKWWCHMNPC LTAFA4 fragment 3 (SEQ ID NO: 52) 29EGEDCKVLPD YSGWSCSSGN KVKTTKVTR

[0594] MED17.11 cells resuspended in differentiation induction medium in the same manner as in Example 1 were seeded at 1 Х 10 per well of a 96-well cell culture plate. 4After dispensing 100 μL of cells / mL, 5 μM TAFA4 peptide fragments 1-3 were treated, and the same volume of dimethyl-sulfoxide (DMSO, Sigma) was treated as a control, and cultured in a 37°C incubator (5% CO2). After culturing for 4 days, neurite length and branch points were analyzed using Incucyte. A total of three replicate experiments were performed, and both neurite length and branch points tended to increase in the TAFA4 fragment 2 (F2) and TAFA4 fragment 3 (F3) treatment groups. (Fig. 11a and b).

[0595]

[0596] The species sequences of TAFA4 fragment 2 (F2) and fragment 3 (F3) are shown in Tables 4 and 5 below.

[0597] Amino acid sequences of TAFA4 fragment 2 by species Species (scientific name) SEQ ID NOSEQUENCEa.a. Human (Homo sapiens), Monkey (Macaca fascicularis), Pig (Sus scrofa) SEQ ID 75GQVAGTTRAQPSCVEASIVIQKWWCHMNPCL31 Rabbit (Oryctolagus cuniculus) SEQ ID 76GQVAGTTQAQPSCVEASIVAQKWWCHMNPCL31 Rat (Rattus norvegicus) SEQ ID 77GQVAGTTRAQPSCVEASIVIEKWWCHMDPCL31 Mouse (Mus musculus) SEQ ID 78GQVAGTTRAQPSCVEAAIVIEKWWCHMNPCL31 Chicken (Gallus gallus) SEQ ID 79GQVAGTTRAQPSCVEASIVLQKWWCHMNPCL31 Komodo Dragon (Varanus komodoensis) SEQ ID NO: 80GQVAGTTRAQPSCVEAAIVVQKWWCHMNPCL31Wall lizard (Podarcis muralis) SEQ ID NO: 81GQVAGTTRAQPSCVEAAIVIQKWWCHMNPCL31Fence lizard (Sceloporus undulatus) SEQ ID NO: 82GQVAGTTRSQPSCVEAAIVIQKWWCHMNPCL31Gecko (Gekko japonicus) SEQ ID NO: 83GQVAGTTRTQPSCVEAAIVIQKWWCQMSPCL31Frog (Xenopus tropicalis) SEQ ID NO: 84GQVAGTTRAQPSCVEASIVIQKWWCHMNPCM31Fish (Danio rerio) SEQ ID NO: 85GQVAGTTRAQPSCVEASIVLQKWWCQMHPCL31

[0598] The sequence reflecting the above fragment 2 and its interspecies variation is as shown in SEQ ID NO: 147.

[0599] Amino acid sequences by species of TAFA4 fragment 3 Species (scientific name) SEQ ID NOSEQUENCE a.a. Human (Homo sapiens), Monkey (Macaca fascicularis), Pig (Sus scrofa), Rabbit (Oryctolagus cuniculus) SEQ ID 52 EGEDCKVLPDYSGWSCSSGNKVKTTKVTR29 Rat (Rattus norvegicus), Mouse (Mus musculus) SEQ ID 53 EGEDCKVLPDSSGWSCSSGNKVKTTKVTR29 Chicken (Gallus gallus), Wall lizard (Podarcis muralis) SEQ ID 54 DGEDCKVLPDYSGWSCSSGNKVKTTKVTR29 Komodo dragon (Varanus komodoensis), Fence lizard (Sceloporus undulatus) SEQ ID 55EGEECKVLPDYSGWSCSSGNKVKTTKVTR29Gecko (Gekko japonicus) SEQ ID NO: 56EGEECKVLPDSSGWSCSSGNKVKTTKVTR29Frog (Xenopus tropicalis) SEQ ID NO: 57EGEECKVLPDLTGWSCSSGNKIKTTKVTR29Fish (Danio rerio) SEQ ID NO: 58DGEECKALPDLTGWSCSTGNKVKTTKVTR29

[0600] The sequence reflecting the above fragment 3 and its interspecies variation is as shown in SEQ ID NO: 144.

[0601]

[0602] [Example 4] Efficacy Evaluation of Subdivided TAFA4 Peptide Fragments

[0603] Based on Example 3, where TAFA4 fragment 2 (F2) and TAFA4 fragment 3 (F3) were effective, further subdivided TAFA4 peptide fragments (TAFA4 fragments 4-8, F4-F8) were synthesized as shown in Table 6 to identify key TAFA4 peptide fragments exhibiting biological efficacy. Peptide synthesis was commissioned to AppClone. The location of each fragment is as shown in Figure 12.

[0604]

[0605] Amino acid sequence of TAFA4 peptide fragments 4-8 Peptide name Length (aa) Amino acid sequence (Sequence, N'→C') TAFA4 fragment 4 (SEQ ID NO: 149) 20HQIKQGTCEV VAVHRCCNKNTAFA4 fragment 5 (SEQ ID NO: 59) 32RIEERSQTVK CSCFPGQVAG TTRAQPSCVE ASTAFA4 fragment 6 (SEQ ID NO: 150) 17FPGQVAGTTR AQPSCVETAFA4 fragment 7 (SEQ ID NO: 28) 22IVIQKWWCHM NPCLEGEDCK VLTAFA4 fragment 8 (SEQ ID NO: 151) 21PDYSGWSCSS GNKVKTTKVT R

[0606] In the same manner as in Example 4, 5 μM TAFA4 peptide fragment and the same volume of DMSO were treated and cultured in a 37°C incubator (5% CO2). After culturing for 4 days, cells were photographed using Incucyte (Satorius), and neurite length and branch points were analyzed. Two replicate experiments were performed, and statistical significance was confirmed through one-way ANOVA (Dunnett's test post hoc).

[0607] The neurite length tended to increase in TAFA4 fragments 5 (F5) and 7 (F7), and in particular, TAFA4 fragment 5 (F5) showed a statistically significant increase (Fig. 13a).

[0608] In the case of branch points, an increasing trend was confirmed only in TAFA4 fragments 5 (F5) and 7 (F7) (Fig. 13b).

[0609]

[0610] Comparison of the interspecies sequence homology of TAFA4 fragment 5 (F5) and fragment 7 (F7) is shown in Tables 7 and 8 below.

[0611]

[0612] Interspecies homology comparison of TAFA4 peptide fragment 5 Species (scientific name) SEQ ID NO Amino acid sequence (Sequence, N'→C') Sequence identity Human (Homo sapiens), Monkey (Macaca fascicularis), Pig (Sus scrofa), Rat (Rattus norvegicus), Chicken (Gallus gallus), Frog (Xenopus tropicalis) Sequence number 59 RIEERSQTVK CSCFPGQVAG TTRAQPSCVE AS 100.0% Rabbit (Oryctolagus cuniculus) Sequence number 60 RIEERSQTVK CSCFPGQVAG TTQAQPSCVE AS 96.9% Mouse (Mus musculus), Wall lizard (Podarcis muralis) Sequence number 61 RIEERSQTVK CSCFPGQVAG TTRAQPSCVE AA 96.9% Komodo dragon (Varanus komodoensis) SEQ ID NO: 62RIEERSQTVK CSCLPGQVAG TTRAQPSCVE AA93.8% Fence lizard (Sceloporus undulatus) SEQ ID NO: 63RIEERSQTVK CSCFPGQVAG TTRSQPSCVE AA93.8% Gecko (Gekko japonicus) SEQ ID NO: 64RIEELSQTGN CSCLPGQVAG TTRTQPSCVE AA81.3% Fish (Danio rerio) SEQ ID NO: 65KIEERSQTVK CSCFPGQVAG TTRAQPSCVE AS96.9%

[0613] Interspecies homology comparison of TAFA4 peptide fragment 7 Species (scientific name) SEQ ID NO Amino acid sequence (Sequence, N'→C' ) Sequence identity Human (Homo sapiens), Monkey (Macaca fascicularis), Pig (Sus scrofa) SEQ ID NO 28 IVIQKWWCHM NPCLEGEDCK VL 100.0% Rabbit (Oryctolagus cuniculus) SEQ ID NO 29 IVAQKWWCHM NPCLEGEDCK VL 95.5% Rat (Rattus norvegicus) SEQ ID NO 30 IVIEKWWCHM DPCLEGEDCK VL 90.9% Mouse (Mus musculus) SEQ ID NO 31 IVIEKWWCHM NPCLEGEDCK VL 95.5% Chicken (Gallus gallus) SEQ ID NO 32 IVLQKWWCHM NPCLDGEDCK VL90.9% Komodo dragon (Varanus komodoensis) SEQ ID NO: 33 IVVQKWWCHM NPCLEGEECK VL90.9% Wall lizard (Podarcis muralis) SEQ ID NO: 34 IVIQKWWCHM NPCLDGEDCK VL95.5% Fence lizard (Sceloporus undulatus) SEQ ID NO: 35 IVIQKWWCHM NPCLEGEECK VL95.5% Gecko (Gekko japonicus) SEQ ID NO: 36 IVIQKWWCQM SPCLEGEECK VL86.4% Frog (Xenopus tropicalis) (SEQ ID NO: 32) SEQ ID NO: 37 IVIQKWWCHM NPCMEGEECK VL90.9% Fish (Danio rerio) SEQ ID NO: 38 IVLQKWWCQM HPCLDGEECK AL72.7%

[0614]

[0615] [Example 5] Evaluation of the efficacy of TAFA4 and its fragments in improving neuropathic pain in vivo.

[0616] We previously demonstrated that rhTAFA4 and TAFA4 fragments 2, 3, 5, and 7 (F2, F3, F5, and F7) increased neurite length and branching points in MED17.11 cells. To confirm their efficacy in vivo, we evaluated the efficacy of rhTAFA4 and TAFA4 fragments in a neuropathic pain model.

[0617] Six-week-old mice were anesthetized, and the skin between the gluteus maximus and biceps femoris muscles of the left hind paw was incised to expose the sciatic nerve. The exposed nerve was separated from the surrounding tissue, and the proximal portion where it divides into the tibial, peroneal, and sural nerves was ligated three times with 6-0 silk (AILEE) at 0.5-1 mm intervals. The incised skin was then sutured, and the mice were allowed to recover for one week. After one week, the level of pain was assessed using the vonfrey test. The vonfrey test was performed by applying each monofilament six times to the mid-left hind paw, starting with a minimal force (0.008 g). If the mouse did not exhibit pain responses (e.g., lifting, brushing, or licking), the monofilament was stimulated with increasingly stronger forces. The g-value of the monofilament that elicited a pain response in three or more of the six stimulations was recorded to determine the 50% threshold.

[0618] Pain caused by sciatic nerve ligation was confirmed, and the degree of pain relief was evaluated by administering rhTAFA4 and TAFA4 fragments. rhTAFA4 and TAFA4 fragments 1, 2, 3, 5, 6, and 7 (F1, F2, F3, F5, F6, and F7) were diluted in PBS to 200 μg / mL and injected intrathecally at the L4-L5 level at 10 μL (2 μg) per mouse. One hour after administration, the vonfrey test was performed again to evaluate the degree of pain relief.

[0619] In the control group administered with the same amount of PBS, the pain caused by nerve ligation persisted, confirming the occurrence of mechanical allodynia. However, in the group administered with rhTAFA4, pain was confirmed to be relieved. In addition, the group administered with TAFA4 fragments 1 and 6 (F1 and F6) showed persistent pain, whereas the group administered with TAFA4 fragments 2, 3, 5, and 7 (F2, F3, F5, and F7), which showed a clear tendency to increase the length and branching of neurites, showed pain relief (Fig. 14).

[0620] Fragments 2 and 5, which exhibited the above efficacy, share the GQVAGTTRAQPSCVEAS sequence (17 amino acids, referred to as fragment 2.5), and fragments 3 and 7 share the EGEDCKVL sequence (8 amino acids, referred to as fragment 3.7). Therefore, the increase in the length and branching point of the neurite of Examples 1-4 and the pain relief effect of this Example are believed to be the effects of a polypeptide containing the sequence of fragment 2.5 or the sequence of fragment 3.7, which are the common sequences of the fragments.

[0621] The sequence of the above TAFA4 fragment 2.5 was compared with the sequences of interspecies mutants of TAFA 1 to 3, and it was confirmed that there was only a difference of 1 to 4 amino acid sequences from human TAFA4 fragment 2.5, confirming that the homology was very high.

[0622] In addition, the sequence of the above TAFA4 fragment 3.7 was compared with the interspecies mutant sequences of TAFA 1 to 3, and it was confirmed that there was only a difference of 1 to 3 amino acid sequences from human TAFA4 fragment 3.7, confirming that fragment 3.7 also had a very high degree of homology.

[0623] The results of comparing the sequence of the above TAFA4 with the interspecies mutant sequences of TAFA 1 to 3 are as shown in Tables 9 (fragment 2.5) and 10 (fragment 3.7) below.

[0624]

[0625] TAFA protein fragment 2.5 and its interspecies variant sequences exhibiting the effect TAFA Species SEQ ID NO Sequence, N'→C' TAFA4 HumanMonkeyPigRatChickenFrogFish SEQ ID NO 15 GQVAG TTRAQPSCVE ASRabbit SEQ ID NO 16 GQVAG TTQAQPSCVE ASMouseKomodo dragonWall lizard SEQ ID NO 17 GQVAG TTRAQPSCVE AAFence lizard SEQ ID NO 18 GQVAG TTRSQPSCVE AAGecko SEQ ID NO 19 GQVAG TTRTQPSCVE AATAFA1 HumanMonkeyPigRabbitRatMouseChickenKomodo dragonWall lizardFence lizardFrog SEQ ID NO 20 GKVAG TTRNRPSCVDASFish SEQ ID NO 21 GKVAG TTRNKPSCVDASGecko SEQ ID NO 22GKVAG TTRNRPSCVD--TAFA2HumanMonkeyPigRabbitRatMouseChickenKomodo dragonWall lizardFence lizardGeckoFishSequence number 23GQVAG TTRAAPSCVDASFrogSequence number 24GQVAG TTRATPSCVDASTAFA3HumanMonkeyPigRabbitRatMouseSequence number 25GQVAG TTRAKPSCVDASCickenKomodo dragonFence lizardFrogFishSequence number 26GQVAG TTRAAPSCVDASWall lizardSequence number 27GQVAG TTHAAPSCVDAS

[0626] Amino acids with differences are shown in bold. The sequence reflecting fragment 2.5 and its interspecies variation is as shown in SEQ ID NO: 145.

[0627]

[0628] TAFA protein fragment 3.7 showing effect and its interspecies variant sequences TAFA Species SEQ ID NO Sequence, N'→C' TAFA 4 HumanMonkeyPigRabbitRatMouse SEQ ID NO 1 EGEDCK VLChickenWall lizard SEQ ID NO 2 DGEDCK VLKomodo dragonFence lizardGeckoFrog SEQ ID NO 3 EGEECK VLFish SEQ ID NO 4 DGEECKALTAFA 1 HumanMonkeyPigRabbitRatMouseChickenKomodo dragonWall lizardFence lizardFrogFish SEQ ID NO 5 EGEECKTLTAFA 2 HumanMonkeyPigRabbitRatMouseChickenKomodo dragonWall lizardFence lizardGecko SEQ ID NO 6 EGEECK VLFrog SEQ ID NO 7 EGEECKILFish SEQ ID NO 8 DGEECK VLTAFA3HumanMonkeyPigRabbitSequence number 9PGEECK VLRatMouseSequence number 10LGEECK VLChickenSequence number 11AGEECK VLKomodo dragonWall lizardFence lizardSequence number 12EGEECK VLFrogSequence number 13EGEDCK VLFishSequence number 14DGEECK VL

[0629] Amino acids with differences are shown in bold. The sequence reflecting fragment 3.7 and its interspecies variation is as shown in SEQ ID NO: 142.

[0630]

[0631] [Example 6] Homology comparison of TAFA4 peptide fragment and TAFA 1 to 3 peptide fragments

[0632] In Example 3, it was confirmed that the interspecies homology of the TAFA4 fragment was 70% or more (F5 81.3% or more, F7 72.7% or more). It is known that there is high sequence identity between TAFA4 and the TAFA protein group. Therefore, it was thought that the common sequence of various TAFA peptide fragments would have the effect of increasing neurite length and branch point, and the sequence homology (Tables 11 to 16) was compared between TAFA1, TAFA2 and TAFA3 peptide fragments of vertebrate mammals (Human, Monkey, Pig, Rabbit, Rat, Mouse), birds (Chicken), reptiles (Komodo dragon, Wall lizard, Fence lizard, Gecko), amphibians (Frog) and fish and human TAFA4 fragment 5 (F5) and fragment 7 (F7), and the consensus sequence was confirmed.

[0633]

[0634] Homology comparison of human TAFA4 fragment 5 and TAFA1 peptide fragment 5 Species (scientific name) SEQ ID NO Amino acid sequence (Sequence, N'→C') Sequence identity Human (Homo sapiens), Monkey (Macaca fascicularis), Pig (Sus scrofa), Rabbit (Oryctolagus cuniculus), Rat (Rattus norvegicus), Mouse (Mus musculus), Chicken (Gallus gallus), Komodo dragon (Varanus komodoensis), Wall lizard (Podarcis muralis), Fence lizard (Sceloporus undulatus), Frog (Xenopus tropicalis) Sequence number 66 RIEERSQTVK CSCLPGKVAG TTRNRPSCVD AS84.4% Gecko (Gekko japonicus) Sequence number 67 RIEERSQTVK CSCLPGKVAG TTRNRPSCVD --78.1% Fish (Danio rerio) Sequence number 68KIEERSQTVK CSCLPGKVAG TTRNKPSCVD AS81.3%

[0635] Homology comparison of human TAFA4 fragment 5 and TAFA2 peptide fragment 5 Species (scientific name) SEQ ID NO Amino acid sequence (Sequence, N'→C') Sequence identity Human (Homo sapiens), Monkey (Macaca fascicularis), Pig (Sus scrofa), Rabbit (Oryctolagus cuniculus), Rat (Rattus norvegicus), Mouse (Mus musculus), Chicken (Gallus gallus), Komodo dragon (Varanus komodoensis), Wall lizard (Podarcis muralis), Fence lizard (Sceloporus undulatus), Gecko (Gekko japonicus), Fish (Danio rerio) Sequence number 69 KIEERSQTVK CSCFPGQVAG TTRAAPSCVD AS90.6% Frog (Xenopus tropicalis) Sequence number 70 KIEERSQTVK CSCFPGQVAG TTRATPSCVD AS90.6%

[0636] Homology comparison of human TAFA4 fragment 5 and TAFA3 peptide fragment 5 Species (scientific name) SEQ ID NO Amino acid sequence (Sequence, N'→C') Sequence identity Human (Homo sapiens), Monkey (Macaca fascicularis), Pig (Sus scrofa) Sequence number 71 RIEERSQTVK CSCFSGQVAG TTRAKPSCVD AS 90.6% Rabbit (Oryctolagus cuniculus), Rat (Rattus norvegicus), Mouse (Mus musculus) Sequence number 72 RIEERSQTVK CSCLSGQVAG TTRAKPSCVD AS 87.5% Chicken (Gallus gallus), Komodo dragon (Varanus komodoensis), Fencee lizard (Sceloporus undulatus), Frog (Xenopus tropicalis), Fish (Danio rerio) Sequence number 73KIEERSQTVK CSCFPGQVAG TTRAAPSCVD AS90.6%Wall lizard (Podarcis muralis)SEQ ID NO. 74KIEERSQTVK CSCFPGQVAG TTHAAPSCVD AS87.5%

[0637] The sequence reflecting the above fragment 5 and its interspecies variation is as shown in SEQ ID NO: 146.

[0638] Homology comparison of human TAFA4 fragment 7 and TAFA1 peptide fragment 7 Species (scientific name) SEQ ID NO Amino acid sequence (Sequence, N'→C') Sequence identity Human (Homo sapiens), Monkey (Macaca fascicularis), Pig (Sus scrofa), Rabbit (Oryctolagus cuniculus), Rat (Rattus norvegicus), Mouse (Mus musculus), Chicken (Gallus gallus), Komodo dragon (Varanus komodoensis), Wall lizard (Podarcis muralis), Fence lizard (Sceloporus undulatus), Frog (Xenopus tropicalis), Fish (Danio rerio) Sequence number 39 IVIGKWWCEM EPCLEGEECK TL77.3%

[0639] Homology comparison of human TAFA4 fragment 7 and TAFA2 peptide fragment 7 Species (scientific name) SEQ ID NO Amino acid sequence (Sequence, N'→C') Sequence identity Human (Homo sapiens), Monkey (Macaca fascicularis), Pig (Sus scrofa), Rabbit (Oryctolagus cuniculus), Rat (Rattus norvegicus), Mouse (Mus musculus), Chicken (Gallus gallus), Komodo dragon (Varanus komodoensis), Wall lizard (Podarcis muralis), Fence lizard (Sceloporus undulatus) Sequence number 40 IVEQKWWCHM QPCLEGEECK VL 8 6.4% Gecko (Gekko japonicus) Sequence number 41 IVEQKWWCQM QPCLEGEECK VL 8 1.8% Frog (Xenopus tropicalis) Sequence number 42 IVEQKWWCHM QPCLEGEECK IL81.8%Fish (Danio rerio)Sequence number 43IVAQKWWCQM QPCMDGEECK VL72.7%

[0640] Homology comparison of human TAFA4 fragment 7 and TAFA3 peptide fragment 7 Species (scientific name) Length (aa) Amino acid sequence (Sequence, N'→C') Sequence identity Human (Homo sapiens), Monkey (Macaca fascicularis), Rabbit (Oryctolagus cuniculus) Sequence number 44 IVLQRWWCQM EPCLPGEECK VL 7 2.7% Pig (Sus scrofa) Sequence number 45 IVLQKWWCQM EPCLPGEECK VL 7 7.3% Rat (Rattus norvegicus), Mouse (Mus musculus) Sequence number 46 IVLQKWWCQM EPCLLGEECK VL 7 7.3% Chicken (Gallus gallus) Sequence number 47 IVLQKWWCQM EPCLAGEECK VL 7 7.3% Komodo dragon (Varanus komodoensis), Fencee lizard (Sceloporus undulatus) SEQ ID NO: 48IVLQKWWCQM QPCLEGEECK VL81.8% Wall lizard (Podarcis muralis) SEQ ID NO: 49IVLQKWWCQM QPCVEGEECK VL77.3% Frog (Xenopus tropicalis) SEQ ID NO: 50IVIQKWWCQM EPCLEGEDCK VL90.9% Fish (Danio rerio) SEQ ID NO: 51IVAQKWWCQM QPCVDGEECK VL72.7%

[0641] The sequence reflecting the above fragment 7 and its interspecies variation is as shown in SEQ ID NO: 143.

[0642] Additionally, the interspecies sequences of fragment 3 of TAFA1 to 3 (Table 17) and the interspecies sequences of fragment 2 (Table 18) are as follows:

[0643]

[0644] Interspecific sequence of fragment 3 of TAFA1 to 3 TAFA species SEQ ID NO Sequence, N'→ C' TAFA1 Human, Monkey, Pig, Rabbit, Rat, Mouse, Chicken, Frog (isoform 2) SEQ ID NO 152 EGEECKTLPDNSGWMCATGNKIKTTR-IHTAFA1 Komodo dragon, Wall lizard, Fence lizard SEQ ID NO 153 EGEECKTLPDNSGWMCATGNKVKTTR-IHTAFA1 Frog (isoform 1) SEQ ID NO 154 EGEECKTLPDNSGWMCATGNKIKTTR-FQTAFA1 Fish (isoform 1) SEQ ID NO 155 EGEECKTLPDNSGWMCYSGNKIKTTRGNITAFA1 Fish (isoform 2) SEQ ID NO 156 EGEECKTLPDNSGWMCYSGNKIKTTRNTHTAFA2 Human, Monkey, Pig, Rabbit, Rat (isoform 2), Mouse SEQ ID NO: 157EGEECKVLPDRKGWSCSSGNKVKTTRVTHTAFA2Rat(isoform1), Chicken(form1_) SEQ ID NO: 158EGEECKVLPDRKGWSCSSGNKVKTTRANVTAFA2Chicken(isoform2), Wall lizard(2), Fence lizard SEQ ID NO: 159EGEECKVLPDRKGWSCSSGNKVKTTRVTRTAFA2Komodo dragon SEQ ID NO: 160EGEECKVLPDRKGWSCSSGNKVKTTRVSKTAFA2Wall Lizard SEQ ID NO: 161EGEECKVLPDRKGWSCSSGNKVKTTRM--TAFA2Gecko SEQ ID NO: 162EGEECKVLPDRKGWSCSSGNKVKTTRAN-TAFA2Frog SEQ ID NO: 163EGEECKILPDQKGWSCASGNKVKTTKVTRTAFA2Fish SEQ ID NO: 164DGEECKVLPDLKGWSCSTGNKVKTTKVTRTAFA3Human, Monkey, Pig,Rabbit SEQ ID NO: 165PGEECKVLPDLSGWSCSSGHKVKTTKVTRTAFA3Rat SEQ ID NO: 166LGEECKVLPDLSGWSCSRGHKVKTTKVTRTAFA3Mouse SEQ ID NO: 167LGEECKVLPDLSGWSCSSGHKVKTTKVTRTAFA3Chicken SEQ ID NO: 168AGEECKVLPDLSGWSCSSGNKVKTTKVTRTAFA3Komodo dragon, Wall lizard, Fencee lizard SEQ ID NO: 169EGEECKVLPDLSGWSCSTGNKVKTTKVTRTAFA3Frog SEQ ID NO: 170EGEDCKVLPDLSGWSCSTGNKVKTTKVTRTAFA3Fish SEQ ID NO: 171DGEECKVLPDLTGWSCSTGNKVKTTKVTR,

[0645] Interspecific sequence of fragment 2 of TAFA1 to 3 TAFA species SEQ ID NO Sequence, N'→ C' TAFA1 Human, Monkey, Pig, Rabbit, Rat, Mouse, Chicken, Komodo dragon, Wall lizard, Fence lizard, Frog SEQ ID 172 GKVAGTTRNRPSCVDASIVIGKWWCEMEPCL TAFA1 Fish SEQ ID 173 GKVAGTTRNKPSCVDASIVIGKWWCEMEPCL TAFA2 Human, Monkey, Pig, Rabbit, Rat, Mouse, Chicken, Komodo dragon, Wall lizard, Fence Lizard SEQ ID NO: 174GQVAGTTRAAPSCVDASIVEQKWWCHMQPCLTAFA2Gecko SEQ ID NO: 175GQVAGTTRAAPSCVDASIVEQKWWCQMQPCLTAFA2Frog SEQ ID NO: 176GQVAGTTRATPSCVDASIVEQKWWCHMQPCLTAFA2Fish SEQ ID NO: 177GQVAGTTRAAPSCVDASIVAQKWWCQMQPCMTAFA3Human, Monkey, Rabbit SEQ ID NO: 178GQVAGTTRAKPSCVDASIVLQRWWCQMEPCLTAFA3Pig, Rat, Mouse SEQ ID NO: 179GQVAGTTRAKPSCVDASIVLQKWWCQMEPCLTAFA3Chicken SEQ ID NO: 180GQVAGTTRAAPSCVDASIVLQKWWCQMEPCLTAFA3Komodo dragon, Fencee lizard SEQ ID NO: 181GQVAGTTRAAPSCVDASIVLQKWWCQMQPCLTAFA3Wall lizard SEQ ID NO: 182GQVAGTTHAAPSCVDASIVLQKWWCQMQPCVTAFA3Frog SEQ ID NO: 183GQVAGTTRAAPSCVDASIVIQKWWCQMEPCLTAFA3Fish SEQ ID NO: 184GQVAGTTRAAPSCVDASIVAQKWWCQMQPCV

[0646] All publications, patents, patent applications, and other documents cited in this application are incorporated herein by reference in their entirety for all purposes, even if each individual publication, patent, patent application, or other document was individually indicated to be incorporated by reference for all purposes.

[0647] While the present disclosure has been described with reference to the aforementioned aspects, those skilled in the art will appreciate that various modifications and variations are possible by adding, altering, deleting, or inserting components within the scope of the present disclosure as defined in the claims. It will be understood that such modifications and variations fall within the scope of the present disclosure.

Claims

1. A polypeptide having the ability to increase the length of neurites or branch points, The above polypeptide is (i) consisting of a sequence of 8 to 61 amino acids, and (ii) A polypeptide characterized by comprising a sequence having at least 50% sequence identity to the amino acid sequence of SEQ ID NO: 1 or the amino acid sequence of SEQ ID NO:

15.

2. In the first paragraph, the amino acid sequence of the polypeptide is characterized by including the amino acid sequence of the following general formula 1 (from the N-terminus to the C-terminus): <General formula 1> X1-GE-X2-CK-X3-L In the above general formula 1 X1 is E, D, P, L or A, X2 is D or E, and X3 is T, V, I, or A.

3. A polypeptide according to claim 2, characterized in that the amino acid sequence of the polypeptide comprises the amino acid sequence of SEQ ID NO:

142.

4. A polypeptide according to claim 2, characterized in that the amino acid sequence of the polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 14.

5. A polypeptide according to claim 2, characterized in that the amino acid sequence of the polypeptide consists of 8 to 43 amino acid sequences.

6. In the first paragraph, a polypeptide characterized in that the amino acid sequence of the polypeptide comprises an amino acid sequence of the following general formula 2 (from the N-terminus to the C-terminus): <General formula 2> IV-X4-X5-X6-WWC-X7-M-X8-PC-X9-X1-GE-X2-CK-X3-L In the above general formula 2 X1 is E, D, P, L or A, X2 is D or E, X3 is T, V, I or A, X4 is I, E, A, L or V, X5 is Q, E or G, X6 is K or R, X7 is E, H or Q, X8 is E, Q, N, D, S or H, and X9 is L, M or V.

7. A polypeptide according to claim 6, characterized in that the amino acid sequence of the polypeptide comprises the amino acid sequence of SEQ ID NO:

143.

8. A polypeptide according to claim 6, characterized in that the amino acid sequence of the polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 28 to 51.

9. In the first paragraph, the amino acid sequence of the polypeptide is characterized by including the amino acid sequence of the following general formula 3 (from the N-terminus to the C-terminus): <General formula 3> X1-GE-X2-CK-X3-LPD-X4-X5-GWSCS-X6-GNK-X7-KTTKVTR In the above general formula 3 X1 is E, D, P, L or A, X2 is D or E, X3 is T, V, I or A, X4 is Y, S or L, X5 is S or T, X6 is S or T, and X7 is V or I.

10. A polypeptide according to claim 9, characterized in that the amino acid sequence of the polypeptide comprises the amino acid sequence of SEQ ID NO:

144.

11. A polypeptide according to claim 9, characterized in that the amino acid sequence of the polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 52 to 58.

12. In the first paragraph, a polypeptide characterized in that the amino acid sequence of the polypeptide comprises an amino acid sequence of the following general formula 4 (from the N-terminus to the C-terminus): <General Formula 4> G-X1-VAGTT-X2-X3-X4-PSCV-X5-X6-X7 In the above general formula 4 X1 is Q or K, X2 is R, H or Q, X3 is A, N, S or T, X4 is R, A, Q, K, or T, X5 is D or E, X6 is A or not present, and X7 is either S, A or non-existent.

13. A polypeptide according to claim 12, characterized in that the amino acid sequence of the polypeptide comprises the amino acid sequence of SEQ ID NO:

145.

14. A polypeptide according to claim 12, characterized in that the amino acid sequence of the polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 15 to 27.

15. A polypeptide according to claim 12, characterized in that the amino acid sequence of the polypeptide consists of a sequence of 15 to 46 amino acids.

16. In the first paragraph, a polypeptide characterized in that the amino acid sequence of the polypeptide comprises the amino acid sequence of the following general formula 5 (from the N-terminus to the C-terminus): <General Formula 5> X8-IEE-X9-SQT-X10-X11-CSC-X12-X13-G-X1-VAGTT-X2-X3-X4-PSCV-X5-X6-X7 In the above general formula 5 X1 is Q or K, X2 is R, H or Q, X3 is A, N, S or T, X4 is R, A, Q, K, or T, X5 is D or E, X6 is A or does not exist, X7 is S, A or not present, X8 is R or K, X9 is R or L, X10 is V or G, X11 is K or N, X12 is F or L, and X13 is P or S.

17. A polypeptide according to claim 16, characterized in that the amino acid sequence of the polypeptide comprises the amino acid sequence of SEQ ID NO:

146.

18. A polypeptide according to claim 16, characterized in that the amino acid sequence of the polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 59 to 74.

19. In the first paragraph, a polypeptide characterized in that the amino acid sequence of the polypeptide comprises the amino acid sequence of the following general formula 6 (from the N-terminus to the C-terminus): <General Formula 6> G-X1-VAGTT-X2-X3-X4-PSCV-X5-X6-X7-IV-X8-X9-KWWC-X10-M-X11-PC-X12 In the above general formula 6 X1 is Q or K, X2 is R, H or Q, X3 is A, N, S or T, X4 is R, A, Q, K or T, X5 is D or E, X6 is A or does not exist, X7 is S, A or not present, X8 is I, A, V or L, X9 is Q or E, X10 is H or Q, X11 is N, D, S or H, and X12 is L or M.

20. A polypeptide according to claim 19, characterized in that the amino acid sequence of the polypeptide comprises the amino acid sequence of SEQ ID NO:

147.

21. A polypeptide according to claim 19, characterized in that the amino acid sequence of the polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 75 to 85.

22. A polypeptide according to claim 1, characterized in that the amino acid sequence of the polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 152 to 184.

23. A nucleic acid molecule encoding a polypeptide according to any one of claims 1 to 22.

24. A vector comprising the nucleic acid molecule of item 23.

25. A recombinant viral particle comprising the vector and capsid protein of claim 24.

26. A recombinant viral particle according to claim 25, characterized in that the virus is AAV.

27. A cell containing the vector of item 24.

28. Cells transformed with the vector of Article 24.

29. A composition comprising a polypeptide according to any one of claims 1 to 22, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

30. A composition according to claim 29, characterized in that the composition is a pharmaceutical composition.

31. A composition according to claim 30, characterized in that the pharmaceutical composition is a composition for preventing or treating retinal neuropathies.

32. A composition according to claim 31, characterized in that the retinal neurodegenerative disease comprises retinopathy, choroidal neovascularization, macular disease, macular degeneration, retinal degeneration, macular edema, retinal edema, macular edema, retinal edema, color vision abnormality, retinal cell degeneration, retinal vascular occlusion, retinal detachment, hereditary retinal disease, or a combination thereof.

33. A composition according to claim 30, characterized in that the pharmaceutical composition is a composition for preventing or treating neuropathic pain.

34. A composition according to claim 33, wherein the neuropathic pain is characterized by allodynia, hyperalgesia, hyperesthesia or dysphagia.

35. A composition according to claim 33, characterized in that the neuropathic pain is central neuropathic pain or peripheral neuropathic pain.

36. A method for producing a composition, comprising the step of producing a composition comprising a polypeptide of any one of claims 1 to 22, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

37. A therapeutic use for the manufacture of a medicament comprising a polypeptide according to any one of claims 1 to 22, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

38. A method for preventing or treating a disease or disorder in a subject in need thereof, comprising administering to the subject a composition comprising a polypeptide according to any one of claims 1 to 22, a nucleic acid molecule encoding the polypeptide, a vector comprising the nucleic acid molecule, a recombinant viral particle comprising the vector and a capsid protein, a cell comprising the vector, a cell transformed with the vector, or a combination thereof.

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