Uses of anti-FAM19A5 antibodies

Abstract

To provide a pharmaceutical composition for treating retinopathy (e.g., diabetic retinopathy) and / or maculopathy (e.g., age-related macular degeneration) in a subject who needs the composition.SOLUTION: Provided is a pharmaceutical composition containing an antibody which specifically binds to FAM19A5 protein or an antigen-binding portion thereof (anti FAM19A5 antibody), a polynucleotide encoding the anti FAM19A5 antibody, or a vector including a polynucleotide thereof, and is used for improving B-wave retinal potential on a retina compared to a standard.SELECTED DRAWING: None

Description

Detailed description of the invention

[0001] [Technical field] Cross-references to related applications This PCT application claims priority to U.S. Extraordinary Application No. 62 / 746,194, filed on 16 October 2018, which is incorporated herein by reference in its entirety.

[0002] References for electronically submitted sequence catalogs The contents of the sequence listing electronically submitted with this application as an ASCII text file (name: 3763.012PC01_SeqListing_ST25.txt; size: 261,609 bytes; and generation date: October 13, 2019) are incorporated herein by reference in their entirety.

[0003] This disclosure relates to a method for treating retinal diseases (e.g., diabetic retinopathy) and / or macular degeneration (e.g., age-related macular degeneration) in a subject (e.g., human) using an antagonist to a family member A5 (FAM19A5) protein having sequence similarity 19, and to a composition comprising said antagonist. [Background technology] Diabetic retinopathy (DR), also known as diabetic eye disease, is caused by damage to the small blood vessels and nerve cells of the retina. DR is the leading cause of vision loss worldwide among the working-age population (e.g., 20-74 years). Lee R., et al., Eye Vis (Lond) 2:17 (2015). As the expected lifespan of people with diabetes increases, the incidence of DR has surged over the past few years. Generally, about 50% of people with diabetes experience DR progression 10 years after the onset of diabetes, 70% after 20 years, and 90% after 30 years. The number of people with diabetes worldwide in the 20-79 age group is expected to nearly double by 2030, suggesting a further increase in the number of individuals diagnosed with DR. Nentwich MM and Ulbig MW, World J Diabetes 6(3):489-499 (2015).

[0004] Several treatment options are available that may help manage DR-related symptoms, but they do not cure the disease itself and may have undesirable side effects. For example, laser therapy can cause significant inconvenience during treatment, permanent retinal scarring, and decreased peripheral vision, color vision, and night vision. See worldweb.aao.org / munnerlyn-laser-surgery-center / laser-treatment-of-proliferative-nonproliferative- for more information. For this reason, alternative treatment options for diabetic retinopathy are currently desired.

[0005] Age-related macular degeneration (AMD) is another retinal disease that affects the eye, more specifically the central part of the eye (i.e., the macula). AMD is thought to be caused by hardening of the arteries that supply nutrients to the retina, depriving the retinal tissue of oxygen and leading to a decline in central vision. Gheorghe A., et al., Rom J Ophthalmol 59(2):74-77 (2015). AMD is the most common type of macular degeneration, affecting approximately 11 million individuals in the United States alone, and has a prevalence of 170 million people worldwide. AMD is the leading cause of visual impairment in industrialized countries and the third leading cause globally. Pennington KLand De Angelis M., Eye Vis (Lond) 3:34 (2016). Aging is the greatest risk factor, and the prevalence of AMD in the United States is projected to increase to 22 million by 2050, while the global prevalence is projected to increase to 288 million by 2040. Wong WL, et al., Lancet Glob Health 2:el06-ell6 (2014).

[0006] AMD, when diagnosed early, can be managed with a variety of treatment options, including vitamin supplements (e.g., antioxidants), dietary changes (e.g., low-fat diets), anti-angiogenic drugs (e.g., anti-VEGF antibodies), laser therapy, surgery (e.g., submacular surgery), and retinal repositioning. Holz FG, et al., J Clin Invest 124(4):1430-1438 (2014). However, there is no known cure for AMD, and most treatment options are not feasible for all AMD patients. For example, many patients who develop choroidal neovascularization (CNV) (i.e., wet AMD) are not eligible for laser therapy because the CNV is too large or its location cannot be accurately determined. Moreover, even with treatment, many AMD patients experience progressive central vision loss. Virgili G. and Bini A., Cochrane Database Syst Rev 3:CD004763 (2007). [Overview of the prefecture] [Problems the invention aims to solve] Therefore, there is a need for more effective and comprehensive treatment options for AMD. [Means for solving the problem] This specification provides a method for improving the retinal potential in the retina of a subject of interest, comprising administering an antagonist ("FAM19A5 antagonist") to a family member A5 ("FAM19A5") protein having sequence similarity 19, compared to a baseline (e.g., the relevant value for the subject before administration of the FAM19A5 antagonist or for a subject without administration of the FAM19A5 antagonist). In some embodiments, the improvement in the retinal potential includes an increase in wave A, wave B, oscillatory potential, or any combination thereof.

[0007] In some embodiments, the subject has a disease or condition related to decreased retinoid potential in the retina. In certain embodiments, the disease or condition is diabetic retinopathy. In other embodiments, the disease or condition is age-related macular degeneration of the retina.

[0008] This specification also provides a method for treating retinal and / or macular diseases in a subject requiring treatment, comprising administering a FAM19A5 antagonist to the subject.

[0009] In some embodiments, the retinal disease is diabetic retinal disease. In some embodiments, the diabetic retinal disease is nonproliferative diabetic retinal disease (NPDR). In other embodiments, the diabetic retinal disease is proliferative diabetic retinal disease. In certain embodiments, the diabetic retinal disease is associated with decreased retinal potential, pericyte loss, acellular capillary formation, vascular congestion, or any combination thereof within the retina of the subject.

[0010] In some embodiments, the macular disease is diabetic macular edema. In certain embodiments, the diabetic macular edema is associated with decreased retinoid potential, perivascular cell loss, acellular capillary formation, vascular congestion, or any combination thereof within the retina of the subject.

[0011] In some embodiments, the FAM19A5 antagonist improves the electroretinometric potential of a control subject compared to a baseline (e.g., the relevant value of a subject before administration of the FAM19A5 antagonist or a subject without administration of the FAM19A5 antagonist). In certain embodiments, the improvement in electroretinometric potential includes an increase in A waves, B waves, rhythmic ripples, or any combination thereof. In some embodiments, the FAM19A5 antagonist reduces the loss of intraretinal perivascular cells in the subject. In some embodiments, the FAM19A5 antagonist reduces and / or inhibits intraretinal acellular capillary formation in the subject. In other embodiments, the FAM19A5 antagonist reduces and / or inhibits vascular congestion within the retinal nerve fiber layer of the subject.

[0012] The disclosure also provides a method for treating macular degeneration in a subject requiring treatment, the method comprising administering an antagonist to the FAM19A5 protein to the subject.

[0013] In some embodiments, the macular degeneration is age-related macular degeneration. In certain embodiments, the age-related macular degeneration is dry (atrophic) macular degeneration. In other embodiments, the age-related macular degeneration is wet (neovascular or exudative) macular degeneration. In certain embodiments, the age-related macular degeneration is (i) early AMD, (ii) mid-stage AMD, or (iii) late or progressive AMD (geographic atrophy). In other embodiments, the age-related macular degeneration is associated with the accumulation or aggregation of intraretinal drusen, neovascularization, decreased retinoid potential, astrocytosis, or any combination thereof.

[0014] In some embodiments, the FAM19A5 antagonist reduces and / or inhibits angiogenesis in the retina of a subject, as evidenced by a decrease in CD31 and / or VEGF expression. In certain embodiments, the FAM19A5 antagonist can reduce and / or inhibit angiogenesis in the retina of mice with laser-induced choroidal angiogenesis, as evidenced by a decrease in CD31 and / or VEGF expression. In other embodiments, the FAM19A5 antagonist reduces the expression of platelet-derived growth factor (PDGF) in the retinal nerve fiber layer and ganglion cell layer of a control subject compared to a baseline (e.g., the relevant value of the subject before administration of the FAM19A5 antagonist or the subject without administration of the FAM19A5 antagonist). In other embodiments, the FAM19A5 antagonist improves the electroretinomytectomy of a control subject compared to a baseline (e.g., the relevant value of the subject before administration of the FAM19A5 antagonist or the subject without administration of the FAM19A5 antagonist). In some embodiments, the improvement in electroretinomytectomy includes an increase in the B wave. In certain embodiments, the FAM19A5 antagonist can reduce and / or inhibit the accumulation or aggregation of target intraretinal drusen.

[0015] In some embodiments, the FAM19A5 antagonist is an antisense oligonucleotide, siRNA, shRNA, miRNA, dsRNA-targeted FAM19A5, aptamer, PNA, or a vector comprising the same.

[0016] In some embodiments, the FAM19A5 antagonist is an antibody or its antigen-binding portion that specifically binds to the FAM19A5 protein ("anti-FAM19A5 antibody"). In certain embodiments, the anti-FAM19A5 antibody is: (a) when measured by enzyme-linked immunosorbent assay (ELISA) D (b) The property of binding to soluble human FAM19A5 when measured by ELISA; (b) K D The property of binding to membrane-bound human FAM19A5 with a concentration of 10 nM or less; or (c) exhibiting one of either (a) or (b).

[0017] In some embodiments, the anti-FAM19A5 antibody cross-competes with a reference antibody selected from the group consisting of antibodies in Tables 2 to 5 in order to bind to the human FAM19A5 epitope. In a particular embodiment, the reference antibody comprises heavy chain CDR1, CDR2 and CDR3 and light chain CDR1, CDR2 and CDR3, wherein (i) the heavy chain CDR1 comprises the amino acid sequence shown in SEQ ID NO: 14, the heavy chain CDR2 comprises the amino acid sequence shown in SEQ ID NO: 15, the heavy chain CDR3 comprises the amino acid sequence shown in SEQ ID NO: 16, the light chain CDR1 comprises the amino acid sequence shown in SEQ ID NO: 26, the light chain CDR2 comprises the amino acid sequence shown in SEQ ID NO: 27, and the light chain CDR3 comprises the amino acid sequence shown in SEQ ID NO: 28; or (ii) the heavy chain CDR1 comprises the amino acid sequence shown in SEQ ID NO: 17, the heavy chain CDR2 comprises the amino acid sequence shown in SEQ ID NO: 18, the heavy chain CDR3 comprises the amino acid sequence shown in SEQ ID NO: 19, the light chain CDR1 comprises the amino acid sequence shown in SEQ ID NO: 29, the light chain CDR2 comprises the amino acid sequence shown in SEQ ID NO: 30, and the light chain CDR3 comprises the amino acid sequence shown in SEQ ID NO: 31.

[0018] In some embodiments, the anti-FAM19A5 antibody specifically binds to the same FAM19A5 epitope as a reference antibody selected from the group consisting of antibodies in Tables 2 to 5. In a particular embodiment, the reference antibody comprises heavy chain CDR1, CDR2 and CDR3 and light chain CDR1, CDR2 and CDR3, wherein (i) the heavy chain CDR1 comprises the amino acid sequence shown in SEQ ID NO: 14, the heavy chain CDR2 comprises the amino acid sequence shown in SEQ ID NO: 15, the heavy chain CDR3 comprises the amino acid sequence shown in SEQ ID NO: 16, the light chain CDR1 comprises the amino acid sequence shown in SEQ ID NO: 26, the light chain CDR2 comprises the amino acid sequence shown in SEQ ID NO: 27, and the light chain CDR3 comprises the amino acid sequence shown in SEQ ID NO: 28; or (ii) the heavy chain CDR1 comprises the amino acid sequence shown in SEQ ID NO: 17, the heavy chain CDR2 comprises the amino acid sequence shown in SEQ ID NO: 18, the heavy chain CDR3 comprises the amino acid sequence shown in SEQ ID NO: 19, the light chain CDR1 comprises the amino acid sequence shown in SEQ ID NO: 29, the light chain CDR2 comprises the amino acid sequence shown in SEQ ID NO: 30, and the light chain CDR3 comprises the amino acid sequence shown in SEQ ID NO: 31.

[0019] In some embodiments, the anti-FAM19A5 antibody specifically binds to at least one FAM19A5 epitope, which is sequence number 6 or sequence number 9.

[0020] In some embodiments, the anti-FAM19A5 antibody comprises heavy chain CDR1, CDR2 and CDR3 and light chain CDR1, CDR2 and CDR3, wherein (i) the heavy chain CDR1 comprises a CDR1 selected from the group consisting of CDR1 shown in Table 2; (ii) the heavy chain CDR2 comprises a CDR2 selected from the group consisting of CDR2 shown in Table 2; (iii) the heavy chain CDR3 comprises a CDR3 selected from the group consisting of CDR3 shown in Table 2; (iv) the light chain CDR1 comprises a CDR1 selected from the group consisting of CDR1 shown in Table 3; (v) the light chain CDR2 comprises a CDR2 selected from the group consisting of CDR2 shown in Table 3; and / or (vi) the light chain CDR3 comprises a CDR3 selected from the group consisting of CDR3 shown in Table 3.

[0021] In some embodiments, the anti-FAM19A5 antibody comprises heavy chain CDR1, CDR2 and CDR3 and light chain CDR1, CDR2 and CDR3, wherein (i) the heavy chain CDR1 comprises the amino acid sequence shown in SEQ ID NO: 14 or SEQ ID NO: 17; (ii) the heavy chain CDR2 comprises the amino acid sequence shown in SEQ ID NO: 15 or SEQ ID NO: 18; (iii) the heavy chain CDR3 comprises the amino acid sequence shown in SEQ ID NO: 16 or SEQ ID NO: 19; (iv) the light chain CDR1 comprises the amino acid sequence shown in SEQ ID NO: 26 or SEQ ID NO: 29; (v) the light chain CDR2 comprises the amino acid sequence shown in SEQ ID NO: 27 or SEQ ID NO: 30; and / or (vi) the light chain CDR3 comprises the amino acid sequence shown in SEQ ID NO: 28 or SEQ ID NO: 31.

[0022] In some embodiments, the anti-FAM19A5 antibody comprises heavy chain CDR1, CDR2, and CDR3 and light chain CDR1, CDR2, and CDR3, wherein the heavy chain CDR1, CDR2, and CDR3 each comprise SEQ ID NOs. 14, 15, and 16, and the light chain CDR1, CDR2, and CDR3 each comprise SEQ ID NOs. 26, 27, and 28. In other embodiments, the anti-FAM19A5 antibody comprises heavy chain CDR1, CDR2, and CDR3 and light chain CDR1, CDR2, and CDR3, wherein the heavy chain CDR1, CDR2, and CDR3 each comprise SEQ ID NOs. 17, 18, and 19, and the light chain CDR1, CDR2, and CDR3 each comprise SEQ ID NOs. 29, 30, and 31.

[0023] In some embodiments, the anti-FAM19A5 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein (i) the VH comprises the amino acid sequence shown in SEQ ID NO: 36 or SEQ ID NO: 37; or (ii) the VL comprises the amino acid sequence shown in SEQ ID NO: 40 or SEQ ID NO: 41. In a particular embodiment, the VH comprises the amino acid sequence of SEQ ID NO: 36, and the VL comprises the amino acid sequence shown in SEQ ID NO: 40. In other embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 37, and the VL comprises the amino acid sequence shown in SEQ ID NO: 41.

[0024] In some embodiments, the anti-FAM19A5 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH contains an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% identical to the amino acid sequence shown in SEQ ID NO: 36 or SEQ ID NO: 37. In some embodiments, the anti-FAM19A5 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VL contains an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% identical to the amino acid sequence shown in SEQ ID NO: 40 or SEQ ID NO: 41.

[0025] In some embodiments, the anti-FAM19A5 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein (i) the VH comprises an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% the same as the amino acid sequence shown in SEQ ID NO: 36 or SEQ ID NO: 37; and (ii) the VL comprises an amino acid sequence that is at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% the same as the amino acid sequence shown in SEQ ID NO: 40 or SEQ ID NO: 41.

[0026] In some embodiments, the anti-FAM19A5 antibody comprises Fab, Fab', F(ab')2, Fv, or single-chain Fv(scFv). In certain embodiments, the anti-FAM19A5 antibody is selected from the group consisting of IgG1, IgG2, IgG3, IgG4, and their variants. In some embodiments, the anti-FAM19A5 antibody is IgG2, IgG4, or a combination thereof. In other embodiments, the anti-FAM19A5 antibody comprises an IgG2 / IgG4 isotype antibody. In some embodiments, the anti-FAM19A5 antibody comprises a constant region without Fc function. In some embodiments, the anti-FAM19A5 antibody is a chimeric antibody, a human antibody, or a humanized antibody.

[0027] In some embodiments, the FAM19A5 antagonist is linked to a molecule having a second binding site to form a bispecific molecule. In some embodiments, the FAM19A5 antagonist is linked to an agent to form an immunoconjugate.

[0028] In some embodiments, the FAM19A5 antagonists disclosed herein are formulated with a pharmaceutically acceptable carrier. In some embodiments, the FAM19A5 antagonist is administered intraocularly. In certain embodiments, the intraocular administration includes intravitreal administration. In some embodiments, the FAM19A5 antagonist is administered intravenously.

[0029] In some embodiments, the methods disclosed herein further include administering a therapeutic agent. In some embodiments, the subject treated in this disclosure is a human being.

[0030] This disclosure also provides a pharmaceutical composition for the prevention or treatment of retinal and / or macular diseases, comprising a pharmaceutically effective amount of an antagonist against the FAM19A5 protein. [Brief description of the drawing] Figures 1A, 1B, and 1C show the effect of anti-FAM19A5 antibody on retinoid potential recovery in 25-week-old db / db mice. These mice were treated with either human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clones "1-65" or "3-2"). Normal (i.e., non-diabetic) mice were used as the control group. Mice in each group were treated with three different flash intensities (-1, 0.2, and 0.8 log cds / m²). 2 After exposure to white light, wave A (μV) (Figure 1A), wave B (μV) (Figure 1B), and rhythmic small wave (μV) (Figure 1C) were measured. Data are expressed as mean ± SD.

[0031] Figures 2A and 2B demonstrate the protective effect of anti-FAM19A5 antibody on retinal microvascular tissue in 25-week-old db / db mice. These mice were treated with either human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clone "1-65" or "3-2"). Normal (i.e., non-diabetic) healthy mice were used as the control group. Figure 2A shows micrographs (100× magnification) of representative retinal regions of mice after periodic acid Schiff (PAS) staining for each of the different treatment groups: (i) normal group (i.e., non-diabetic) (upper left panel), (ii) human IgG control group (upper right panel), and (iii) anti-FAM19A5 antibody (clone 1-65, lower left panel; clone 3-2, lower right panel). Black arrows indicate endothelial cells, white arrows indicate perivascular cells, and gray arrows indicate acellular capillaries. Figure 2B shows a comparison of the ratio of endothelial cells to perivascular cells ("E / P ratio") observed within the entire retinal region analyzed in mice from the different treatment groups. To determine the E / P ratio, the number of endothelial cells and perivascular cells was calculated in each photograph. Data are expressed as mean ± SD. "***" on the bars indicates a statistically significant difference (p<0.001) compared to the IgG treatment group.

[0032] Figure 2C shows a comparison of the number of acellular capillaries observed within the analyzed whole retinal region of mice from different treatment groups. Data are expressed as mean ± SD. An asterisk (*) on the bar indicates a statistically significant difference (p<0.05) compared to the IgG treatment group.

[0033] Figure 2D shows a comparison of observed acellular capillary lengths. Data are expressed as mean ± SD. "**" and "***" on the bars indicate statistically significant differences compared to the IgG treatment group (p<0.01 and p<0.001, respectively).

[0034] Figure 3 shows the effect of anti-FAM19A5 antibody treatment on intraretinal vascular congestion in 25-week-old db / db mice using H&E staining. The mice were treated with human IgG control antibody ("IgG", upper right panel) or anti-FAM19A5 antibody (clone "1-65", lower left panel or clone "3-2", lower right panel). Normal (i.e., diabetes-free) healthy mice (upper left panel) were used as the control group. Arrowheads indicate examples of vascular congestion. The different retinal layers shown include (i) ganglion cell layer (GCL), (ii) internal plexiform layer (IPL), (iii) inner nuclear layer (INL), (iv) outer nuclear layer (ONL), and (v) retinal pigment epithelium (PE).

[0035] Figure 4 shows the retinal cell protective effect of anti-FAM19A5 antibody treatment in 25-week-old db / db mice using TUNEL analysis. The mice were treated with human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clones "1-65" or "3-2"). Normal (i.e., diabetes-free) healthy mice were used as a control group. In the middle column ("TUNEL"), red arrowheads indicate retinal cells undergoing cell death. The intensity of TUNEL staining correlates with the degree of cell death (e.g., decreased intensity indicates partial cell death). The different retinal layers shown include (i) ganglion cell layer (GCL), (ii) inner nucleus layer (INL), (iii) outer nucleus layer (ONL), and (iv) retinal pigment epithelium (PE).

[0036] Figure 5 shows the inhibitory effect of anti-FAM19A5 antibody on retinal neovascularization in 25-week-old db / db mice using immunohistochemistry staining for CD31 and VEGF expression. The mice were treated with human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clones "1-65" or "3-2"). Normal (i.e., diabetes-free) healthy mice were used as the control group. Arrowheads indicate positive CD31 (top row) or positive VEGF (bottom row) staining.

[0037] Figure 6A shows a photograph of an image-guided laser system like the one used in Example 8. Figure 6B shows an example of a photograph of a retina exposed to the laser from the image-guided laser system shown in Figure 6A.

[0038] Figures 7A and 7B show the inhibitory effect of anti-FAM19A5 antibody (clone 3-2) on choroidal neovascularization (CNV) using fluorescein angiography (FFA) analysis. CNV-inducible mice treated with human IgG antibody were used as a control group. Figure 7A shows representative retinal images of mice treated with human IgG antibody (top row) or anti-FAM19A5 (clone 3-2) (bottom row) on the day of CNV induction ("day 0") and 13 days after CNV induction, showing four wet AMD-like lesions (indicated by arrows in the left column). Figure 7B provides a comparison of corrected total fluorescence (CTF) values ​​for CNV-inducible mice treated with human IgG antibody ("IgG") or anti-FAM19A5 antibody (clone "3-2") 13 days after CNV induction. Data are presented as means and individually. "*" indicates a statistically significant difference compared to the IgG-treated group (p<0.05).

[0039] Figures 8A, 8B, and 8C demonstrate the inhibitory effect of anti-FAM19A5 antibody (clone 3-2) on choroidal neovascularization (CNV) using optical coherence tomography (OCT) analysis. CNV-inducible mice administered with human IgG antibody were used as a control group. Figure 8A provides representative retinal images showing four wet AMD-like lesions (labeled 1, 2, 3, and 4) in mice treated with human IgG antibody (top row) or anti-FAM19A5 (clone 3-2) (bottom row) 13 days after CNV induction. The columns labeled 1, 2, 3, and 4 provide OCT images of the lesions shown in the leftmost image. Figure 8B shows the scale bars used to quantify the size of the lesions. Figure 8C provides a comparison of the size of CNV lesions observed 13 days after CNV induction in mice treated with human IgG control antibody or anti-FAM19A5 antibody (clone 3-2). Data are presented as averages and individually. "***" indicates a statistically significant difference (p<0.001) compared to the IgG treatment group.

[0040] Figure 9 shows the effect of anti-FAM19A5 antibody on the recovery of B-wave values ​​in CNV-inducing mice. The CNV-inducing mice were treated with either human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clone "3-2"). Normal (i.e., non-diabetic) mice were used as the control group. Data are expressed as mean ± SD. "*" indicates a statistically significant difference compared to the IgG-treated group (p<0.05).

[0041] Figures 10A and 10B show the inhibitory effect of anti-FAM19A5 antibody (clone 3-2) on choroidal neovascularization (CNV). Figure 10A identifies different retinal layers using H&E analysis (left image) or OCT analysis (right image). These shown layers include: (i) retinal nerve fiber layer / ganglion cell layer (RNFL / GCL); (ii) inner plexiform layer; (iii) inner plexiform layer; (iv) outer plexiform layer; (v) outer plexiform layer; (vi) photoreceptor inner / outer segment (IS / OS); and (vii) retinal pigment epithelium (RPE). Figure 10B shows representative H&E images of the relevant cell layers of the mouse retina in different groups: (i) normal healthy animals, (ii) CNV-induced mice treated with human IgG antibody, and (iii) CNV-induced mice treated with anti-FAM19A5 antibody. Arrowheads indicate areas of choroidal neovascularization.

[0042] Figure 11 shows the inhibitory effect of anti-FAM19A5 antibody on PDGF expression in both the nerve fiber layer and ganglion cell layer of the retina, as demonstrated by IHC analysis. CNV-inducing mice were treated with human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clone "3-2"). Normal (i.e., non-diabetic) mice were used as the control group. Arrowheads indicate positive PDGF staining.

[0043] Figure 12 shows the inhibitory effect of anti-FAM19A5 antibody on oligomeric Aβ and drusen formation in APP / PS-1 / tau gene-transfected mice (dementia model) using immunofluorescence microscopy. The APP / PS-1 / tau gene-transfected mice were either untreated (middle row) or treated (bottom row) with anti-FAM19A5 antibody. Untreated (naive) wild-type C57BL / 6 mice were used as the negative control group (top row). The column labeled "H33342" (nucleic acid staining) shows cells within retinal tissue. The column labeled "QD525-oAβ" shows oligomeric Aβ. The column labeled "647 Alexa Phalloidin" shows drusen formation, with drusen illustrated by arrows. The column labeled "merged" shows a composite image of the top three columns for each treatment group.

[0044] Figures 13A–13C show a comparison of the retinal microvascular protective effects of anti-FAM19A5 antibodies (clones 1–30) after intravitreous or intravenous administration in a mouse model of diabetic retina. The mice were administered either (i) human IgG control antibody ("human IgG"), (ii) aflibercept, or (iii) anti-FAM19A5 antibody intravitreously ("IVT"), or (iv) anti-FAM19A5 antibody intravenously ("IV"). Normal (i.e., non-diabetic) healthy mice were used as the control group. Figure 13A provides micrographs (100× magnification) of representative retinal regions of mice in each of the different treatment groups after periodate Schiff (PAS) staining. Black arrows indicate endothelial cells, white arrows indicate perivascular cells, and black arrows indicate acellular capillaries. Figure 13B provides a comparison of the number of acellular capillaries observed within the analyzed overall retinal region of mice from the different treatment groups. Figure 13C provides a comparison of the ratio of endothelial cells to perivascular cells ("E / P ratio") observed within the analyzed whole retinal region of mice from different treatment groups. To determine the E / P ratio, the number of endothelial cells and perivascular cells was calculated in each photograph. In Figures 13B and 13C, data are expressed as mean ± SD. "*" and "***" on the bars indicate statistically significant differences compared to untreated mice (p<0.05 and p<0.001, respectively). "##" and "###" on the bars indicate statistically significant differences compared to the human IgG antibody treatment group (p<0.01 and p<0.001, respectively). "†††" on the bars indicate a statistically significant difference compared to the aflibercept treatment group (p<0.001). "§" on the bars indicates a statistically significant difference compared to the IVT group (p<0.05).

[0045] Figure 14 shows the inhibitory effect of anti-FAM19A5 antibody on intraretinal vascular congestion formation in diabetic retinal mice after intravenous and intravitreous administration. The treatment groups are the same as those described in Figures 13A to 13C. Arrowheads indicate examples of vascular congestion.

[0046] Figure 15 shows the retinal cell protective effect of anti-FAM19A5 antibody measured using TUNEL analysis upon intravitreal or intravenous administration. The treatment groups are the same as those described in FIGS. 13A to 13C. In the middle column (“TUNEL”), the arrowheads indicate retinal cells in which cell death is progressing. The intensity of TUNEL staining is correlated with the degree of cell death (e.g., a decrease in intensity indicates some cell death).

[0047] FIGS. 16A and 16B show the inhibitory effect of anti-FAM19A5 antibody on retinal neovascularization in diabetic retinopathy mice after intravitreal and intravenous administration. The treatment groups are the same as those described in FIGS. 13A to 13C. Neovascularization was measured using VEGF (FIG. 16A) and CD31 (FIG. 16B) expression. The arrowheads indicate positive VEGF or CD31 staining. [Mode for Carrying Out the Invention] To facilitate understanding of the disclosure provided herein, a number of terms and phrases are defined. Additional definitions are set forth throughout the detailed description.

[0048] <I. Definitions>[ Throughout the present disclosure, the term “one” or “a” entity refers to one or more of that entity, e.g., “one antibody” is understood to represent one or more antibodies. Thus, the terms “one” (or “a”), “one or more,” and “at least one” are used herein with the same meaning.

[0049] Furthermore, as used herein, “and / or” should be considered to specifically disclose one of each of two identified features or components together with the other feature or component, or on its own. Accordingly, the term “and / or” as used herein in phrases such as “A and / or B” is intended to include “A and B,” “A or B,” “A” (alone), and “B” (alone). Similarly, the term “and / or” as used in phrases such as “A, B and / or C” is intended to include each of the following embodiments: A, B and C; A, B or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0050] In this specification, if an embodiment is described using the term “including,” it is understood that other similar embodiments described in terms of “being” and / or “essentially being” are also provided.

[0051] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as they would be commonly understood by a person of the ordinary skill in the art relating to this disclosure. 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 a majority of the terms used herein to a person of the skill.

[0052] Units, prefixes, and symbols are shown in the forms recognized by the Systeme International de Unites (SI). Numerical ranges include the digits that limit the range. Unless otherwise indicated, amino acid sequences are written from left to right, from amino to carboxy. The titles provided herein are not limitations on the various aspects of the disclosure, and may refer to the specification as a whole. Accordingly, the terms defined below are more fully defined by referring to the specification as a whole.

[0053] In this specification, the term “approximately” is used to mean roughly, to a certain extent, or within a given range. When the term “approximately” is used with a numerical range, it modifies the range by extending the boundaries above and below the stated numerical value. Generally, the term “approximately” can change the numerical values ​​above and below the specified value by, for example, a 10% change above or below (higher or lower).

[0054] As used herein, the term “retinal disease” refers to a disease, inflammation, or injury of the retina (i.e., the tissue surrounding the inner surface of the back of the eyeball that captures images passing through the cornea and lens).

[0055] As used herein, the term “diabetic retinopathy” (DR) refers to retinal diseases induced by complications associated with diabetes. Depending on the severity of the disease, DR may be asymptomatic, cause mild vision problems, or lead to blindness. DR is a result of microvascular retinal changes. Hyperglycemia-induced intramural perivascular cell death and basement membrane thickening can lead to vascular wall failure. This damage alters the formation of the blood-retinal barrier, making retinal vessels more permeable. Perivascular cell death can be induced when hyperglycemia sustainably activates protein kinase C-δ (PKC-δ encoded by Prkcd) and p38 mitogen-activated protein kinase (MAPK), increasing the expression of Src homology-2 domain-containing phosphatase-1 (SHP-1), a protein tyrosine phosphatase, as a previously unknown target of PKC-δ signaling. This signaling cascade can lead to PDGF receptor dephosphorylation and a decrease in downstream signaling from this receptor, potentially resulting in “perivascular cell death.” Small blood vessels, such as those in the eye, can be particularly vulnerable to dysregulation of blood glucose. Excessive accumulation of glucose and / or fructose can damage small blood vessels in the retina.

[0056] Diabetic retinopathy (DR) can be classified into two distinct stages. (Wu L., el al., World J Diabetes 4(6):290-294(2013)). Stage 1, non-proliferative diabetic retinopathy (NPDR), is associated with early diabetic retinopathy. NPDR is generally asymptomatic or associated with mild vision distortion caused by blood vessels leaking into surrounding tissues. The only way to examine NPDR is through fundus photography, which allows observation of microaneurysms (tiny bulges in the artery walls where blood accumulates). If left untreated, DR patients may progress to a more serious stage 2 called proliferative diabetic retinopathy (PDR). PDR is characterized by abnormal neovascularization (new blood vessel formation) that ruptures and bleeds, causing visual field blurring. Other symptoms of PDR include floating spots or dark lines in the field of vision ("floaters"), changes in visual acuity, color vision impairment, dark or empty areas in the field of vision, pain, variability in visual acuity, and complete vision loss.

[0057] The term “diabetic retinopathy” includes, but is not limited to, any type of diabetic retinopathy, including nonproliferative diabetic retinopathy (NPDR), proliferative diabetic retinopathy (PDR), diabetic macular disease, and diabetic macular edema.

[0058] In some embodiments, PDR occurs after the onset of NPRD (for example, initially diagnosed as NPRD, with the disease progressing to PDR). In other embodiments, PDR occurs independently of NPRD. As used herein, the term “diabetic retinopathy” also includes all types of diabetic retinopathy and any and all symptoms of diabetic retinopathy, regardless of cause. 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 injury.

[0059] As used herein, the term “macular disease” refers to any pathological condition of the macula, the central region of the retina associated with highly sensitive and accurate vision. In some embodiments, the terms “macular disease” and “retinal disease” are used interchangeably (i.e., when only the macula is affected). In some embodiments, the macular disease is diabetic macular disease.

[0060] "Diabetic macular disease" occurs when the macula is affected by retinal changes caused by diabetes. The term refers to two distinct eye diseases: diabetic macular edema and diabetic ischemic macular disease. These two types of macular disease often coorbid; that is, people with macular edema often also have ischemic macular disease. Ischemic macular disease occurs with macular edema and can occur even when macular edema is mild. In some embodiments, retinal changes associated with diabetic macular disease include decreased retinal potential, loss of perivascular cells, acellular capillary formation, vascular congestion, vascular dysfunction, vascular leakage, vascular occlusion, tissue swelling (edema), tissue ischemia, or any combination thereof within the retina of the subject.

[0061] As used herein, the term "acellular capillary" refers to a capillary-sized vessel tube that does not contain a nucleus anywhere along its length.

[0062] As used herein, the term “vascular congestion” refers to a type of vascular injury that is an important factor in the pathogenesis of various eye diseases disclosed herein (e.g., diabetic macular edema). Vascular congestion is associated with the accumulation of fluid (e.g., intravascular blood) within vascular tissues. In some embodiments, vascular congestion may be caused by hyperglycemia (i.e., hyperglycemia).

[0063] As used herein, the term “macular degeneration” refers to any number of disorders and symptoms in which the macula degenerates or loses its functional activity. Such degeneration or loss of functional activity may result, for example, from cell death, decreased cell proliferation, loss of normal biological function, or a combination thereof. Macular degeneration may lead to and / or manifest as changes in the structural integrity of cells and / or the extracellular matrix of the macula, changes in the composition of normal cells and / or the 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, and / or capillary endothelial cells. Age-related macular degeneration is the most common type of macular degeneration, but the term “macular degeneration” does not necessarily exclude macular degeneration in patients other than the elderly. Non-restrictive examples of macular degeneration include: age-related macular degeneration (wet or dry); Best macular dystrophy, Sorsby fundus dystrophy, Mallatia leventinese, Doyne honeycomb retinal dystrophy, Stargardt disease (also known as Stargardt macular dystrophy, adolescent macular degeneration, or fundus flavimaculatus), and macular degeneration associated with pigment epithelial detachment.

[0064] As used herein, the term “age-related macular degeneration” (AMD) generally refers to a retinal disease affecting older adults, associated with central vision loss due to damage to the central part of the retina (i.e., the macula). AMD is generally characterized by the gradual accumulation or aggregation of yellowish, insoluble extracellular deposits called intramacula 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 gradually worsen the macula and lead to central vision loss. As used herein, the term “macula” refers to the central part of the retina responsible for central, high-resolution color vision.

[0065] Several theories have been proposed, including oxidative stress, mitochondrial dysfunction, and inflammatory processes, but the pathogenesis of age-related macular degeneration (AMD) is not well understood. An imbalance between the production and degradation of damaged cellular components leads to the accumulation of harmful products, such as intracellular lipofuscin and extracellular drusen. Early atrophy is distinguished by areas of thinning or decolorization of the retinal pigment epithelium (RPE), which precedes geographic atrophy in the early stages of AMD. In the progressive stages of AMD, RPE atrophy (geographic atrophy) and / or the development of new blood vessels (angiogenesis) lead to photoreceptor death and central vision loss. In dry (non-exudative) AMD, cellular debris called drusen accumulates between the retina and choroid, causing retinal atrophy and scarring. In the more severe wet (exudative) AMD, blood vessels grow in the choroid behind the retina (angiogenesis), leading to leakage of exudate and fluid, and potentially inducing bleeding.

[0066] Depending on the extent of the 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 several small drusen (e.g., less than approximately 63 microns in diameter) or several medium-sized drusen (e.g., approximately 63–124 microns in diameter). In the early stage, patients do not experience vision loss and have no apparent symptoms. Intermediate AMD is characterized by the presence of many medium-sized drusen or one or more large drusen (e.g., greater than approximately 125 microns in diameter). During this stage, some patients may begin to experience a faint spot in the center of their visual field. Advanced or late AMD is characterized by damage to a large area of ​​retinal tissue, resulting in a central blind spot and ultimately central vision loss. Based on the type of injury (e.g., presence or absence of angiogenesis), advanced or late-stage AMD can be further classified into two subtypes: (i) geographic atrophy (also known as atrophic AMD) and (ii) wet AMD (also known as neovascular or exudative AMD).

[0067] 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 and wet AMD. As used herein, the term “age-related macular degeneration” also includes all types of age-related macular degeneration and any and all symptoms of age-related macular degeneration, regardless of the cause. Non-limiting examples of symptoms associated with macular degeneration (e.g., age-related macular degeneration) include: loss of central vision, distortion, decreased contrast sensitivity, blurred vision, difficulty adapting to low light, sudden onset, rapid worsening of symptoms, and decreased color vision. In some embodiments, macular degeneration (e.g., age-related macular degeneration) may cause macular edema (i.e., swelling of the macular edema due to the accumulation of fluid and protein deposits on or below the macula).

[0068] 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 except wet (neovascular) AMD. This includes not only early and mid-stage AMD, but also the advanced stage known as geographic atrophy. Patients with dry AMD tend to have minimal symptoms in the earlier stages; visual loss occurs more frequently when symptoms progress to geographic atrophy.

[0069] 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 and is the most advanced form of AMD. In wet AMD, blood vessels grow from choroidal capillaries and, in some cases, from the underlying retinal pigment epithelium (choroidal neovascularization or neovascularization) due to defects in Bruch's membrane. The organization of serous or hemorrhagic exudates from these vessels can lead to the formation of fibrovascular scarring in the macular region, along with associated neuroretinal degeneration, detachment and rupture of the retinal pigment epithelium, vitreous hemorrhage, and permanent damage to central vision.

[0070] As used herein, the term “angiogenesis” refers to the abnormal growth of new blood vessels in different parts of the eye that can induce bleeding and cause 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 containing connective tissue, located 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 may impair visual function through blood and lipid leakage. As used herein, the term “retinal neovascularization” refers to the abnormal development, proliferation, and / or growth of blood vessels in the upper or inner part of the retina, for example, on the retinal surface. Retinal neovascularization can occur in many retinal diseases associated with retinal ischemia and / or inflammatory diseases (e.g., diabetic retinopathy, sickle cell retinopathy, Eales' disease, ocular ischemic syndrome, carotid cavernous fistula, familial exudative vitreoretinopathy, hyperviscosity syndrome, idiopathic arteriolar occlusion, radiation retinopathy, retinal vein occlusion, retinal artery occlusion, retinal embolism, birdshot retinochoroidopathy, retinal vasculitis, papilosarcoma, toxoplasmosis and uveitis, choroidal melanoma, chronic retinal detachment, anterior ischemic optic neuropathy (AION), non-arteritic anterior ischemic optic neuropathy (NAION), and incontinentia pigmenti). Methods for detecting angiogenesis are publicly known in the industry and include, but are not limited to, measuring the expression of CD31 (platelet endothelial cell adhesion molecule, also known as PECAM-1) and vascular endothelial growth factor (VEGF) in tissue. For example, see Schluter A., ​​et al., BMC Cancer 18(1):272 (2018).

[0071] The retinal pigment epithelium (RPE) is a single layer of epithelial cells located on the back of the vertebrate eye, between the choroidal blood supply (choroidal capillary layer) and the neuroretina. The RPE acts as a component of the blood-retinal barrier, and RPE cells play a crucial role in maintaining the visual cycle, phagocytosis of photoreceptor external segments, and the transport of nutrients, metabolic waste, ions, and fluids between the distal retina and the choroidal capillary layer.

[0072] The term “intraocular” refers to the inside or lower part of the ocular tissue. As used herein, “intraocular administration” refers to any administration that can deliver a composition (e.g., the anti-FAM19A5 antibody disclosed herein) to the sub-Tenon, subconjunctival, suprachoroidal, intravitreous, and similar locations of the eye. In some embodiments, intraocular administration includes intravitreous administration.

[0073] The term “ocular condition” refers to any disease, illness, or symptom that affects or is associated with the eye or a part or area of ​​the eye, such as retinal disease. In some embodiments, ocular conditions treatable with the present disclosure include diabetic retinopathy and age-related macular degeneration. Other ocular conditions treatable with the compositions and methods disclosed herein include diabetic macular edema, anterior ischemic optic neuropathy (AION), non-arteritic anterior ischemic optic neuropathy (NAION), retinal vein occlusion, retinal artery occlusion, or a combination thereof.

[0074] As used herein, the term “eye” includes the eyeball and the tissues and fluids that make up the eyeball (e.g., sclera, cornea, anterior chamber and posterior chamber, iris / pupil, lens, vitreous fluid, retina, fovea, macula and choroid), the muscles around the eye (e.g., sclerotropin and rectus muscles), and parts of the optic nerve inside or adjacent to the eyeball.

[0075] The term “optic nerve injury” refers to any change in the normal structure or function of the optic nerve. Changes in the normal structure or function of the optic nerve may result from any disease, disorder, or injury, including glaucoma. Changes in the normal function of the optic nerve include any changes in the optic nerve’s proper functional capacity, such as its ability to transmit visual information from the retina to the brain. Functional changes may manifest spontaneously, such as visual field loss, central vision damage, or abnormal color vision. Examples of structural changes include retinal nerve fiber loss, optic nerve cupping, and / or cell loss from the retinal ganglion cell layer of the retina. As used herein, “optic nerve injury” may include optic nerve injury to one or both of the optic nerves of the subject.

[0076] As used herein, the terms “treat,” “treating,” and “treatment” refer to any type of intervention or process performed on a subject with the aim of reversing, alleviating, improving, inhibiting, delaying, or preventing the progression, development, severity, or recurrence of a disease-related syndrome, complication, symptom, or biochemical sign, or the administration of an activating agent to a subject. Treatment may be performed on a subject with the disease or on a subject without the disease (e.g., for preventative purposes).

[0077] As used herein, “administration” means the physical introduction of a therapeutic agent or a composition containing a therapeutic agent into a subject using any of the various methods and delivery systems known to those skilled in the art. Preferred routes of administration for the antibodies described herein include intravenous, intraperitoneal, intramuscular, subcutaneous, spinal, intravitreous, or other parenteral administration routes, such as by injection or infusion. As used herein, “parenteral administration” generally means, but is not limited to, methods of administration other than intestinal and topical administration by injection, and includes intravenous, intraperitoneal, intramuscular, intraarterial, intrathecal, intralymphatic, intralesional, intracapsular, intraorbital, intracardiac, intratracheal, subcutaneous, subepidermal, intravitreous, intra-articular, subcapsular, subarachnoid, intraspinal, epidural, and intrasternal injections and infusions, as well as intra vivo electroporation. In contrast, the antibodies described herein may be administered via parenteral routes, such as topical, epidermal, or mucosal administration routes, such as intranasal, oral, vaginal, rectal, sublingual, or topical. The administration may also be carried out, for example, once, multiple times, and / or over one or more extended periods.

[0078] As used herein, the term “therapeutic dose” refers to the amount of a drug, alone or in combination with other therapeutic agents, that is effective in “treating” the disease or disorder in question, or in reducing the risk, potential, likelihood, or incidence of the disease or disorder (e.g., glaucoma). “Therapeutic dose” includes the amount of a drug or therapeutic agent that provides some improvement or practical benefit to a subject who has or is at risk of having the disease or disorder (e.g., glaucoma as disclosed herein). Thus, “therapeutic dose” is the amount that reduces the risk, potential, likelihood, or incidence of the disease or disorder, or provides some relief or reduction, or reduces at least one indicator (e.g., increased inflammation), or reduces at least one clinical symptom of the disease or disorder.

[0079] As used herein, the term “subject” includes any human or non-human animal. The term “non-human animal” includes all vertebrates, such as mammals and non-human primates, as well as non-mammals such as sheep, dogs, cattle, chickens, amphibians, and reptiles.

[0080] The term “family with sequence similarity 19, member A5” or “FAM19A5” refers to proteins belonging to the TAFA family (also known as the FAM19 family), which consists of five highly homologous proteins. See Tang TY et al., Genomics 83(4):727-34 (2004). These proteins contain conserved cysteine ​​residues and are loosely related to macrophageous inflammatory protein 1-alpha (MIP-1-alpha), a type of CC-chemokine family protein. The aforementioned TAFA proteins are expressed in specific regions of the brain and spinal cord. These proteins are thought to be produced and secreted by adult neural stem cells during neurogenesis. FAM19A5 is also known as TAFA5 or chemokine-like protein TAFA-5.

[0081] FAM19A5 is considered crucial for the development, differentiation, and formation of the complete central nervous system. FAM19A5 also plays a vital role in the pathogenesis of many central nervous system injuries and / or degenerative brain diseases (e.g., Huntington's disease, Parkinson's disease, Alzheimer's disease, spinal cord injury, stroke, and brain tumors). Upon central nervous system injury, neural stem cells produce FAM19A5, which differentiates normal astrocytes into reactive astrocytes. These reactive astrocytes (along with microglia) express various extracellular matrix components (ECMs, e.g., proteoglycans) and can induce the formation of glial scars, which surround the injured area of ​​the central nervous system like a network, preventing neuronal regeneration. Antagonists of FAM19A5 (e.g., anti-FAM19A5 antibodies) can be used to prevent and / or treat central nervous system injuries and / or diseases. See U.S. Patent No. 9,579,398.

[0082] However, recent studies suggest that FAM19A5 antagonists may not be a viable treatment option for specific diseases, particularly those associated with vascular abnormalities. Wang et al., Circulation 138(1):48-63 (2018). Wang et al. showed that adipose tissue in normal, healthy individuals produces FAM19A5, which transmits signals that inhibit the proliferation and migration of vascular smooth muscle cells (VSMCs) and weaken post-intimal fusion through sphingosine-1-phosphate receptor 2 (S1PR2). However, in obese individuals, FAM19A5 expression (both mRNA and protein) is significantly reduced in adipose tissue. Wang et al. concluded that this reduction in FAM19A5 activity accelerates the progression of cardiovascular disease observed in many obese individuals.

[0083] In humans, the gene encoding FAM19A5 is located on chromosome 22. There are three isoforms of human FAM19A5 (UniProt:Q7Z5A7) that are produced by alternative splicing: isoform 1 (UniProt:Q7Z5A7-1) consisting of 132 amino acids, isoform 2 (UniProt:Q7Z5A7-2) consisting of 125 amino acids, and isoform 3 (UniProt:Q7Z5A7-3) consisting of 53 amino acids. Human FAM19A5 protein is thought to exist in membrane-bound and soluble (secreted) forms. Isoform 1 is thought to be a membrane protein with a single transmembrane region. Isoform 2, reported as a secreted protein (soluble) in Tang TY et al., Genomics 83(4):727-34 (2004), contains a signal peptide at amino acid positions 1-25. Isoform 3 is predicted based on EST data. Next are the amino acid sequences of three known human FAM19A5 isotypes.

[0084] (I) Isotype 1 (UniProt:Q7Z5A7-1, transmembrane protein): This isotype was selected as the standard sequence.

[0085] MAPSPRTGSR QDATALPSMS STFWAFMILA SLLIAYCSQL AAGTCEIVTL DRDSSQPRRT IARQTARCAC RKGQIAGTTR ARPACVDARI IKTKQWCDML PCLEGEGCDL LINRSGWTCT QPGGRIKTTT VS(Sequence ID 1) (II) Isotype 2 (UniProt: Q7Z5A7-2, soluble protein): MQLLKALWAL AGAALCCFLV LVIHAQFLKE GQLAAGTCEI VTLDRDSSQP RRTIARQTAR CACRKGQIAG TTRARPACVD ARIIKTKQWC DMLPCLEGEG CDLLINRSGW TCTQPGGRIK TTTVS(Sequence ID 2) (III) Isotype 3 (UniProt: Q7Z5A7-3): MYHHREWPAR IIKTKQWCDM LPCLEGEGCD LLINRSGWTC TQPGGRIKTT TVS(Sequence ID 3) The term “FAM19A5” includes any variant or isotype of FAM19A5 spontaneously expressed by cells. Therefore, the antibodies described herein may cross-react with different isotypes within the same species (e.g., different isotypes of human FAM19A5) or with FAM19A5 from non-human species (e.g., mouse FAM19A5). In contrast, the antibodies may be specific to human FAM19A5 and may not exhibit any cross-reaction with different species. FAM19A5 or any variant and isotype thereof may be isolated from cells or tissues that spontaneously express them, or produced by recombination. The polynucleotide encoding human FAM19A5 has GenBank accession number BC039396 and the following sequence:

[0086] [Table 1]

[0087] The term “antagonist against FAM19A5 protein” refers to all antagonists that suppress and / or neutralize the expression and / or activity of the FAM19A5 protein. Such antagonists may be peptides, nucleic acids, or compounds. More specifically, the antagonist may be an antisense oligonucleotide, siRNA, shRNA, miRNA, dsRNA, aptamer, PNA (peptide nucleic acid), or vector containing the same that targets FAM19A5. In some embodiments, the antagonist may be an antibody or its antigen-binding moiety that specifically binds to the FAM19A5 protein.

[0088] The terms “antibody” and “antibodies” are industry terms, interchangeable herein, and refer to molecules having an antigen-binding site that specifically binds to an antigen. The terms used herein include whole antibodies and any antigen-binding fragments thereof (i.e., “antigen-binding portions”) or single chains. In one embodiment, “antibody” refers to a glycoprotein or its antigen-binding portion comprising at least two heavy (H) chains and two light (L) chains linked to each other by disulfide bonds. In yet another embodiment, “antibody” refers to a single-chain antibody comprising a single variable domain, e.g., a VHH domain. Each heavy chain consists of a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region. In a particular spontaneous antibody, the heavy chain constant region consists of three domains CH1, CH2, and CH3. In a particular spontaneous antibody, each light chain consists of a light chain variable region (abbreviated herein as VL) and a light chain constant region. The light chain constant region consists of one domain CL.

[0089] The VH and VL regions can be further subdivided into highly variable regions called complementarity-determining regions (CDRs), which contain more conserved regions called framework regions (FRs). Each of the VH and VL regions consists of three CDRs and four FRs arranged in the following order from the amino terminus to the carboxyl terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. The variable regions of the heavy and light chains contain binding domains that interact with antigens. The constant regions of the antibody can mediate the binding of immunoglobulins to various cells of the immune system (e.g., effector cells) and to host tissues or factors, including the first component (Clq) of the classical complement system.

[0090] The term “Kabat numbering” and similar terms are recognized in this industry and refer to a system for numbering amino acid residues in the heavy and light chain variable regions of an antibody or its antigen-binding moiety. In certain embodiments, the CDR of an antibody may be determined according to the Kabat numbering system (see, for example, Kabat EA & Wu TT (1971) Ann NY Acad Sci 190:382-391 and Kabat EA et al, (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, USD Department of Health and Human Services, NIH Publication No. 91-3242). Using the Kabat numbering system, the intramolecular CDRs of the antibody heavy chain are typically located at amino acid positions 31-35 (CDR1) (which may include one or two additional amino acids after position 35 (referred to as 35A and 35B in the Kabat numbering system)), amino acid positions 50-65 (CDR2), and amino acid positions 95-102 (CDR3). Using the Kabat numbering system, the intramolecular CDRs of the antibody light chain are typically located at amino acid positions 24-34 (CDR1), amino acid positions 50-56 (CDR2), and amino acid positions 89-97 (CDR3). In certain embodiments, the CDRs of the antibodies described herein are determined by the Kabat numbering system.

[0091] The terms “Kabat-style amino acid position numbering,” “Kabat position,” and their grammatical variations refer to the numbering system used for the heavy chain variable domain or light chain variable domain of antibody compilations in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991). Using this numbering system, the actual linear amino acid sequence can contain additional amino acids corresponding to the shortening or insertion of the FW or CDR of the variable domain. For example, the heavy chain variable domain can include a single amino acid inserted after H2 residue 52 (residue 52a according to Kabat) and residues inserted after heavy chain FW residue 82 (e.g., residues 82a, 82b, and 82c according to Kabat). See Table 1B.

[0092] [Table 2]

[0093] The Kabat numbering of residues for a given antibody can be determined by alignment in regions of homology between the “standard” Kabat-numbered sequence and the antibody sequence. Alternatively, Chothia refers to the position of the structural loop (Chothia and Lesk, J.Mol.Biol.196:901-917(1987)). When numbered using the Kabat numbering convention, the end of the Chothia CDR-H1 loop varies between H32 and H34 depending on the length of the loop (this is because the Kabat numbering system places insertions at H35A and H35B; if neither 35A nor 35B is present, the loop ends at 32; if only 35A is present, the loop ends at 33; if both 35A and 35B are present, the loop ends at 34). The AbM hypervariable region represents the negotiation between Kabat CDR and Chothia structural loops and is used by Oxford Molecular's AbM antibody modeling software.

[0094] IMGT (ImMunoGeneTics) also provides a numbering system for immunoglobulin variable regions, including CDRs. For example, see Lefranc, MP et al., Dev. Comp. Immunol. 27:55-77 (2003), which is included as reference herein. The IMGT numbering system is based on the alignment, structural data, and characterization of over 5,000 sequences, facilitating the comparison of variable and CDR regions across all species. According to the IMGT numbering system, VH-CDR1 is located at positions 26-35, VH-CDR2 at positions 51-57, VH-CDR3 at positions 93-102, VL-CDR1 at positions 27-32, VL-CDR2 at positions 50-52, and VL-CDR3 at positions 89-97.

[0095] For all heavy chain constant region amino acid positions discussed in this disclosure, the numbering follows the EU index first described in Edelman et al., 1969, Proc. Natl. Acad. Sci. USA 63(1):78-85), which described the amino acid sequence of myeloma protein EU, the first sequenced human IgG1. Edelman et al.'s EU index is also presented in Kabat et al., 1991, Sequences of Proteins of Immunological Interest, 5th Ed., United States Public Health Service, National Institutes of Health, Bethesda. Accordingly, the phrases “the EU index presented in Kabat” or “Kabat’s EU index” and “positions following the EU index presented in Kabat…” and their grammatical variations refer to the residue numbering system based on the human IgG1 EU antibody by Edelman et al. as presented in Kabat 1991.

[0096] The numbering system used for the amino acid sequences of the variable domain (both heavy and light chains) and the constant region of the light chain is the one presented in Kabat 1991.

[0097] Antibodies may have any type of immunoglobulin molecule (e.g., IgG, IgE, IgM, IgD, IgA, or IgY), any subtype (e.g., IgD, IgG2, IgG3, IgG4, IgA1, or IgA2), or any subtype (e.g., IgG1, IgG2, IgG3, and IgG4 in humans; and IgG1, IgG2a, IgG2b, and IgG3 in mice). Immunoglobulins, such as IgG1, exist in a variety of allotypes that differ from each other at a maximum of several amino acids. Antibodies disclosed herein may consist of any of the commonly known isotypes, subtypes, or allotypes. In certain embodiments, the antibodies disclosed herein are IgG1, IgG2, IgG3, or IgG4 subtypes or any hybrids thereof. In certain embodiments, the antibodies have a human IgG1 subtype or a human IgG2 or human IgG4 subtype.

[0098] "Antibodies" include, for example, spontaneously occurring and non-spontaneously occurring antibodies; monoclonal and polyclonal antibodies; chimeric and humanized antibodies; human and non-human antibodies; totally synthetic antibodies; single-chain antibodies; monospecific antibodies; multispecific antibodies (including bispecific antibodies); tetramer antibodies containing two heavy chains and two light chain molecules; antibody light chain monomers; antibody heavy chain monomers; antibody light chain dimers; antibody heavy chain dimers; antibody light chain-antibody heavy chain pairs; intrabody antibodies; heteroconjugate antibodies; monovalent antibodies; single-chain antibodies; camelized antibodies; affybody antibodies; anti-idiotype (anti-Id) antibodies (e.g., anti-anti-Id antibodies); and single-domain antibodies (sdAb) containing a binding molecule consisting of a single monomer variable antibody domain (e.g., VH domain or VL domain) that is fully antigen-binding (Harmen MM and Haard HJAppl Microbiol Biotechnol. 77(1):13-22 (2007)).

[0099] As used herein, the term “antigen-binding portion” of an antibody refers to one or more fragments of an antibody that possess the ability to specifically bind to an antigen (e.g., human FAM19A5). Such “fragments” are, for example, about 8 to about 1500 amino acids long, preferably about 8 to about 745 amino acids long, preferably about 8 to about 300 amino acids, for example, about 8 to about 200 amino acids or about 10 to about 50 or 100 amino acids long. It has been shown that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody. Examples of binding fragments included in the term “antigen-binding moiety” of antibodies, for example, the anti-FAM19A5 antibody disclosed herein, include: (i) a Fab fragment which is a monovalent fragment consisting of VL, VH, CL, and CH1 domains; (ii) a F(ab')2 fragment which is a bivalent fragment containing two Fab fragments linked by a disulfide linkage at a hinge region; (iii) an Fd fragment consisting of VH and CH1 domains; (iv) an Fv fragment consisting of the VL and VH domains of a single arm of the antibody and a disulfide-linked Fvs(sdFv); (v) a dAb fragment consisting of a VH domain (Ward et al., (1989) Nature 341:544-546); and (vi) a separated complementarity-determining region (CDR) or (vii) a combination of two or more separated CDRs which may be joined by a synthetic linker. Furthermore, the two domains of the Fv fragment, VL and VH, are encoded by separate genes, but these can be joined by a synthetic linker that can be used by recombinant methods to create a single protein chain (known as single-chain Fv (scFv)) in which the VL and VH regions pair up to form a monovalent molecule; see, for example, Bird et al., (1988) Science 242:423-426; and Huston et al., (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883. Such single-chain antibodies are also included in the term "antigen-binding portion" of an antibody. These antibody fragments are obtained using prior art known to those skilled in the art, and the fragments are screened for usefulness in the same manner as intact antibodies. Antigen-binding portions can be generated by recombinant DNA technology or by enzymatic or chemical cleavage of intact immunoglobulins.

[0100] The terms “variable region” and “variable domain” as used herein are synonymous and are universal in this industry. The variable region typically refers to a part of an antibody, generally a part of the light chain or heavy chain. Typically, the mature heavy chain contains approximately 110-120 amino acids at the amino terminus, and the mature light chain contains approximately 90-115 amino acids. These vary widely in sequence among antibodies and are used in the binding and specificity of a particular antibody to a particular antigen. The sequence variability is concentrated in a region called the complementarity-determining region (CDR), while a more highly conserved region within the variable domain is called the framework region (FR).

[0101] While not limited to specific mechanisms or theories, the CDRs of the light and heavy chains are thought to be primarily responsible for antigen-antibody interaction and specificity. In certain embodiments, the variable region is a human variable region. In certain embodiments, the variable region includes a rodent or murid CDR and a human framework region (FR). In certain embodiments, the variable region is a primate (e.g., non-human primate) variable region. In certain embodiments, the variable region includes a rodent or murid CDR and a primate (e.g., non-human primate) framework region (FR).

[0102] As used herein, the term “heavy chain (HC)” can refer, when used in relation to an antibody, to any distinct type of IgG based on the amino acid sequence of the constant domain, such as IgG1, IgG2, IgG3, and IgG4, which give rise to the IgA, IgD, IgE, IgG, and IgM types of antibodies, such as alpha (α), delta (δ), epsilon (ε), gamma (γ), and mu (μ).

[0103] As used herein, the term “light chain (LC),” when used in relation to antibodies, may refer to any distinct type based on the amino acid sequence of the constant domain, such as kappa (κ) and lambda (λ). Light chain amino acid sequences are well known in the art. In certain embodiments, the light chain is a human light chain.

[0104] The terms "VL" and "VL domain" are used interchangeably to refer to the variable region of the antibody light chain.

[0105] The terms "VH" and "VH domain" are used interchangeably to refer to the variable region of the antibody's heavy chain.

[0106] The terms “constant region” or “constant domain” as used herein are interchangeable and have the common meanings of the art. The constant domain is the carboxyl terminus of the light and / or heavy chain that does not directly participate in the binding of the antibody to the antigen, for example, but can exhibit various effector functions such as interaction with the Fc receptor. The constant region of an immunoglobulin molecule generally has a more conserved amino acid sequence than the immunoglobulin variable domain.

[0107] The “Fc region” (fragment-crystallizable region), “Fc domain,” or “Fc” refers to the C-terminal region of an antibody heavy chain that mediates the binding of immunoglobulins to host tissues or factors, including binding to Fc receptors located on various cells of the immune system (e.g., effector cells) or to the first component (C1q) of the classical complement system. Thus, the Fc region includes the constant region of the antibody excluding the first constant region immunoglobulin domain (e.g., CH1 or CL). In IgG, IgA, and IgD antibody isotypes, the Fc region contains two identical protein fragments derived from the second (CH2) and third (CH3) constant domains of the two heavy chains of the antibody; the IgM and IgE Fc regions contain three heavy chain constant domains (CH domains 2-4) in their respective polypeptide chains. In the case of IgG, the Fc region includes hinges between immunoglobulin domains Cγ2 and Cγ3 and between Cγ1 and Cγ2. While the boundaries of the Fc region of immunoglobulin heavy chains can vary, the human IgG heavy chain Fc region is generally limited to the extension from the amino acid residue at position C226 or P230 (or the amino acid between these two) to the carboxyl terminus of the heavy chain, although the numbering follows the EU index, as in Kabat. The CH2 domain of the human IgG Fc region is extended from approximately amino acid 231 to approximately amino acid 340, and the CH3 domain is located on the C-terminal side of the Cm domain in the Fc region, i.e., from approximately amino acid 341 to approximately amino acid 447 of IgG. As used herein, the Fc region may be a native sequence Fc containing any allogeneic variant, or a mutant Fc (e.g., a non-spontaneous Fc). Fc may also mean that this region is isolated or in relation to an Fc-containing protein polypeptide such as an “Fc-region-containing binding protein” also called an “Fc fusion protein” (e.g., an antibody or immunoadhesion).

[0108] A “natural sequence Fc region” or “natural sequence Fc” contains the same amino acid sequence as a naturally occurring Fc region. Natural sequence human Fc regions include not only the natural sequence human IgG1 Fc region; the natural sequence human IgG2 Fc region; the natural sequence human IgG3 Fc region; and the natural sequence human IgG4 Fc region, but also their naturally occurring variants. Natural sequence Fc includes various allogeneic forms of Fc (see, for example, Jefferis et al. (2009) mAbs 1:1; Vidarsson G. et al. Front Immunol. 5:520 (published online October 20, 2014)).

[0109] An “Fc receptor” or “FcR” is a receptor that binds to the Fc region of immunoglobulins. FcRs that bind to IgG antibodies include the Fcγ family of receptors, allele variants of these receptors, and other spliced ​​forms. The Fcγ family consists of three activating receptors (mouse FcγRI, FcγRIII, and FcγRIV; human FcγRIA, FcγRIIA, and FcγRIIIA) and one inhibitory receptor (FcγRIIB). Human IgG1 binds to most human Fc receptors and derives the strongest Fc effector function. Human IgG1 can be considered equivalent to mouse IgG2a in relation to the type of activating Fc receptor it binds to. Conversely, human IgG4 derives the least Fc effector function (see Vidarsson G. et al. Front Immunol. 5:520 (online published October 20, 2014)).

[0110] The constant region may be manipulated, for example, by recombinant technology, to remove one or more effector functions. “Effector function” refers to the interaction between the antibody Fc region and an Fc receptor or ligand, or the biochemical reaction therefrom. Exemplary “effector functions” include FcγR-mediated effector functions such as C1q binding, complement-dependent cytotoxicity (CDC), Fc receptor binding, ADCC, and antibody-dependent cell-mediated phagocytosis (ADCP), and downward regulation of cell surface receptors (e.g., B cell receptors; BCRs). Such effector functions generally require the Fc region to be combined with a binding domain (e.g., an antibody variable domain). Therefore, the terminology... A “steady-state region without Fc function” includes a steady-state region in which one or more effector functions mediated by the Fc region are reduced or absent.

[0111] The effector function of an antibody can be reduced or avoided by different approaches. The effector function of an antibody can be reduced or avoided by using antibody fragments lacking an Fc region (e.g., Fab, F(ab')2, single-chain Fv(scFv), or sdAb consisting of monomeric VH or VL domains). Alternatively, so-called aglycosylated antibodies can be produced by reducing the effector function of an antibody by removing a sugar linked to a specific residue in the Fc region while retaining other valuable properties of the Fc region (e.g., long half-life and heterodimerization). Aglycosylated antibodies can be produced, for example, by deleting or altering the residue to which the sugar is attached, by enzymatically removing the sugar, by producing the antibody in cells cultured in the presence of a glycosylation inhibitor, or by expressing the antibody in cells that cannot glycosylate proteins (e.g., bacterial host cells). See, for example, U.S. Patent Publication No. 20120100140. Another approach involves utilizing the Fc region of IgG subtypes with reduced effector function; for example, IgG2 and IgG4 antibodies are characterized by having lower levels of Fc effector function compared to IgG1 and IgG3. The residue closest to the hinge region in the CH2 domain of the Fc portion is responsible for the antibody's effector function, and it contains a binding site that largely overlaps with C1q (complement) and the IgG-Fc receptor (FcγR) on effector cells of the innate immune system (Vidarsson G. et al. Front Immunol. 5:520 (published online October 20, 2014)). Therefore, antibodies with reduced or absent Fc effector function can be produced, for example, by creating a chimeric Fc region containing the CH2 domain of an IgG4 isotype IgG antibody and the CH3 domain of an IgG1 isotype IgG antibody, or a chimeric Fc region containing the hinge region of IgG2 and the CH2 region of IgG4 (see, e.g., Lau C. et al. J.Immunol. 191:4769-4777 (2013)), or by generating an Fc region with a mutation that alters Fc effector function, for example, by reducing or eliminating Fc function. Such mutated Fc regions are known in the art.For example, see U.S. Patent Publication No. 20120100140, whose disclosure is included herein by reference in whole, and the U.S. and PCT applications cited therein, as well as An et al., mAbs 1:6, 572-579 (2009).

[0112] The terms “hinge,” “hinge domain,” “hinge region,” or “antibody hinge region” refer to the domain of the heavy chain constant region, including the upper, middle, and lower parts of the hinge, formed by the conjugation of the CH1 domain with the CH2 domain (Roux et al., J.Immunol. 1998 161:4083). The hinge provides antibody binding and a change in the level of flexibility between effector regions, and also provides a site for intermolecular disulfide bonding between the two heavy chain constant regions. As disclosed herein, the hinge begins at Glu216 and ends at Gly237 for all IgG isotypes (Roux et al., 1998 J Immunol 161:4083). The sequences of the wild-type IgG1, IgG2, IgG3, and IgG4 hinges are known to the art. For example, see Kabat EA et al., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, USD Department of Health and Human Services, NIH Publication No. 91-3242; Vidarsson G. et al., Front Immunol. 5:520 (published online October 20, 2014).

[0113] The term “CH1 domain” refers to the heavy chain constant region that hinges the variable domain within the heavy chain constant domain. As used herein, the CH1 domain begins at A118 and ends at V215. The term “CH1 domain” includes not only the wild-type CH1 domain but also its naturally occurring variants (e.g., allogeneic variants). The CH1 domain sequences of IgG1, IgG2, IgG3, and IgG4 (including wild-type and allogeneic variants) are publicly known in the art (e.g., Kabat EA et al., (1991) and Vidarsson G. et al., Front Immunol. 5:520 (online publication October 20, 2014)). Exemplary CH1 domains include, for example, CH1 domains with mutations that alter the half-life of antibodies described in U.S. Patent Publication No. 20120100140 and the U.S. patents, publications, and PCT publications cited therein.

[0114] The term “CH2 domain” refers to the heavy chain constant region that hinges to the CH3 domain in the heavy chain constant domain. As used herein, the CH2 domain begins at P238 and ends at K340. The term “CH2 domain” includes not only the wild-type CH2 domain but also its naturally occurring variants (e.g., allogeneic variants). The CH2 domain sequences of IgG1, IgG2, IgG3, and IgG4 (including wild-type and allogeneic variants) are publicly known in the art (e.g., Kabat EA et al., (1991) and Vidarsson G. et al., Front Immunol. 5:520 (online publication October 20, 2014)). Exemplary CH2 domains include, for example, CH2 domains with mutations that alter the biological activity of antibodies described in U.S. Patent Publication No. 20120100140 and the U.S. patents, publications, and PCT publications cited therein, e.g., half-life and / or reduced Fc effector function.

[0115] The term “CH3 domain” refers to the C-terminal heavy chain constant region relative to the CH2 domain in the heavy chain constant domain. As used herein, the CH3 domain begins at G341 and ends at K447. The term “CH3 domain” includes not only the wild-type CH3 domain but also its naturally occurring variants (e.g., allogeneic variants). The CH3 domain sequences of IgG1, IgG2, IgG3, and IgG4 (including wild-type and allogeneic variants) are publicly known in the art (e.g., Kabat EA et al., (1991) and Vidarsson G. et al., Front Immunol. 5:520 (online publication October 20, 2014)). Exemplary CH3 domains include, for example, CH3 domains with mutations that alter the biological activity, e.g., half-life, of antibodies described in U.S. Patent Publication No. 20120100140 and the U.S. patents, publications, and PCT publications cited herein.

[0116] As used herein, “isotype” refers to antibody types encoded by heavy chain constant region genes (e.g., IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2, IgD, and IgE antibodies).

[0117] An "allogeneic variant" refers to a spontaneously occurring variant within a specific isotype group that differs by a certain number of amino acids (see, e.g., Jefferis et al., (2009) mAbs 1:1). The antibodies described herein may have any allogeneic variant. Allogeneic variants of IgG1, IgG2, IgG3, and IgG4 are publicly known in the art. See, for example, Kabat EA et al., (1991); Vidarsson G. et al., Front Immunol. 5:520 (published online October 20, 2014); and Lefranc MP, mAbs 1:4, 1-7 (2009).

[0118] The terms “antibody that recognizes an antigen” and “antibody that is specific to an antigen” are used interchangeably in this specification with the term “antibody that specifically binds to an antigen.”

[0119] As used herein, an "isolated antibody" means an antibody substantially free of other antibodies having different antigen specificities (e.g., an isolated antibody that specifically binds to FAM19A5 is substantially free of antibodies that specifically bind to antigens other than FAM19A5). However, an isolated antibody that specifically binds to an epitope of FAM19A5 can have cross-reactivity with other FAM19A5 proteins of different species.

[0120] "Binding affinity" generally means the sum of the strengths of non-covalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). Unless otherwise indicated, "binding affinity" as used herein means intrinsic binding affinity that reflects a 1:1 interaction between the members of a binding pair (e.g., an antibody and an antigen). The affinity of molecule X for partner Y can generally be expressed by the dissociation constant (K D ). Affinity can be measured and / or expressed in a number of ways known in the art, including, but not limited to, the equilibrium dissociation constant (K D ) and the equilibrium association constant (K A ). The K D is calculated from the quotient of k off / k on and is expressed in molar concentration (M), and K A is calculated from the quotient of k on / k off . k on means, for example, the association rate constant of an antibody for an antigen, and k off means, for example, the dissociation of an antibody from an antigen. k on and k off can be determined by techniques known to those skilled in the art, such as immunoassays (e.g., enzyme-linked immunosorbent assay (ELISA)), BIACORE TM or kinetic exclusion analysis (KinExA®).

[0121] The terms “specifically binding,” “specifically recognizing,” “specific binding,” “selective binding,” and “selectively binding” as used herein are similar terms in relation to antibodies and refer to molecules (e.g., antibodies) that bind to antigens (e.g., epitopes or immune complexes), and binding is understood by those skilled in the art. For example, molecules that specifically bind to antigens are used in immunoassays, BIACORE, etc. TM KinExA (registered trademark) 3000 devices (Sapidyne When determined by Instruments, Boise, ID or other analytical methods known to the art, it can typically bind to other peptides or polypeptides with lower affinity. In certain embodiments, a molecule that specifically binds to an antigen may, when this molecule binds to another antigen, result in K A Compared to this, K is at least 2logs, 2.5logs, 3logs, 4logs or more and even larger. A It then binds to the antigen.

[0122] Antibodies are typically 10 -5 ~10 -11 Dissociation constants (K) less than or equal to M D It specifically binds to these cognate antigens with a high affinity, as reflected by ). -4 Any K greater than M D This is generally considered to mean nonspecific binding. As used herein, an antibody that "specifically binds" to an antigen refers to an antibody that binds to the antigen and substantially the same antigen with high affinity, for example, a BIACORE using the aforementioned antigen. TM When determined by immunoassay (e.g., ELISA) or surface plasma resonance (SPR) technology using 2000 instruments, 10 -7 M or less, preferably 10 -8 M or less, more preferably 10 -9 M or less, most preferably 10 -8 M~10 -10 K below M D This means that the antibody possesses the necessary properties, but it does not bind to unrelated antigens with high affinity.

[0123] As used herein, “antigen” refers to any naturally occurring or synthetic immunogenic substance, such as a protein, peptide, or hapten. The antigen may be FAM19A5 or a fragment thereof.

[0124] As used herein, “epitope” is an industry term referring to a localized region of an antigen to which an antibody can specifically bind. An epitope may be, for example, an adjacent amino acid of a polypeptide (linear or adjacent epitope), or an epitope may be, for example, a polypeptide or an aggregate of two or more non-adjacent regions of a polypeptide (stereomorphic, nonlinear, discontinuous, or non-adjacent epitope). Epitopes formed from adjacent amino acids are typically maintained upon exposure to a denaturing solvent, though not always, whereas epitopes formed by tertiary folding are typically lost upon treatment with a denaturing solvent. An epitope typically contains at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 20 amino acids within a specific spatial stereomorphic structure. Methods for determining which epitopes bind to a given antibody (i.e., epitope mapping) are well known in the art and include, for example, immunoblotting and immunoprecipitation analysis for testing duplicate or adjacent peptides (e.g., FAM19A5) for reactivity with a given antibody (e.g., anti-FAM19A5 antibody). Methods for determining the spatial stereomorphism of epitopes include art in the art and art described herein, such as X-ray crystallography, two-dimensional nuclear magnetic resonance and HDX-MS (see, for example, Epitope Mapping Protocols in Methods in Molecular Biology, Vol. 66, GEMorris, Ed. (1996)).

[0125] In certain embodiments, the epitope to which the antibody binds may be determined, for example, by NMR spectroscopy, X-ray diffraction crystallography studies, ELISA analysis, hydrogen / deuterium exchange combined with mass spectrometry (e.g., liquid chromatography-electron atomization mass spectrometry), array-based oligopeptide scanning analysis, and / or mutagenesis mapping (e.g., site-directed mutagenesis mapping). In the case of X-ray crystallography, determination may be achieved using any method known to the art (see, for example, Giege R et al, (1994) Acta Crystallogr D Biol Crystallogr 50(Pt4):339-350; McPherson A (1990) Eur J Biochem 189:1-23; Chayen NE (1997) Structure 5:1269-1274; McPherson A (1976) J Biol Chem 251:6300-6303). Antibody-antigen crystals can be studied using well-known X-ray diffraction techniques, such as X-PLOR (Yale University, 1992, distributed by Molecular Simulations, Inc.; e.g., Meth Enzymol (1985) volumes 114 & 115, eds Wyck). off HWet al.,; US2004 / 0014194 reference) and BUSTER (Bricogne G. (1993) Acta Crystallogr D Biol Crystallogr 49(Pt1):37-60; Bricogne G. (1997) Meth Enzymol 276A:361-423, ed Carter CW; Roversi It can be refined using computer software such as P. et al., (2000) Acta Crystallogr D Biol Crystallogr 56 (Pt 10):1316-1323). Mutagenic mapping studies can be performed using any method known to those skilled in the art. For descriptions of mutagenic techniques, including alanine scanning mutagenic techniques, see, for example, Champe M. et al., (1995) J Biol Chem 270:1388-1394 and Cunningham BC & Wells JA (1989) Science 244:1081-1085.

[0126] The term "epitope mapping" refers to the process of identifying molecular determinants for antibody-antigen recognition.

[0127] In relation to two or more antibodies, the term “binding to the same epitope” means that the antibodies bind to the same segment of amino acid residues, as determined by a predetermined method. Techniques for determining whether an antibody binds to the “same epitope on FAM19A5” as the antibody described herein include epitope mapping methods, such as X-ray analysis of the antigen:antibody complex crystals and hydrogen / deuterium exchange mass spectrometry (HDX-MS) that provide atomic resolution of the epitope. Other methods involve monitoring the binding of antibodies to antigen fragments or mutated variants of the antigen, where binding loss due to deformation of amino acid residues in the antigen sequence is generally considered an indication of the epitope component. Computerized combination methods for epitope mapping are also available. These methods depend on the ability of the antibody of interest to affinity-separate specific short peptides from a combination phage display peptide library. Antibodies having the same VH and VL or the same CDR1, 2, and 3 sequences are expected to bind to the same epitope.

[0128] An antibody that “competes with other antibodies for binding to a target” refers to an antibody that (partially or completely) inhibits the target binding of the other antibody. Whether two antibodies compete with each other for binding to a target, that is, whether one antibody inhibits the target binding of the other antibody, and to what extent, can be determined using known competition experiments. In certain embodiments, one antibody competes with another antibody for target binding, inhibiting this binding by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%. The level of inhibition or competition may differ depending on whether the antibody is a “blocking antibody” (i.e., a cold antibody that is first cultured with the target). Competitive analysis can be performed as described in Chapter 11 of “Using Antibodies” by Ed Harlow and David Lane, Cold Spring Harbor Protoc; 2006; doi: 10.1101 / pdb.prot 4277 or Ed Harlow and David Lane, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA 1999. Competitive antibodies bind to the same epitope, a duplicate epitope, or an adjacent epitope (e.g., as demonstrated by steric hindrance).

[0129] Other competitive binding analyses include solid-phase direct or indirect radioimmunoassay (RIA), solid-phase direct or indirect enzyme immunoassay (EIA), sandwich competitive analysis (see Stahli et al., Methods in Enzymology 9:242 (1983)); solid-phase direct biotin-avidin EIA (see Kirkland et al., J.Immunol. 137:3614 (1986)); solid-phase direct labeling analysis, solid-phase direct labeling sandwich analysis (see Harlow and Lane, Antibodies: A Laboratory Manual, Cold Spring Harbor Press (1988)); solid-phase direct labeling RIA using 1-125 labeling (see Morel et al., Mol.Immunol. 25(1):7 (1988)); solid-phase direct biotin-avidin EIA (see Cheung et al., Virology This includes 176:546 (1990) and directly labeled RIA (see Moldenhauer et al., Scand. J. Immunol. 32:77 (1990)).

[0130] A “bispecific” or “bifunctional” antibody is an artificial hybrid antibody in which two different heavy / light chain pairs have two different binding sites. Bispecific antibodies can be produced by various methods, including hybridoma fusion or Fab' fragment linking. See, for example, Songsivilai & Lachmann, Clin. Exp. Immunol. 79:315-321 (1990); Kostelny et al., J. Immunol. 148, 1547-1553 (1992).

[0131] As used herein, “monoclonal antibody” refers to an antibody that exhibits single-binding specificity and affinity to a specific epitope, or an antibody composition in which all antibodies exhibit single-binding specificity and affinity to a specific epitope. Therefore, the term “human monoclonal antibody” refers to an antibody or antibody composition that exhibits single-binding specificity and has variable and selective constant regions derived from human germline immunoglobulin sequences. In some embodiments, human monoclonal antibodies are produced, for example, by hybridomas containing B cells obtained from transgenic non-human animals, such as transgenic mice, that have a genome containing human heavy chain and light chain transposable genes fused to immortalized cells.

[0132] As used herein, the term “recombinant human antibody” includes all human antibodies manufactured, expressed, produced or isolated by recombinant means, such as (a) antibodies isolated from animals (e.g., mice) that have been transchromosomal to human immunoglobulin genes, or hybridomas produced therefrom; (b) antibodies isolated from host cells transformed to express antibodies, such as transfectomas; (c) antibodies isolated from recombinant human antibody libraries; and (d) antibodies manufactured, expressed, produced or isolated by any other means involving splicing human immunoglobulin gene sequences to other DNA sequences. Such recombinant human antibodies utilize specific human germline immunoglobulin sequences encoded by germline genes, but include variable and constant regions, such as subsequent rearrangements and mutations that occur during antibody maturation. As is well known in the industry (see, for example, Lonberg (2005) Nature Biotech. 23(9):1117-1125), the variable region contains antigen-binding domains encoded by various genes that are rearranged to form antibodies specific to foreign antigens. In addition to rearrangement, the variable region can be further modified by numerous single-amino acid changes (known as somatic mutations or hypermutations) to increase the affinity of antibodies to foreign antigens. The constant region will change in response to additional reactions with the antigen (i.e., isotypic changes). Therefore, rearranged and somatically mutated nucleic acid molecules encoding light-chain and heavy-chain immunoglobulin polypeptides in response to an antigen cannot have sequence identity with the original nucleic acid molecule, but instead will be substantially identical or similar (i.e., at least 80% identical).

[0133] A “human antibody (HuMAb)” refers to an antibody having a variable region in which both the framework and CDR region are derived from human germline immunoglobulin sequences. Furthermore, if the antibody contains a constant region, the constant region is also derived from a human germline immunoglobulin sequence. Antibodies described herein may contain amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by in vitro random or site-directed mutagenesis or in vivo somatic mutation). However, the term “human antibody” as used herein does not include antibodies in which a CDR sequence derived from the germline of another mammalian species, such as mouse, has been transplanted into a human framework sequence. The terms “human” antibody and “completely human” antibody are used interchangeably.

[0134] A “humanized” antibody is an antibody in which some, most, or all of the amino acids outside the CDR domain of a non-human antibody are replaced with corresponding amino acids derived from human immunoglobulins. In some forms of the humanized form of an antibody, some, most, or all of the amino acids outside the CDR domain are replaced with amino acids from human immunoglobulins, while some, most, or all of the amino acids inside one or more CDR regions remain unchanged. Small additions, deletions, insertions, substitutions, or modifications of amino acids are acceptable as long as they do not eliminate the antibody's ability to bind to a specific antigen. “Humanized” antibodies maintain antigen specificity similar to that of the primitive antibody.

[0135] A "chimeric antibody" refers to an antibody in which the variable region originates from one species and the constant region originates from another species, such as an antibody in which the variable region originates from a mouse antibody and the constant region originates from a human antibody.

[0136] As used herein, the term “cross-reactivity” refers to the ability of the antibodies disclosed herein to bind to FAM19A5 of different species. For example, an antibody disclosed herein that binds to human FAM19A5 may also bind to FAM19A5 of other species (e.g., mouse FAM19A5). As used herein, cross-reactivity may be measured by detecting specific reactivity with an antigen purified by binding analysis (e.g., SPR, ELISA), or by binding to or functionally interacting with cells that physiologically express FAM19A5. Methods for determining cross-reactivity include standard binding analyses described herein, e.g., BIACORE TM BIACORE using the 2000 SPR machine (Biacore AB, Uppsala, Sweden) TM This includes surface plasma resonance (SPR) analysis or fluid cell counting techniques.

[0137] When the term “spontaneous” as disclosed herein is applied to a subject, it means the fact that the subject can be found in nature. For example, polypeptides or polynucleotide sequences present in organisms (including viruses) that are separable from a natural source and not intentionally altered by humans in a laboratory are spontaneous.

[0138] A "polypeptide" refers to a chain containing at least two consecutively linked amino acid residues, with no upper limit on the length of the chain. One or more amino acid residues within a protein may, but are not limited to, undergo deformations such as glycosylation, phosphorylation, or disulfide bond formation. A "protein" may contain one or more polypeptides.

[0139] As used in this invention, the term "nucleic acid molecule" includes DNA molecules and RNA molecules. Nucleic acid molecules can be single-stranded or double-stranded, and may be cDNA.

[0140] As used herein, the term “vector” refers to a nucleic acid molecule capable of transporting other ligated nucleic acids. One type of vector is a “plasmid,” which means a circular double-stranded DNA loop to which additional DNA fragments can be ligated. Another type of vector is a viral vector, to which additional DNA fragments can be ligated into a viral genome. Certain vectors can self-replicate in the host cell into which they are introduced (e.g., bacterial vectors and episomal mammalian vectors of bacterial replication origin). Other vectors (e.g., non-episomal mammalian vectors) can integrate into the host cell's genome upon introduction into the host cell, thereby replicating along the host genome. Furthermore, certain vectors can direct the expression of genes to which they are operationally ligated. Such vectors are referred herein as “recombinant expression vectors” (or simply “expression vectors”). Generally, expression vectors that are useful in recombinant DNA technology are often in the form of plasmids. In this specification, “plasmid” and “vector” may be used interchangeably, as plasmids are the most commonly used form of vector. However, this also includes other forms of expression vectors that perform equivalent functions, such as viral vectors (e.g., replication-defective retroviruses, adenoviruses, and adeno-associated viruses).

[0141] As used in this invention, the term “recombinant host cell” (or simply “host cell”) refers to a cell containing nucleic acids that do not naturally exist within the cell, and may be a cell into which a recombinant expression vector has been introduced. Such a term should be understood to refer not only to specific target cells but also to the offspring of such cells. Although such offspring are not actually identical to the parent cells, as mutations and environmental influences may cause specific deformations in the next generation, they are still included within the scope of the term “host cell” as used herein.

[0142] As used herein, the term "linked" refers to the association of two or more molecules. The linkage can be a covalent linkage or a non-covalent linkage. The linkage can also be a genetic linkage (i.e., a recombinant fusion). Such linkages can be achieved using various techniques recognized in the art, such as chemical conjugation and recombinant protein production.

[0143] As used herein, the term "therapeutically effective amount" means an amount of a drug alone or in combination with other therapeutic agents effective to "treat" a disease or disorder in a subject, or to reduce the risk, potentiality, likelihood or occurrence of a disease or disorder (e.g., diabetic retinopathy or age-related macular degeneration). A "therapeutically effective amount" includes an amount of a drug or therapeutic agent (e.g., an anti-FAM19A5 antibody disclosed herein) that provides some improvement or benefit to a subject having or at risk of having a disease or disorder (e.g., diabetic retinopathy or age-related macular degeneration). Thus, a "therapeutically effective amount" is an amount that reduces the risk, potentiality, likelihood or occurrence of a disease or disorder, or provides some alleviation, mitigation and / or reduction of at least one indicator and / or reduction of at least one clinical symptom of a disease or disorder (e.g., diabetic retinopathy or age-related macular degeneration).

[0144] <II. Methods of Treating Retinopathy and / or Macular Degeneration> This specification discloses methods of treating eye diseases in a subject in need thereof. More specifically, the methods disclosed herein can be used to treat retinopathies, particularly diabetic retinopathy. The methods disclosed herein can also be used to treat macular degenerations, particularly age-related macular degeneration. As described below, eye disorders such as diabetic retinopathy and age-related macular degeneration are associated with a decrease in retinal potential. Thus, this specification also provides methods of improving retinal potential in a subject in need thereof (e.g., a subject having diabetic retinopathy or age-related macular degeneration).

[0145] In some embodiments, the method disclosed herein comprises administering to the subject an antagonist to the FAM19A5 protein ("FAM19A5 antagonist"). In some embodiments, the FAM19A5 antagonist is an antisense oligonucleotide, siRNA, shRNA, miRNA, dsRNA-targeted FAM19A5, an aptamer, PNA, or a vector comprising the same. In other embodiments, the FAM19A5 antagonist is an antibody or an antigen-binding moiety thereof that specifically binds to the FAM19A5 protein ("anti-FAM19A5 antibody"), a polynucleotide encoding the anti-FAM19A5 antibody, or a vector comprising the polynucleotide. In some embodiments, the anti-FAM19A5 antibody binds to the FAM19A5 protein and reduces FAM19A5 activity.

[0146] Subjects with retinal diseases (e.g., diabetic retinopathy) and / or macular degeneration (e.g., age-related macular degeneration) may exhibit one or more of the following features in the retina: (i) decreased electroretinography, (ii) perivascular cell loss, (iii) increased acellular capillary formation, (iv) increased vascular congestion (e.g., in the nerve fiber layer of the retina), (v) increased cell death (e.g., in the nerve fiber layer and / or inner core layer of the retina), (vi) increased retinal and / or choroidal neovascularization, and (vii) increased astrocytocyte formation (e.g., evidenced by increased PDGF staining in the nerve fiber layer and / or ganglion cell layer of the retina). See Examples 6-10.

[0147] Therefore, regardless of any single theory, the FAM19A5 antagonists disclosed herein (e.g., anti-FAM19A5 antibodies) can treat retinal diseases and / or macular degeneration by reducing, mitigating, and / or reversing one or more of the characteristics described above. In some embodiments, the FAM19A5 antagonist is, for example, a FAM19A5 antagonist that can neutralize the FAM19A5 protein. In other embodiments, the FAM19A5 antagonist is an anti-FAM19A5 antibody. In some embodiments, the anti-FAM19A5 antibody is a neutralizing antibody.

[0148] In some embodiments, FAM19A5 antagonists can improve the electroretinogram (EEG) in subjects (e.g., subjects with diabetic retinopathy or age-related macular degeneration). In certain embodiments, the EEG improvement includes an increase in values ​​for wave A, wave B, and / or rhythmic wave patterns compared to a baseline (e.g., the corresponding values ​​for subjects with DR or AMD who have not been treated with the FAM19A5 antagonists disclosed herein). In some embodiments, the values ​​for wave A, wave B, and / or rhythmic wave patterns are at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% or more compared to the baseline.

[0149] In some embodiments, the FAM19A5 antagonists disclosed herein (e.g., anti-FAM19A5 antibodies) can reduce or prevent the loss of intraretinal perivascular cells in subjects (e.g., subjects with diabetic retinopathy or age-related macular degeneration). In certain embodiments, the loss of intraretinal perivascular cells is reduced by at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to a baseline (e.g., the relevant value for DR or AMD subjects not treated with the FAM19A5 antagonists disclosed herein).

[0150] In some embodiments, the FAM19A5 antagonists of this disclosure can reduce and / or inhibit intraretinal acellular capillary formation in subjects (e.g., subjects with diabetic retinopathy or age-related macular degeneration). In certain embodiments, the acellular capillary formation is reduced and / or inhibited by at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to a baseline (e.g., the relevant value for DR or AMD subjects not treated with the FAM19A5 antagonists disclosed herein).

[0151] In some embodiments, FAM19A5 antagonists (e.g., anti-FAM19A5 antibodies) can reduce and / or inhibit intraretinal vascular congestion in subjects (e.g., subjects with diabetic retinopathy or age-related macular degeneration). In certain embodiments, the intraretinal vascular congestion is reduced and / or inhibited by at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to a baseline (e.g., the corresponding value in a DR or AMD subject untreated with a FAM19A5 antagonist disclosed herein).

[0152] In some embodiments, the FAM19A5 antagonists disclosed herein can reduce and / or inhibit cell death of retinal cells in a subject (e.g., a subject with diabetic retinopathy or age-related macular degeneration). In certain embodiments, the number of retinal cells undergoing such cell death is reduced by at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to a baseline (e.g., the relevant value for a DR or AMD subject not treated with the FAM19A5 antagonists disclosed herein).

[0153] In some embodiments, FAM19A5 antagonists can reduce and / or inhibit intraretinal neovascularization (e.g., retinal and / or choroidal neovascularization) in subjects (e.g., subjects with diabetic retinopathy or age-related macular degeneration). In certain embodiments, intraretinal neovascularization (e.g., retinal and / or choroidal neovascularization) is reduced and / or inhibited by at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to a baseline (e.g., the corresponding value for DR or AMD subjects not treated with the FAM19A5 antagonists disclosed herein).

[0154] In some embodiments, FAM19A5 antagonists reduce and / or inhibit intraretinal PDGF in subjects (e.g., subjects with diabetic retinopathy or age-related macular degeneration). In certain embodiments, intraretinal PDGF expression is reduced and / or inhibited by at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to a baseline (e.g., the relevant value for DR or AMD subjects untreated with the FAM19A5 antagonists disclosed herein). In some embodiments, the reduction and / or inhibition of PDGF expression is associated with reduced intraretinal astrocytocyte formation in the subjects.

[0155] In some embodiments, the term “criteria” as used herein refers to the subject(s) who have not been administered with the composition disclosed herein (e.g., a FAM19A5 antagonist, e.g., an anti-FAM19A5 antibody) (e.g., a subject with diabetic retinopathy or age-related macular degeneration). The term “criteria” may also refer to the same subject(s) before administration with the composition disclosed herein (e.g., a subject with diabetic retinopathy or age-related macular degeneration). In certain embodiments, the term “criteria” refers to the average of a population of subjects(s) (e.g., a subject with diabetic retinopathy or age-related macular degeneration).

[0156] In some embodiments, the retinal diseases treatable by this disclosure include diabetic retinal diseases. In certain embodiments, diabetic retinal diseases are non-proliferative diabetic retinal diseases (NPDRs). In other embodiments, diabetic retinal diseases are proliferative diabetic retinal diseases (PDRs). In some embodiments, diabetic retinal diseases are diabetic macular diseases. In other embodiments, diabetic retinal diseases are diabetic macular edema. In some embodiments, diabetic retinal diseases are any retinal diseases associated with intraretinal ischemic injury.

[0157] In some embodiments, the macular degeneration treatable by the present disclosure includes age-related macular degeneration (AMD). In some embodiments, the age-related macular degeneration is early-stage AMD. In other embodiments, the age-related macular degeneration is mid-stage AMD. In other embodiments, the age-related macular degeneration is late-stage or progressive AMD (i.e., geographic atrophy). In some embodiments, the age-related macular degeneration is dry (non-exudative) AMD. In other embodiments, the age-related macular degeneration is wet (neovascular or exudative) AMD.

[0158] In some embodiments, the subjects that can be treated by this method are non-human animals such as rats or mice. In other embodiments, the subjects that can be treated by the method disclosed herein are humans.

[0159] In some embodiments, the FAM19A5 antagonists disclosed herein (e.g., anti-FAM19A5 antibodies), bispecific molecules, immunoconjugates, or compositions thereof are delivered to a subject by intraocular administration or by any other administration capable of delivering the compositions disclosed herein (e.g., FAM19A5 antagonists) to sub-Tenon's capsule, subconjunctival, suprachoroidal, intravitreous, and similar locations within the eye. In some embodiments, intraocular administration includes intravitreous administration.

[0160] The dosage for administering the FAM19A5 antagonists or compositions thereof disclosed herein (e.g., anti-FAM19A5 antibodies) is in the range of about 0.0001 to 100 mg / kg.

[0161] In some embodiments, the FAM19A5 antagonist or composition thereof can be administered in combination with one or more additional agents (e.g., therapeutic agents), such as standard therapies, for treating retinal diseases (e.g., diabetic retinopathy) and / or macular degeneration (e.g., age-related macular degeneration), such as (i) corticosteroids, (ii) anti-angiogenic inhibitors (e.g., anti-VEGF inhibitors), (iii) laser therapy (e.g., laser photocoagulation), (iv) antioxidants. Other currently available therapeutic options that can be used in combination with the FAM19A5 antagonist or composition thereof include, but are not limited to, surgery (e.g., vitrectomy) and modulation of modifiable risk factors (e.g., reduction of glucose and cholesterol levels, smoking cessation).

[0162] <III.FAM19A5 Antagonist> In some embodiments, the FAM19A5 antagonist useful in the present disclosure is an antisense oligonucleotide, siRNA, shRNA, miRNA, dsRNA, aptamer, PNA (peptide nucleic acid) or a vector containing the same that specifically targets FAM19A5. In other embodiments, the FAM19A5 antagonist is an antibody or antigen-binding portion thereof that specifically binds to the FAM19A5 protein (“anti-FAM19A5 antibody”), a polynucleotide encoding the anti-FAM19A5 antibody or a vector containing the polynucleotide thereof.

[0163] Antibodies useful in the methods disclosed herein include monoclonal antibodies characterized by certain functional features or properties. For example, the antibodies specifically bind to human FAM19A5, including soluble FAM19A5 and membrane-bound FAM19A5. In addition to specifically binding to soluble and / or membrane-bound human FAM19A5, the antibodies described herein also (a) bind to soluble human FAM19A5 with a K D of 10 nM or less; (b) bind to membrane-bound human FAM19A5 with a K D of 10 nM or less; or both (a) and (b).

[0164] In some embodiments, the anti-FAM19A5 antibody or antigen-binding portion thereof has a high affinity, e.g., a KD 10 -7 M or less, 10 -8 M or less, 10 -9 M (1nM) or less, 10 -10 M (10nM) or less, 10 -11 M or less or 10 -12 M or less, for example, 10 -12 M~10 -7 M, 10 -11 M~10 -7 M, 10 -10 M~10 -7 M or 10 -9 M~10 -7 M, for example 10 -12 M, 5×10 -12 M, 10 -11 M, 5×10 -11 M, 10 -10 M, 5×10 -10 M, 10 -9 M, 5×10 -9 M, 10 -8 M, 5×10 -8 M, 10 -7 M or 5×10 -7 M specifically binds to soluble human FAM19A5 or membrane-bound human FAM19A5. Standard analytical methods for evaluating the binding ability of antibodies to various species of human FAM19A5 are known in the art, including, for example, ELISA, Western blotting, and RIA. Preferred analytical methods are described in detail in the Examples section. The binding kinetics (e.g., binding affinity) of the antibody are determined by ELISA, BIACORE TM It may be evaluated by analysis or by standard analytical methods well known in the industry, such as KINEXA®.

[0165] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety, when determined by, for example, ELISA, is K D 10 -7 M or less, 10 -8 M (10nM) or less, 10 -9 M (1nM) or less, 10 -10 M or less, 10 -12 M~10 -7 M, 10 -11 M~10 -7 M, 10-10 M to 10 -7 M, 10 -9 M to 10 -7 M, or 10 -8 M to 10 -7 is M and binds to soluble human FAM19A5. In some embodiments, the anti - FAM19A5 antibody or antigen - binding portion thereof has a K D that is 10 nM or less, such as 0.1 - 10 nM, 0.1 - 5 nM, 0.1 - 1 nM, 0.5 - 10 nM, 0.5 - 5 nM, 0.5 - 1 nM, 1 - 10 nM, 1 - 5 nM, or 5 - 10 nM, and binds to soluble FAM19A5. In some embodiments, the anti - FAM19A5 antibody or antigen - binding portion thereof has a K D that is about 1 pM, 2 pM, 3 pM, 4 pM, 5 pM, 6 pM, 7 pM, 8 pM, 9 pM, 10 pM, 20 pM, 30 pM, 40 pM, 50 pM, 60 pM, 70 pM, 80 pM, 90 pM, 100 pM, 200 pM, 300 pM, 400 pM, 500 pM, 600 pM, 700 pM, 800 pM, or 900 pM, or about 1 nM, 2 nM, 3 nM, 4 nM, 5 nM, 6 nM, 7 nM, 8 nM, or 9 nM, or about 10 nM, 20 nM, 30 nM, 40 nM, 50 nM, 60 nM, 70 nM, 80 nM, or 90 nM, and specifically binds to soluble human FAM19A5.

[0166] In some embodiments, the anti - FAM19A5 antibody or antigen - binding portion thereof has a K D that is 10 -7 M or less, 10 -8 M (10 nM) or less, 10 -9 M (1 nM) or less, 10 -10 M or less, 10 -12 M to 10 -7 M, 10 -11 M to 10 -7 M, 10 -10 M to 10 -7 M, 10 -9 M to 10 -7 M or 10 -8 M to 10 -7M binds to membrane-bound human FAM19A5. In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety is determined by ELISA to be K D It specifically binds to membrane-bound human FAM19A5 with a K content of 10 nM or less, for example, 0.1-10 nM, 0.1-5 nM, 0.1-1 nM, 0.5-10 nM, 0.5-5 nM, 0.5-1 nM, 1-10 nM, 1-5 nM, or 5-10 nM. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety, when determined by ELISA, is K D It binds to membrane-bound human FAM19A5 in concentrations of approximately 1 pM, 2 pM, 3 pM, 4 pM, 5 pM, 6 pM, 7 pM, 8 pM, 9 pM, 10 pM, 20 pM, 30 pM, 40 pM, 50 pM, 60 pM, 70 pM, 80 pM, 90 pM, 100 pM, 200 pM, 300 pM, 400 pM, 500 pM, 600 pM, 700 pM, 800 pM or 900 pM, or approximately 1 nM, 2 nM, 3 nM, 4 nM, 5 nM, 6 nM, 7 nM, 8 nM or 9 nM, or approximately 10 nM, 20 nM, 30 nM, 40 nM, 50 nM, 60 nM, 70 nM, 80 nM or 90 nM.

[0167] In some embodiments, an anti-FAM19A5 antibody or its antigen-binding moiety useful in the methods disclosed herein cross-compete with an anti-FAM19A5 antibody containing a CDR or variable region disclosed herein for binding to (or inhibiting) the human FAM19A5 epitope.

[0168] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety inhibits the binding of a reference antibody comprising heavy chain CDR1, CDR2, and CDR3, and light chain CDR1, CDR2, and CDR3, wherein (i) the heavy chain CDR1, CDR2, and CDR3 of the reference antibody each contain the amino acid sequences of SEQ ID NO: 11, SEQ ID NO: 12, and SEQ ID NO: 13, and the light chain CDR1, CDR2, and CDR3 of the reference antibody each contain the amino acid sequences of SEQ ID NO: 23, SEQ ID NO: 24, and SEQ ID NO: 25; (ii) the heavy (iii) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 14, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 15, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 16, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 26, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 27, the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 28; (iii) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 17, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 18, the (iv) The heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 19, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 29, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 30, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 31; (iv) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 20, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 21, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 22, and the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 32. (v) The light chain CDR2 contains the amino acid sequence of SEQ ID NO: 33, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 34; (v) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 89, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 90, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 91, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 92, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 93, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 94;(vi) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 95, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 96, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 97, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 98, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 99, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 100; (vii) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 101, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: (viii) The heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 103, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 104, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 105, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 106; (viii) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 107, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 108, and the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 109 (ix) The light chain CDR1 contains the amino acid sequence of SEQ ID NO: 110, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 111, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 112; (ix) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 113, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 114, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 115, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 116, and the light chain CDR2 contains SEQ ID NO: (x) The light chain CDR3 contains the amino acid sequence of SEQ ID NO: 117, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 118; (x) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 119, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 120, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 121, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 122, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 123, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 124;(xi) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 125, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 126, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 127, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 128, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 129, the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 130; (xii) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 131, the heavy chain CDR2 contains (xiii) The heavy chain CDR3 contains the amino acid sequence of sequence number 133, the light chain CDR1 contains the amino acid sequence of sequence number 134, the light chain CDR2 contains the amino acid sequence of sequence number 135, and the light chain CDR3 contains the amino acid sequence of sequence number 136; (xiii) The heavy chain CDR1 contains the amino acid sequence of sequence number 137, the heavy chain CDR2 contains the amino acid sequence of sequence number 138, and the heavy chain CDR3 contains the amino acid sequence of sequence number 139 (xiv) The light chain CDR1 contains the amino acid sequence of SEQ ID NO: 140, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 141, the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 142; (xiv) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 143, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 144, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 145, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 146, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: (xv) The light chain CDR3 contains the amino acid sequence of SEQ ID NO. 147, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO. 148; or (xv) the heavy chain CDR1 contains the amino acid sequence of SEQ ID NO. 149, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO. 150, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO. 151, the light chain CDR1 contains the amino acid sequence of SEQ ID NO. 152, the light chain CDR2 contains the amino acid sequence of SEQ ID NO. 153, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO. 154.

[0169] In some embodiments, the reference antibody is (a) heavy chain and light chain variable region sequences including SEQ ID NOs. 35 and 39, respectively; (b) heavy chain and light chain variable region sequences including SEQ ID NOs. 36 and 40, respectively; (c) heavy chain and light chain variable region sequences including SEQ ID NOs. 37 and 41, respectively; (d) heavy chain and light chain variable region sequences including SEQ ID NOs. 38 and 42, respectively; (e) heavy chain and light chain variable region sequences including SEQ ID NOs. 155 and 166, respectively; (f) heavy chain and light chain variable region sequences including SEQ ID NOs. 156 and 167, respectively; (g) heavy chain and light chain variable region sequences including SEQ ID NOs. 157 and 168, respectively; (h) heavy chain and light chain variable region sequences including SEQ ID NOs. 158 and (i) Heavy chain and light chain variable region sequences including sequence numbers 159 and 170, respectively; (j) Heavy chain and light chain variable region sequences including sequence numbers 160 and 171, respectively; (k) Heavy chain and light chain variable region sequences including sequence numbers 161 and 172, respectively; (l) Heavy chain and light chain variable region sequences including sequence numbers 162 and 173, respectively; (m) Heavy chain and light chain variable region sequences including sequence numbers 163 and 174, respectively; (n) Heavy chain and light chain variable region sequences including sequence numbers 164 and 175, respectively; or (o) Heavy chain and light chain variable region sequences including sequence numbers 165 and 176, respectively.

[0170] In certain embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety inhibits the binding of such reference antibodies to human FAM19A5 by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%. Competitive antibodies bind to the same epitope, overlapping epitope, or adjacent epitope (e.g., demonstrated by steric hindrance). Whether two antibodies compete with each other for binding to a target can be determined using competition experiments known in the art, such as RIA and EIA.

[0171] In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety binds to the same FAM19A5 epitope as the reference antibody disclosed herein, which comprises heavy chain CDR1, CDR2 and CDR3, and light chain CDR1, CDR2 and CDR3, wherein (i) the heavy chain CDR1 comprises the amino acid sequence of SEQ ID NO: 11, the heavy chain CDR2 comprises the amino acid sequence of SEQ ID NO: 12, the heavy chain CDR3 comprises the amino acid sequence of SEQ ID NO: 13, the light chain CDR1 comprises the amino acid sequence of SEQ ID NO: 23, and the light chain CDR2 comprises the amino acid sequence of SEQ ID NO: 24 (ii) The light chain CDR3 contains an amino acid sequence, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 25; (ii) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 14, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 15, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 16, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 26, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 27, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 28; (iii) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 17 (iv) The heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 18, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 19, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 29, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 30, the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 31; (iv) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 20, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 21, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 22, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: (v) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO. 32, the light chain CDR2 contains the amino acid sequence of SEQ ID NO. 33, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO. 34; (v) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO. 89, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO. 90, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO. 91, the light chain CDR1 contains the amino acid sequence of SEQ ID NO. 92, the light chain CDR2 contains the amino acid sequence of SEQ ID NO. 93, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO. 94;(vi) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 95, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 96, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 97, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 98, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 99, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 100; (vii) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 101, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: (viii) The heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 103, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 104, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 105, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 106; (viii) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 107, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 108, and the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 109 (ix) The light chain CDR1 contains the amino acid sequence of SEQ ID NO: 110, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 111, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 112; (ix) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 113, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 114, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 115, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 116, and the light chain CDR2 contains SEQ ID NO: (x) The light chain CDR3 contains the amino acid sequence of SEQ ID NO: 117, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 118; (x) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 119, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 120, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 121, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 122, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 123, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 124;(xi) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 125, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 126, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 127, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 128, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 129, the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 130; (xii) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 131, the heavy chain CDR2 contains (xiii) The heavy chain CDR3 contains the amino acid sequence of sequence number 133, the light chain CDR1 contains the amino acid sequence of sequence number 134, the light chain CDR2 contains the amino acid sequence of sequence number 135, and the light chain CDR3 contains the amino acid sequence of sequence number 136; (xiii) The heavy chain CDR1 contains the amino acid sequence of sequence number 137, the heavy chain CDR2 contains the amino acid sequence of sequence number 138, and the heavy chain CDR3 contains the amino acid sequence of sequence number 139 (xiv) The light chain CDR1 contains the amino acid sequence of SEQ ID NO: 140, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: 141, the light chain CDR3 contains the amino acid sequence of SEQ ID NO: 142; (xiv) The heavy chain CDR1 contains the amino acid sequence of SEQ ID NO: 143, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO: 144, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO: 145, the light chain CDR1 contains the amino acid sequence of SEQ ID NO: 146, the light chain CDR2 contains the amino acid sequence of SEQ ID NO: (xv) The light chain CDR3 contains the amino acid sequence of SEQ ID NO. 147, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO. 148; or (xv) the heavy chain CDR1 contains the amino acid sequence of SEQ ID NO. 149, the heavy chain CDR2 contains the amino acid sequence of SEQ ID NO. 150, the heavy chain CDR3 contains the amino acid sequence of SEQ ID NO. 151, the light chain CDR1 contains the amino acid sequence of SEQ ID NO. 152, the light chain CDR2 contains the amino acid sequence of SEQ ID NO. 153, and the light chain CDR3 contains the amino acid sequence of SEQ ID NO. 154.

[0172] In some embodiments, the reference antibody is (a) heavy chain and light chain variable region sequences including SEQ ID NOs. 35 and 39, respectively; (b) heavy chain and light chain variable region sequences including SEQ ID NOs. 36 and 40, respectively; (c) heavy chain and light chain variable region sequences including SEQ ID NOs. 37 and 41, respectively; (d) heavy chain and light chain variable region sequences including SEQ ID NOs. 38 and 42, respectively; (e) heavy chain and light chain variable region sequences including SEQ ID NOs. 155 and 166, respectively; (f) heavy chain and light chain variable region sequences including SEQ ID NOs. 156 and 167, respectively; (g) heavy chain and light chain variable region sequences including SEQ ID NOs. 157 and 168, respectively; (h) heavy chain and light chain variable region sequences including SEQ ID NOs. 158 and (i) Heavy chain and light chain variable region sequences including sequence numbers 159 and 170, respectively; (j) Heavy chain and light chain variable region sequences including sequence numbers 160 and 171, respectively; (k) Heavy chain and light chain variable region sequences including sequence numbers 161 and 172, respectively; (l) Heavy chain and light chain variable region sequences including sequence numbers 162 and 173, respectively; (m) Heavy chain and light chain variable region sequences including sequence numbers 163 and 174, respectively; (n) Heavy chain and light chain variable region sequences including sequence numbers 164 and 175, respectively; or (o) Heavy chain and light chain variable region sequences including sequence numbers 165 and 176, respectively.

[0173] Techniques for determining whether two antibodies bind to the same epitope include, for example, epitope mapping methods such as X-ray analysis of antigen:antibody complex crystals and hydrogen / deuterium exchange mass spectrometry (HDX-MS) that provide atomic resolution of the epitope; methods for monitoring the binding of antibodies to antigen fragments or mutated variants of antigens, where binding loss due to deformation of amino acid residues within the antigen sequence is usually considered an indication of the epitope component; and computational combinatorial methods for epitope mapping.

[0174] An anti-FAM19A5 antibody or its antigen-binding moiety useful in the methods disclosed herein can bind to at least one epitope of mature human FAM19A5, as determined, for example, by the binding of the antibody to a fragment of human FAM19A5. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds to a fragment located within the amino acid sequence of TLDRDSSQPRRTIARQTARC (amino acid residues 42-61 of SEQ ID NO: 6 or SEQ ID NO: 2), for example, an epitope having at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 amino acids of SEQ ID NO: 6. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds to SEQ ID NO: 6 with one or more amino acids corresponding to amino acid residues 46-51 (i.e., DSSQPR), for example amino acid residues 46, 50 and 52 (i.e., D---PR), for example amino acid residues 46, 47, 48 and 50 (i.e., DSS-P) of SEQ ID NO: 2. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds to a fragment located within the amino acid sequence of CDMLPCLEGEGCDLLINRSG (amino acids 90-109 of SEQ ID NO: 9 or SEQ ID NO: 2), for example, an epitope having at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 amino acids of SEQ ID NO: 9. In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety binds to one or more amino acid residues 99-107 (i.e., EGCDLLINR), for example amino acid residues 102, 103, 105 and 107 (i.e., DL-IR), for example amino acid residues 99, 100, 102, 103, 105 and 107 (i.e., EG-DL-IR), for example amino acid residues 99, 100 and 107 of SEQ ID NO: 4 (i.e., EG------R), at SEQ ID NO: 9.

[0175] In some embodiments, the at least one epitope has the same amino acid sequence as SEQ ID NO: 6 by at least 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100%. In some embodiments, the at least one epitope has the same amino acid sequence as SEQ ID NO: 9 by at least 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100%.

[0176] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds only to the human FAM19A5 epitope of SEQ ID NOs. 5, 6, 7, 8, 9, or 10, or to fragments located within the amino acid sequence of SEQ ID NOs. 5, 6, 7, 8, 9, or 10, for example, to an epitope having 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 amino acids of SEQ ID NOs.

[0177] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety of the Disclosure binds to SEQ ID NO: 6 or its native stereomorphic form (i.e., undenatured) fragment. In some embodiments, the anti-FAM19A5 antibody binds to SEQ ID NO: 9 or its native stereomorphic form (i.e., undenatured) fragment. In other embodiments, the at least one epitope has the same amino acid sequence as SEQ ID NO: 5 by at least 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100%. The epitope has the same amino acid sequence as SEQ ID NO: 10 by at least 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least 99%, or about 100%. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds to both glycosylated and unglycosylated human FAM19A5.

[0178] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety further binds to one or more additional FAM19A5 epitopes. Thus, a specific anti-FAM19A5 antibody or its antigen-binding moiety binds to the epitope of SEQ ID NO: 6 and the additional epitope, or to the epitope of SEQ ID NO: 9 and the additional epitope. Other anti-FAM19A5 antibodies or their antigen-binding moieties can bind to the epitopes of SEQ ID NO: 5, SEQ ID NO: 9, and the additional epitopes. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds to the epitope of SEQ ID NO: 6, the epitope of SEQ ID NO: 10, and the additional epitopes.

[0179] In some embodiments, the one or more additional FAM19A5 epitopes are selected from QLAAGTCEIVTLDR (SEQ ID NO: 5, Epitope F1), TLDRDSSQPRRTIARQTARC (SEQ ID NO: 6, Epitope F2), TARCACRKGQIAGTTRARPA (SEQ ID NO: 7, Epitope F3), ARPACVDARIIKTKQWCDML (SEQ ID NO: 8, Epitope F4), CDMLPCLEGEGCDLLINRSG (SEQ ID NO: 9, Epitope F5), or NRSGWTCTQPGGRIKTTTVS (SEQ ID NO: 10, Epitope F6), or fragments located within the amino acid sequence of SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, or SEQ ID NO: 10, or any combination thereof. A fragment located within the amino acid sequence of SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, or SEQ ID NO: 10 includes fragments having any 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 amino acids from SEQ ID NO: 5, SEQ ID NO: 6 and SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, or SEQ ID NO: 10. In some embodiments, the one or more additional FAM19A5 epitopes are selected from SEQ ID NO: 5, 6, 7, 8, 9, or 10, or fragments located within the amino acid sequence of SEQ ID NO: 5, 6, 7, 8, 9, or 10, for example, fragments having 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 amino acids from SEQ ID NO: 5, 6, 7, 8, 9, or 10, or any combination thereof. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety of the Disclosure binds to one or more of the aforementioned additional epitopes in their native stereomorphic form (i.e., undenatured). In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds to both one or more of the aforementioned glycosylated and unglycosylated FAM19A5 epitopes.

[0180] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety of the Disclosure binds to at least one FAM19A5 epitope identified as EP2, EP4, and / or EP8, wherein EP2 contains, is essentially composed of, or consists of the amino acid DSSQP (SEQ ID NO: 66), EP4 contains, is essentially composed of, or consists of the amino acid ARCACRK (SEQ ID NO: 68), and EP8 contains, is essentially composed of, or consists of the amino acid TCTQPGGR (SEQ ID NO: 72). In some embodiments, the at least one epitope has the same amino acid sequence as EP2, EP4, or EP8 by at least 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100%. In some embodiments, only the anti-FAM19A5 antibody or its antigen-binding moiety binds to EP2. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds to EP4 and EP8.

[0181] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds to at least one FAM19A5 epitope identified as EP6, EP7, or EP8, wherein EP6 contains the amino acid KTKQWCDML (SEQ ID NO: 70), EP7 contains the amino acid GCDLINR (SEQ ID NO: 71), and EP8 contains the amino acid TCTQPGGR (SEQ ID NO: 72). In some embodiments, the at least one epitope has the same amino acid sequence as EP6, EP7, or EP8 by at least 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100%. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds only to EP6, EP7, or EP8. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds to EP6, EP7, and EP8. In some embodiments, the anti-FAM9A5 antibody or its antigen-binding moiety binds to EP7 and EP8. In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety binds to EP7.

[0182] In some embodiments, the anti-FAM19A5 antibody or antigen-binding portion thereof binds to one or more FAM19A5 epitopes selected from the group consisting of SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, and any combination thereof.

[0183] In some embodiments, the present disclosure provides an antibody or antigen-binding portion thereof that binds to FAM19A5 (e.g., human FAM19A5) with an affinity that is 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or higher compared to other proteins in the FAM19A family when measured, for example, by immunoassay (e.g., ELISA), surface plasmon resonance or size exclusion chromatography. In certain embodiments, the present disclosure provides an antibody or antigen-binding portion thereof that binds to FAM19A5 (e.g., human FAM19A5) without cross-reactivity with further other proteins in the FAM19A family when measured, for example, by immunoassay.

[0184] In some embodiments, the anti-FAM19A5 antibody is not a natural antibody or a naturally occurring antibody. For example, the anti-FAM19A5 antibody has post-translational modifications that are different from those of naturally occurring antibodies, such as having more or fewer or different types of post-translational modifications.

[0185] <IV. Exemplary Anti-FAM19A5 Antibodies> The specific antibodies usable in the methods disclosed herein are antibodies having the CDR and / or variable region sequences disclosed herein, for example, not only monoclonal antibodies but also antibodies having at least 80% identity (e.g., at least 85%, at least 90%, at least 95%, or 99% identity) to these variable region or CDR sequences. The amino acid sequences for the VH and VL CDRs for the different anti-FAM19A5 antibodies are provided in Tables 2 and 3, respectively. The CDRs for the following antibodies were identified using the Kabat numbering system (see above): 1-65, 3-2, 2-13, 1-28, P2-C12, 13B4, 13F7, 15A9, Pl-A03, P1-A08, P1-F02, P2-A01, P2-A03, P2-F07, P2-F11, SS01-13, SS0l-13-s5, and S5-2.GKNG. The CDRs for the following antibodies were identified using the IMGT numbering system (see above): 1-7A-IT, Low-PI, 1-30, 1-17, 1-32, 4-11, 6-10, 2-13D, 2-13D-37, 2-13D-37-1.5W-41, and 2-13D-37-3W-16. The VH and VL amino acid sequences for the individual anti-FAM19A5 antibodies in this disclosure are provided in Tables 4 and 5, respectively.

[0186] [Table 3]

[0187] JPEG0007868876000004.jpg210169

[0188] JPEG0007868876000005.jpg152169

[0189] [Table 4]

[0190] JPEG0007868876000007.jpg220169

[0191] JPEG0007868876000008.jpg141169

[0192] [Table 5]

[0193] JPEG0007868876000010.jpg221169

[0194] JPEG0007868876000011.jpg164169

[0195] [Table 6]

[0196] JPEG0007868876000013.jpg218169

[0197] JPEG0007868876000014.jpg184169

[0198] Accordingly, this specification provides an isolated anti-FAM19A5 antibody or its antigen-binding moiety comprising a heavy chain and a light chain variable region, wherein the heavy chain variable region comprises the amino acid sequence of SEQ ID NOs: 35-38, 155-165, or 232-240 (see Table 4). In another embodiment, the isolated anti-FAM19A5 antibody or its antigen-binding moiety comprises a CDR of a heavy chain variable region selected from the group consisting of SEQ ID NOs: 35-38, 155-165, or 232-240 (see Table 4).

[0199] This specification also includes heavy chain and light chain variable regions, the light chain variable region providing an anti-FAM19A5 antibody or its antigen-binding moiety comprising the amino acid sequence of SEQ ID NOs: 39-42, 166-176, or 241-250 (see Table 5). In other embodiments, the isolated anti-FAM19A5 antibody or its antigen-binding moiety comprises a CDR of a light chain variable region selected from the group consisting of SEQ ID NOs: 39-42, 166-176, or 241-250 (see Table 5).

[0200] In a particular embodiment, the isolated anti-FAM19A5 antibody or its antigen-binding moiety includes a heavy chain variable region CDR selected from the group consisting of SEQ ID NOs: 35-38, 155-165, or 232-240, and a light chain variable region CDR selected from the group consisting of SEQ ID NOs: 39-42, 166-176, or 241-250.

[0201] This specification also includes heavy chain and light chain variable regions, (i) the heavy chain variable region includes the amino acid sequence of SEQ ID NO: 37 and the light chain variable region includes the amino acid sequence of SEQ ID NO: 39; (ii) the heavy chain variable region includes the amino acid sequence of SEQ ID NO: 36 and the light chain variable region includes the amino acid sequence of SEQ ID NO: 40; (iii) the heavy chain variable region includes the amino acid sequence of SEQ ID NO: 37 and the light chain variable region includes the amino acid sequence of SEQ ID NO: 41; (iv) the heavy chain variable region includes the amino acid sequence of SEQ ID NO: 38 and the light chain variable region is (v) the heavy chain variable region includes the amino acid sequence of SEQ ID NO: 42; (vi) the heavy chain variable region includes the amino acid sequence of SEQ ID NO: 155, and the light chain variable region includes the amino acid sequence of SEQ ID NO: 166; (vi) the heavy chain variable region includes the amino acid sequence of SEQ ID NO: 156, and the light chain variable region includes the amino acid sequence of SEQ ID NO: 167; (vii) the heavy chain variable region includes the amino acid sequence of SEQ ID NO: 157, and the light chain variable region includes the amino acid sequence of SEQ ID NO: 168; (viii) the heavy chain variable region includes the amino acid sequence of SEQ ID NO: 158, and the light chain variable (ix) The variable region includes the amino acid sequence of SEQ ID NO: 169; (x) The heavy chain variable region includes the amino acid sequence of SEQ ID NO: 159, and the light chain variable region includes the amino acid sequence of SEQ ID NO: 170; (xi) The heavy chain variable region includes the amino acid sequence of SEQ ID NO: 160, and the light chain variable region includes the amino acid sequence of SEQ ID NO: 171; (xii) The heavy chain variable region includes the amino acid sequence of SEQ ID NO: 161, and the light chain variable region includes the amino acid sequence of SEQ ID NO: 172; (xii) The heavy chain variable region includes the amino acid sequence of SEQ ID NO: 162, and the The present invention provides an anti-FAM19A5 antibody or its antigen-binding moiety, wherein the light chain variable region comprises the amino acid sequence of SEQ ID NO: 173; (xiii) the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 163 and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 174; (xiv) the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 164 and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 175; and (xv) the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 165 and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 176.In some embodiments, the isolated anti-FAM19A5 antibody or its antigen-binding fragment comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises the VH sequence shown in Table 4 and the VL comprises the VL sequence shown in Table 5.

[0202] This specification provides isolated anti-FAM19A5 antibodies or their antigen-binding moieties, comprising a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region contains at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% the same amino acid sequence as the amino acid sequences shown in SEQ ID NOs. 35-38, 155-165, or 232-240 (see Table 4).

[0203] This specification also provides isolated anti-FAM19A5 antibodies or their antigen-binding moieties, comprising heavy chain variable regions and light chain variable regions, wherein the light chain variable regions contain at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% the same amino acid sequence as the amino acid sequences shown in SEQ ID NOs. 39-42, 166-176, or 241-250 (see Table 5).

[0204] This specification also provides isolated anti-FAM19A5 antibodies or their antigen-binding moieties, comprising heavy chain and light chain variable regions, wherein the heavy chain variable regions contain at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% the same amino acid sequence as the amino acid sequence shown in SEQ ID NOs. 35-38, 155-165, or 232-240 (see Table 4), and the light chain variable regions contain at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% the same amino acid sequence as the amino acid sequence shown in SEQ ID NOs. 39-42, 166-176, or 241-250 (see Table 5).

[0205] In some embodiments, this disclosure includes: (a) Heavy chain and light chain variable region sequences containing sequence numbers 35 and 39, respectively; (b) Heavy chain and light chain variable region sequences containing sequence numbers 36 and 40, respectively; (c) Heavy chain and light chain variable region sequences containing sequence numbers 37 and 41, respectively; (d) Heavy chain and light chain variable region sequences containing sequence numbers 38 and 42, respectively; (e) Heavy chain and light chain variable region sequences containing sequence numbers 155 and 166, respectively; (f) Heavy chain and light chain variable region sequences containing sequence numbers 156 and 167, respectively; (g) Heavy chain and light chain variable region sequences containing sequence numbers 157 and 168, respectively; (h) Heavy chain and light chain variable region sequences containing sequence numbers 158 and 169, respectively; (i) Heavy chain and light chain variable region sequences containing sequence numbers 159 and 170, respectively; (j) Heavy chain and light chain variable region sequences containing sequence numbers 160 and 171, respectively; (k) Heavy chain and light chain variable region sequences containing sequence numbers 161 and 172, respectively; (l) Heavy chain and light chain variable region sequences containing sequence numbers 162 and 173, respectively; (m) Heavy chain and light chain variable region sequences containing sequence numbers 163 and 174, respectively; (n) Heavy chain and light chain variable region sequences containing sequence numbers 164 and 175, respectively; (o) Heavy chain and light chain variable region sequences containing sequence numbers 165 and 176, respectively; (p) Heavy and light chain variable region sequences containing sequence numbers 232 and 241, respectively; (q) Heavy chain and light chain variable region sequences containing sequence numbers 233 and 242, respectively; (r) Heavy chain and light chain variable region sequences containing sequence numbers 234 and 242, respectively; (s) Heavy chain and light chain variable region sequences containing sequence numbers 235 and 243, respectively; (t) Heavy chain and light chain variable region sequences containing sequence numbers 236 and 244, respectively; (u) Heavy chain and light chain variable region sequences containing sequence numbers 236 and 245, respectively; (v) Heavy chain and light chain variable region sequences containing sequence numbers 236 and 246, respectively; (w) Heavy chain and light chain variable region sequences containing sequence numbers 236 and 247, respectively; (x) Heavy chain and light chain variable region sequences containing sequence numbers 236 and 248, respectively; (y) Heavy chain and light chain variable region sequences containing sequence numbers 236 and 249, respectively; (z) Heavy and light chain variable region sequences containing sequence numbers 237 and 250, respectively; (aa) Heavy chain and light chain variable region sequences containing sequence numbers 238 and 250, respectively; (bb) Heavy and light chain variable region sequences containing sequence numbers 239 and 250, respectively; or (cc) Provides an isolated anti-FAM19A5 antibody or its antigen-binding moiety, each containing heavy chain and light chain variable region sequences, respectively, including sequence numbers 240 and 250.

[0206] In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety is (i) heavy chain CDR1, CDR2 and CDR3 of 2-13 or a combination thereof and / or light chain CDR1, CDR2 and CDR3 of 2-13 or a combination thereof; (ii) heavy chain CDR1, CDR2 and CDR3 of 3-2 or a combination thereof and / or light chain CDR1, CDR2 and CDR3 of 3-2 or any combination thereof; (iii) heavy chain CDR1, CDR2 and CDR3 of 1-65 or a combination thereof and / or light chain CDR1, CDR2 and (iv) Heavy chains CDR1, CDR2 and CDR3 of 1-28 or combinations thereof and / or light chains CDR1, CDR2 and CDR3 of 1-28 or any combinations thereof; (v) Heavy chains CDR1, CDR2 and CDR3 of P2-C12 or combinations thereof and / or light chains CDR1, CDR2 and CDR3 of P2-C12 or any combinations thereof; (vi) Heavy chains CDR1, CDR2 and CDR3 of 13B4 or combinations thereof and / or light chains CDR1, CDR2 and CDR3 of 13B4 or any combinations thereof Any combination; (vii) Heavy chains CDR1, CDR2, and CDR3 of 13F7 or any combination thereof and / or light chains CDR1, CDR2, and CDR3 of 13F7 or any combination thereof; (viii) Heavy chains CDR1, CDR2, and CDR3 of 15A9 or any combination thereof and / or light chains CDR1, CDR2, and CDR3 of 15A9 or any combination thereof; (ix) Heavy chains CDR1, CDR2, and CDR3 of P1-A03 or any combination thereof and / or light chains CDR1, CDR2, and CDR3 of P1-A03 or any combination thereof (x) Heavy chains CDR1, CDR2, and CDR3 of P1-A08 or any combination thereof and / or light chains CDR1, CDR2, and CDR3 of P1-A08 or any combination thereof; (xi) Heavy chains CDR1, CDR2, and CDR3 of P1-F02 or any combination thereof and / or light chains CDR1, CDR2, and CDR3 of P1-F02 or any combination thereof; (xii) Heavy chains CDR1, CDR2, and CDR3 of P2-A01 or any combination thereof and / or light chains CDR1, CDR2, and CDR3 of P2-A01 or any combination thereof;(xiii) Heavy chain CDR1, CDR2, and CDR3 of P2-A03 or any combination thereof and / or Light chain CDR1, CDR2, and CDR3 of P2-A03 or any combination thereof; (xiv) Heavy chain CDR1, CDR2, and CDR3 of P2-F07 or any combination thereof and / or Light chain CDR1, CDR2, and CDR3 of P2-F07 or any combination thereof; (xv) Heavy chain CDR1, CDR2, and CDR3 of P2-F11 or any combination thereof and / or Light chain CDR1, CDR2, and CDR3 of F2-F11 or any combination thereof; (x vi) Heavy chains CDR1, CDR2, and CDR3 of SS01-13 or any combination thereof and / or light chains CDR1, CDR2, and CDR3 of SS01-13 or any combination thereof; (xvii) Heavy chains CDR1, CDR2, and CDR3 of SS01-13-s5 or any combination thereof and / or light chains CDR1, CDR2, and CDR3 of SS01-13-s5 or any combination thereof; (xviii) Heavy chains CDR1, CDR2, and CDR3 of S5-2.GKNG or any combination thereof and / or light chains CDR1, CDR2, and CD of S5-2.GKNG R3 or any combination thereof; (xix) Heavy chain CDR1, CDR2 and CDR3 of 1-7A-IT or any combination thereof and / or Light chain CDR1, CDR2 and CDR3 of 1-7A-IT or any combination thereof; (xx) Heavy chain CDR1, CDR2 and CDR3 of Low-PI or any combination thereof and / or Light chain CDR1, CDR2 and CDR3 of Low-PI or any combination thereof; (xxi) Heavy chain CDR1, CDR2 and CDR3 of 1-30 or any combination thereof and / or Light chain CDR1, CDR2 and CDR3 of 1-30 or Any combination of these; (xxii) Heavy chains CDR1, CDR2 and CDR3 of 1-17 or any combination thereof and / or light chains CDR1, CDR2 and CDR3 of 1-17 or any combination thereof; (xxiii) Heavy chains CDR1, CDR2 and CDR3 of 1-32 or any combination thereof and / or light chains CDR1, CDR2 and CDR3 of 1-32 or any combination thereof; (xxiv) Heavy chains CDR1, CDR2 and CDR3 of 4-11 or any combination thereof and / or light chains CDR1, CDR2 and CDR3 of 4-11 or any combination thereof;(xxv) Heavy chains CDR1, CDR2 and CDR3 of 6-10 or combinations thereof and / or light chains CDR1, CDR2 and CDR3 of 6-10 or any combination thereof; (xxvi) Heavy chains CDR1, CDR2 and CDR3 of 2-13D or combinations thereof and / or light chains CDR1, CDR2 and CDR3 of 2-13D or any combination thereof; (xxvii) Heavy chains CDR1, CDR2 and CDR3 of 2-13D-37 or combinations thereof and / or light chains CDR1, CDR Includes (xxviii) heavy chain CDR1, CDR2, and CDR3 of (xxviii)2-13D-37-1.5W-41 or combinations thereof and / or light chain CDR1, CDR2, and CDR3-41 of (xxviii)2-13D-37-1.5W-41 or any combinations thereof; or (xxiv) heavy chain CDR1, CDR2, and CDR3 of (xxiv)2-13D-37-3W-16 or combinations thereof and / or light chain CDR1, CDR2, and CDR3 of (xxiviv)2-13D-37-3W-16 or combinations thereof. The amino acid sequences of VH CDR1, CDR2, and CDR3 for different anti-FAM19A5 antibodies disclosed herein are provided in Table 2. The amino acid sequences of VL CDR1, CDR2, and CDR3 for different anti-FAM19A5 antibodies disclosed herein are provided in Table 3.

[0207] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding portion of the present disclosure is: (a) VH CDR1 containing the amino acid sequence of SEQ ID NO: 11; and / or (b) VH CDR2 containing the amino acid sequence of SEQ ID NO: 12; and / or (c) Contains VH CDR3 containing the amino acid sequence of SEQ ID NO: 13

[0208] In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety comprises one, two, or three of the VH CDRs.

[0209] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety is: (a) VL CDR1 containing the amino acid sequence of SEQ ID NO: 23; and / or (b) VL CDR2 containing the amino acid sequence of SEQ ID NO: 24; and / or (c) Contains VL CDR3 containing the amino acid sequence of SEQ ID NO: 25

[0210] In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety comprises one, two, or three of the VL CDRs.

[0211] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety is: (a) VH CDR1 containing the amino acid sequence of SEQ ID NO: 11; (b) VH CDR2 containing the amino acid sequence of SEQ ID NO: 12; (c) VH CDR3 containing the amino acid sequence of SEQ ID NO: 13; (d) VL CDR1 containing the amino acid sequence of SEQ ID NO: 23; (e) VL CDR2 containing the amino acid sequence of SEQ ID NO: 24; and / or (f) Contains VL CDR3 containing the amino acid sequence of SEQ ID NO: 25

[0212] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety is: (a) VH CDR1 containing the amino acid sequence of SEQ ID NO: 14; (b) VH CDR2 containing the amino acid sequence of SEQ ID NO: 15; and / or (c) Contains VH CDR3 containing the amino acid sequence of SEQ ID NO: 16

[0213] In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety comprises one, two, or three of the VL CDRs.

[0214] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety specifically binds to human FAM19A5: (a) VL CDR1 containing the amino acid sequence of SEQ ID NO: 26; (b) VL CDR2 containing the amino acid sequence of SEQ ID NO: 27; and / or (c) Contains VL CDR3 containing the amino acid sequence of SEQ ID NO: 28

[0215] In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety comprises one, two, or three of the VL CDRs.

[0216] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety specifically binds to human FAM19A5: (a) VH CDR1 containing the amino acid sequence of SEQ ID NO: 14; (b) VH CDR2 containing the amino acid sequence of SEQ ID NO: 15; (c) VH CDR3 containing the amino acid sequence of SEQ ID NO: 16; (d) VL CDR1 containing the amino acid sequence of SEQ ID NO: 26; (e) VL CDR2 containing the amino acid sequence of SEQ ID NO: 27; and / or (f) Contains VL CDR3 containing the amino acid sequence of SEQ ID NO: 28

[0217] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety described herein specifically binds to human FAM19A5: (a) VH CDR1 containing the amino acid sequence of SEQ ID NO: 17; (b) VH CDR2 containing the amino acid sequence of SEQ ID NO: 18; and / or (c) Contains VH CDR3 with the amino acid sequence of SEQ ID NO: 19

[0218] In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety comprises one, two, or three of the VH CDRs.

[0219] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety specifically binds to human FAM19A5: (a) VL CDR1 containing the amino acid sequence of SEQ ID NO: 29; (b) VL CDR2 containing the amino acid sequence of SEQ ID NO: 30; and / or (c) Contains VL CDR3 containing the amino acid sequence of SEQ ID NO: 31

[0220] In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety comprises one, two, or three of the VL CDRs.

[0221] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety specifically binds to human FAM19A5: (a) VH CDR1 containing the amino acid sequence of SEQ ID NO: 17; (b) VH CDR2 containing the amino acid sequence of SEQ ID NO: 18; (c) VH CDR3 containing the amino acid sequence of SEQ ID NO: 19; (d) VL CDR1 containing the amino acid sequence of SEQ ID NO: 29; (e) VL CDR2 containing the amino acid sequence of SEQ ID NO: 30; and / or (f) Contains VL CDR3 containing the amino acid sequence of SEQ ID NO: 31

[0222] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety of the present disclosure specifically binds to human FAM19A5: (a) VH CDR1 containing the amino acid sequence of SEQ ID NO: 20; (b) VH CDR2 containing the amino acid sequence of SEQ ID NO: 21; and / or (c) Contains VH CDR3 with the amino acid sequence of SEQ ID NO: 22

[0223] In a particular embodiment, the anti-FAM19A5 antibody or its antigen-binding moiety comprises one, two, or three of the VH CDRs.

[0224] In some embodiments, the anti-FAM19A5 antibody or its antigen-binding moiety specifically binds to human FAM19A5: (a) VL CDR1 containing the amino acid sequence of SEQ ID NO: 32; (b) VL CDR2 containing the amino acid sequence of SEQ ID NO: 33; and / or (c) It comprises a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 34.

[0225] In certain embodiments, the anti-FAM19A5 antibody or antigen-binding portion thereof comprises any one, two or three of the foregoing VL CDRs.

[0226] In some embodiments, the anti-FAM19A5 antibody or antigen-binding portion thereof specifically binds to human FAM19A5: (a) A VH CDR1 comprising the amino acid sequence of SEQ ID NO: 20; (b) A VH CDR2 comprising the amino acid sequence of SEQ ID NO: 21; (c) A VH CDR3 comprising the amino acid sequence of SEQ ID NO: 22; (d) A VL CDR1 comprising the amino acid sequence of SEQ ID NO: 32; (e) A VL CDR2 comprising the amino acid sequence of SEQ ID NO: 33; and / or (f) A VL CDR3 comprising the amino acid sequence of SEQ ID NO: 34.

[0227] In certain embodiments, the anti-FAM19A5 antibody or antigen-binding portion thereof comprises one, two, three, four, five or six of the foregoing CDRs.

[0228] The VH domain or one or more CDRs thereof described herein can be linked to a constant domain for forming a heavy chain, such as a full-length heavy chain. Similarly, the VL domain or one or more CDRs thereof described herein can be linked to a constant domain for forming a light chain, such as a full-length light chain. The full-length heavy chain and full-length light chain combine to generate a full-length antibody.

[0229] Accordingly, in certain embodiments, this specification provides antibodies comprising an antibody light chain and a heavy chain, for example, a separate light chain and a heavy chain. In relation to the light chain, in certain embodiments, the light chain of the antibody described herein is a kappa light chain. In other certain embodiments, the light chain of the antibody described herein is a lambda light chain. In yet another certain embodiment, the light chain of the antibody described herein is a human kappa light chain or a human lambda light chain. In certain embodiments, the antibody described herein that specifically binds to a FAM19A5 polypeptide (e.g., human FAM19A5) comprises a light chain comprising any VL or VL CDR amino acid sequence described herein, wherein the constant region of the light chain comprises the amino acid sequence of the human kappa light chain constant region. In certain embodiments, the antibody described herein that specifically binds to a FAM19A5 polypeptide (e.g., human FAM19A5) comprises a light chain comprising a VL or VL CDR amino acid sequence described herein, wherein the constant region of the light chain comprises the amino acid sequence of the human lambda light chain constant region. Non-restrictive examples of human constant region sequences are described in the art. See, for example, U.S. Patent No. 5,693,780 and Kabat EA et al, (1991) above.

[0230] In relation to the heavy chain, in some embodiments, the heavy chain of the antibody described herein may be an alpha (α), delta (δ), epsilon (ε), gamma (γ), or mu (μ) heavy chain. In other specific embodiments, the heavy chain of the antibody described herein may include a human alpha (α), delta (δ), epsilon (ε), gamma (γ), or mu (μ) heavy chain. In one embodiment, the antibody described herein that specifically binds to FAM19A5 (e.g., human FAM19A5) comprises a heavy chain containing a VH or VH CDR amino acid sequence disclosed herein, wherein the constant region of the heavy chain comprises the amino acid sequence of the human gamma (γ) heavy chain constant region. In other embodiments, the antibody described herein that specifically binds to FAM19A5 (e.g., human FAM19A5) comprises a heavy chain containing a VH or VH CDR amino acid sequence disclosed herein, wherein the constant region of the heavy chain comprises the amino acids of a human heavy chain described herein or known in the art. Non-restrictive examples of human constant region sequences are described in the art. See, for example, U.S. Patent No. 5,693,780 and Kabat EA et al., (1991) above.

[0231] In some embodiments, the antibodies described herein comprise a VL domain and a VH domain comprising a VH or VH CDR and a VL and VL CDR as described herein, wherein the constant region comprises the amino acid sequence of the constant region of an IgG, IgE, IgM, IgD, IgA or IgY immunoglobulin molecule or a human IgG, IgE, IgM, IgD, IgA or IgY immunoglobulin molecule. In other specific embodiments, the antibodies described herein that specifically bind to FAM19A5 (e.g., human FAM19A5) comprise a VL domain and a VH domain comprising any amino acid sequence described herein, wherein the constant region comprises the amino acid sequence of the constant region of an IgG, IgE, IgM, IgD, IgA or IgY immunoglobulin molecule and any subtype of immunoglobulin molecule (e.g., IgG1, IgG2, IgG3, IgG4, IgAl and IgA2). In some embodiments, the constant region includes the amino acid sequence of the constant region of natural human IgG, including subtypes (e.g., IgG1, IgG2, IgG3, or IgG4) and allogeneic types (e.g., Glm, G2m, G3m, and nG4m) and their variants. See, for example, Vidarsson G. et al. Front Immunol. 5:520 (published online October 20, 2014) and Jefferis R. and Lefranc MP, mAbs 1:4, 1-7 (2009). In some embodiments, the constant region includes the amino acid sequence of the constant region of human IgG1, IgG2, IgG3, or IgG4 or their variants.

[0232] In certain embodiments, the anti-FAM19A5 antibodies disclosed herein or their antigen-binding moieties lack Fc effector function, such as complement-dependent cytotoxicity (CDC) and / or antibody-dependent cellular phagocytic activity (ADCP). Effector function is mediated by the Fc region, and the residue in the CH2 domain of the Fc region closest to the hinge region contains a binding site that largely overlaps with Clq (complement) and the IgG-Fc receptor (FcγR) on effector cells of the innate immune system, thus contributing to the antibody's effector function. Furthermore, IgG2 and IgG4 antibodies have lower levels of Fc effector function than IgG1 and IgG3 antibodies. The effector function of antibodies is achieved by (1) using antibody fragments lacking an Fc region (e.g., Fab, F(ab')2, single-chain Fv(scFv), or sdAb consisting of monomeric VH or VL domains); (2) generating glycation-free antibodies by, for example, deleting or altering sugar-attached residues, enzymatically removing sugar, generating antibodies in cells cultured in the presence of glycosylation inhibitors, or expressing antibodies in cells incapable of protein glycosylation (e.g., bacterial host cells, see, for example, U.S. Patent Publication No. 20120100140); and (3) using Fc regions of IgG subtypes with reduced effector function (e.g., regions of IgG2 and IgG4 antibodies or chimeric Fc regions containing the CH2 domain of IgG2 and IgG4 antibodies, see, for example, U.S. Patent Publication No. 20120100140 and Lau C. et al.). See al. J. Immunol. 191:4769-4777 (2013); and (4) it may be reduced or avoided by different approaches known to the art, including generating Fc regions with mutations that reduce or eliminate Fc function. See, for example, U.S. Patent Publication No. 20120100140 and the U.S. and PCT applications cited therein and An et al. mAbs 1:6,572-579 (2009).

[0233] Thereby, in some embodiments, the anti-FAM19A5 antibody or antigen-binding portion thereof disclosed herein is an sdAb consisting of Fab, Fab’, F(ab’)2, Fv, single-chain Fv (scFv), or monomeric VH or VL domains. Such antibody fragments are well known in the art and are described above.

[0234] In some embodiments, the anti-FAM19A5 antibody or antigen-binding portion thereof disclosed herein comprises an Fc region with reduced or absent Fc effector function. In some embodiments, the constant region comprises the amino acid sequence of the Fc region of human IgG2 or IgG4. In some embodiments, the anti-FAM19A5 antibody has an IgG2 / IgG4 isotype. In some embodiments, the anti-FAM19A5 antibody comprises a chimeric Fc region comprising the CH2 domain of an IgG antibody of the IgG4 isotype and the CH3 domain of an IgG antibody of the IgG1 isotype, or a chimeric Fc region comprising the hinge region of IgG2 and the CH2 region of IgG4, or an Fc region having a mutation with reduced or absent Fc effector function. Fc regions with reduced or absent Fc effector function include those known in the art. See, for example, Lau C. et al, J. Immunol. 191:4769-4777 (2013); An et al, mAbs 1:6,572-579 (2009); and U.S. Patent Publication No. 20120100140 and the U.S. patents, published patent applications, and PCT published applications cited therein. Also, Fc regions with reduced or absent Fc effector function can be readily made by one of ordinary skill in the art.

[0235] <V. Nucleic Acid Molecules> Further embodiments described herein relate to one or more nucleic acid molecules encoding any one of the antibodies or antigen-binding moieties described herein. Such nucleic acids may exist in whole cell, cell lysate, or in partially purified or substantially pure form. Nucleic acids are “separated” or “substantially pure” when purified from other cellular components or other contaminants, such as other cellular nucleic acids (e.g., other chromosomal DNA, e.g., chromosomal DNA linked to DNA isolated from nature) or proteins, by standard techniques including alkali / SDS treatment, CsCl banding, column chromatography, restriction enzymes, agarose gel electrophoresis, and other techniques known to the art. See F. Ausubel, elal., ed. (1987) Current Protocols in Molecular Biology, Greene Publishing and Wiley Interscience, New York. The nucleic acids described herein may be, for example, DNA or RNA, and may or may not contain intron sequences. In certain embodiments, such nucleic acids are cDNA molecules.

[0236] The nucleic acids described herein can be obtained using standard molecular biology techniques. In the case of antibodies expressed by hybridomas (e.g., hybridomas produced from transgenic mice carrying human immunoglobulin genes, as further described below), the cDNA encoding the light and heavy chains of the antibody produced by the hybridoma can be obtained by standard PCR amplification or cDNA cloning techniques. In the case of antibodies obtained from an immunoglobulin gene library (e.g., using phage display techniques), the nucleic acids encoding the antibody can be recovered from the library.

[0237] The specific nucleic acid molecules encode the VH and VL sequences of various anti-FAM19A5 antibodies described herein. Exemplary DNA sequences encoding the VH sequences of such antibodies are shown in SEQ ID NOs: 43-46 and 177. Exemplary DNA sequences encoding the VL sequences of such antibodies are shown in SEQ ID NOs: 47-50 and 178.

[0238]

Table 7

[0239] JPEG0007868876000016.jpg219169

[0240] JPEG0007868876000017.jpg210169

[0241]

Table 8

[0242] JPEG0007868876000019.jpg228169

[0243] The method for producing an anti-FAM19A5 antibody disclosed in this specification can include expressing, together with the signal peptide, the relevant heavy and light chains of the antibody, for example, SEQ ID NO: 43 and 47, SEQ ID NO: 44 and 48, SEQ ID NO: 45 and 49, SEQ ID NO: 46 and 50, SEQ ID NO: 177 and 178, in a cell line containing the nucleotide sequences encoding the heavy and light chains. Host cells containing these nucleotide sequences are included herein.

[0244] When DNA fragments encoding VH and VL segments are obtained, these DNA fragments can be further manipulated by standard recombinant DNA techniques, for example, the variable region genes can be converted into full-length antibody chain genes, Fab fragment genes or scFv genes. In these manipulations, the VL or VH-coding DNA fragments are operably linked to other DNA fragments encoding further other proteins such as antibody constant regions or flexible linkers. The term "operably linked" used in connection with this is intended to mean that two DNA fragments are joined and the amino acid sequences encoded by the two DNA fragments remain in-frame.

[0245] Isolated DNA encoding the VH region can be converted into a full-length heavy chain gene by operatively ligating the VH-coding DNA to another DNA molecule encoding the heavy chain constant region (hinge, CH1, CH2, and / or CH3). The sequences of human heavy chain constant region genes are publicly known in the art (see, for example, Kabat, EA, el al. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, US Department of Health and Human Services, NIH Publication No. 91-3242), and DNA fragments containing these regions can be obtained by standard PCR amplification. The heavy chain constant region may be the IgG1, IgG2, IgG3, IgG4, IgA, IgE, IgM, or IgD constant region, for example, the IgG2 and / or IgG4 constant region. In the case of Fab fragment heavy chain genes, the VH-coding DNA can be operatively ligated to another DNA molecule encoding only the heavy chain CH1 constant region.

[0246] Isolated DNA encoding the VL region can be converted into a full-length light chain gene (and even a Fab light chain gene) by operatively ligating the VL-coding DNA to another DNA molecule encoding the light chain constant region (CL). The sequences of human light chain constant region genes are publicly known in the industry (e.g., Kabat, EA, et al. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, US Department of Health and Human Services, NIH Publication No. 91-3242), and DNA fragments containing these regions can be obtained by standard PCR amplification. The light chain constant region may be a kappa or lambda constant region.

[0247] To generate scFv antibodies, the VH- and VL-coding DNA fragments are operatively linked to a flexible linker, for example, another fragment encoding the amino acid sequence (Gly4-Ser)3, and the VH and VL sequences can be expressed as adjacent single-chain proteins with the VL and VH regions linked by the flexible linker (see, for example, Bird et al., (1988) Science 242:423-426; Huston et al., (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883; McCafferty et al., (1990) Nature 348:552-554).

[0248] In some embodiments, the disclosure provides a vector comprising an isolated nucleic acid molecule containing a nucleotide sequence encoding an antibody or its antigen-binding portion. In other embodiments, the vector can be used for gene therapy.

[0249] Vectors suitable for this disclosure include expression vectors, viral vectors, and plasmid vectors. In one embodiment, the vector is a viral vector.

[0250] As used herein, an expression vector refers to any nucleic acid structure containing the elements necessary for the transcription and translation of an inserted coding sequence, or, in the case of an RNA viral vector, the elements necessary for replication and translation upon introduction into a suitable host cell. Expression vectors may include plasmids, phagemids, viruses, and derivatives thereof.

[0251] The expression vectors described herein may include polynucleotides encoding an antibody or its antigen-binding moiety as described herein. In one embodiment, the coding sequence for the antibody or its antigen-binding moiety is operably ligated to an expression regulatory sequence. The two nucleic acid sequences used herein are operably ligated when they are covalently bonded in such a manner that each component nucleic acid sequence maintains its function. The coding sequence and the gene expression regulatory sequence are operably ligated when they are covalently bonded such that the expression or transcription and / or translation of the coding sequence is under the influence or control of the gene expression regulatory sequence. The two DNA sequences are operably ligated when the promoter induction in the 5' gene expression sequence causes transcription of the coding sequence, and the binding properties between the two DNA sequences do not (1) introduce a frame-shift mutation, (2) interfere with the ability of the promoter region to direct transcription of the coding sequence, or (3) interfere with the ability of the corresponding RNA transcript to be translated into a protein. In cases where a gene expression sequence can affect the transcription of a coding nucleic acid sequence, the gene expression sequence is operably linked to the coding nucleic acid sequence, and the resulting transcript is translated into the desired antibody or its antigen-binding portion.

[0252] Viral vectors contain, but are not limited to, the nucleic acid sequences of viruses such as: retroviruses, e.g., Moloney mouse leukemia virus, Harvey mouse sarcoma virus, mouse mammary cancer virus, and Rous sarcoma virus; lentiviruses; adenoviruses; adeno-associated viruses; SV40 virus; polyomaviruses; Epstein-Barr virus; papillomavirus; herpesviruses; vaccinia virus; infantile paralysis virus; and RNA viruses such as retroviruses. Other vectors known in the art can readily be used. Certain viral vectors are based on non-cellular eukaryotic viruses in which non-essential genes are replaced with genes of interest. Non-cellular viruses include retroviruses, whose life cycle involves reverse transcription of genomic viral RNA into DNA, followed by proviral integration into host cell DNA. Retroviruses have been approved for human gene therapy trials. Most useful are retroviruses with replication defects (i.e., they can direct the synthesis of the desired protein but cannot produce infectious particles). Such genetically modified retroviral expression vectors are generally useful for highly efficient gene transduction in vivo. Standard protocols for producing retroviruses with replication defects (including the steps of contaminating a plasmid with exogenous genetic material, transjecting a packaged cell line using the plasmid, producing recombinant retroviruses using the packaged cell line, recovering viral particles from tissue culture medium, and infecting target cells with the viral particles) are provided in Kriegler, M., Gene Transfer and Expression, A Laboratory Manual, WH Freeman Co., New York (1990) and Murry, ET, Methods in Molecular Biology, Vol. 7, Humana Press, Inc., Cliffton, NJ (1991).

[0253] In one embodiment, the virus is an adeno-associated virus, which is a double-stranded DNA virus. The adeno-associated virus may be engineered to have replication defects and can infect a wide range of cell types and species. It also has advantages such as thermal and lipid solvent stability; high transduction frequency in various cell lineages, including hematopoietic cells; lack of inhibition of co-infection; and the ability to induce a variety of transductions. It has been reported that the adeno-associated virus can be integrated into human cell DNA in a site-specific manner, thereby minimizing the possibility of inducing insertional mutations and the variability of the inserted gene expression characteristics of retroviral infection. Furthermore, wild-type adeno-associated virus infection has been tracked in tissue culture for more than 100 passages without selective pressure, which means that adeno-associated virus genome integration is a relatively stable reaction. The adeno-associated virus may also act extrachromosomally.

[0254] In other embodiments, the vector is derived from a lentivirus. In specific embodiments, the vector is a recombinant lentivirus vector capable of infecting non-dividing cells.

[0255] Lentiviral genomes and proviral DNA generally contain three genes, gag, pol, and env, which have two long terminal repeat (LTR) sequences on both sides, as found in retroviruses. The gag gene encodes internal structure (matrix, capsid, and nucleocapsid) proteins; the pol gene encodes RNA-directed DNA polymerase (reverse transcriptase), proteases, and integrases; and the env gene encodes viral coat glycoproteins. The 5' and 3' LTRs play a role in promoting virion RNA transcription and polyadenylation. LTRs contain all other cis-acting sequences necessary for viral replication. Lentiviruses also possess additional genes, including vif, vpr, tat, rev, vpu, nef, and vpx (in HIV-1, HIV-2, and / or SIV).

[0256] The sequences necessary for reverse transcription of the genome (tRNA primer binding site) and for efficiently encapsulating viral RNA in the particle (Psi site) are adjacent to the 5'LTR. If the sequences necessary for encapsulation (or packaging retroviral RNA into infectious virions) are missing from the viral genome, the cis defect prevents the encapsulation of genomic RNA.

[0257] However, the resulting mutants remain capable of directing the synthesis of all virion proteins. This disclosure provides a method for producing recombinant lentiviruses capable of infecting non-dividing cells, comprising transjecting suitable host cells using two or more vectors possessing packaging function, namely gag, pol, and env, as well as rev and tat. As disclosed below, vectors lacking the functional tat gene are suitable for specific applications. For example, a first vector may provide nucleic acids encoding viral gag and viral pol, and other vectors may provide nucleic acids encoding viral env to produce packaging cells. In this specification, introducing a vector providing heterologous genes identified as transfer vectors into packaging cells yields producer cells that release infectious viral particles carrying the exogenous gene of interest.

[0258] According to the vector and exogenous gene configuration described above, the second vector can provide a nucleic acid encoding a viral coat (env) gene. The env gene can be derived from virtually any suitable virus, including retroviruses. In some embodiments, the env protein is an amphibian coat protein that enables transduction of human and other species cells.

[0259] Examples of retrovirus-derived env genes include, but are not limited to, Moloney's mouse leukemia virus (MoMuLV or MMLV), Harvey's mouse sarcoma virus (HaMuSV or HSV), mouse mammary cancer virus (MuMTV or MMTV), gibbon ape leukemia virus, human immunodeficiency virus (HIV), and Rous sarcoma virus. Other env genes, such as vesicular stomatitis virus (VSV) protein G (VSV G), hepatitis viruses, and influenza genes may also be used.

[0260] The vector providing the viral env nucleic acid sequence is operably associated with the regulatory sequences described elsewhere in this specification.

[0261] In certain embodiments, the vector comprises a lentiviral vector in which the HIV toxicity genes env, vif, vpr, vpu, and nef are deleted without impairing the vector's ability to transduce non-dividing cells.

[0262] In some embodiments, the vector comprises a lentiviral vector containing a deletion in the U3 region of the 3'LTR. The deletion in the U3 region may be a total deletion or a partial deletion.

[0263] In some embodiments, the lentiviral vector of this disclosure, comprising the FVIII nucleotide sequence described herein, may be transfused using (a) a first nucleotide sequence comprising the gag, pol, or gag and pol genes, and (b) a second nucleotide sequence comprising a heterologous env gene; in this case, the lentiviral vector lacks the functional tat gene. In other embodiments, the cells are further transfused using a fourth nucleotide sequence comprising the rev gene. In certain embodiments, the lentiviral vector lacks a functional gene selected from vif, vpr, vpu, vpx, and nef or a combination thereof.

[0264] In certain embodiments, the lentiviral vector comprises one or more nucleotide sequences encoding a gag protein, a Rev response element, a central polypurine tract (cPPT), or any combination thereof.

[0265] Examples of lentiviral vectors are disclosed in W09931251, W09712622, W09817815, W09817816 and W09818934, the entire contents of which are hereby incorporated by reference.

[0266] Other vectors include plasmid vectors. Plasmid vectors are extensively described in the art and are well known to those skilled in the art. See, for example, Sambrook el al., Molecular Cloning: A Laboratory Manual, Second Edition, Cold Spring Harbor Laboratory Press, 1989. In recent years, plasmid vectors have been shown to be particularly advantageous for delivering genes to cells in vivo because they replicate within the host genome and cannot integrate into the host genome. However, these plasmids having a promoter compatible with the host cell can express peptides from genes operably encoded within the plasmid. Some commonly available and commercially available plasmids include pBR322, pUCl8, pUCl9, various pcDNA plasmids, pRC / CMV, various pCMV plasmids, pSV40 and pBlueScript. Additional examples of specific plasmids are pcDNA3.1, catalog number V79020; pcDNA3.1 / hygro, catalog number V87020; pcDNA4 / myc-His, catalog number V86320; and pBudCE4.l, catalog number V53220 (all from Invitrogen (Carlsbad, CA)). Other plasmids are well known to those skilled in the art. Furthermore, plasmids can be custom designed using standard molecular biology techniques to remove and / or add specific fragments of DNA.

[0267] <VI. Antibody Production>Antibodies or fragments thereof that immunospecifically bind to FAM19A5 (e.g., human FAM19A5) can be produced by any method known in the art for antibody synthesis, such as chemical synthesis or recombinant expression techniques. Unless otherwise specified, the methods disclosed herein utilize prior art in molecular biology, microbiology, genetic analysis, recombinant DNA, organic chemistry, biochemistry, PCR, oligonucleotide synthesis and modification, nucleic acid hybridization, and related fields of the art of the art. These techniques are fully described, for example, in the references referenced herein.For example, Maniatis T et al., (1982) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press; Sambrook J et al., (1989), Molecular Cloning: A Laboratory Manual, Second Edition, Cold Spring Harbor Laboratory Press; Sambrook J et al., (2001) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; Ausubel FM et al. al.,Current Protocols in Molecular Biology,John Wiley & Sons(1987 and annual updates);Current Protocols in Immunology,John Wiley & Sons(1987 and annual updates)Gait(ed.)(1984)Oligonucleotide Synthesis:A Practical Approach,IRL Press;Eckstein(ed.)(1991)Oligonucleotides and Analogues:A Practical Approach,IRL Press;Birren B et Please refer to al., (eds.) (1999) Genome Analysis: A Laboratory Manual, Cold Spring Harbor Laboratory Press.

[0268] In certain embodiments, the antibodies described herein are antibodies (e.g., recombinant antibodies) produced, expressed, generated, or isolated by any means, including synthesis, or generation using genetic manipulation of DNA sequences. In certain embodiments, such antibodies contain sequences (e.g., DNA sequences or amino acid sequences) that do not naturally exist within the antibody germline repertoire of an in vivo animal or mammal (e.g., human).

[0269] <VII. Pharmaceutical Compositions> This specification provides compositions comprising an antibody or an antigen-binding portion thereof described herein, having a desired degree of purity in a physiologically acceptable carrier, excipient, or stabilizer (Remington’s Pharmaceutical Sciences (1990) Mack Publishing Co., Easton, PA). The acceptable carrier, excipient, or stabilizer is non-toxic to the recipient at the dosage and concentrations employed, and includes buffers such as phosphates, citrates and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (such as octadecyl dimethyl benzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins such as serum albumin, gelatin or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine or lysine; monosaccharides, disaccharides and other carbohydrates including glucose, mannose or dextrin; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salt-forming counterions such as sodium; metal complexes (e.g., Zn-protein complexes); and / or nonionic surfactants such as TWEEN TM , PLURONICS TM or polyethylene glycol (PEG).

[0270] In some embodiments, the pharmaceutical composition comprises a pharmaceutically acceptable carrier containing an antibody or its antigen-binding moiety, a bispecific molecule, or an immune complex as described herein. In some embodiments, the pharmaceutical composition comprises a pharmaceutically acceptable carrier containing an effective amount of the antibody or its antigen-binding moiety as described herein and optionally one or more additional prophylactic or therapeutic agents. In some embodiments, the antibody is the sole active ingredient contained in the pharmaceutical composition. The pharmaceutical compositions described herein may be useful in reducing FAM19A5 activity and treating eye diseases, disorders, or symptoms such as glaucoma.

[0271] Pharmacokinetically acceptable carriers used in parenteral formulations include aqueous vehicles, non-aqueous vehicles, antimicrobial agents, isotonic agents, buffers, antioxidants, topical anesthetics, suspensions and dispersants, emulsifiers, metal ion sequestering or chelating agents, and other pharmacokinetically acceptable substances. Examples of aqueous vehicles include sodium chloride injection, intravenous infusion, isotonic dextrose injection, sterile water injection, and dextrose and lactate intravenous infusion. Non-aqueous parenteral vehicles include plant-derived fixative oils, cottonseed oil, corn oil, sesame oil, and peanut oil. Antimicrobial agents at bacteriostatic or fungiostatic concentrations may be added to parenteral formulations packaged in multi-dose containers containing phenol or cresol, mercury-containing substances, benzyl alcohol, chlorobutanol, methyl and propyl p-hydroxybenzoates, thimerosal, benzalkonium chloride, and benzethonium chloride. Isotonic agents include sodium chloride and dextrose. The buffering agent contains phosphate and citrate. The antioxidant contains sodium bisulfate. The topical anesthetic contains procaine hydrochloride. The suspension and dispersing agent contains sodium carboxymethylcellulose, hydroxypropyl methylcellulose, and polyvinylpyrrolidone. The emulsifier contains Polysorbate 80 (TWEEN® 80). The metal ion sequestering or chelating agent contains EDTA. The pharmaceutical carrier also contains ethyl alcohol, polyethylene glycol, and propylene glycol for water-miscible vehicles; and sodium hydroxide, hydrochloric acid, citric acid, or lactic acid for pH adjustment.

[0272] The pharmaceutical compositions disclosed herein can be formulated for any route of administration to a subject. In some embodiments, the pharmaceutical compositions disclosed herein can be administered via any route capable of delivering the composition to sub-Tenon's capsule, subconjunctival, superchoroidal, intravitreous, and similar locations in the eye. In certain embodiments, the pharmaceutical compositions disclosed herein are delivered to a subject by intraocular administration. In some embodiments, intraocular administration includes intravitreous administration. Parenteral administration, characterized by subcutaneous, intramuscular, intraperitoneal, or intravenous injection, is also considered herein. Injectable preparations are conventional forms and may be manufactured as liquid solutions or suspensions, solid forms suitable for liquid solutions or suspensions before injection, or as emulsions. Injectable preparations, solutions, and emulsions also contain one or more excipients. Suitable excipients include, for example, water, saline, dextrose, glycerol, or ethanol. Furthermore, if necessary, the administered pharmaceutical composition may also contain small amounts of non-toxic adjuncts such as wetting or emulsifying agents, pH buffers, stabilizers, solubility enhancers, and formulations such as sodium acetate, sorbitan monolaurate, triethanolamine oleate, and cyclodextrin.

[0273] Antibody parenteral administration formulations include sterile, dried, soluble products such as freeze-dried powders that can be immediately compounded with a solvent immediately before use, including immediate sterile solutions for injection and tablets for subcutaneous injection; immediate sterile suspensions for injection; sterile, dried, insoluble products that can be immediately compounded with a vehicle immediately before use; and sterile emulsions. The solutions may be aqueous or non-aqueous.

[0274] For intravenous administration, suitable carriers include physiological saline or phosphate-buffered saline (PBS) and a solution containing thickeners and solubilizers such as glucose, polyethylene glycol, polypropylene glycol, and mixtures thereof.

[0275] Topical mixtures containing antibodies are prepared as described for topical and systemic administration. The resulting mixtures may be solutions, suspensions, emulsions, etc., and can be formulated into creams, gels, ointments, emulsions, solutions, elixirs, lotions, suspensions, tinctures, pastes, foams, aerosols, irrigation solutions, sprays, suppositories, bandages, skin patches, or any other dosage form suitable for topical administration.

[0276] The antibodies or antigen-binding moieties described herein can be formulated, for example, as topical aerosols for inhalation (see, for example, U.S. Patents 4,044,126,4414,209 and 4,364,923, which describe steroid delivery aerosols useful for the treatment of inflammatory diseases, particularly asthma). These formulations for airway administration may be in the form of aerosols for sprayers or solutions, or as fine powders for inhalation, and may be used alone or in combination with an inactive carrier such as lactose. In such cases, the particles of the formulation may have a diameter of less than 50 microns in one embodiment and less than 10 microns in another embodiment.

[0277] The antibodies or antigen-binding moieties described herein can be formulated for topical or local use, such as topical application to the skin and mucous membranes, including the eyes, in the form of gels, creams, and lotions, and for application to the eyes or intracerebral or intraspinal applications. Topical administration is also considered for transdermal delivery and for application to the eyes, mucous membranes, or inhalation therapy. Nasal solutions of the antibodies can be administered alone or in combination with other pharmaceutically acceptable excipients.

[0278] Transdermal patches containing ionography and electrophoresis devices are well known to those skilled in the art and can be used to administer antibodies. For example, such patches are disclosed in U.S. Patent Nos. 6,267,983, 6,261,595, 6,256,533, 6,167,301, 6,024,975, 6,010,715, 5,985,317, 5,983,134, 5,948,433 and 5,860,957.

[0279] In some embodiments, the pharmaceutical composition comprising the antibody or its antigen-binding moiety described herein is a lyophilized powder that can be reconstituted for administration as a solution, emulsion, or other mixture. The lyophilized powder can also be reconstituted and formulated as a solid or gel. The lyophilized powder is obtained by dissolving the antibody or its antigen-binding moiety, or a pharmaceutically acceptable derivative thereof, in a suitable solvent. In some embodiments, the lyophilized powder is sterile. The solvent may contain excipients that improve the stability or other pharmacological components of the powder or the reconstituted solution prepared from the powder. Available excipients include, but are not limited to, dextrose, sorbitol, fructose, corn syrup, xylitol, glycerin, glucose, sucrose, or other suitable formulations. The solvent may also contain buffers such as citrate, sodium phosphate, or potassium phosphate, or in one embodiment other such buffers with a substantially neutral pH. The solution is then sterile filtered and lyophilized under standard conditions known to those skilled in the art to provide the desired dosage form. In one embodiment, the obtained solution can be distributed into a lyophilized vial. Each vial may contain a single dose or multiple doses of the compound. The lyophilized powder can be stored under suitable conditions, such as approximately 4°C to room temperature.

[0280] Such lyophilized powders are reconstituted with sterile water for injection to provide dosage forms for parenteral administration. For reconstitution, the lyophilized powder is added to sterile water or another suitable carrier. The exact amount varies depending on the selected compound. Such amounts can be determined empirically.

[0281] The antibodies or their antigen-binding moieties, bispecific molecules, or immune complexes described herein, and other compositions provided herein, may also be formulated to target specific tissues, receptors, or other body regions to be treated. Many such targeting methods are well known to those skilled in the art. In this specification, any such targeting method is considered for use in the present compositions. For non-restrictive examples of targeting methods, see, for example, U.S. Patents 6,316,652, 6,274,552, 6,271,359, 6,253,872, 6,139,865, 6,131,570, 6,120,751, 6,071,495, 6,060,082, 6,048,736, 6,039,975, 6,004,534, 5,985,307, 5,972,366, 5,900,252, 5,840,674, 5,759,542 and 5,709,874. In certain embodiments, the vectors or antigen-binding moieties described herein can be used to treat glaucoma and / or reduce, improve, or inhibit glaucoma-related inflammation.

[0282] Compositions used for in vivo administration may be sterile. This can be easily achieved, for example, by filtration using a sterile filtration membrane.

[0283] <VIII.キット> This specification provides kits comprising one or more antibodies or antigen-binding moieties thereof as described herein. In certain embodiments, this specification provides pharmaceutical packs or kits comprising one or more containers filled with one or more components of the pharmaceutical compositions described herein, such as one or more antibodies or antigen-binding moieties provided herein, and optionally instructions for use. In some embodiments, the kit contains the pharmaceutical compositions described herein and any prophylactic or therapeutic agents as described herein.

[0284] <Examples> (Example 1) Evaluation of electroretinopathy after in vivo administration of anti-FAM19A5 antibody in a mouse model of diabetic retinopathy. To begin evaluating the efficacy of anti-FAM19A5 antibody therapy for diabetic retinopathy, type 2 diabetic db / db mice (B6.BKS(D)-Leprdb / j) (25 weeks old) were used. Briefly, after stabilizing the mice for one week, human IgG control antibody or anti-FAM19A5 antibody (1-65 or 3-2 clones) was administered intravitreously to the mice using a micropump system (7 doses in total, 4 μg / eye every 2 weeks). Normal (i.e., wild-type) mice without diabetes were used as the control group. Next, electroretinography (ERG) was used to evaluate the retinoid potential.

[0285] The electroretinogram (ERG) is a diagnostic test that measures the electrical activity (i.e., electroretinography) generated by various cells in the retina, including photoreceptors (rodular and cone cells), internal retinal cells (bipolar and axonal cells), and ganglion cells. Generally, in response to a bright flash of light, the ERG of a healthy eye will show a complex waveform consisting of an initial negative deflection ("wave A") followed by a gradient of increasingly larger high-frequency oscillations known as "rhythmic scotopic waves" (OP), and then a B wave with a faster peak. The A wave is influenced by the collective response of rodular cells, and the dark-adapted (scotopic) B wave is influenced by the response of bipolar rodular cells. The OP occurs in the inner plexiform layer where bipolar, axonal, and ganglion cells interact. See Wilsey et al., Curr Opin Ophthalmol 27(2):118-124 (2016). By evaluating these values, the health status of the various cells found in the retina can be assessed.

[0286] To evaluate the electroretinocyte electroencephalography (ERG), the pupils of animals in different treatment groups were dilated by topical treatment with 2.5% phenylephrine hydrochloride. The animals were then dehaired and anesthetized (after administration of ketamine (80 mg / kg) and xylazine (16 mg / kg), followed by 0.5% paracaine), and electrodes were attached to the skin, tail, and cornea. A single flash of white light (0.9 log cd sec / m) was used. 2The retina was stimulated with ) . To evaluate the electrical activity inside the eyes of different animals (i.e., "potential retinopathy"), the amplitude values ​​of wave A (from the trough of wave A to the baseline), wave B (from the trough of wave A to the peak of wave B), and rhythmic ovulation (OP) (amplitude and duration of small oscillations observed from the ascending limb of wave B) were measured.

[0287] As shown in Figures 1A to 1C, mice administered with the anti-FAM19A5 antibody (clone 1-65 or 3-2) exhibited improved A-waves, B-waves, and rhythmic ripples compared to mice administered with the control group IgG antibody. This indicates that animals administered with the anti-FAM19A5 antibody recovered normal retinal function.

[0288] (Example 2) Evaluation of morphological changes after anti-FAM19A5 antibody treatment in a mouse model of diabetic retinal disease. To evaluate whether anti-FAM19A5 antibody treatment caused morphological changes in the retina, the type 2 diabetic db / db mice from Example 1 were sacrificed, and both eyes were removed from the animals.

[0289] After excision, one eye from each animal was fixed in 10% neutral formalin for one day. The cornea, lens, and sclera were then removed, and the remaining ocular tissue was washed five times with distilled water for one hour with shaking. After the final wash, the distilled water was removed, and the ocular tissue was cultured for two hours in 3% trypsin dissolved in 0.1 M Tris buffer at 37°C. Retinal vessels were then isolated from the remaining retinal tissue and attached to slides for staining with periodate Schiff (PAS) to evaluate retinal microvessels against characteristic markers of early diabetic retinopathy (e.g., perivascular cell loss and acellular capillary formation).

[0290] The other eye of each animal was pre-fixed in 4% glutaraldehyde and then fixed in 10% formalin for 3 days. The eyes were then placed in disposable cassettes and embedded in paraffin blocks. The frozen tissue was then thinned and placed on slides for further analysis. H&E, TUNEL, and IHC analyses were performed to analyze vascular congestion, vascular markers (CD31 and VEGF), and retinal cell protective effects.

[0291] Figure 2A provides micrographs (100× magnification) of representative retinal regions from animals belonging to different treatment groups. The endothelial cell-to-perivascular cell ratio (E / P ratio) and the number and length of acellular capillaries detected in mice treated with anti-FAM19A5 antibody (clones 1-65 or 3-2) were all similar to those of normal, healthy animals. See Figures 2B-2D. In contrast, mice treated with human IgG control antibody had a much higher E / P ratio, suggesting loss of perivascular cells. See Figure 2B. Human IgG control mice also had a greater number of acellular capillaries (Figure 2C), and the acellular capillaries were much longer (Figure 2D). These results suggest that anti-FAM19A5 antibody can prevent pericellular cell loss and protect retinal microvessels from acellular capillary formation in a diabetic retinal disease model.

[0292] Furthermore, as shown in Figure 3, mice treated with anti-FAM19A5 antibody (bottom two panels) also showed significantly reduced vascular congestion in the retinal nerve fiber layer compared to mice administered with human IgG control antibody (top right panel). TUNEL analysis confirmed that both the nerve fiber layer and the inner core layer of the retina of mice treated with anti-FAM19A5 antibody had fewer cells undergoing cell death compared to the corresponding layers of the retina of mice treated with human IgG control antibody. See Figure 4. This suggests that anti-FAM19A5 antibody can protect retinal cells from damage caused by diabetes.

[0293] Finally, as shown in Figure 5, the nerve fiber layer of mice treated with human IgG control antibody showed extensive CD31 and VEGF staining, indicating neovascularization, a characteristic of a more advanced stage of diabetic retina. In contrast, mice treated with anti-FAM19A5 antibody (clones 1-65 or 3-2) showed significantly reduced CD31 and VEGF staining. This result suggests that anti-FAM19A5 antibody can effectively block angiogenesis in a model of diabetic retina.

[0294] In short, the data demonstrates that the anti-FAM19A5 antibody improves most of the underlying conditions associated with retinal diseases such as diabetic retinopathy, and that the disease is treatable.

[0295] (Example 3) Evaluation of the effect of anti-FAM19A5 antibodies on choroidal angiogenesis in a mouse model of age-related macular degeneration. To evaluate the efficacy of anti-FAM19A5 antibody therapy for age-related macular degeneration, a mouse-induced choroidal neovascularization (CNV) model was used. More specifically, the CNV model is an animal model of human wet age-related macular degeneration. Briefly, C57BL / 6 mice were stabilized for one week, and then the pupils of the animals' eyes were dilated (by local treatment with 2.5% phenylephrine hydrochloride). The animals were then intraperitoneally treated with 2% fluorescein (Sigma) to stain the blood vessels. Approximately 3-5 minutes after fluorescein injection, the mice were anesthetized, and wet AMD-like lesions were induced in four regions of the optic nerve using Phoenix green laser photocoagulation (240mW, 70ms duration). See Figure 6. Subsequently, on the day of CNV induction (i.e., day 0) and on days 3 and 7 after CNV induction, human IgG control antibody and anti-FAM19A5 antibody were administered intravitreously to mice (4 μg / eye). On day 13 after CNV induction, the degree of choroidal neovascularization was measured using fluorescein angiography (FFA) and the ImageJ program, and the corrected total fluorescence (CTF = integrated density - (area of ​​selected lesion × average fluorescence of background interpretation)) value was calculated. In addition, retinal tomography was performed using image-guided optical coherence tomography (OCT) equipment to measure the degree of neovascular lesions. Neovascular lesions were quantified using the ImageJ program.

[0296] As shown in Figure 7A, the fluorescence intensity observed in the four laser-exposed regions (indicated by arrows) immediately after CNV induction and before antibody treatment (day 0, left column) was similar in the two treatment groups. However, mice treated with anti-FAM19A5 antibody on day 13 after CNV induction had statistically lower CTF values ​​compared to mice treated with human IgG control antibody. See Figure 7B. This effect was confirmed by both image-guided optical coherence tomography (OCT) (see Figures 8A-8C) and H&E staining (see Figure 10B). These results demonstrate that anti-FAM19A5 antibody can inhibit and / or reduce laser-induced choroidal angiogenesis.

[0297] (Example 4) Evaluation of electroretinoid potential difference after in vivo administration of anti-FAM19A5 antibody in a mouse model of age-related macular degeneration. To further evaluate the efficacy of anti-FAM19A5 antibody therapy for age-related macular degeneration, electroretinography (ERG) was performed on mice from Example 3. The pupils of animals in different treatment groups were briefly dilated by topical treatment with 2.5% phenylephrine hydrochloride. The animals were then anesthetized, and electrodes were attached to the skin, tail, and cornea. A single flash of white light (0.9 log cd sec / m²) was used. 2 The retina was stimulated with ). To evaluate the electrical activity inside the eyes of different animals (i.e., "potential retinal difference"), the B-wave value was measured as described in Example 1.

[0298] As shown in Figure 9, CNV-inducing mice treated with the anti-FAM19A5 antibody had improved B-wave retinal potential compared to mice treated with human IgG antibodies. This result suggests that administration of the anti-FAM19A5 antibody to CNV-inducing mice restored normal retinal function.

[0299] (Example 5) Evaluation of morphological changes after anti-FAM19A5 antibody treatment in a mouse model of age-related macular degeneration. Next, to evaluate whether administration of anti-FAM19A5 antibody caused intraretinal morphological changes, CNV-inducing mice from Examples 3 and 4 were sacrificed, and both eyes were extracted from the animals. The eyes were then fixed in 10% neutral formalin for 48 hours. Subsequently, the fixed eyes were embedded in paraffin blocks, and H&E and IHC analysis were performed. Using H&E, the inhibitory effect of the anti-FAM19A5 antibody on choroidal angiogenesis, as shown in Example 3, was confirmed. Platelet-derived growth factor (PDGF) expression was stained using IHC. PDGF is known to be secreted from retinal neurons and plays an important role in astrocytic cell formation and choroidal angiogenesis.

[0300] As shown in Figures 10A and 10B, choroidal angiogenesis in CNV-induced mice treated with anti-FAM19A5 antibody was significantly reduced in size up to 13 days after CNV induction compared with the human IgG-treated group. This result confirms that the initial data (see Example 3) demonstrate the inhibitory effect of anti-FAM19A5 on choroidal angiogenesis. Furthermore, as shown in Figure 11, CNV-induced mice treated with anti-FAM19A5 antibody showed significantly less PDGF staining in both the nerve fiber layer and ganglion cell layer of the retina compared with the IgG-treated group.

[0301] (Example 6) Evaluation of the inhibitory effect of anti-FAM19A5 antibodies on drsen generation. As mentioned above, drusen are small yellow or white deposits of extracellular material (mainly proteins and lipids) that accumulate between Brooks' membrane and the retinal pigment epithelium (RPE) of the eye. To evaluate whether administration of anti-FAM19A5 antibody can inhibit drusen formation, male APP / PS-1 / tau-generated dementia mice (C57BL / 6; 4 months old) were used. Anti-FAM19A5 antibody was briefly administered intravenously to the mice at a dose of 0.7 mg / kg twice a week for a total of 8 weeks. Age-matched, untreated, unadministered wild-type C57BL / 6 and APP / PS-1 / tau-generated mice were used as control groups. Eight weeks after initial antibody administration, the animals were sacrificed and their eyes were extracted. After harvesting, the eyes were fixed in 4% paraformaldehyde (PFA) at 4°C for 24 hours. Subsequently, the eye was placed in a plastic mold with OCT compound (Lot no. 686794, Sigma, USA) and frozen. The frozen eye tissue was then thinned (25 μm thick) using a microtome (Cryostats CM1860, Leica, Germany) at -20°C. The thinned tissue sections were placed on slides and stained with oligomeric Aβ and Alexa 647 phalloidin to observe drusen formation.

[0302] As shown in Figure 12, in untreated APP / PS-1 / tau transgenic mice, oligomeric Aβ was strongly expressed around visual pigment cells, while drusen (verified by positive phalloidin staining) was strongly expressed around retinal tissue, cone cells, and ganglion cell layers. In contrast, APP / PS-1 / tau transgenic mice treated with anti-FAM19A5 antibody showed negligible oligomeric Aβ and drusen staining, similar to untreated wild-type C57BL / 6 control mice.

[0303] These results confirm that the anti-FAM19A5 antibody can inhibit oligomeric Aβ and drusen formation in the retina, including in surrounding tissues.

[0304] In short, the data above demonstrates that the anti-FAM19A5 antibody improves most of the underlying conditions associated with macular degeneration, such as age-related macular degeneration, and can treat the disease.

[0305] (Example 7) Evaluation of the inhibitory effect of anti-FAM19A5 antibody on diabetic retinopathy after intravenous administration. To evaluate the therapeutic effect of anti-FAM19A5 antibodies on diabetic retinopathy after intravenous administration, type 2 diabetic db / db mice (e.g., the mice described in Example 1) were used. After briefly stabilizing the animals, they were treated with one of the following: (i) human IgG antibody, (ii) aflibercept (EYLEA, Regeneron Pharmaceuticals), or (iii) anti-FAM19A5 antibody (clones 1-30). Human IgG antibody and aflibercept were used as negative and positive control groups, respectively. An untreated group (i.e., animals without diabetic retinopathy) was used as an additional control group. Different treatment formulations were administered intravitreously into the vitreous cavity of mice using a micropump system (2 μL / eye every 2 weeks for a total of 7 doses) or intravenously by tail vein injection (100 μL every 2 weeks for a total of 7 doses). The treatment groups are listed in Table 8 below.

[0306] [Table 9]

[0307] For example, the overall health status of the animals was periodically monitored by measuring serum blood glucose levels and / or body weight. After the last dose, the mice were euthanized and both eyes were removed from the animals. After removal, the eyes were prepared on OCT blocks and fixed for trypsinogenesis analysis, hematoxylin-eosin staining (H&E staining), TUNEL analysis, and immunofluorescence staining, as described in Example 2 and described in more detail below. Trypsinogenesis analysis was performed to evaluate microvessels against perivascular cell loss and acellular capillary formation (i.e., markers of early diabetic retinal disease). H&E staining, TUNEL analysis, and immunofluorescence staining were performed to analyze vascular congestion, vascular markers (CD31 and VEGF), and retinal cell protective effects.

[0308] H&E analysis The frozen tissue was thinned and placed on a slide (see Example 2), and the slide was dried at room temperature for about 1 hour. Next, the slide was stained with hematoxylin for 3-5 minutes and washed with HCl solution for 30 seconds. Then, the slide was immersed in eosin solution for 1 minute and washed again with HCl solution. Next, the slide was washed with 80%-100% ethanol for 3 minutes and reacted with carboxylene and xylene for 5 minutes each. The stained slides were observed using a microscope (NanoZoomer 2.0 RS, Hamamatsu).

[0309] TUNEL analysis Slides with frozen tissue sections mounted (see Example 2) were dried at room temperature for approximately 1 hour. Once dried, the slides were immersed in 1% paraformaldehyde for 10 minutes, followed by two washes with PBS for 5 minutes each. The slides were then immersed in equilibrium buffer for 10 minutes. Subsequently, the tissue samples were treated with TdT enzyme in a humidified chamber at 37°C for 2 hours. The slides were then washed four times with PBS for 2 minutes each. The stained slides were allowed to dry completely at room temperature. Once dried, the tissue samples were stained with DAPI and observed under a fluorescence microscope (LEICA DM 2500).

[0310] Immunofluorescence analysis Slides with frozen tissue sections mounted (see Example 2) were dried at room temperature for approximately 1 hour. Once dried, the tissue samples were treated with 5% (v / v) Triton X-100 solution dissolved in PBS and washed three times with PBS for 5 minutes each. The tissues were then blocked for 1 hour with normal donkey serum, 3% (v / v) BSA, and 0.5% Triton X-100 solution. Subsequently, the tissues were stained overnight at 4°C with anti-VEGF and / or anti-CD31 primary antibody (diluted 1:300 in PBS with 3% (v / v) BSA and 0.5% Triton X-100 solution). The following day, the samples were washed three times with PBS for 5 minutes each. The tissue samples were then reacted with secondary antibody (Alexa Fluor 555 donkey anti-rabbit IgG, diluted 1:500 in PBS with 3% (v / v) BSA and 0.5% Triton X-100 solution) at room temperature for 1 hour. Next, the samples were washed three times with PBS for 5 minutes each, stained with DAPI, and observed under a fluorescence microscope (LEICA DM2500).

[0311] result As shown in Table 9 (below), Figures 13A and 13C (consistent with the data provided in Example 2), the E / P ratio (i.e., the ratio of perivascular cells to endothelial cells) in animals treated with anti-FAM19A5 antibody (both intravenous and intravitreal administration groups) was lower than in animals treated with anti-human IgG alone ("negative control group"). Anti-FAM19A5 administration also showed a greater effect on the E / P ratio compared to the positive control group (i.e., animals treated with aflibercept), and was similar to the ratio observed in untreated (i.e., healthy) animals. The greatest effect was observed with intravitreal administration, but noteworthy therapeutic effects were also observed with intravenous administration. Similar results were observed when the number of acellular capillaries was measured in the eyes of the animals (see Table 9 and Figure 13B).

[0312] [Table 10]

[0313] The route of administration is considered to have little effect on the ability of the anti-FAM19A5 antibody to reduce vascular congestion and protect retinal cells from diabetic damage (see Figure 14). As shown in Figure 15, TUNEL analysis showed that in both the nerve fiber layer and inner core layer of the retina of mice treated intravenously with anti-FAM19A5 antibody, there were fewer cells undergoing cell death in the same layers of the retina of mice treated with human IgG control antibody. The effect was similar to that observed in animals treated intravitreously with anti-FAM19A5 antibody (see Figures 14 and 15, IVT vs. IV). Similar results were observed when evaluating the expression of CD31 and VEGF (labelers for neovascularization). In both animals treated intravitreously or intravenously with anti-FAM19A5 antibody, CD31 and VEGF expression was significantly reduced compared to the control group (i.e., animals treated with human IgG control antibody alone) (see Figures 16A and 16B).

[0314] In short, the data above confirms the results of Example 2 (i.e., intravitreal administration) and demonstrates that anti-FAM19A5 antibodies have a therapeutic effect on diabetic retinopathy after intravenous and intravitreal administration.

[0315] The detailed descriptive portions other than the summary and abstract should be understood to be used for interpreting the claims. The summary and abstract represent one or more exemplary aspects of the disclosure as considered by the inventor, but not all exemplary aspects, and are not intended to limit the disclosure or the attached claims in any way.

[0316] The contents of this disclosure have been described above using functional components that illustrate the embodiment of specific functions and their relationships. The boundaries of these functional components are arbitrarily defined herein for the sake of clarity. Alternative boundaries may be defined, provided that the specific functions and their relationships are appropriately represented.

[0317] The above description of specific embodiments is intended to fully represent the general attributes of the Disclosure so that others, by applying their knowledge of the art, may readily modify and / or adapt such specific embodiments for various applications without excessive experimentation, without exceeding the general concepts of the Disclosure. Accordingly, such adaptations and modifications are intended to be equivalent to the embodiments disclosed herein, based on the teachings and guidance presented herein. It will be understood that the words or terms used herein are intended to be descriptive, not restrictive, so that they may be interpreted by those skilled in the art in light of the teachings and guidance of the invention.

[0318] The scope and breadth of this disclosure should not be limited by any of the exemplary embodiments described above, but should be defined solely by the claims and equivalents set forth below.

[0319] All publications, patents, patent applications, internet sites, and accession numbers / database sequences (both polynucleotide and polypeptide sequences) cited herein are incorporated herein in whole for all purposes to the same extent as if each individual publication, patent, patent application, internet site, or accession number / database sequence were specifically and individually indicated as being included for reference. [Brief explanation of the drawing]

[0320] [Figure 1A] Figure 1A shows the effect of anti-FAM19A5 antibody on retinoid potential recovery in 25-week-old db / db mice. These mice were treated with either human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clones "1-65" or "3-2"). Normal (i.e., non-diabetic) mice were used as the control group. After exposing each group of mice to white light at three different flash intensities (-1, 0.2, and 0.8 log cds / m2), wave A (μV) (Figure 1A), wave B (μV) (Figure 1B), and rhythmic wave (μV) (Figure 1C) were measured. Data are expressed as mean ± SD. [Figure 1B] Figure 1B shows the effect of anti-FAM19A5 antibody on retinoid potential recovery in 25-week-old db / db mice. These mice were treated with either human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clones "1-65" or "3-2"). Normal (i.e., non-diabetic) mice were used as the control group. After exposing each group of mice to white light at three different flash intensities (-1, 0.2, and 0.8 log cds / m2), wave A (μV) (Figure 1A), wave B (μV) (Figure 1B), and rhythmic wave (μV) (Figure 1C) were measured. Data are expressed as mean ± SD. [Figure 1C] Figure 1C shows the effect of anti-FAM19A5 antibody on retinoid potential recovery in 25-week-old db / db mice. These mice were treated with either human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clones "1-65" or "3-2"). Normal (i.e., non-diabetic) mice were used as the control group. After exposing each group of mice to white light at three different flash intensities (-1, 0.2, and 0.8 log cds / m2), wave A (μV) (Figure 1A), wave B (μV) (Figure 1B), and rhythmic wave (μV) (Figure 1C) were measured. Data are expressed as mean ± SD. [Figure 2A]Figure 2A shows the protective effect of anti-FAM19A5 antibody on retinal microvascular tissue in 25-week-old db / db mice. These mice were treated with either human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clone "1-65" or "3-2"). Normal (i.e., non-diabetic) healthy mice were used as the control group. Figure 2A shows micrographs (100× magnification) of representative retinal regions of mice after periodic acid Schiff (PAS) staining for each of the different treatment groups: (i) normal group (i.e., non-diabetic) (upper left panel), (ii) human IgG control group (upper right panel), and (iii) anti-FAM19A5 antibody (clone 1-65, lower left panel; clone 3-2, lower right panel). Black arrows indicate endothelial cells, white arrows indicate perivascular cells, and gray arrows indicate acellular capillaries. Figure 2B shows a comparison of the ratio of endothelial cells to perivascular cells ("E / P ratio") observed within the entire retinal region analyzed in mice from the different treatment groups. To determine the E / P ratio, the number of endothelial cells and perivascular cells was calculated in each photograph. Data are expressed as mean ± SD. "***" on the bars indicates a statistically significant difference (p<0.001) compared to the IgG treatment group. [Figure 2B]Figure 2B shows the protective effect of anti-FAM19A5 antibody on retinal microvascular tissue in 25-week-old db / db mice. These mice were treated with either human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clone "1-65" or "3-2"). Normal (i.e., non-diabetic) healthy mice were used as the control group. Figure 2A shows micrographs (100× magnification) of representative retinal regions of mice after periodic acid Schiff (PAS) staining for each of the different treatment groups: (i) normal group (i.e., non-diabetic) (upper left panel), (ii) human IgG control group (upper right panel), and (iii) anti-FAM19A5 antibody (clone 1-65, lower left panel; clone 3-2, lower right panel). Black arrows indicate endothelial cells, white arrows indicate perivascular cells, and gray arrows indicate acellular capillaries. Figure 2B shows a comparison of the ratio of endothelial cells to perivascular cells ("E / P ratio") observed within the entire retinal region analyzed in mice from the different treatment groups. To determine the E / P ratio, the number of endothelial cells and perivascular cells was calculated in each photograph. Data are expressed as mean ± SD. "***" on the bars indicates a statistically significant difference (p<0.001) compared to the IgG treatment group. [Figure 2C] Figure 2C shows a comparison of the number of acellular capillaries observed within the analyzed whole retinal region of mice from different treatment groups. Data are expressed as mean ± SD. An asterisk (*) on the bar indicates a statistically significant difference (p<0.05) compared to the IgG treatment group. [Figure 2D] Figure 2D shows a comparison of observed acellular capillary lengths. Data are expressed as mean ± SD. "**" and "***" on the bars indicate statistically significant differences compared to the IgG treatment group (p<0.01 and p<0.001, respectively). [Figure 3]Figure 3 shows the effect of anti-FAM19A5 antibody treatment on intraretinal vascular congestion in 25-week-old db / db mice using H&E staining. The mice were treated with human IgG control antibody ("IgG", upper right panel) or anti-FAM19A5 antibody (clone "1-65", lower left panel or clone "3-2", lower right panel). Normal (i.e., diabetes-free) healthy mice (upper left panel) were used as the control group. Arrowheads indicate examples of vascular congestion. The different retinal layers shown include (i) ganglion cell layer (GCL), (ii) internal plexiform layer (IPL), (iii) inner nuclear layer (INL), (iv) outer nuclear layer (ONL), and (v) retinal pigment epithelium (PE). [Figure 4] Figure 4 shows the retinal cell protective effect of anti-FAM19A5 antibody treatment in 25-week-old db / db mice using TUNEL analysis. The mice were treated with human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clones "1-65" or "3-2"). Normal (i.e., diabetes-free) healthy mice were used as a control group. In the middle column ("TUNEL"), red arrowheads indicate retinal cells undergoing cell death. The intensity of TUNEL staining correlates with the degree of cell death (e.g., decreased intensity indicates partial cell death). The different retinal layers shown include (i) ganglion cell layer (GCL), (ii) inner nucleus layer (INL), (iii) outer nucleus layer (ONL), and (iv) retinal pigment epithelium (PE). [Figure 5] Figure 5 shows the inhibitory effect of anti-FAM19A5 antibody on retinal neovascularization in 25-week-old db / db mice using immunohistochemistry staining for CD31 and VEGF expression. The mice were treated with human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clones "1-65" or "3-2"). Normal (i.e., diabetes-free) healthy mice were used as the control group. Arrowheads indicate positive CD31 (top row) or positive VEGF (bottom row) staining. [Figure 6A] Figure 6A shows a photograph of an image-guided laser system like the one used in Example 8. [Figure 6B]Figure 6B shows an example of a photograph of a retina exposed to a laser from the image-guided laser system shown in Figure 6A. [Figure 7A] Figure 7A shows the inhibitory effect of anti-FAM19A5 antibody (clone 3-2) on choroidal neovascularization (CNV) using fluorescein angiography (FFA) analysis. CNV-inducible mice treated with human IgG antibody were used as a control group. Figure 7A shows representative retinal images of mice treated with human IgG antibody (top row) or anti-FAM19A5 (clone 3-2) (bottom row) on the day of CNV induction ("day 0") and 13 days after CNV induction, showing four wet AMD-like lesions (indicated by arrows in the left column). Figure 7B provides a comparison of corrected total fluorescence (CTF) values ​​for CNV-inducible mice treated with human IgG antibody ("IgG") or anti-FAM19A5 antibody (clone "3-2") 13 days after CNV induction. Data are presented as means and individually. "*" indicates a statistically significant difference compared to the IgG-treated group (p<0.05). [Figure 7B] Figure 7B shows the inhibitory effect of anti-FAM19A5 antibody (clone 3-2) on choroidal neovascularization (CNV) using fluorescein angiography (FFA) analysis. CNV-inducible mice treated with human IgG antibody were used as a control group. Figure 7A shows representative retinal images of mice treated with human IgG antibody (top row) or anti-FAM19A5 (clone 3-2) (bottom row) on the day of CNV induction ("day 0") and 13 days after CNV induction, showing four wet AMD-like lesions (indicated by arrows in the left column). Figure 7B provides a comparison of corrected total fluorescence (CTF) values ​​for CNV-inducible mice treated with human IgG antibody ("IgG") or anti-FAM19A5 antibody (clone "3-2") 13 days after CNV induction. Data are presented as means and individually. "*" indicates a statistically significant difference compared to the IgG-treated group (p<0.05). [Figure 8A]Figure 8A shows the inhibitory effect of anti-FAM19A5 antibody (clone 3-2) on choroidal neovascularization (CNV) using optical coherence tomography (OCT) analysis. CNV-inducible mice administered with human IgG antibody were used as a control group. Figure 8A provides representative retinal images showing four wet AMD-like lesions (labeled 1, 2, 3, and 4) in mice treated with human IgG antibody (top row) or anti-FAM19A5 (clone 3-2) (bottom row) 13 days after CNV induction. The columns labeled 1, 2, 3, and 4 provide OCT images of the lesions shown in the leftmost image. Figure 8B shows the scale bars used to quantify the size of the lesions. Figure 8C provides a comparison of the size of CNV lesions observed 13 days after CNV induction in mice treated with human IgG control antibody or anti-FAM19A5 antibody (clone 3-2). Data are presented as averages and individually. "***" indicates a statistically significant difference (p<0.001) compared to the IgG treatment group. [Figure 8B] Figure 8B shows the inhibitory effect of anti-FAM19A5 antibody (clone 3-2) on choroidal neovascularization (CNV) using optical coherence tomography (OCT) analysis. CNV-inducible mice administered with human IgG antibody were used as a control group. Figure 8A provides representative retinal images showing four wet AMD-like lesions (labeled 1, 2, 3, and 4) in mice treated with human IgG antibody (top row) or anti-FAM19A5 (clone 3-2) (bottom row) 13 days after CNV induction. The columns labeled 1, 2, 3, and 4 provide OCT images of the lesions shown in the leftmost image. Figure 8B shows the scale bars used to quantify the size of the lesions. Figure 8C provides a comparison of the size of CNV lesions observed 13 days after CNV induction in mice treated with human IgG control antibody or anti-FAM19A5 antibody (clone 3-2). Data are presented as averages and individually. "***" indicates a statistically significant difference (p<0.001) compared to the IgG treatment group. [Figure 8C]Figure 8C shows the inhibitory effect of anti-FAM19A5 antibody (clone 3-2) on choroidal neovascularization (CNV) using optical coherence tomography (OCT) analysis. CNV-inducible mice administered with human IgG antibody were used as a control group. Figure 8A provides representative retinal images showing four wet AMD-like lesions (labeled 1, 2, 3, and 4) in mice treated with human IgG antibody (top row) or anti-FAM19A5 (clone 3-2) (bottom row) 13 days after CNV induction. The columns labeled 1, 2, 3, and 4 provide OCT images of the lesions shown in the leftmost image. Figure 8B shows the scale bars used to quantify the size of the lesions. Figure 8C provides a comparison of the size of CNV lesions observed 13 days after CNV induction in mice treated with human IgG control antibody or anti-FAM19A5 antibody (clone 3-2). Data are presented as averages and individually. "***" indicates a statistically significant difference (p<0.001) compared to the IgG treatment group. [Figure 9] Figure 9 shows the effect of anti-FAM19A5 antibody on the recovery of B-wave values ​​in CNV-inducing mice. The CNV-inducing mice were treated with either human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clone "3-2"). Normal (i.e., non-diabetic) mice were used as the control group. Data are expressed as mean ± SD. "*" indicates a statistically significant difference compared to the IgG-treated group (p<0.05). [Figure 10A]Figure 10A shows the inhibitory effect of anti-FAM19A5 antibody (clone 3-2) on choroidal neovascularization (CNV). Figure 10A identifies different retinal layers using H&E analysis (left image) or OCT analysis (right image). These shown layers include: (i) retinal nerve fiber layer / ganglion cell layer (RNFL / GCL); (ii) inner plexiform layer; (iii) inner plexiform layer; (iv) outer plexiform layer; (v) outer plexiform layer; (vi) photoreceptor inner / outer segment (IS / OS); and (vii) retinal pigment epithelium (RPE). Figure 10B shows representative H&E images of the relevant cell layers of the mouse retina in different groups: (i) normal healthy animals, (ii) CNV-induced mice treated with human IgG antibody, and (iii) CNV-induced mice treated with anti-FAM19A5 antibody. Arrowheads indicate areas of choroidal neovascularization. [Figure 10B] Figure 10B shows the inhibitory effect of anti-FAM19A5 antibody (clone 3-2) on choroidal neovascularization (CNV). Figure 10A identifies different retinal layers using H&E analysis (left image) or OCT analysis (right image). These shown layers include: (i) retinal nerve fiber layer / ganglion cell layer (RNFL / GCL); (ii) inner plexiform layer; (iii) inner plexiform layer; (iv) outer plexiform layer; (v) outer plexiform layer; (vi) photoreceptor inner / outer segment (IS / OS); and (vii) retinal pigment epithelium (RPE). Figure 10B shows representative H&E images of the relevant cell layers of the mouse retina in different groups: (i) normal healthy animals, (ii) CNV-induced mice treated with human IgG antibody, and (iii) CNV-induced mice treated with anti-FAM19A5 antibody. Arrowheads indicate areas of choroidal neovascularization. [Figure 11] Figure 11 shows the inhibitory effect of anti-FAM19A5 antibody on PDGF expression in both the nerve fiber layer and ganglion cell layer of the retina, as demonstrated by IHC analysis. CNV-inducing mice were treated with human IgG control antibody ("IgG") or anti-FAM19A5 antibody (clone "3-2"). Normal (i.e., non-diabetic) mice were used as the control group. Arrowheads indicate positive PDGF staining. [Figure 12]Figure 12 shows the inhibitory effect of anti-FAM19A5 antibody on oligomeric Aβ and drusen formation in APP / PS-1 / tau gene-transfected mice (dementia model) using immunofluorescence microscopy. The APP / PS-1 / tau gene-transfected mice were either untreated (middle row) or treated (bottom row) with anti-FAM19A5 antibody. Untreated (naive) wild-type C57BL / 6 mice were used as the negative control group (top row). The column labeled "H33342" (nucleic acid staining) shows cells within retinal tissue. The column labeled "QD525-oAβ" shows oligomeric Aβ. The column labeled "647 Alexa Phalloidin" shows drusen formation, with drusen illustrated by arrows. The column labeled "merged" shows a composite image of the top three columns for each treatment group. [Figure 13A]Figure 13A shows a comparison of the retinal microvascular protective effects of anti-FAM19A5 antibodies (clones 1-30) after intravitreous or intravenous administration in a mouse model of diabetic retina. The mice were administered either (i) human IgG control antibody ("human IgG"), (ii) aflibercept, or (iii) anti-FAM19A5 antibody intravitreously ("IVT"), or (iv) anti-FAM19A5 antibody intravenously ("IV"). Normal (i.e., non-diabetic) healthy mice were used as the control group. Figure 13A provides micrographs (100× magnification) of representative retinal regions of mice in each of the different treatment groups after periodate Schiff (PAS) staining. Black arrows indicate endothelial cells, white arrows indicate perivascular cells, and black arrows indicate acellular capillaries. Figure 13B provides a comparison of the number of acellular capillaries observed within the analyzed overall retinal region of mice from the different treatment groups. Figure 13C provides a comparison of the ratio of endothelial cells to perivascular cells ("E / P ratio") observed within the analyzed whole retinal region of mice from different treatment groups. To determine the E / P ratio, the number of endothelial cells and perivascular cells was calculated in each photograph. In Figures 13B and 13C, data are expressed as mean ± SD. "*" and "***" on the bars indicate statistically significant differences compared to untreated mice (p<0.05 and p<0.001, respectively). "##" and "###" on the bars indicate statistically significant differences compared to the human IgG antibody treatment group (p<0.01 and p<0.001, respectively). "†††" on the bars indicate a statistically significant difference compared to the aflibercept treatment group (p<0.001). "§" on the bars indicates a statistically significant difference compared to the IVT group (p<0.05). [Figure 13B]Figure 13B shows a comparison of the retinal microvascular protective effects of anti-FAM19A5 antibodies (clones 1-30) after intravitreous or intravenous administration in a mouse model of diabetic retina. The mice were administered either (i) human IgG control antibody ("human IgG"), (ii) aflibercept, or (iii) anti-FAM19A5 antibody intravitreously ("IVT"), or (iv) anti-FAM19A5 antibody intravenously ("IV"). Normal (i.e., non-diabetic) healthy mice were used as the control group. Figure 13A provides micrographs (100× magnification) of representative retinal regions of mice in each of the different treatment groups after periodate Schiff (PAS) staining. Black arrows indicate endothelial cells, white arrows indicate perivascular cells, and black arrows indicate acellular capillaries. Figure 13B provides a comparison of the number of acellular capillaries observed within the analyzed overall retinal region of mice from the different treatment groups. Figure 13C provides a comparison of the ratio of endothelial cells to perivascular cells ("E / P ratio") observed within the analyzed whole retinal region of mice from different treatment groups. To determine the E / P ratio, the number of endothelial cells and perivascular cells was calculated in each photograph. In Figures 13B and 13C, data are expressed as mean ± SD. "*" and "***" on the bars indicate statistically significant differences compared to untreated mice (p<0.05 and p<0.001, respectively). "##" and "###" on the bars indicate statistically significant differences compared to the human IgG antibody treatment group (p<0.01 and p<0.001, respectively). "†††" on the bars indicate a statistically significant difference compared to the aflibercept treatment group (p<0.001). "§" on the bars indicates a statistically significant difference compared to the IVT group (p<0.05). [Figure 13C]Figure 13C shows a comparison of the retinal microvascular protective effects of anti-FAM19A5 antibodies (clones 1-30) after intravitreous or intravenous administration in a mouse model of diabetic retina. The mice were administered either (i) human IgG control antibody ("human IgG"), (ii) aflibercept, or (iii) anti-FAM19A5 antibody intravitreously ("IVT"), or (iv) anti-FAM19A5 antibody intravenously ("IV"). Normal (i.e., non-diabetic) healthy mice were used as the control group. Figure 13A provides micrographs (100× magnification) of representative retinal regions of mice in each of the different treatment groups after periodate Schiff (PAS) staining. Black arrows indicate endothelial cells, white arrows indicate perivascular cells, and black arrows indicate acellular capillaries. Figure 13B provides a comparison of the number of acellular capillaries observed within the analyzed overall retinal region of mice from the different treatment groups. Figure 13C provides a comparison of the ratio of endothelial cells to perivascular cells ("E / P ratio") observed within the analyzed whole retinal region of mice from different treatment groups. To determine the E / P ratio, the number of endothelial cells and perivascular cells was calculated in each photograph. In Figures 13B and 13C, data are expressed as mean ± SD. "*" and "***" on the bars indicate statistically significant differences compared to untreated mice (p<0.05 and p<0.001, respectively). "##" and "###" on the bars indicate statistically significant differences compared to the human IgG antibody treatment group (p<0.01 and p<0.001, respectively). "†††" on the bars indicate a statistically significant difference compared to the aflibercept treatment group (p<0.001). "§" on the bars indicates a statistically significant difference compared to the IVT group (p<0.05). [Figure 14] Figure 14 shows the inhibitory effect of anti-FAM19A5 antibody on intraretinal vascular congestion formation in diabetic retinal mice after intravenous and intravitreous administration. The treatment groups are the same as those described in Figures 13A to 13C. Arrowheads indicate examples of vascular congestion. [Figure 15]Figure 15 shows the retinal cell protective effect of anti-FAM19A5 antibody, measured using TUNEL analysis after intravitreous or intravenous administration. The treatment groups are the same as those described in Figures 13A to 13C. In the middle column ("TUNEL"), arrowheads indicate retinal cells undergoing cell death. The intensity of TUNEL staining correlates with the degree of cell death (for example, decreased intensity indicates partial cell death). [Figure 16A] Figure 16A shows the inhibitory effect of anti-FAM19A5 antibody on intraretinal neovascularization in diabetic retinal mice after intravitreous and intravenous administration. The treatment groups are the same as those described in Figures 13A to 13C. Angiogenesis was measured using VEGF (Figure 16A) and CD31 (Figure 16B) expression. Arrowheads indicate positive VEGF or CD31 staining. [Figure 16B] Figure 16B shows the inhibitory effect of the anti-FAM19A5 antibody on intraretinal neovascularization in diabetic retinal mice after intravitreous and intravenous administration. The treatment groups are the same as those described in Figures 13A to 13C. Angiogenesis was measured using VEGF (Figure 16A) and CD31 (Figure 16B) expression. Arrowheads indicate positive VEGF or CD31 staining.

Claims

1. A pharmaceutical composition for reducing or inhibiting intraretinal neovascularization in the target area, It contains a FAM19A5 antagonist for the family member A5 ("FAM19A5") protein, which has sequence similarity 19. The FAM19A5 antagonist is an antibody that specifically binds to the FAM19A5 protein or its antigen-binding portion ("anti-FAM19A5 antibody"), a polynucleotide encoding the anti-FAM19A5 antibody, or a vector containing the polynucleotide. The anti-FAM19A5 antibody comprises heavy chain CDR1, CDR2 and CDR3 and light chain CDR1, CDR2 and CDR3. (i) The heavy chain CDR1 includes the amino acid sequence shown in SEQ ID NO: 14, the heavy chain CDR2 includes the amino acid sequence shown in SEQ ID NO: 15, the heavy chain CDR3 includes the amino acid sequence shown in SEQ ID NO: 16, the light chain CDR1 includes the amino acid sequence shown in SEQ ID NO: 26, the light chain CDR2 includes the amino acid sequence shown in SEQ ID NO: 27, and the light chain CDR3 includes the amino acid sequence shown in SEQ ID NO: 28; (ii) The heavy chain CDR1 includes the amino acid sequence shown in SEQ ID NO: 17, the heavy chain CDR2 includes the amino acid sequence shown in SEQ ID NO: 18, the heavy chain CDR3 includes the amino acid sequence shown in SEQ ID NO: 19, the light chain CDR1 includes the amino acid sequence shown in SEQ ID NO: 29, the light chain CDR2 includes the amino acid sequence shown in SEQ ID NO: 30, the light chain CDR3 includes the amino acid sequence shown in SEQ ID NO: 31; or, (iii) A pharmaceutical composition in which the heavy chain CDR1 contains the amino acid sequence shown in SEQ ID NO: 212, the heavy chain CDR2 contains the amino acid sequence shown in SEQ ID NO: 213, the heavy chain CDR3 contains the amino acid sequence shown in SEQ ID NO: 16, the light chain CDR1 contains the amino acid sequence shown in SEQ ID NO: 222, the light chain CDR2 contains the amino acid sequence shown in SEQ ID NO: 225, and the light chain CDR3 contains the amino acid sequence shown in SEQ ID NO:

224.

2. The pharmaceutical composition according to claim 1, wherein the neovascularization is retinal neovascularization or choroidal neovascularization.

3. The anti-FAM19A5 antibody comprises a heavy chain variable region (VH) and a light chain variable region (VL), (i) The VH comprises the amino acid sequence of SEQ ID NO: 36, and the VL comprises the amino acid sequence of SEQ ID NO: 40; (ii) The VH comprises the amino acid sequence of SEQ ID NO: 37, and the VL comprises the amino acid sequence of SEQ ID NO: 41; or, (iii) The pharmaceutical composition according to claim 1, wherein VH comprises the amino acid sequence of SEQ ID NO: 236 and VL comprises the amino acid sequence of SEQ ID NO:

245.

4. The pharmaceutical composition according to claim 1, wherein the anti-FAM19A5 antibody comprises Fab, Fab', F(ab')2, Fv, or single-chain Fv(scFv).

5. The pharmaceutical composition according to claim 1, wherein the anti-FAM19A5 antibody is selected from the group consisting of IgG1, IgG2, IgG3, IgG4 and their variants.

6. The pharmaceutical composition according to claim 5, wherein the anti-FAM19A5 antibody is IgG2, IgG4, or a combination thereof.

7. The pharmaceutical composition according to claim 6, wherein the anti-FAM19A5 antibody comprises an IgG2 / IgG4 isotype antibody.

8. The pharmaceutical composition according to claim 1, wherein the anti-FAM19A5 antibody includes a constant region that lacks Fc function.

9. The pharmaceutical composition according to claim 1, wherein the anti-FAM19A5 antibody is a chimeric antibody, a human antibody, or a humanized antibody.

10. The pharmaceutical composition according to claim 1, wherein the anti-FAM19A5 antibody is linked to a molecule having a second binding site to form a bispecific molecule.

11. The pharmaceutical composition according to claim 1, wherein the anti-FAM19A5 antibody is linked to the activator to form an immunozygote.

12. The pharmaceutical composition according to claim 1, further comprising administering a therapeutic agent.

13. The pharmaceutical composition according to claim 1, wherein the anti-FAM19A5 antibody is formed into a dosage form together with a pharmaceutically acceptable carrier.

14. The pharmaceutical composition according to claim 1, wherein the pharmaceutical composition is administered by intraocular administration.

15. The pharmaceutical composition according to claim 14, wherein the intraocular administration includes intravitreal administration.

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