Antibodies targeting angpt2 for the treatment of fibrotic diseases, in particular kidney diseases, conditions or injuries associated with kidney fibrosis

Abstract

Provided are antibodies targeting ANGPT2 for use in treating or preventing a fibrotic disease, particularly a kidney disease, disorder or injury, in a subject, wherein the kidney disease, disorder or injury is associated with kidney fibrosis.

Description

Technical Field

[0001] This invention relates to antibodies targeting ANGPT2 for the treatment or prevention of fibrotic diseases in subjects, particularly kidney diseases, conditions, or injuries, wherein the kidney diseases, conditions, or injuries are associated with kidney fibrosis. Background Technology

[0002] Chronic kidney disease (CKD) affects approximately 13% of the global population and is a leading cause of death from cardiovascular disease. While a variety of conditions, including diabetes and hypertension, can lead to CKD, the common pathogenic pathways ultimately shared in both animal models and patients include endothelial dysfunction, capillary desquamation, tubular atrophy, and renal fibrosis. Because there is currently no cure and relatively few therapies can slow progression, there is an urgent need to identify and develop new and effective treatments.

[0003] Angiopoietin (ANGPT) is a class of angiogenesis factors composed of three ANGPTs (ANGPT1, ANGPT2, and ANGPT3 / ANGPT4 (mice / human, respectively)). These ANGPTs share multiple cellular functions related to cell survival, proliferation, migration, and inflammation. ANGPTs exert their physiological and pathological effects through the TIE tyrosine kinase receptor. The ANGPT-TIE2 signaling pathway is involved in developmental and tumor-induced angiogenesis and is also involved in many disease environments, such as vascular diseases, systemic inflammation, and cancer. In the context of the kidney, ANGPT signaling is involved in the regulation of renal vascular development and responses to injury.

[0004] Endothelial growth factor 2 (TIE2), also known as TEK, is a tyrosine kinase receptor primarily expressed on endothelial cells. TIE2 signaling promotes vascular rest and reduces vascular permeability, inflammation, and endothelial cell apoptosis. TIE2 activity is regulated by the angiopoietin (ANGPT) ligands ANGPT1 and ANGPT2, as well as vascular endothelial tyrosine phosphatase (VEPTP).

[0005] ANGPT2-binding antibodies have been reported (e.g., US 7,658,924, US 8,987,420, and US 11,498,962), primarily in the context of cancer therapeutics. These include peptide-Fc fusion proteins; trobanib (also known as AMG386); and monoclonal antibodies targeting ANGPT2; MEDI3617, LY3127804, and nevaxumab (also known as REGN910) which neutralize the interaction between ANGPT2 and TIE2 (Saharinen et al., 2017). However, their therapeutic efficacy in kidney disease and other fibrotic diseases remains unknown.

[0006] Currently, there are no effective treatments for renal fibrosis. Current approaches include renin-angiotensin system (RAS) blockade, SGLT2 inhibition, GLP-1 receptor agonists, atrasentan (endothelin-1 blocker), tolvaptan (vasopressin receptor 2 antagonist), and fenelone (nonsteroidal anti-mineralocorticoid) (Huang et al., 2023). Nevertheless, fibrosis is a prominent symptom, thus there is an urgent need to explore innovative approaches to treat fibrotic diseases, particularly renal fibrosis. Summary of the Invention

[0007] As outlined above, TIE2-activating therapeutic agents have shown great potential for treating fibrotic diseases, such as kidney disease. However, certain drawbacks are associated with the use of these TIE2-activating therapeutic agents, limiting their clinical application, such as limitations related to the production, storage, half-life, specificity, and efficacy of these compounds. Therefore, there remains a need for TIE2-activating therapeutic agents with differentiated characteristics for the treatment of fibrotic diseases, such as kidney disease.

[0008] ANGPT2-binding antibodies (such as ABTAA) have shown great therapeutic potential in cancer, sepsis, and neovascular age-related macular degeneration; however, it remains unclear whether ABTAA is effective in treating the complex pathophysiology associated with fibrotic diseases, such as kidney diseases, which are associated with renal fibrosis.

[0009] The inventors have discovered that the ANGPT2-binding and TIE2-activating antibody ABTAA has therapeutic efficacy in treating fibrotic diseases, particularly kidney diseases (wherein the kidney diseases are associated with renal fibrosis), as outlined herein.

[0010] Examples of this invention demonstrate the unexpected therapeutic effects of ABTAA-induced ANGPT2 and TIE2 activation in an in vivo model of kidney disease, where ABTAA prevented the development of renal tubulointerstitial fibrosis compared to IgG-injected mice and WT mice. In contrast, TIE2... iECKO The disease was exacerbated in mice, and they exhibited significantly more tubulointerstitial fibrosis compared to WT mice and mice treated with ABTAA. These therapeutic effects of ABTAA on kidney disease were also observed in long-term models of kidney disease with higher clinical relevance.

[0011] Examples of this invention also demonstrate the unexpected therapeutic effects of ABTAA-induced ANGPT2 and TIE2 activation in an in vivo model of early diabetic complications, where ABTAA normalized plasma blood urea nitrogen levels, indicating a protective effect against nephropathy. ABTAA also significantly maintained capillary density in the renal cortex and preserved normal fenestrations in peritubular capillaries, which are crucial for renal function. Regarding cardiomyopathy, ABTAA treatment prevented elevation of plasma creatine kinase, indicating a protective effect against myocardial injury. Furthermore, ABTAA significantly reduced vascular degeneration in the retina (i.e., the number of empty collagen sheaths), indicating preservation of capillary structure.

[0012] Examples of the present invention also demonstrate the unexpected therapeutic effects of ABTAA-induced ANGPT2 and TIE2 activation in an in vivo model of coagulation disorders, wherein ABTAA completely blocked TNFα-induced shortening of tail bleeding time, indicating a protective effect against coagulation disorders.

[0013] Importantly, ABTAA exhibits an unexpected dual effect: normalizing pathologically unstable blood vessels and preventing the progression of fibrosis in kidney disease. This avoids the adverse effects associated with TIE2 overactivation, such as vasodilation and venous malformations (Vikkula et al., 1996). This antifibrotic effect of ABTAA has not been previously reported. Furthermore, ABTAA is advantageous as a therapeutic agent for fibrotic diseases (e.g., kidney disease) due to its long half-life of 198 hours in circulation, compared to 30 minutes for other TIE2 activators such as COMP-ANGPT1 (Han et al., 2016).

[0014] This invention relates to antibodies or antigen-binding fragments thereof that bind to ANGPT2 and, via ANGPT2, to the TIE2 receptor for the treatment or prevention of fibrotic diseases. In some embodiments, this invention relates to antibodies or antigen-binding fragments thereof that bind to ANGPT2 and, via ANGPT2, to the TIE2 receptor for the treatment or prevention of fibrosis. In some embodiments, this invention relates to antibodies or antigen-binding fragments thereof that bind to ANGPT2 and, via ANGPT2, to the TIE2 receptor for the treatment or prevention of kidney disease, condition, or injury in a subject, wherein the kidney disease, condition, or injury is associated with kidney fibrosis.

[0015] In one respect, the antibody or its antigen-binding fragment is characterized by binding to sites from position 336 to position 434 of angiopoietin-2 (ANGPT2) (SEQ ID NO: 30).

[0016] On the other hand, antibodies or their antigen-binding fragments contain:

[0017] a) A complementarity-determining region (CDR) of the heavy chain variable region, comprising: HCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0018] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0019] On the other hand, the antibody or its antigen-binding fragment comprises a variable heavy chain of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 31 or SEQ ID NO: 33, and a variable light chain of SEQ ID NO: 7, SEQ ID NO: 11, SEQ ID NO: 32, SEQ ID NO: 34 or SEQ ID NO: 38.

[0020] In some preferred embodiments, the antibody or its antigen-binding fragment comprises the variable heavy chain of SEQ ID NO: 33 and the variable light chain of SEQ ID NO: 34.

[0021] In some preferred embodiments, the antibody or its antigen-binding fragment comprises: a complementarity-determining region (CDR) of the heavy chain variable region, the CDR comprising: HCDR1, the HCDR1 comprising the amino acid sequence of SEQ ID NO: 27; HCDR2, the HCDR2 comprising the amino acid sequence of SEQ ID NO: 37; and HCDR3, the HCDR3 comprising the amino acid sequence of SEQ ID NO: 29; and a complementarity-determining region (CDR) of the light chain variable region, the CDR comprising: LCDR1, the LCDR1 comprising the amino acid sequence of SEQ ID NO: 35; LCDR2, the LCDR2 comprising the amino acid sequence of SEQ ID NO: 36; and LCDR3, the LCDR3 comprising the amino acid sequence of SEQ ID NO: 26.

[0022] In some embodiments, the antibody or antigen-binding fragment thereof described herein is encoded by a nucleotide sequence as described in U.S. Patent No. 9,994,632 B2.

[0023] On the other hand, the present invention relates to the following method:

[0024] a) Reduce or inhibit human angiopoietin-2 (ANGPT2) signaling, and / or induce TIE2 activation, and / or induce TIE2 phosphorylation in fibrotic diseases;

[0025] b) To normalize pathological blood vessels and treat and alleviate fibrotic diseases, and prevent the progression of fibrotic diseases;

[0026] c) Treat and / or reduce fibrotic disease and / or prevent the progression of fibrotic disease;

[0027] d) Prevent or treat inadequate perfusion in subjects, where the disease, condition, or injury is associated with fibrosis;

[0028] e) Prevent or reduce capillary sparsity associated with a subject’s disease, condition, or injury, wherein the disease, condition, or injury is associated with fibrosis; and / or

[0029] This includes administering an antibody or its antigen-binding fragment to a subject in need, wherein the antibody or its antigen-binding fragment comprises:

[0030] a) A complementarity-determining region (CDR) of the heavy chain variable region, comprising: HCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0031] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0032] On the other hand, the present invention relates to the following method:

[0033] a) Reduce or inhibit human angiopoietin-2 (ANGPT2) signaling in diseased kidneys and / or induce TIE2 activation and / or induce TIE2 phosphorylation, wherein the diseased kidney is a fibrotic kidney;

[0034] b) To normalize pathological blood vessels and treat and alleviate fibrosis in diseased kidneys, and prevent the progression of fibrosis in diseased kidneys;

[0035] c) Treat and / or reduce fibrosis in the diseased kidney and / or prevent the progression of fibrosis in the diseased kidney;

[0036] d) Prevent or treat renal hypoperfusion in subjects where kidney disease, condition, or injury is associated with renal fibrosis;

[0037] e) Prevent or reduce capillary sparsity associated with kidney disease, condition, or injury in the subject, wherein the kidney disease, condition, or injury is associated with fibrosis; and / or

[0038] f) Prevent or treat a reduction in peritubular capillary fenestrations associated with kidney disease, condition or injury in the subject, where the kidney disease, condition or injury is associated with renal fibrosis.

[0039] This includes administering an antibody or its antigen-binding fragment to a subject in need, wherein the antibody or its antigen-binding fragment comprises:

[0040] a) A complementarity-determining region (CDR) of the heavy chain variable region, comprising: HCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0041] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26. Attached Figure Description

[0042] Figure 1 (A) Schematic diagram of ABTAA administration and experimental setup, and (B) Inducible conditioned mouse strain.

[0043] Figure 2 ANGPT2 staining in the renal cortex of a UUO kidney after unilateral ureteral obstruction (UUO). Confocal imaging of the vascular system labeled with Cdh5-TdTomato in a UUO kidney 3 days later. Scale bar = 50µm, original magnification × 400.

[0044] Figure 3 UUO reduces TIE2 activation. WT mice and TIE2 iECKO (T2) KO TIE2 ELISA of kidney tissue homogenates from CL and 3-day-old UUO mice. Data are expressed as mean (SD), and n is represented by a point in the figure. #### P<0.0001, compared with WT UUO using Student's t-test (unpaired, two-tailed) within the same group. ****P<0.0001, compared with WT CL using one-way ANOVA followed by Bonferroni post-hoc test.

[0045] Figure 4 ABTAA-induced TIE2 activation reduced UUO-induced renal perfusion. Renal perfusion in UUO-induced and CL-induced kidneys was measured 2 days after intravenous injection of the circulatory marker MicroMarker and contrast imaging ultrasound in ABTAA-treated (ABT) mice and Veptp. iECKO (Ve KO Mice, TIE2 iECKO (T2) KO Mice and Pdgfb iKO (Pb) KO Results for mice and controls are expressed as UUO / CL. Data are expressed as mean (SD), and n is represented by a point in the figure. # P<0.05, ### P < 0.001, compared with WT / IgG UUO within the same group using Student's t-test (unpaired, two-tailed). **P < 0.01, ****P < 0.0001, compared with WT / IgG CL using one-way ANOVA followed by Bonferroni post-hoc test.

[0046] Figure 5 ABTAA-induced TIE2 activation alleviated UUO-induced capillary sparsity.

[0047] (A) Mice treated with ABTAA, Veptp iECKO Mice, TIE2 iECKO Mice and Pdgfb iKO Immunohistochemistry and quantification of endothelial markers, including (B) endothelial mucin and (C) podocyte marker proteins, were performed on the renal cortex of mice and their controls at 3 days of UUO. Scale bar = 50 µm, original magnification × 400. Five images of the renal cortex were taken from each mouse, and the mean was calculated. Data are expressed as mean (SD), and n in the figure is represented by a point. # P<0.05, ## P < 0.01, compared with WT / IgG UUO within the same group using Student's t-test (unpaired, two-tailed). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, compared with WT / IgG CL using one-way ANOVA followed by Bonferroni post-hoc test.

[0048] Figure 6 ABTAA-induced TIE2 activation alleviated UUO-induced endothelial injury. (A, B) Based on ABTAA-treated (ABT) mice, Veptp iECKO (VeKO Mice and TIE2 iECKO (T2) KO Microscopic images of peritubular capillaries in the kidneys of 3-day-old UUO mice, with semi-quantitative grading of capillary lengths with fenestrations. Scores: 0: 0%–5%, 1: 6%–25%, 2: 26%–50%, 3: 51%–75%, and 4: 76%–100%. Scale bar = 2µm. Data are expressed as mean (SD), and n is represented by a point in the figure. *P<0.01, **P<0.01, ***P<0.001, ****P<0.0001, compared with WT / IgG CL using one-way ANOVA followed by Bonferroni post-hoc test. # P<0.05, ## P<0.01, ### P<0.001, compared with WT / IgG UUO in the same group using Student's t-test (unpaired, two-tailed).

[0049] Figure 7 ABTAA-induced TIE2 activation alleviated UUO-induced renal tubular interstitial fibrosis. (AC) in ABTAA-treated (ABT) mice and Veptp iECKO (Ve KO Mice, TIE2 iECKO (T2) KO Mice and Pdgfb iKO (Pb) KO Immunohistochemistry and quantification of fibrosis markers (aSMA and vimentin) in the renal cortex of mice and controls after 3 days of UUO. Scale bar = 50µm, original magnification × 400. Five images of the renal cortex from each mouse were used for quantification, and the average value was taken. (D) Expression of Col1a1 in the kidneys of ABTAA-treated mice. (E) Veptp iECKO Expression of Pdgfrb in the kidneys of mice. (F) TIE2 iECKO Gene expression of Col1a1, Tagln, and Fn1 in the kidneys of mice with 3-day-old UUO. Data are presented as mean (SD), and n is represented by a dot in the figure. **P<0.01, ***P<0.001, ****P<0.0001, compared with WT / IgG CL using one-way ANOVA followed by Bonferroni post-hoc test. # P<0.05, ## P<0.01, ### P<0.001, #### P<0.0001, compared with WT / IgG UUO in the same group using Student's t-test (unpaired, two-tailed).

[0050] Figure 8 ABTAA-induced TIE2 activation alleviated UUO-induced renal tubular injury. (A) Renal tubular segments with pathological vacuoles are marked with white arrows in toluidine blue-stained kidney sections from 3-day-old UUO patients. (B) ABTAA-treated (ABT) mice and Veptp iECKO (Ve KO Mice and TIE2 iECKO (T2) KO Quantitative analysis was performed on renal tubular segments exhibiting vacuolation in mice. Data are expressed as mean (SD), and n is represented by a point in the figure. # P<0.05, compared with WT / IgG UUO in the same group using Student's t-test (unpaired, two-tailed).

[0051] Figure 9 ABTAA-induced TIE2 activation alleviated UUO-induced renal tubular injury and reduced PDGFB expression. (AC) ABTAA-treated (ABT) mice and Veptp iECKO (VeKO) mice and Pdgfb iKO (Pb) KO Expression of Pdgfb mRNA (A, C) and PDGFB protein (B) in the kidneys of 3-day-old UUO mice. *P<0.05, ***P<0.001, ****P<0.0001, compared with WT / IgG CL using one-way ANOVA followed by Bonferroni post-hoc test. # P<0.05, ## P<0.01, #### P<0.0001, compared with WT / IgG UUO in the same group using Student's t-test (unpaired, two-tailed).

[0052] Figure 10 Delayed initiation of ABTAA treatment reduces UUO-induced damage. (A) Schematic diagram of ABTAA administration used to evaluate kidney damage in UUO at 10 days. (BD) ABTAA treatment in mice (ABT) and TIE2. iECKO (T2) KO Immunohistochemistry was performed on the renal cortex of 10-day-old UUO kidneys of mice to detect renal tubulointerstitial fibrosis (aSMA, vimentin). Scale bar = 50 µm, original magnification × 400. (E) Protein concentration of PDGFB in the kidneys of 10-day-old ABTAA-treated (ABT) mice. Data are expressed as mean (SD), and n is represented by a point in the figure. ***P < 0.001, ****P < 0.0001, compared with WT / IgG CL using one-way ANOVA followed by Bonferroni post-hoc test. #P<0.05, ## P<0.01, compared with WT / IgG UUO in the same group using Student's t-test (unpaired, two-tailed).

[0053] Figure 11 ABTAA therapy has a protective effect against early diabetic complications. (A) ANGPT2 staining in the glomeruli of diabetic BTBRob / ob mice compared with non-diabetic mice (BTBRLean). (B) Preliminary results from the first BTBRob / ob study: treatment with 25 mg / kg ABTAA (ABT) or IgG compared with non-diabetic controls (ND). Mice were treated from 4 weeks of age and observed until 11 weeks of age. (C) Renal function (nephropathy) was evaluated by plasma blood urea nitrogen (BUN). (D) Capillary density of peritubular capillaries in the renal cortex was quantified by endothelial mucin (EMCN) staining. (E) Peritubulular capillary health was evaluated by scoring the normal fenestration pattern in electron microscopy images. (F) Plasma creatine kinase was used as a marker of cardiac injury. (G) Capillaries and basement membranes were stained with Pecam1 and type IV collagen, respectively, to evaluate retinopathy. Degenerated blood vessels and type IV collagen “vacuum” (i.e., where capillaries are lost) were quantified. Data are presented as mean (SD). *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001, compared with non-diabetic mice (ND) or as indicated, were analyzed using one-way ANOVA followed by Bonferroni post-hoc tests. # P<0.05, Student's t-test (unpaired, two-tailed) was used to compare ob / ob mice treated with ABX and IgG.

[0054] Figure 12 ABTAA therapy has a protective effect in coagulation disorders. (A) TNFα triggers the release of ANGPT2 in mouse plasma. (B) TNFα-induced coagulation disorders treated with ABX using a tail hemorrhage model. Data are expressed as mean (SD). **P<0.01, ****P<0.0001, compared with control Student's t-test (unpaired, two-tailed). Detailed Implementation

[0055] definition

[0056] As used herein, unless the context clearly states otherwise, the singular forms “a / an” and “the” may contain plural references. Thus, for example, a reference to “antibody” includes multiple such constructs.

[0057] The term "some implementations" may include one or more implementations.

[0058] As used herein, “antibody” (immunoglobulin) can be an oligoclonal antibody, polyclonal antibody, monoclonal antibody (including full-length monoclonal antibody), camelified antibody, chimeric antibody, CDR transplanted antibody, multispecific antibody, bispecific antibody, catalytic antibody, chimeric antibody, humanized antibody, fully human antibody, anti-idiotype antibody, and one or more antibodies capable of being labeled in soluble or bound form, as well as fragments, variants, or derivatives thereof, which may exist alone or in combination with other amino acid sequences provided by known techniques. Antibodies can be derived from any species. Antibodies comprise a polypeptide or polypeptide group consisting of at least one binding domain formed by the folding of a polypeptide chain having a three-dimensional binding space whose inner surface shape and charge distribution are complementary to the characteristics of the antigenic determinants of the antigen. Antibodies are typically in tetrameric form, comprising two pairs of identical polypeptide chains, each pair having a “light” chain and a “heavy” chain. Variable regions of each light / heavy chain pair form antibody binding sites. Natural antibodies are typically heterotetrameric glycoproteins of approximately 150,000 Daltons, composed of two identical light (L) chains and two identical heavy (H) chains. Each light chain is linked to the heavy chain by a covalent disulfide bond, the number of which varies among heavy chains of different immunoglobulin isotypes. Each heavy and light chain also has regularly spaced intrachain disulfide bridges. Each heavy chain has a variable domain (VH) at one end, followed by multiple constant domains. Each light chain has a variable domain (VL) at one end and a constant domain at the other end; the constant domain of the light chain is aligned with the first constant domain of the heavy chain, and the variable domain of the light chain is aligned with the variable domain of the heavy chain. Based on the amino acid sequence of the constant region of the light chain, the light chain is classified as either a λ chain or a κ chain. The variable domain of the κ light chain may also be referred to as VK in this document. The term "variable region" may also be used to describe the variable domain of either the heavy or light chain. It is believed that specific amino acid residues form an interface between the variable domains of the light and heavy chains. The variable region of each light chain / heavy chain pair forms an antibody binding site.

[0059] The term "antibody fragment" refers to any derivative of an antibody that is less than its full length. In one aspect, an antibody fragment retains at least a substantial portion of the specific binding capacity of the full-length antibody, particularly as a binding partner. Examples of antibody fragments include, but are not limited to, Fab, Fab', F(ab')2, scFv, Fv, dsFv biantibodies, and Fd fragments. Antibody fragments can be produced in any manner. For example, antibody fragments can be produced enzymatically or chemically by fragmentation of a complete antibody, or they can be produced recombinantly from a gene encoding a portion of the antibody sequence. Alternatively, antibody fragments can be produced wholly or partially synthetically. Antibody fragments may comprise single-chain antibody fragments. In another embodiment, the fragment may comprise, for example, multiple chains linked together by disulfide bonds. The fragment may also comprise a multimolecular complex. Functional antibody fragments typically contain at least about 50 amino acids, and more typically will contain at least about 200 amino acids.

[0060] The term "monoclonal antibody" refers to an antibody derived from a single cell clone (including any eukaryotic or prokaryotic cell clone) or bacteriophage clone, and not to the method of producing the antibody. Therefore, the term "monoclonal antibody" is not limited to antibodies produced through hybridoma technology.

[0061] As used herein, the term "ANGPT2-binding antibody" refers to an antibody that binds to ANGPT2, thereby altering the biological activity of ANGPT2, and is used interchangeably with "anti-ANGPT2 antibody," "ANGPT2-binding antibody," and "antibody that specifically binds to ANGPT2." As used herein, "antibody" refers to an immunoglobulin molecule that is immunoreactive to a specific antigen, meaning a protein molecule that acts as a receptor and specifically recognizes an antigen, and may include polyclonal antibodies, monoclonal antibodies (monoclonal antibodies), whole antibodies, and antibody fragments. Furthermore, antibodies may include chimeric antibodies (e.g., humanized mouse antibodies) and bivalent or bispecific molecules (e.g., bispecific antibodies), biantibodies, triantibodies, and tetraantibodies. In some preferred embodiments, the antibodies described herein are whole antibodies.

[0062] As used herein, the term "ABTAA" refers to a group of antibodies that bind to angiopoietin-2 and activate TIE2. These antibodies bind to ANGPT-2 and, via ANGPT2, to the TIE2 receptor, thereby activating the TIE2 receptor. ABTAA is used interchangeably with "ABT," "ABTAA," and "ABX." This group includes the anti-ANGPT2 antibody in US 11,498,962 and the anti-ANGPT2 antibody in US 2015 / 337033 A1.

[0063] As used herein, the term "pathological vascularity" refers to a pathological condition or pathophysiological process in which blood vessels exhibit abnormal or pathological features that deviate from normal vascular structure and function. These abnormal vessels may be associated with a variety of medical conditions, including but not limited to cancer, inflammatory conditions, vascular malformations, and certain eye diseases. Characteristics of pathological vascularity may include excessive angiogenesis, irregular branching, increased permeability, loss of fenestration, loss of capillary density, and impaired structural integrity.

[0064] As used herein, the term "endothelial dysfunction" refers to a pathological condition or pathophysiological process characterized by impaired or abnormal function of the endothelium (the lining of blood vessels). Endothelial dysfunction involves dysregulation of various physiological processes, including the control of vascular tone, blood coagulation, and inflammatory responses. It is associated with an imbalance in the production and availability of endothelial-derived factors, including but not limited to ANGPT2, thrombomodulin, nitric oxide, and endothelin, which play crucial roles in vascular homeostasis.

[0065] As used herein, the term “fibrotic disease” refers to any disease or condition caused by or associated with fibrosis. As used herein, the terms “fibrotic disease” and “fibrosis” are used interchangeably. Fibrosis is a pathological condition or pathophysiological process characterized by the excessive accumulation of extracellular matrix (ECM) components, resulting in the replacement of normal tissue structures with fibrous tissue. This process leads to the disruption of normal organ function and can affect a variety of organs, including the kidneys and heart. Fibrtic diseases involve a regulatory imbalance in ECM production and degradation and are often associated with chronic inflammation, tissue damage, and abnormal wound healing responses. Fibrosis can be associated with several chronic conditions, including type 2 diabetes. Fibrosis can also be induced by, spread by, and / or associated with coagulation disorders. As used herein, the term “fibrotic tissue” refers to any tissue associated with fibrosis. Fibrtic tissue can be identified by its typically firm, fibrotic, thickened, and inflexible nature compared to healthy tissue. Fibrtic tissue may also be referred to herein as “disease tissue.”

[0066] Fibrosis is defined as the replacement of functional tissue parenchyma by the extracellular matrix (ECM) and is used interchangeably with “sclerosis” and / or “scarring.” For example, fibrosis in transplanted kidneys primarily affects the tubulointerstitium, which comprises 90% of the kidney volume. Therefore, the terms “fibrosis” and “tubulointerstitial fibrosis / interstitial fibrosis / tubular atrophy (IFTA)” are used interchangeably in this document. IFTA can result from various kidney injuries, such as ischemia-reperfusion injury, rejection, infection, diabetes, and drug treatments (e.g., anticalcineurin agents).

[0067] As used herein, the term "renal fibrosis" refers to a pathological condition or pathophysiological process characterized by the excessive accumulation of extracellular matrix (ECM) proteins (including but not limited to collagen, smooth muscle actin, and vimentin) within renal tissue. This fibrotic process involves the activation of myofibroblasts, leading to scar tissue formation and disruption of the normal structure of the kidney. Renal fibrosis is commonly associated with various chronic kidney diseases and represents a progressive and irreversible condition that can result in impaired kidney function.

[0068] As used herein, the term "kidney disease, condition, or injury, wherein the kidney disease, condition, or injury is associated with renal fibrosis" refers to any condition in which renal fibrosis is a symptom or cause of the condition, or a condition that may be exacerbated by the development of renal fibrosis, or whose progression is associated with the progression of renal fibrosis. Therefore, conditions associated with renal fibrosis can provide therapeutic benefits by preventing, inhibiting, and / or delaying the progression of renal fibrosis. Conditions associated with renal fibrosis include, but are not limited to, diabetic nephropathy, chronic kidney disease, end-stage renal disease, systemic lupus erythematosus, vasculitis, IgA nephropathy, other autoimmune diseases, paraprotein diseases, and diabetes. Since chronic kidney disease associated with renal fibrosis is a very important risk factor for cardiovascular disease, it will be apparent to those skilled in the art that therapeutic agents that prevent or reduce renal fibrosis will have beneficial effects on systemic heart and vascular disease.

[0069] As used herein, the term "renal tubulointerstitial fibrosis" refers to a pathological condition or pathophysiological process characterized by the excessive accumulation of fibrous tissue (including but not limited to collagen, smooth muscle actin, and vimentin) within the tubulointerstitial region of the kidney. This fibrotic process involves the activation of myofibroblasts and the deposition of extracellular matrix, leading to structural changes, tissue scarring, and impaired renal function. Renal tubulointerstitial fibrosis is commonly associated with a variety of kidney conditions, including chronic kidney disease and inflammatory conditions.

[0070] As used herein, the term "glomerulosclerosis" refers to a pathological condition or pathophysiological process characterized by intraventricular scarring and excessive accumulation of extracellular matrix (including, but not limited to, collagen and smooth muscle actin) in the glomerular region. This scarring process involves the activation of mesangial cells and the deposition of extracellular matrix, leading to structural changes, tissue scarring, and impaired renal function. Glomerulosclerosis is commonly associated with a variety of kidney conditions, including chronic kidney disease.

[0071] As used herein, the term "capillary sparsity" refers to a pathological condition or pathophysiological process characterized by a reduction in the density of microvessels within a specific capillary network of a given tissue or organ. This phenomenon involves a quantifiable decrease in the number of capillaries, leading to impaired microcirculation perfusion, tissue hypoxia, and physiological dysfunction. Capillary sparsity can manifest in a variety of biological contexts and is associated with pathological states involving structural alterations in microvascular structure, thereby adversely affecting tissue perfusion and homeostasis.

[0072] As used herein, the term "renal tubular atrophy" refers to a pathological condition or pathophysiological process characterized by degeneration and necrosis of the renal tubular epithelial cells within the nephron. This process involves the loss of integrity and functional capacity of the renal tubular cells and is typically observed in the context of chronic kidney disease and various renal pathologies. Renal tubular atrophy significantly leads to the destruction of normal renal histological structure, impairs renal tubular integrity, and is an indicator of the progression of kidney damage.

[0073] As used herein, the term "renal tubular injury" refers to a pathological condition or pathophysiological process characterized by detrimental alterations to the structural and functional integrity of the renal tubules within the nephron. This type of injury encompasses morphological changes in renal tubular epithelial cells, impaired cellular integrity, and functional disturbances. Renal tubular injury is a marker of various renal pathological conditions, including acute kidney injury (AKI) and chronic kidney disease (CKD).

[0074] As used in this article, "oxidative stress" refers to the imbalance between the production and manifestation of excessive levels of molecular oxygen or reactive oxygen species (ROS) and the biological system's ability to rapidly detoxify reactive intermediates or repair the resulting damage via endogenous antioxidant systems. Disruptions to the normal redox state of tissues can cause toxic effects by producing peroxides and free radicals that damage all components of cells, including proteins, lipids, and DNA, through mechanisms including lipid peroxidation, protein oxidation and aggregation, and DNA damage. Some reactive oxidants can even act as messengers through a phenomenon known as redox signaling.

[0075] As used in this article, "hypoxia" refers to a pathological condition or pathophysiological process in which the kidney or a region of the kidney lacks adequate oxygen supply at the tissue level. Such damage may result from insufficient renal vascular perfusion, loss of capillary density, or altered capillary function. Hypoxia is a hallmark of various renal pathologies, including chronic kidney disease (CKD).

[0076] As used herein, the terms “treatment” or “treating” are methods used to obtain beneficial or desired outcomes, including clinical outcomes. Beneficial or desired clinical outcomes may include, but are not limited to: reduction or improvement of one or more symptoms or conditions; reduction of the severity of the disease or condition; stabilization of the disease or condition state (i.e., no worsening); prevention of the disease or condition; delay or slowing of the progression of the disease or condition; improvement or remission of the disease state; and remission (whether partial or complete), whether detectable or undetectable.

[0077] As used herein, the term "chronic kidney disease" (CKD) refers to kidney damage, including but not limited to tubulointerstitial fibrosis, glomerulosclerosis, and / or an estimated glomerular filtration rate (eGFR) of less than 60 ml / min per 1.73 square meters, lasting for 90 days or longer. It is a state of progressive loss of kidney function that ultimately leads to the need for renal replacement therapy (dialysis or transplantation). CKD has its general meaning in this field and is used to classify a wide range of conditions affecting the kidneys, damage to the renal parenchyma, and loss of functional nephrons or glomeruli. Examples of causes of CKD include, but are not limited to, cardiovascular disease, hypertension, diabetes, glomerulonephritis, polycystic kidney disease, and kidney graft rejection.

[0078] As used in this article, the term "coagulation disorder" refers to a pathological condition or pathophysiological process in which the blood clotting system is impaired, which may lead to the formation of blood clots / thrombi.

[0079] Anti-ANGPT2 antibody

[0080] This invention relates to antibodies or antigen-binding fragments that bind to ANGPT2 and, via ANGPT2, to the TIE2 receptor for the treatment or prevention of fibrotic diseases. In some embodiments, this invention relates to antibodies or antigen-binding fragments that bind to ANGPT2 and, via ANGPT2, to the TIE2 receptor for the treatment or prevention of kidney disease, condition, or injury in a subject, wherein the kidney disease, condition, or injury is associated with kidney fibrosis.

[0081] In one embodiment, the antibody or its antigen-binding fragment is characterized by binding to sites from position 336 to position 434 of angiopoietin-2 (ANGPT2) (SEQ ID NO: 30). In another embodiment, the binding site is about 2 to about 20 consecutive amino acids long, such as about 2 to about 18, 5 to about 20, or about 10 to about 20, such as about 15 to 20.

[0082] In one embodiment, the antibody or its antigen-binding fragment is characterized by binding to the epitope of human angiopoietin-2 (ANGPT2) at SEQ ID NO: 9 or SEQ ID NO: 30.

[0083] In one embodiment, the antibody or its antigen-binding fragment comprises:

[0084] a) A complementarity-determining region (CDR) of the heavy chain variable region, comprising: HCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0085] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0086] In one embodiment, the antibody or its antigen-binding fragment comprises:

[0087] a) The complementarity-determining region (CDR) of the heavy chain variable region, comprising: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0088] b) The complementarity-determining region (CDR) of the light chain variable region, comprising: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3; or the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0089] In one embodiment, the antibody or its antigen-binding fragment comprises:

[0090] The variable heavy chains of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 31 or SEQ ID NO: 33, and the variable light chains of SEQ ID NO: 7, SEQ ID NO: 11, SEQ ID NO: 32, SEQ ID NO: 34 or SEQ ID NO: 38.

[0091] In one embodiment, the antibody or its antigen-binding fragment comprises:

[0092] a) The complementarity-determining region (CDR) of the heavy chain variable region, comprising: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; and

[0093] b) The CDR of the light chain variable region, comprising: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3.

[0094] In one embodiment, the antibody or its antigen-binding fragment comprises a variable heavy chain of SEQ ID NO: 8 or SEQ ID NO: 10 and a variable light chain of SEQ ID NO: 7 or SEQ ID NO: 11.

[0095] In one embodiment, the antibody or its antigen-binding fragment is characterized by binding to the epitope of human angiopoietin-2 (ANGPT2) SEQ ID NO: 9.

[0096] In one embodiment, the antibody or its antigen-binding fragment comprises:

[0097] a) The complementarity-determining region (CDR) of the heavy chain variable region, comprising: the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0098] b) The complementarity-determining region (CDR) of the light chain variable region, which comprises the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0099] In one embodiment, the antibody or its antigen-binding fragment comprises a variable heavy chain of SEQ ID NO: 31 or SEQ ID NO: 33 and a variable light chain of SEQ ID NO: 32 or SEQ ID NO: 34 or SEQ ID NO: 38.

[0100] In one embodiment, the antibody or its antigen-binding fragment is characterized by binding to the epitope of human angiopoietin-2 (ANGPT2) at SEQ ID NO: 30.

[0101] The amino acid sequences of the antibodies or antigen-binding fragments disclosed herein may be conservedly substituted. "Conservative substitution" refers to a modification of a polypeptide, including replacing at least one amino acid with an amino acid whose biochemical properties are similar to those of the corresponding polypeptide, without resulting in loss of biological or biochemical function. "Conservative amino acid substitution" refers to a substitution in which an amino acid residue is replaced by an amino acid residue having a similar side chain. Categories of amino acid residues having similar side chains are defined in the art. These categories include amino acids with basic side chains (e.g., lysine, arginine, histidine), amino acids with acidic side chains (e.g., aspartic acid, glutamic acid), amino acids with uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), amino acids with nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), amino acids with β-branched side chains (e.g., threonine, valine, isoleucine), and amino acids with aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). It is anticipated that the antibodies of the present invention will retain their activity while having conserved amino acid substitutions.

[0102] In this invention, the antibody or its antigen-binding fragment is characterized by comprising a complementarity-determining region (CDR) of an antibody generated from: a cell line deposited on January 30, 2018, with accession number KCLRF-BP-00417 at the Korean Cell Bank (KCLB) of the Cancer Institute, Seoul National University College of Medicine, Korea (address: 28 Yongon-dong, Chongno-Gu, Seoul, 110-744, Korea); or a hybridoma deposited on April 23, 2013, with accession number KCLRF-BP-00295 at the Korean Cell Bank (address: Yongon-dong, Chongno-gu, Seoul, South Korea).

[0103] The sequences of the antibodies or antigen-binding fragments thereof of the present invention may differ from the sequences provided in this application. For example, the amino acid sequences may differ from those described above, wherein: (a) the variable region may be separated from the constant domain of the light chain; (b) the amino acids may differ from those described above without significantly affecting the chemical properties of the residues (so-called conserved substitution); and (c) the amino acids may differ from those described above by a given percentage of homology, for example, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% homology. Alternatively, the nucleic acid encoding the antibody may: (a) be separated from the constant domain of the light chain; (b) differ from the nucleic acid described above without altering the residues encoded therein; or (c) differ from the nucleic acid described above by a given percentage of homology, for example, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% homology.

[0104] When making conserved alterations to amino acid sequences, the hydrophilicity index of the amino acids can be considered. The importance of the hydrophilicity index of amino acids in conferring biological functions of protein-protein interactions is generally known in the art. It is accepted that the relative hydrophilicity of amino acids contributes to the secondary structure of the resulting protein, which in turn defines the protein's interactions with other molecules, such as enzymes, substrates, receptors, DNA, antibodies, antigens, etc.

[0105] It is also understood in the art that similar amino acids can be substituted efficiently based on hydrophilicity. For example, the maximum local average hydrophilicity of a protein (determined by the hydrophilicity of its adjacent amino acids) is related to the protein's biological properties. It should be understood that an amino acid can be substituted by another amino acid having similar hydrophilicity, producing a biologically or immunologically modified protein. In such modifications, amino acid substitutions with hydrophilicity values ​​within + / -2 are preferred, those within + / -1 are particularly preferred, and those within + / -0.5 are even more particularly preferred.

[0106] As outlined above, amino acid substitutions are typically based on the relative similarity of the substituents in the amino acid side chains, such as their hydrophobicity, hydrophilicity, charge, size, etc. Exemplary substitutions taking into account the aforementioned properties are well known to those skilled in the art and include arginine and lysine; glutamic acid and aspartic acid; serine and threonine; glutamine and asparagine; and valine, leucine, and isoleucine.

[0107] In this disclosure, the antibody or its antigen-binding fragment can bind to human and / or mouse ANGPT2. In one embodiment, the antibody or its antigen-binding fragment binds to human ANGPT2. In one embodiment, the antibody or its antigen-binding fragment binds to mouse ANGPT2.

[0108] In one embodiment, the antibody or its antigen-binding fragment is polyclonal or monoclonal. In one embodiment, the antigen-binding fragment is scFv or Fab.

[0109] In this disclosure, the antibody or its antigen-binding fragment may be a humanized antibody or a human antibody.

[0110] A complete antibody has a structure comprising two full-length light chains and two full-length heavy chains, wherein each light chain is linked to a corresponding heavy chain via disulfide bonds. A complete antibody may include IgA, IgD, IgE, IgM, and / or IgG, where IgG is a subtype and may include IgG1, IgG2, IgG3, and / or IgG4. In one embodiment, the antibody is the human subtype IgG1.

[0111] The heavy chain constant region has γ, μ, α, δ, and ε types, and is subclassed into γ1, γ2, γ3, γ4, α1, and α2. The light chain constant region has κ and λ types. The basic tetrachain antibody unit is a heterotetrameric glycoprotein composed of two identical light chains (L) and two identical heavy chains (H). The light chain has a variable region (VL) at the N-terminus and a constant region at the other end. The heavy chain has a variable region (VH) at the N-terminus, three constant regions (CH) for the α and γ chains, and four CH regions for the μ and ε isotypes. The term "variable" means that specific portions of the variable region differ significantly in sequence between antibodies. The V region mediates antigen binding and defines the specificity of a particular antibody for its specific antigen. In both the light and heavy chain variable regions, variability is concentrated in three segments called "hypervariable regions (HVR)," or CDRs. The more highly conserved portions of the variable regions are called "frame regions (FR)." The variable regions of the heavy chain and the variable regions of the light chain have FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4 structures from the N end to the C end.

[0112] As used herein, the term “heavy chain” encompasses both the full-length heavy chain and its fragments, the full-length heavy chain containing a variable domain (VH) and three constant domains (CH1, CH2, and CH3), the variable domain containing an amino acid sequence having a variable region sequence sufficient to confer antigen specificity.

[0113] As used herein, the term “light chain” encompasses both the full-length light chain and its fragments, which contain a variable domain (VL) and a constant domain (CL), the variable domain containing an amino acid sequence having a variable region sequence sufficient to confer antigen specificity.

[0114] ANGPT2-TIE2 signal transduction

[0115] ANGPT2 is known to also act as an agonist to induce TIE2 receptor activation in several processes, including lymphangiogenesis and maintenance, and is therefore thought to perform various functions depending on the context. Pathological expression of ANGPT2 disrupts these processes.

[0116] This invention provides antibodies or antigen-binding fragments thereof that bind to ANGPT2. In some embodiments, the antibodies or antigen-binding fragments thereof exhibit a dual function of binding to and neutralizing ANGPT2 and activating the TIE2 receptor. In fact, in some embodiments, the antibodies or antigen-binding fragments thereof:

[0117] a) Reduce or inhibit human angiopoietin-2 (ANGPT2); and / or

[0118] b) Inducing TIE2 activation; and

[0119] c) Inducing TIE2 phosphorylation.

[0120] In some implementations, the antibody or antigen-binding fragment is present in the diseased kidney:

[0121] a. Decrease or inhibit human angiopoietin-2 (ANGPT2); and / or

[0122] b. Inducing TIE2 activation; and

[0123] c. Induces TIE2 phosphorylation.

[0124] In some implementations, the antibody or antigen-binding fragment is present in the fibrotic tissue:

[0125] a. Decrease or inhibit human angiopoietin-2 (ANGPT2); and / or

[0126] b. Inducing TIE2 activation; and

[0127] c. Induces TIE2 phosphorylation.

[0128] In one embodiment, the induction of TIE2 activation depends on ANGPT2. In one embodiment, the antibody or its antigen-binding fragment normalizes pathological vascularity and / or prevents the progression of fibrosis.

[0129] Another advantage of this invention is that the antibodies described herein that bind to ANGPT2 and activate TIE2 depend on the presence of ANGPT2. Therefore, TIE2 is activated only at sites where ANGPT2 levels are elevated. Consequently, the antibodies described herein will only activate TIE2 in diseased and / or damaged tissues, thereby reducing off-target effects.

[0130] ANGPT2-TIE2 signaling plays a role in various pathophysiological processes associated with fibrosis. ANGPT2-TIE2 signaling also plays a role in various renal pathophysiological processes.

[0131] In one embodiment, an antibody or its antigen-binding fragment prevents or mitigates inadequate perfusion (e.g., renal perfusion associated with a fibrotic disease, such as kidney disease, condition, or injury).

[0132] In one embodiment, the antibody or its antigen-binding fragment prevents or mitigates capillary sparsity associated with fibrotic diseases, such as kidney disease, condition, or injury. In one embodiment, the prevention or mitigation of capillary sparsity is evaluated via the quantification of endothelial markers such as endothelial mucin or podocyte marker proteins.

[0133] In one implementation, the antibody or its antigen-binding fragment prevents or reduces the reduction of peritubular capillary fenestrations associated with kidney disease, condition, or injury.

[0134] In one embodiment, an antibody or its antigen-binding fragment prevents or mitigates fibrosis, such as fibrosis associated with kidney disease, condition, or injury. In some embodiments, fibrosis is tubulointerstitial fibrosis and / or glomerular sclerosis. In some embodiments, prevention or mitigation of fibrosis is evaluated via the quantification of fibrosis markers such as α-smooth muscle actin (aSMA), vimentin, type I collagen (Col1), or platelet-derived growth factor receptor-β (PDGFR-β).

[0135] In one embodiment, the antibody or its antigen-binding fragment described herein is used in a kidney disease, condition, or injury induced, spread, and / or associated with an increased extracellular matrix, such as α-smooth muscle actin (aSMA), vimentin, type I collagen (Col1), or platelet-derived growth factor receptor-β (PDGFR-β).

[0136] Pharmaceutical Composition

[0137] As used herein, the term "pharmaceutical composition" refers to a combination of an active agent (e.g., the composition of the present invention) with an inert or active carrier, making the composition particularly suitable for in vivo or in vitro diagnostic or therapeutic use. The term "pharmaceutical composition" can also refer to a formulation containing the composition of the present invention and / or an agent for treating or preventing fibrotic diseases (e.g., kidney disease, condition, or injury) in a subject, wherein the kidney disease, condition, or injury is associated with kidney fibrosis, and the formulation is in a form suitable for administration to a subject. In another embodiment, the present invention relates to a pharmaceutical composition comprising an antibody or an antigen-binding fragment thereof as an active ingredient. The pharmaceutical composition is characterized by containing a pharmaceutically effective amount of an antibody or an antigen-binding fragment thereof according to the present invention, and a pharmaceutically acceptable carrier and / or excipient.

[0138] In one embodiment, the present invention relates to a pharmaceutical composition comprising an antibody or an antigen-binding fragment thereof according to any one of the foregoing embodiments for treating or preventing fibrotic diseases. In some embodiments, the disease is a kidney disease, wherein the kidney disease, condition, or injury is associated with kidney fibrosis.

[0139] The pharmaceutical composition may also contain a small molecule inhibitor or a vascular endothelial growth factor (VEGF) antagonist used in chemotherapy. The VEGF antagonist may be an anti-VEGF antibody, a VEGF inhibitory fusion protein, or a small molecule kinase inhibitor. It may also prove effective, particularly by combining the antibody of this disclosure or its antigen-binding fragment with other therapies targeting different aspects of ANGPT2 function. In another embodiment, the antibody of this disclosure or its antigen-binding fragment may be conjugated with at least one reagent to form an antibody conjugate to increase the efficacy of the antibody molecule as a diagnostic or therapeutic agent.

[0140] This invention includes antibodies and their antigen-binding fragments, compositions, and methods for treating or preventing fibrotic diseases in subjects. This invention also includes antibodies and their antigen-binding fragments, compositions, and methods for treating or preventing kidney diseases, conditions, or injuries in subjects. Administration of the antibodies and their antigen-binding fragments and / or compositions according to the invention will generally be performed via any common route. This includes, but is not limited to, parenteral, orthotopic, intradermal, subcutaneous, intramuscular, intraperitoneal, intranasal, or intravenous injection. Additional formulations suitable for other routes of administration include oral formulations. Oral formulations include such commonly used excipients, such as pharmaceutical-grade mannitol, lactose, starch, magnesium stearate, sodium saccharin, cellulose, magnesium carbonate, etc. These compositions are in the form of solutions, suspensions, tablets, pills, capsules, sustained-release formulations, or powders, and contain about 10% to about 95%, preferably about 25% to about 70% of the active ingredient.

[0141] Typically, the compositions of the present invention are administered in a manner compatible with dosage forms and at a therapeutically effective amount. The amount to be administered depends on the subject to be treated. The precise amount of active ingredient to be administered depends on the judgment of the practitioner.

[0142] The route of administration can vary widely. Any conventional method for administering antibodies is applicable. These methods are believed to include oral administration on a solid, physiologically acceptable matrix or in a physiologically acceptable dispersion, parenteral administration, administration by injection, etc. The dosage of the pharmaceutical composition will depend on the route of administration and will vary according to the subject's body size and health. Organ function assays, such as renal function assays and / or assessments of fibrotic disease, condition, or injury or its status (e.g., kidney disease, condition, or injury or its status), may be performed after administration, including but not limited to measurements of relevant biomarkers, tissue capillary density, and structural integrity of affected tissues, as well as evaluation of associated complications (e.g., cardiomyopathy, coagulation disorders, and retinopathy).

[0143] The phrase "pharmaceuticalally acceptable" or "pharmacologically acceptable" means a molecular entity or component that does not produce adverse, allergic, or other adverse reactions when administered to animals or humans. As used herein, "pharmaceuticalally acceptable excipients" include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic agents, and absorption-delaying agents. The use of such media and reagents for pharmaceutically active substances is well known in the art. Unless any conventional media or reagent is incompatible with the active ingredient, it is contemplated for use in therapeutic compositions.

[0144] This composition can be formulated in a neutral or salt form. Pharmaceutically acceptable salts include acid addition salts (formed with the free amino group of a protein) that are formed with an inorganic acid (such as, for example, hydrochloric acid or phosphoric acid) or an organic acid (such as acetic acid, oxalic acid, tartaric acid, mandelic acid). Salts formed with a free carboxyl group can also be derived from inorganic bases (such as, for example, sodium hydroxide, potassium hydroxide, ammonium hydroxide, calcium hydroxide, or iron hydroxide) and organic bases (such as isopropylamine, trimethylamine, histidine, procaine, etc.).

[0145] The carrier can also be a solvent or dispersion medium containing, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, liquid polyethylene glycol, etc.), suitable mixtures thereof, and vegetable oils. Microbial activity can be prevented by various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, sorbic acid, thimerosal, etc.

[0146] The effective amount of a therapeutic or prophylactic composition is determined based on the intended target. The term "unit dose" or "dose" refers to a physically discrete unit suitable for use in a subject, each unit containing a predetermined amount of the composition calculated to produce the desired response discussed above regarding its administration, i.e., appropriate routes and protocols. The amount to be administered, depending on both the number of treatments and the unit dose, depends on the desired outcome and / or protection. The precise amount of the composition also depends on the practitioner's judgment and varies from individual to individual. Factors influencing the dosage include the subject's physical and clinical condition, the route of administration, the intended target of treatment (symptom relief rather than cure), and the potency, stability, and toxicity of the particular composition. After formulation, the solution is administered in a manner compatible with the dosage form and in a therapeutically or prophylactically effective amount. The formulation is readily available for administration in a variety of dosage forms.

[0147] Indications / Treatment Plan

[0148] In one embodiment, the antibody or its antigen-binding fragment is administered prior to the onset of fibrotic disease, condition, or injury. In another embodiment, the antibody or its antigen-binding fragment is administered during the onset of fibrotic disease, condition, or injury. In yet another embodiment, the antibody or its antigen-binding fragment is administered after the onset of fibrotic disease, condition, or injury.

[0149] In one embodiment, the antibody or its antigen-binding fragment is administered prior to the onset of kidney disease, condition, or injury. In another embodiment, the antibody or its antigen-binding fragment is administered during the onset of kidney disease, condition, or injury. In yet another embodiment, the antibody or its antigen-binding fragment is administered after the onset of kidney disease, condition, or injury.

[0150] In one embodiment, the kidney disease, condition, or injury is induced by, spread by, and / or associated with endothelial dysfunction. In another embodiment, the kidney disease, condition, or injury is induced by, spread by, and / or associated with renal fibrosis.

[0151] In some implementations, the fibrotic disease is selected from the following: renal fibrosis and myocardial fibrosis.

[0152] In some embodiments, renal fibrosis is tubulointerstitial fibrosis. In some embodiments, renal fibrosis is glomerular sclerosis. In one embodiment, the kidney disease, condition, or injury is induced by, spread by, and / or associated with renal capillary spallation. In one embodiment, the kidney disease, condition, or injury is induced by, spread by, and / or associated with renal tubular atrophy. In one embodiment, the kidney disease, condition, or injury is induced by, spread by, and / or associated with renal tubular injury. In one embodiment, the kidney disease, condition, or injury is induced by, spread by, and / or associated with renal tubular injury. In one embodiment, the kidney disease, condition, or injury is induced by, spread by, and / or associated with oxidative stress.

[0153] In one embodiment, the kidney disease, condition, or injury is acute kidney injury (AKI). In some embodiments, the kidney disease, condition, or injury is acute kidney disease (AKD). In some embodiments, the kidney disease, condition, or injury is chronic kidney disease (CKD). In some embodiments, CKD is induced by, spread by, and / or associated with a coagulation disorder.

[0154] Kidney disease can be divided into acute kidney injury (AKI) and chronic kidney disease (CKD). AKI is a sudden and usually reversible decline in kidney function that occurs within hours to days. It is usually triggered by conditions such as dehydration, infection (sepsis), drug toxicity, or urinary tract obstruction. AKI is characterized by a rapid increase in blood urea nitrogen (BUN) and creatinine levels, and may present with symptoms such as decreased urine output, swelling, and fatigue. Sepsis is the leading cause of AKI. Early intervention can usually restore normal kidney function.

[0155] Chronic kidney disease (CKD) is a progressive and irreversible loss of kidney function that develops over months to years. CKD is often associated with long-term conditions such as diabetes, hypertension, or autoimmune diseases. It progresses in stages based on glomerular filtration rate (GFR) and can lead to complications such as anemia, bone disease, and cardiovascular problems. Symptoms are often subtle in the early stages but may include persistent fatigue, fluid retention, and altered urination patterns. Histologically, CKD is associated with tubulointerstitial fibrosis, while fibrosis is not seen in AKI. As noted above, in some implementations, kidney disease is CKD associated with tubulointerstitial fibrosis. In some implementations, kidney disease is CKD associated with glomerular sclerosis.

[0156] The main differences lie in duration and reversibility: AKD is sudden and potentially reversible, while CKD is progressive and permanent. Both conditions require timely medical attention, but they differ in their management, prognosis, and long-term health impact.

[0157] As demonstrated by the data in this paper, ABTAA exhibits a protective effect in two preclinical models of chronic kidney disease: one model with fibrosis as the clinical endpoint, and the other a type 2 diabetes model with renal and cardiac complications. The effect of ABTAA on fibrosis is novel and unexpected.

[0158] Chronic kidney disease encompasses several clinical kidney diseases as defined by the European Society of Nephrology, all of which are associated with fibrotic / connective tissue components (fibrosis) in the renal tubules or glomeruli (Venkat-Raman et al., NephrolDial Transpl, 2012).

[0159] In one implementation, the kidney disease, condition, or injury is selected from the group comprising: chronic kidney damage, chronic renal failure syndrome, glomerulonephritis, type I diabetes, type II diabetes, ischemic nephropathy, focal segmental glomerulosclerosis, nephrotic syndrome, cardiorenal syndrome, tubulointerstitial disease associated with metabolic disorders, minimal change disease, hypertensive nephropathy, malignant hypertensive nephropathy, IgA nephropathy, IgM nephropathy, mesangial capillary glomerulonephritis, crescentic glomerulonephritis, diffuse intracapillary proliferative glomerulonephritis, mesangial proliferative glomerulonephritis, focal and segmental proliferative glomerulonephritis, systemic vasculitis, pulmonary hemorrhage nephritis syndrome, Dent disease, systemic lupus erythematosus glomerulonephritis syndrome, allergic purpura, systemic sclerosis, scleroderma, nephropathy associated with urinary tract obstruction, tubulointerstitial nephritis, interstitial nephritis, and lithium nephropathy.

[0160] In one implementation, renal fibrosis is induced by, spread by, and / or associated with: chronic kidney damage, chronic renal failure syndrome, glomerulonephritis, type I diabetes, type II diabetes, ischemic nephropathy, focal segmental glomerulosclerosis, nephrotic syndrome, cardiorenal syndrome, tubulointerstitial disease associated with metabolic disorders, minimal change disease, hypertensive nephropathy, malignant hypertensive nephropathy, and IgA nephropathy. Diseases, IgM nephropathy, mesangial capillary glomerulonephritis, crescentic glomerulonephritis, diffuse intracapillary proliferative glomerulonephritis, mesangial proliferative glomerulonephritis, focal and segmental proliferative glomerulonephritis, systemic vasculitis, pulmonary hemorrhage nephritis syndrome, Dent disease, systemic lupus erythematosus glomerulonephritis syndrome, allergic purpura, systemic sclerosis, scleroderma, nephropathy associated with urinary tract obstruction, tubulointerstitial nephritis, interstitial nephritis and / or lithium nephropathy.

[0161] In some implementations, fibrotic diseases (e.g., kidney disease, condition, or injury) are induced by, spread by, and / or associated with type 2 diabetes.

[0162] In some implementations, the subjects had type 2 diabetes. In some implementations, the fibrotic disease was caused by type 2 diabetes.

[0163] In some implementations, ABTAA treatment does not affect the subject's obesity and diabetes status, demonstrating its potential for combination therapy with other drugs targeting obesity and blood glucose levels.

[0164] In some embodiments, the present invention provides an antibody or antigen-binding fragment thereof that binds to ANGPT2 and binds to the TIE2 receptor via ANGPT2 for the treatment or prevention of diabetic complications.

[0165] In some embodiments, the fibrotic disease is myocardial fibrosis. In some embodiments, myocardial fibrosis is induced by, spread from, and / or associated with myocardial infarction, hypertensive heart disease, or cardiomyopathy. In some embodiments, the fibrotic disease is cardiomyopathy. In some embodiments, the cardiomyopathy is hypertrophic cardiomyopathy. In some embodiments, the cardiomyopathy is idiopathic dilated cardiomyopathy.

[0166] In some embodiments, the present invention provides an antibody or an antigen-binding fragment thereof that binds to ANGPT2 and, via ANGPT2, to the TIE2 receptor for the treatment or prevention of coagulation disorders. In some embodiments, the coagulation disorder is caused by or associated with fibrotic disease.

[0167] In some implementations, the methods disclosed in the section entitled “Indications / Treatment” may also include administering the pharmaceutical composition disclosed in the section entitled “Pharmaceutical Compositions” to a subject in need.

[0168] In the context of this invention, the term "treatment or prevention" includes not only the prevention and / or treatment of a disease, but also generally includes preventing the onset of a disease, slowing or reversing the progression of a disease, preventing or slowing the onset of one or more symptoms associated with a disease, reducing and / or alleviating one or more symptoms associated with a disease, reducing the severity and / or duration of a disease and / or any symptoms associated with it, and / or preventing a further increase in the severity of a disease and / or any symptoms associated with it, preventing, reducing or reversing any physiological damage caused by a disease, and any pharmacological effects that are generally beneficial to a patient who is receiving treatment.

[0169] Subjects to be treated can be any warm-blooded animal, but particularly mammals, and even more particularly humans. As will be apparent to those skilled in the art, subjects to be treated will particularly be persons suffering from or at risk of the diseases and conditions mentioned herein.

[0170] Referring to the sections titled "Anti-ANGPT2 Antibody" and "ANGPT2-TIE2 Signaling," in one aspect, the present invention relates to a method for treating fibrotic diseases, conditions, or injuries, comprising administering a therapeutically effective amount of an antibody or antigen-binding fragment. In another aspect, the present invention relates to a method for treating kidney diseases, conditions, or injuries associated with kidney fibrosis, comprising administering a therapeutically effective amount of an antibody or antigen-binding fragment.

[0171] In one aspect, the present invention relates to the preparation of medicaments for treating fibrotic diseases. In another aspect, the present invention relates to the preparation of medicaments for treating kidney diseases, conditions, or injuries, wherein the kidney disease, condition, or injury is associated with kidney fibrosis.

[0172] As used herein, "therapeutic effective amount" means an amount sufficient to treat a disease with a reasonable benefit / risk ratio suitable for medical treatment, and in the case of a composition containing an anti-ANGPT2 antibody. The precise amount can vary depending on a variety of factors, including, but not limited to, the components and physical properties of the therapeutic composition, the intended patient population, individual patient considerations, etc., and can be readily determined by those skilled in the art. When these factors are fully considered, it is important to administer the minimum amount sufficient to achieve the maximum effect without side effects, and this dosage can be readily determined by those skilled in the art.

[0173] The dosage of the pharmaceutical composition of the present invention is not particularly limited, but varies according to various factors including the patient's health status and weight, the severity of the disease, and the type of drug, route of administration, and time of administration. The composition can be administered via routes commonly permitted in mammals (including rats, mice, cattle, humans, etc.), such as oral, rectal, intravenous, subcutaneous, intrarenal, intrauterine, or intracerebrovascular administration in a single dose daily or every other day or weekly.

[0174] On the other hand, the present invention relates to the following method:

[0175] a) Reduce or inhibit human angiopoietin-2 (ANGPT2) signaling in diseased kidneys, and / or induce TIE2 activation, and / or induce TIE2 phosphorylation in fibrotic tissue (e.g., diseased kidneys), wherein the diseased kidneys are fibrotic kidneys;

[0176] b) To normalize pathological blood vessels and treat and reduce fibrosis in tissues (e.g., diseased kidneys) and prevent the progression of fibrosis in tissues (e.g., diseased kidneys);

[0177] c) Treat and / or reduce fibrosis in tissues (e.g., diseased kidneys) and / or prevent the progression of fibrosis in tissues (e.g., diseased kidneys);

[0178] d) Prevent or treat subjects with inadequate perfusion, such as when the perfusion is renal perfusion, the disease is a kidney disease, condition or injury, and is associated with renal fibrosis;

[0179] e) Prevent or reduce capillary sparsity associated with a fibrotic disease (e.g., kidney disease), condition, or injury in the subject, wherein the kidney disease, condition, or injury is associated with fibrosis; and / or

[0180] f) Prevent or treat a reduction in peritubular capillary fenestrations associated with kidney disease, condition or injury in the subject, where the kidney disease, condition or injury is associated with renal fibrosis.

[0181] This includes administering an antibody or its antigen-binding fragment to a subject in need, wherein the antibody or its antigen-binding fragment comprises:

[0182] a) A complementarity-determining region (CDR) of the heavy chain variable region, comprising: HCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0183] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0184] On the other hand, the present invention relates to the following method:

[0185] a) Reduce or inhibit human angiopoietin-2 (ANGPT2) signaling in tissues (e.g., diseased kidneys), and / or induce TIE2 activation and / or induce TIE2 phosphorylation, wherein the diseased kidney is a fibrotic kidney;

[0186] b) To normalize pathological blood vessels and treat and reduce fibrosis in tissues (e.g., diseased kidneys) and prevent the progression of fibrosis in tissues (e.g., diseased kidneys);

[0187] c) Treat and / or reduce fibrosis in tissues (e.g., diseased kidneys) and / or prevent the progression of fibrosis in tissues (e.g., diseased kidneys);

[0188] d) Prevent or treat subjects with inadequate perfusion, such as when the perfusion is renal perfusion, the disease is a kidney disease, condition or injury, and is associated with renal fibrosis;

[0189] e) Prevent or reduce capillary sparsity associated with a fibrotic disease (e.g., kidney disease), condition, or injury in the subject, wherein the kidney disease, condition, or injury is associated with fibrosis; and / or

[0190] f) Prevent or treat a reduction in peritubular capillary fenestrations associated with kidney disease, condition or injury in the subject, where the kidney disease, condition or injury is associated with renal fibrosis.

[0191] This includes administering an antibody or its antigen-binding fragment to a subject in need, wherein the antibody or its antigen-binding fragment comprises:

[0192] a) The complementarity-determining region (CDR) of the heavy chain variable region, comprising: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0193] b) The complementarity-determining region (CDR) of the light chain variable region, comprising: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3; or the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0194] And / or

[0195] The variable heavy chains of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 31 or SEQ ID NO: 33, and the variable light chains of SEQ ID NO: 7, SEQ ID NO: 11, SEQ ID NO: 32, SEQ ID NO: 34 or SEQ ID NO: 38,

[0196] And / or

[0197] a) The complementarity-determining region (CDR) of the heavy chain variable region, comprising: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; and

[0198] b) The CDR of the light chain variable region, comprising: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3.

[0199] And / or

[0200] The variable heavy chain of SEQ ID NO: 8 or SEQ ID NO: 10 and the variable light chain of SEQ ID NO: 7 or SEQ ID NO: 11,

[0201] And / or

[0202] The antibody or its antigen-binding fragment is characterized by binding to the epitope of human angiopoietin-2 (ANGPT2) SEQ ID NO: 9.

[0203] And / or

[0204] The antibody or its antigen-binding fragment comprises:

[0205] a) The complementarity-determining region (CDR) of the heavy chain variable region, comprising: the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0206] b) The complementarity-determining region (CDR) of the light chain variable region, comprising the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0207] And / or

[0208] The antibody or its antigen-binding fragment comprises a variable heavy chain of SEQ ID NO: 31 or SEQ ID NO: 33 and a variable light chain of SEQ ID NO: 32 or SEQ ID NO: 34 or SEQ ID NO: 38.

[0209] And / or

[0210] The antibody or its antigen-binding fragment is characterized by binding to the epitope of human angiopoietin-2 (ANGPT2) SEQ ID NO: 30.

[0211] Regarding the methods described above, in some embodiments, the induction of TIE2 activation is ANGPT2-dependent. In some embodiments, prevention or mitigation of capillary sparsity is evaluated by quantification of endothelial markers such as endothelial mucin or podocyte marker proteins. In some embodiments, fibrosis is tubulointerstitial fibrosis. In some embodiments, fibrosis is glomerular sclerosis. In other embodiments, prevention or mitigation of fibrosis is evaluated by quantification of fibrosis markers such as α-smooth muscle actin (aSMA), vimentin, type I collagen (Col1), or platelet-derived growth factor receptor-β (PDGFR-β).

[0212] Production methods

[0213] On the other hand, the present invention relates to nucleic acids encoding antibodies or antigen-binding fragments thereof. In some embodiments, the nucleic acid may be present in cells, cell lysates, or may also be present in a partially purified or substantially pure form. When the nucleic acid is purified from other cellular components or other contaminants (e.g., other cellular nucleic acids or proteins) using standard techniques, including alkaline / SDS treatment, CsCl banding, column chromatography, agarose gel electrophoresis, and other techniques well known in the art, the nucleic acid is "isolated" or "substantially pure". The nucleic acid of the present invention may be, for example, DNA or RNA, and may or may not include intron sequences.

[0214] In another embodiment, the present invention relates to recombinant expression vectors comprising nucleic acids. For expressing an antibody or a fragment thereof, DNA encoding light and heavy chains of partial or full length is obtained via standard molecular biology techniques (e.g., PCR amplification using a hybridoma expressing the target antibody or cDNA cloning), and the DNA can be "operably bound" to transcriptional and translational control sequences for insertion into the expression vector.

[0215] As used herein, the term "operably bound" indicates that the antibody gene is linked to a vector such that the transcriptional and translational control sequences in the vector have the intended function of controlling the transcription and translation of the antibody gene. The expression vector and expression control sequences are selected to be compatible with the host cells to be used for expression. The antibody light chain gene and antibody heavy chain gene are inserted into separate vectors, or both genes are inserted into the same expression vector. The antibody is inserted into the expression vector using standard methods (e.g., linking the antibody gene fragment to a complementary restriction enzyme site on the vector, or blunt-end ligation in the absence of restriction enzyme sites). In some cases, the recombinant expression vector may encode a signal peptide that promotes the secretion of the antibody chain from the host cell. The antibody chain gene may be cloned into the vector such that the signal peptide binds to the N-terminus of the antibody chain gene according to a framework. The signal peptide may be an immunoglobulin signal peptide or a heterologous signal peptide (i.e., a signal peptide derived from a protein other than an immunoglobulin). Furthermore, the recombinant expression vector contains regulatory sequences that regulate the expression of the antibody chain gene in the host cell. "Regulatory sequences" may include promoters, enhancers, and other expression control elements (e.g., polyadenylation signals) that control the transcription or translation of antibody chain genes. Those skilled in the art will recognize that the design of expression vectors can be varied by altering the regulatory sequences based on factors such as the selection of host cells to be transformed and the protein expression level.

[0216] In yet another embodiment, the present invention relates to cells transformed with a recombinant expression vector. The cells used to generate the antibodies or antigen-binding fragments thereof disclosed herein can be prokaryotes, yeast, or higher eukaryotic cells, but are not limited thereto.

[0217] In some implementations, strains of the genus Bacillus, such as Escherichia coli, Bacillus subtilis and Bacillus thuringiensis, Streptomyces, Pseudomonas, and prokaryotic host cells, such as Proteus mirabilis and Staphylococcus, may be used.

[0218] In some implementations, animal cells may be used. Examples include, but are not limited to, COS-7, BHK, CHO, CHOK1, DXB-11, DG-44, CHO / −DHFR, CV1, COS-7, HEK293, BHK, TM4, VERO, HELA, MDCK, BRL 3A, W138, Hep G2, SK-Hep, MMT, TRI, MRC 5, FS4, 3T3, RIN, A549, PC12, K562, PER.C6, SP2 / 0, NS-0, U20S, or HT1080.

[0219] Nucleic acid or vector is transfected into host cells. For "transfection," various commonly used techniques, such as electrophoresis, calcium phosphate precipitation, DEAE-dextran transfection, liposome transfection, etc., can be used to introduce exogenous nucleic acids (DNA or RNA) into prokaryotic or eukaryotic host cells. Considering applicability to mammalian cells, the antibodies according to the invention can be expressed in eukaryotic cells, preferably in mammalian host cells. As an example, suitable mammalian host cells for antibody expression may include Chinese hamster ovary (CHO) cells (e.g., dhfr-CHO cells used in conjunction with DHFR selection markers), NSO myeloma cells, COS cells, or SP2 cells, etc.

[0220] In another embodiment, the present invention relates to a method for producing an anti-ANGPT2 antibody or an antigen-binding fragment thereof, comprising culturing host cells and expressing the antibody or the antigen-binding fragment thereof. When a recombinant expression vector encoding an antibody gene is introduced into a mammalian host cell, the antibody can be produced by culturing the host cells for a sufficient period of time to allow the antibody to be expressed in the host cells, or more preferably, by culturing for a sufficient period of time to allow the antibody to be secreted into the culture medium in which the host cells are cultured.

[0221] In some implementations, the expressed antibody can be separated from the host cell and purified for homogeneity. Antibody separation or purification can be performed using methods commonly used for protein separation and purification, such as chromatography. Chromatography may include, for example, affinity chromatography, ion exchange chromatography, or hydrophobic chromatography, including protein A columns and protein G columns. In addition to chromatography, antibodies can be separated and purified by combining filtration, ultrafiltration, salting out, dialysis, etc.

[0222] Example

[0223] Materials and methods

[0224] Mice: The inducible mouse strains used in this example are summarized from Figure 1 In section B, the experimental mouse groups used in this embodiment are summarized in Table 1.

[0225] Table 1. Experimental mouse groups .

[0226]

[0227] Floxed Veptp and TIE2 mice were tamponade-induced Cdh5-Cre ERT2 Mice (Pitulescu et al., 2010) were crossbred to produce endothelial cell-specific knockouts of the gene (TIE2, etc.). iECKO and Veptp iECKO The TIE2 mice were also compared with reporter mice Ai14-TdTomato(Gt(ROSA)26Sor) tm14(CAG-tdTomato)Hze JAX strain 007914 (Madisen et al., 2010) were crossed to obtain pedigree markers for endothelial cells. Litter control mice were wt / wt mice with the TIE2 and Veptp alleles (WT). Floxed Pdgfb mice (Pdgfb...) were used as the primary control. tm2Cbet (Enge et al., 2002) and his moxifen-induced Actb-Cre ERT2 Mouse (Tg(CAG-cre / Esr1*)5Amc, JAX stock 004682) (Hayashi et al., 2002) hybridization was used to achieve systemic knockout of Pdgfb (Pdgfb iKO ). Litter control mice were Cre-negative Pdgfb wt / lox or lox / lox mice. At 4 weeks of age (TIE2) iECKO and Veptp iECKO ), or 1 week before the experiment (Pdgfb iKOKnockout was induced by oral gavage of three doses of tamoxifen (2 mg) in peanut oil. Mice were used as a mixed background. Genotyping of mice was performed using primers described in Table 2. Figure 1 A and Figure 11 At the time points indicated in A, mice were injected intraperitoneally with ABTAA (25 mg / kg body weight, in PBS). Control mice were injected with the same dose of human IgG Fc (AG714, Millipore, in PBS). Mice used for ABTAA treatment and for early gene regulation analysis were either internally bred C57BL6 / J mice or WT mice bred as described above. Adult mice (8–16 weeks old) were used in all experiments. CKD was induced by UUO, as previously described (Loganathan et al., 2018; Chevalier et al., 2009). Comparisons were made between the unobstructed contralateral (CL) kidney and the obstructed UUO kidney. Both female and male mice were used in all experiments because the UUO model did not show any sex differences (Yang et al., 2010).

[0228] Table 2: Genotyping Primers

[0229]

[0230] Kidney perfusion measured using contrast imaging ultrasound: Kidney perfusion was measured in isoflurane-anesthetized mice using a Vevo 2100 ultrasound system with an MS250 transducer and contrast imaging software (Visual Sonics, Fujifilm) on a heated platform. Measurements were taken continuously via an anal probe, and mouse body temperature was maintained between 36°C and 37°C. Imaging was performed from the mouse's back, and the ultrasound transducer was fixed in place using a mechanical positioning system. Standard B-mode imaging was used to optimize mouse positioning for simultaneous imaging of both kidneys. 100 µl of a microbubble-based contrast agent (VevoMicroMarker, VS-11913, Fujifilm Visual Sonics) from a vial resuspended in 2 ml saline was injected into the mice via the tail vein. Nonlinear contrast images were acquired using general imaging in a vascular bag at 21 GHz. Contrast intensity of the images was analyzed using Vevo Lab 3.2.6 (Fujifilm Visual Sonics) after manually marking regions of interest (ROIs), i.e., the cortex of both the CL and UUO kidneys. The perfusion of each mouse was normalized to the CL kidney. Five sham-operated WT mice were used as a comparison in the ultrasound images, showing the left kidney normalized to the right kidney.

[0231] Immunohistochemistry: After protein blocking with 0.25% Triton X100 (X0909, DAKO) for 2 hours, immunohistochemistry was performed on floating 100µm thick vibratory microtome sections in well plates. Sections were incubated overnight with primary antibody on a shaker at 4°C. Antibodies are described in Table 3. After washing with PBS containing 0.05% Tween-20, appropriate secondary antibody was added and incubated at room temperature for 2 hours. Cell nuclei were stained with Hoechst 33342 (Thermo Fisher Scientific).

[0232] Table 3: Antibodies

[0233]

[0234] Image analysis: The renal cortex was imaged using a Leica SP8 confocal microscope at an initial magnification of ×400 (5 images / mouse). To assess fibrosis, positive staining for aSMA and vimentin in each image was quantified using the Otzu thresholding method in ImageJ (NIH). Large arteries and glomeruli were excluded. Fibrosis area is expressed as a percentage of the total image area. Vascular area was quantified in the same manner as endothelial mucins and podocyte marker proteins described above and correlated with the total number of nuclei per image. The number of nuclei was counted by using Analyze Particles in ImageJ (NIH) and selecting positive regions of 50 to 800 pixels.

[0235] Quantitative real-time PCR: mRNA was extracted using RNAeasy Micro or Minikit (Qiagen) according to the manufacturer's protocol, followed by cDNA synthesis of 1 µg mRNA using iScript reverse transcription supermix (170-8841, BioRad). Real-time PCR was performed on a CFX-96 real-time PCR system (BioRad) using cDNA, Taqman gene expression premix (4369016, Thermo Fisher Scientific), and probes. Probes are listed in Table 4. Expression results were normalized to endogenous controls Hprt or Gapdh and analyzed using the Livak method (2... -ΔΔCT (To perform relative quantification)

[0236] Table 4: Probes for Real-Time PCR

[0237]

[0238] Transmission electron microscopy: For electron microscopy, kidneys were cut into 1 mm cubes and immersed in 0.1 M phosphate buffer (pH 7.4) in 2.5% glutaraldehyde (Ted Pella) and 1% paraformaldehyde (Merck), and stored at 4°C until further processing. The samples were rinsed in 0.1 M phosphate buffer for 10 min, then incubated in 0.1 M phosphate buffer with 1% osmium tetroxide (TAAB Laboratories Equipment) for 1 h. After rinsing in phosphate buffer, the samples were dehydrated in alcohol and then incubated in propylene oxide (TAAB Laboratories Equipment) for 5 min. The samples were then incubated in Epon resin (TAAB Laboratories Equipment) and propylene oxide (1:1) for 1 h, and then incubated overnight in 100% resin. Subsequently, the samples were encapsulated in fresh resin for 1 h to 2 h, and then heat-cured at 60°C for 48 h. The specimens were cut into semi-thin sections, stained with toluidine blue, and examined under an optical microscope. Ultrathin sections (60 nm to 70 nm) were cut in an EM UC7 microtome (Leica) and placed on a grid. The sections were contrast-stained with 5% uranyl acetate and Reynolds lead citrate and visualized under a Tecnai G2 Spirit BioTwin electron microscope (Thermo Fisher Scientific / FEI) at 80 kV using an Orius SC200 CCD camera and Gatan Digital Micrograph software (both from Gatan Inc.). Micrographs of the peritubular capillaries were taken, and the number of capillary pores was semi-quantitatively graded and expressed as a score between 0 and 4, where 0: 0%–5%, 1: 6%–25%, 2: 26%–50%, 3: 51%–75%, and 4: 76%–100% of the capillary length had pores (i.e., a score of 4 in healthy kidneys). An increased number of vacuoles was observed during electron microscopy, and therefore the number of affected renal tubular segments was quantified on toluidine blue-stained semi-thin sections as correlated with the total number of renal tubular segments in each image taken at the original magnification of ×400. When vacuoles are present throughout the epithelial cells, from the luminal side to the basal side ( Figure 8 In case A), the renal tubular segment is defined as affected.

[0239] Statistical analysis: Data are expressed as geometric mean (SD). Means between groups were compared using unpaired two-tailed Student's t-test (2 groups) or one-way ANOVA with Bonferroni post-hoc multiple comparisons using GraphPad Prism version 10 (GraphPad Software Inc.) (≥3 groups). p < 0.05 was considered statistically significant.

[0240] Example 1 - ABTAA's TIE2 activation improves renal perfusion and prevents capillary sparsity after UUO.

[0241] In the embodiments described herein, the ABTAA antibody is as defined in SEQ ID NOs 33 and 34. .

[0242] Purpose

[0243] To evaluate whether TIE2 activation via ABTAA improves renal perfusion and capillary sparsity in an in vivo model of kidney disease (UUO).

[0244] Materials and methods

[0245] Renal perfusion was evaluated as described above, i.e., by contrast imaging ultrasound following intravenous injection of circulatory vascular markers (MicroMarkers). Capillary density in the renal cortex was assessed by staining for endothelial markers endothelial mucin and podocyte marker proteins (excluding glomeruli).

[0246] result

[0247] Following UUO, the increased ANGPT2 immunoreactivity in the peritubular capillaries of the renal tubules was confirmed. Figure 2 Following UUO, TIE2 levels decreased, and TIE2... iECKO Mice showed a significant decrease in TIE2 ( Figure 3 ).

[0248] Compared with IgG-treated and wild-type (WT) controls, UUO-induced (2-day UUO) reduction in renal perfusion was observed in ABTAA-treated and Veptp-treated controls. iECKO Significant recovery was observed in mice. In contrast, TIE2 showed improvement compared to WT mice. iECKO Significantly lower UUO-induced renal perfusion ( Figure 4 ).

[0249] Compared to CL kidneys, 3-day UUO caused a significant decrease in capillary density. ABTAA-treated mice and Veptp iECKO Mice were significantly protected from this effect of UUO, while capillary sparsity in TIE2 iECKOThe condition was significantly aggravated in mice. (Pdgfb) iKO It has no effect on UUO-induced capillary density. Figure 5 AC).

[0250] in conclusion

[0251] ABTAA significantly restored renal perfusion and capillary sparsity in in vivo models of kidney disease.

[0252] Example 2 - ABTAA-induced TIE2 activation alleviated UUO-induced endothelial damage.

[0253] Purpose

[0254] To assess whether TIE2 activation via ABTAA improves endothelial injury in an in vivo model of kidney disease (UUO).

[0255] Materials and methods

[0256] The fenestrations of the peritubular capillaries are important structures for the reabsorption of water and solutes from the renal tubular system. These fenestrations are reduced in CKD and can be used to estimate capillary dysfunction (Babickova et al., 2000). In 3-day UUO and CL kidneys, the fenestrations of the peritubular capillaries are reduced at TIE2. iECKO Veptp iECKO The results were semi-quantitatively analyzed using transmission electron microscopy in the ABTAA treatment group.

[0257] result

[0258] In all groups, CL kidneys showed continuous fenestrations in the peritubular capillary endothelium. Figure 6 A, Figure 6 B). Compared with IgG treatment and WT control, UUO-induced reduction in fenestration was observed in ABTAA treatment and Veptp control, respectively. iECKO Significant recovery was achieved in mice ( Figure 6 In contrast, TIE2 compared to WT mice... iECKO The window score induced by UUO showed a significant deterioration.

[0259] in conclusion

[0260] In an in vivo model of kidney disease, ABTAA significantly restored renal endothelial damage.

[0261] Example 3 - ABTAA-induced TIE2 activation alleviated UUO-induced renal tubular interstitial fibrosis.

[0262] Purpose

[0263] To evaluate whether TIE2 activation via ABTAA improves renal tubular interstitial fibrosis in an in vivo model of kidney disease (UUO).

[0264] Materials and methods

[0265] Activation of perivascular mesenchymal cells into myofibroblasts is characterized by the expression of α-smooth muscle actin (aSMA) and the preparation of extracellular matrix components, including type I collagen, fibronectin, and vimentin. Renal tubulointerstitial fibrosis (excluding glomeruli) is quantified by staining the renal cortex for aSMA and vimentin.

[0266] result

[0267] Compared with IgG-injected mice and WT mice, UUO-induced renal tubulointerstitial fibrosis was significantly reduced in ABTAA-treated and Veptp-treated mice. iECKO Significantly reduced in mice ( Figure 7 A to Figure 7 C). The reduction in fibrosis is also reflected in ABTAA treatment and Veptp. iECKO UUO-induced decreases in the expression of Col1a1 and Pdgfrb in mice ( Figure 7 D、 Figure 7 E). Pdgfb iKO Mice exhibited significantly reduced renal tubulointerstitial fibrosis, thus supporting the concept of PDGFB as a major mesenchymal activator. In contrast, TIE2... iECKO The disease in mice was aggravated, and renal tubular interstitial fibrosis was significantly increased in the kidneys after 3 days of UUO treatment, along with a significant increase in the expression of Col1a1, Tagln, and Fn1. Figure 7 F).

[0268] in conclusion

[0269] ABTAA significantly reduced renal fibrosis in an in vivo model of kidney disease.

[0270] Example 4 - ABTAA-induced TIE2 activation alleviated UUO-induced renal tubular damage and reduced PDGFB expression.

[0271] Purpose

[0272] To evaluate whether TIE2 activation via ABTAA reduces renal tubular damage in an in vivo model of kidney disease (UUO).

[0273] Materials and methods

[0274] Quantitative analysis of UUO-induced renal tubular vacuoles by transmission electron microscopy ( Figure 8 A) and PDGFB expression were used as readouts for renal tubular injury.

[0275] result

[0276] Compared with IgG-treated mice and WT mice, ABTAA mice and Veptp mice, respectively iECKO The number of cortical tubular segments with UUO-induced pathological vacuolation was significantly reduced in mice. Figure 8 B). In contrast, compared to WT mice, TIE2 iECKO This leads to a significant increase in pathological renal tubular segments after UUO (UUO) Figure 8 B).

[0277] UUO-induced Pdgfb / PDGFB expression in ABTAA-treated mice and Veptp iECKO Significantly reduced in mice ( Figure 9 A, Figure 9 B), thereby supporting vascular protection of the renal tubular system. As expected, Pdgfb iKO Pdgfb mRNA expression was significantly reduced in both CL and UUO kidneys of mice; however, compared with CL kidneys of WT mice, Pdgfb levels were still significantly increased in UUO-treated kidneys after 3 days. Figure 9 C).

[0278] in conclusion

[0279] ABTAA significantly reduced tubular damage in an in vivo model of kidney disease.

[0280] Example 5 - Post-injury treatment with ABTAA to reduce UUO-induced damage.

[0281] Purpose

[0282] To further evaluate the potential of ABTAA treatment, more clinically relevant treatment options for kidney disease were investigated.

[0283] Materials and methods

[0284] Advanced ABTAA treatment was initiated on day 3 post-UUO, and renal analysis was performed on day 10 post-UUO. Figure 10 A). Fibrosis was evaluated by aSMA and vimentin staining, and tubular lesions were evaluated by PDGFB expression.

[0285] result

[0286] As measured by aSMA and vimentin staining, advanced ABTAA treatment significantly reduced UUO-induced renal tubular interstitial fibrosis, while TIE2... iECKO Mice showed a significant increase in fibrosis ( Figure 10 B to Figure 10 D). Tubular lesions as reflected by PDGFB expression were significantly lower in mice treated with ABTAA. Figure 10 E).

[0287] in conclusion

[0288] In more clinically relevant models of kidney disease, ABTAA significantly reduced tubulointerstitial fibrosis and tubular damage in in vivo models of kidney disease.

[0289] Example 6 - ABTAA therapy has a protective effect against early diabetic complications.

[0290] Purpose

[0291] The study investigates the protective effect of ABTAA therapy against early diabetic complications.

[0292] Materials and methods

[0293] Experiments were conducted using the BTBRob / ob (also known as ob / ob) mouse model, which develops type 2 diabetes and several cardiovascular complications. It has been previously shown that this model already exhibits significant capillary density loss in the kidneys at 6 weeks of age (Zhou et al., 2024). The BTBRob / ob model has also developed cardiomyopathy (Ye et al., 2017) and retinopathy (Lee et al., 2018), making it an ideal model for studying therapeutic interventions in diabetic complications. The inventors confirmed the upregulation of ANGPT2 in the kidney tissue of ob / ob mice, as this is required by the ABTAA mechanism of action. Figure 11 A). Non-diabetic heterozygous mice were used as controls. Mice were subcutaneously injected weekly with 25 mg / kg ABTAA or human IgG until 11 weeks of age. Various measurements were performed, including plasma blood urea nitrogen levels, capillary density in the renal cortex (staining with renal cortical endothelial mucin), fenestrations in peritubular capillaries (visible by electron microscopy), and plasma creatine kinase levels. Retinopathy was assessed by staining capillaries with Pecam1 and the basement membrane with collagen IV, followed by quantification of the amount of empty collagen sheaths left upon capillary disappearance.

[0294] result

[0295] ABTAA treatment normalized blood urea nitrogen levels in ob / ob mice, indicating a protective effect against nephropathy. Figure 11 C). Compared with the control, the capillary density in the renal cortex of mice treated with ABTAA was maintained ( Figure 11 D). Furthermore, ABTAA maintains normal fenestrations in the peritubular capillaries, indicating improved renal function. Figure 11 E). In cardiomyopathy, ABTAA prevented the increase of plasma creatine kinase levels, indicating a protective effect against myocardial injury. Figure 11 F). For retinal diseases, ABTAA reduced the number of hollow collagen sheaths in retinal capillaries, demonstrating its protective effect. Figure 11 G).

[0296] in conclusion

[0297] ABTAA therapy effectively prevented early diabetic complications in the BTBRob / ob mouse model, maintained renal function and capillary integrity, and prevented myocardial damage. These findings suggest that ABX has potential therapeutic benefits in managing diabetic complications and may be used in combination with other obesity and hypoglycemic agents.

[0298] Example 7 - ABTAA therapy has a protective effect in coagulation disorders.

[0299] Purpose

[0300] This study investigates whether ABTAA has a protective effect against coagulation disorders.

[0301] Materials and methods

[0302] ANGPT2 inhibits thrombomodulin (TM)-mediated protein C activation (Hultstrom et al., 2022), suggesting an additional role for ANGPT2 in endothelial dysfunction and coagulation disorders, also observed in diabetic patients. The inventors have developed a coagulation disorder model. Mice were injected with the inflammatory cytokine TNFα to trigger the release of endogenous ANGPT2 (…). Figure 12 A). Then, coagulation ability was evaluated by measuring the tail bleeding time.

[0303] result

[0304] Administration of TNFα significantly shortened tail bleeding time, indicating impaired coagulation. However, this effect was completely blocked by ABTAA treatment. Figure 12 B), indicating that ABTAA has a protective effect in coagulation disorders.

[0305] in conclusion

[0306] This study demonstrates that ABTAA has a protective effect against TNFα-induced coagulation disorders. This suggests that ABTAA has potential therapeutic benefits in managing coagulation disorders.

[0307] Sequence Overview

[0308] LCCDR1LCCDR2LCCDR3 SEQIDNO:1SEQIDNO:2SEQIDNO:3 LC CDR1: KASQDVGTAVALC CDR2: WASTRHTLC CDR3: QQYSSYPLT HCCDR1HCCDR2HCCDR3 SEQIDNO:4SEQIDNO:5SEQIDNO:6 HC CDR1: SYWMHHC CDR2: MIDPSDSETRLNQKFKDHC CDR3: RFYYGSDWYFDV LCVR SEQIDNO:7 <![CDATA[DIVMTQSHKFMSTSVGDRVSITC KASQDVGTAVA WYQQKPGQSPKLLIY WASTRHT GVPDRFTGSGSGTDFTLTISNVQSEDLADYFC QQYSSYPLT FGSGTKLEIK]]> HCVR SEQ ID NO:8 <![CDATA[QVQLQQSGPQLVRPGASVKISCKASGYSFT SYWMH WVKQRPGQGLEWIG MIDPSDSETRLN QKFKD KASLTVDKSSSTAYMQLSSPTSGDSAVYYCAR RFYYGSDWYFDV WGAGSTVTVSS]]> Epitope SEQ ID NO:9 QRTWKEYKVGFGNPSGEY 2C8H11VH SEQ ID NO:10 <![CDATA[QVQLVQSGAEVKKPGASVKVSCKASGYTFT SYWMH WVRQAPGQGLEWMG MIDPSDSETRLN QKFKD RVTMTTSTSTTVYMELSSLRSEDTAVYYCAR RFYYGSDWYFDV WGQGTLVTVSS<!-- 28 --> ]]> 2C8H11VL SEQ ID NO:11 <![CDATA[DIQLTQSPSFLSASVGDRVTITC KASQDVGTAVA WYQQKPGKAPKLLIY WASTRHT GVPSRFSGSGSGTEFTLTISSLQPEDFATYYC QQYSSYPLT FGQGTKVEIK]]> TIE2wt forward SEQ ID NO:12 TCCTTGCCGCCAACTTGTAAAC TIE2wt reverse SEQ ID NO:13 AGCAAGCTGACTCCACAGAGAAC TIE2flox forward SEQ ID NO:14 CACTTTCTACTCTGTTGAC TIE2flox reverse SEQ ID NO:15 CCTTACATGTTTTACTAG Cre forward SEQ ID NO:16 ATGTCCAATTTACTGACCG Cre reverse SEQ ID NO:17 CGCCGCATAACCAAGTGAA TdTomato forward SEQ ID NO:18 CTGTTCCTGTACGGCATGG TdTomato reverse SEQ ID NO:19 GGCATTAAAGCAGCGTATCC Veptpflox forward SEQ ID NO:20 GCGTCTATCCAGTGGAGGACTTTC Veptpflox reverse SEQ ID NO:21 CCAGGTGCCGTTCATTCAGC Pdgfbflox forward SEQ ID NO:22 TTTGAAGCGTGCAGAATGCC Pdgfbflox reverse SEQ ID NO:23 GGAACGGATTTTGGAGGTAGTGTC LCDR1 LCDR2 LCDR3 SEQ ID NO:24 SEQ ID NO:25 SEQ ID NO:26 KASQSVSNDVAYASNRYPQQDYSSPWT HCDR1 HCDR2 HCDR3 SEQ ID NO:27 SEQ ID NO:28 SEQ ID NO:29 SDYAWNYINYSGNTDYNPSLKSGNFEGAMDY Epitope SEQ ID NO:30 SQPGNDFSTKDGDNDKCI VH SEQ ID NO:31 DVQLQESGPDLVKPSQSLSLTCTVTGYSITSDYAWNWIRQFPGNKLEWMGYINYSGNTDYNPSLKSRSSITRDTSKNQFFLQLNSVTTGDTATYYCARGNFEGAMDYWGQGTSVTVS VL SEQ ID NO:32 SIVMTQTPKFLLVSAGDRVTITCKASQSVSNDVAWYQQKPGQPKLLIYYASNRYPGVPDRFTGSGYGTDFTFTISTVQAEDLAVYFCQQDYSSPWTFGGGTKLEIK VH SEQ ID NO:33 <![CDATA[QVQLQESGPGLVKPSETLSLTCAVSGYSIT SDYAWN WIRQPPGKGLEWMG KISYSGKTDYN PSLKS RSTISRDTSKNQFSLKLSSVTAADTAVYYCAR GNFEGAMDY WGQGTLVTVSS]]> VL SEQ ID NO:34 <![CDATA[DIQMTQSPSSLSASVGDRVTITC KASQSVSNDVH WYQQKPGKAPKLLIY YASIPYP GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC QQDYSSPWT FGQGTKVEIK]]> LCDR1 LCDR2 LCDR3 SEQ ID NO:35 SEQ ID NO:36 SEQ ID NO:26 <![CDATA[ KASQSVSNDVHYASIPYPQQDYSSPWT ]]> HCDR1 HCDR2 HCDR3 SEQ ID NO:27 SEQ ID NO:37 SEQ ID NO:29 <![CDATA[ SDYAWNKISYSGKTDYNPSLKSGNFEGAMDY <!-- 32 -->]]> Human ANGPT2 SEQ ID NO:39 MWQIVFFTLS CDLVLAAAYN NFRKSMDSIG KKQYQVQHGSCSYTFLLPEMDNCRSSSSPY VSNAVQRDAP LEYDDSVQRLQVLENIMENN TQWLMKLENYIQDNMKKEMV EIQQHAVQNQTAVMIEIGTN LLNQTAEQTR KLTDVEAQVLNQTTRLELQLLEHSLSTNKL EKQILDQTSE INKLQDKNSFLEKKVLAMEDKHIIQLQSIK EEKDQLQVLV SKQNSIIEELEKKIVTATVNNSVLQKQQHD LMETVNNLLT MMSTSNSAKDPTVAKEBQISFRDCAEVFKS GHTTNGIYTL TFPNSTEEIKAYCDMEAGGGGWTIIQRRED GSVDFQRTWK EYKVGFGNPSGEYWLGNEFVSQLTNQQRYV LKIHLKDWEG NEAYSLYEHFYLSSEELNYRIHLKGLTGTA GKISSISQPG NDFSTKDGDNDKCICKCSQMLTGGWWFDAC GPSNLNGMYY PQRQNTNKFNGIKWYYWKGSGYSLKATTMM IRPADF VL SEQ ID NO:38 <![CDATA[DIQMTQSPSSLSASVGDRVTITC KASQSVSNDVH WYQQKPGKAPKLLIY YASIPYP GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC QQDYSSPWT FGQGTKLEIK]]>

[0309] References

[0310] Saharinen, P., Eklund, L. & Alitalo, K. Therapeutic targeting of theangiopoietin-TIE pathway. Nat Rev Drug Discov 16, 635-661 (2017). https: / / doi.org / 10.1038 / nrd.2016.278

[0311] Miikka Vikkula, Laurence M Boon, Kermit L.Carraway Iii, Jennifer TCalvert, A.John Diamonti, Boyan Goumnerov, Krystyna A Pasyk, Douglas AMarchuk, Matthew L Warman, Lewis C Cantley, John B Mulliken, Bjorn R Olsen, Vascular Dysmorphogenesis Caused by an Activating Mutation in the ReceptorTyrosine Kinase TIE2, Cell, Volume 87, Issue 7, 1996, Pages 1181-1190.

[0312] Han S, Lee SJ, Kim KE, Lee HS, Oh N, Park I, Ko E, Oh SJ, Lee YS, KimD, Lee S, Lee DH, Lee KH, Chae SY, Lee JH, Kim SJ, Kim HC, Kim S, Kim SH, KimC, Nakaoka Y, He Y, Augustin HG, Hu J, Song PH, Kim YI, Kim P, Kim I, Koh GY.Amelioration of sepsis by TIE2 activation-induced vascular protection. SciTransl Med.2016 Apr 20;8(335):335ra55. doi: 10.1126 / scitranslmed.aad9260.PMID: 27099174.

[0313] Pitulescu ME, Schmidt I, Benedito R, and Adams RH. Inducible genetargeting in the neonatal vasculature and analysis of retinal angiogenesis inmice. Nat Protoc. 2010;5(9):1518-34.

[0314] Madisen L, Zwingman TA, Sunkin SM, Oh SW, Zariwala HA, Gu H, et al. Arobust and high-throughput Cre reporting and characterization system for thewhole mouse brain. Nat Neurosci. 2010;13(1):133-40.

[0315] Enge M, Bjarnegard M, Gerhardt H, Gustafsson E, Kalen M, Asker N, etal. Endothelium-specific platelet-derived growth factor-B ablation mimicsdiabetic retinopathy. EMBO J. 2002;21(16):4307-16.

[0316] Hayashi S, and McMahon AP. Efficient recombination in diverse tissuesby a tamoxifen-inducible form of Cre: a tool for temporally regulated geneactivation / inactivation in the mouse. Dev Biol. 2002;244(2):305-18.

[0317] Hamilton TG, Klinghoffer RA, Corrin PD, and Soriano P. Evolutionarydivergence of platelet-derived growth factor alpha receptor signalingmechanisms. Molecular and cellular biology. 2003;23(11):4013-25.

[0318] Loganathan K, Salem Said E, Winterrowd E, Orebrand M, He L,Vanlandewijck M, et al. Angiopoietin-1 deficiency increases renal capillaryrarefaction and tubulointerstitial fibrosis in mice. PloS one. 2018;13(1):e0189433.

[0319] Yang HC, Zuo Y, and Fogo AB.Models of chronic kidney disease.Drugdiscovery today Disease models. 2010;7(1-2):13-9.

[0320] Kramann R, Tanaka M, and Humphreys BD. Fluorescence microangiographyfor quantitative assessment of peritubular capillary changes after AKI inmice. J Am Soc Nephrol. 2014;25(9):1924-31.

[0321] Babickova J, Klinkhammer BM, Buhl EM, Djudjaj S, Hoss M, Heymann F,et al. Regardless of etiology, progressive renal disease causesultrastructural and functional alterations of peritubular capillaries. KidneyInt. 2017; 91(1):70-85

[0322] Bolignano D, Lacquaniti A, Coppolino G, Donato V, Campo S, Fazio MR,et al. Neutrophil gelatinase-associated lipocalin (NGAL) and progression ofchronic kidney disease. Clinical journal of the American Society ofNephrology : CJASN. 2009;4(2):337-44.

[0323] Zhou Aa, Jeansson Ma, He L, et al. Renal endothelial single-celltranscriptomics reveals spatiotemporal regulation and divergent roles ofdifferential gene transcription and alternative splicing in murine diabeticnephropathy. Int J Mol Sci. 2024; a, b equal contribution.

[0324] Ye Y, Bajaj M, Yang HC, Perez-Polo JR, Birnbaum Y. SGLT-2 Inhibitionwith Dapagliflozin Reduces the Activation of the Nlrp3 / ASC Inflammasome andAttenuates the Development of Diabetic Cardiomyopathy in Mice with Type 2Diabetes. Further Augmentation of the Effects with Saxagliptin, a DPP4Inhibitor. Cardiovasc Drugs Ther. 2017;31(2):119-132.

[0325] Lee VK, Hosking BM, Holeniewska J, Kubala EC, Lundh von Leithner P,Gardner PJ, Foxton RH, Shima DT. BTBR ob / ob mouse model of type 2 diabetesexhibits early loss of retinal function and retinal inflammation followed bylate vascular changes. Diabetologia. 2018;61(11):2422-2432

[0326] Hultstrom M, Fromell K, Larsson A, Persson B, Nilsson B, Quaggin SE,Betsholtz C, Frithiof R, Lipcsey M, Jeansson M. Angiopoietin-2 Inhibition of Thrombomodulin-Mediated Anticoagulation-A Novel Mechanism That May Contribute to Hypercoagulation in Critically Ill COVID-19 Patients. Biomedicines. 2022;10(6)

[0327] item

[0328] 1. An antibody or an antigen-binding fragment thereof, said antibody or antigen-binding fragment thereof binding to ANGPT2 and via ANGPT2 binding to the TIE2 receptor, for the treatment or prevention of fibrotic diseases.

[0329] 2. An antibody or an antigen-binding fragment thereof, said antibody or antigen-binding fragment thereof binding to ANGPT2 and via ANGPT2 binding to a TIE2 receptor, for treating or preventing a subject’s kidney disease, condition or injury, said kidney disease, condition or injury being associated with kidney fibrosis.

[0330] 3. An antibody or an antigen-binding fragment thereof, said antibody or antigen-binding fragment thereof binding to ANGPT2 and via ANGPT2 binding to the TIE2 receptor, for the treatment or prevention of coagulation disorders.

[0331] 4. An antibody or an antigen-binding fragment thereof, said antibody or antigen-binding fragment thereof binding to ANGPT2 and via ANGPT2 to a TIE2 receptor, for the treatment or prevention of diabetic complications, such as early diabetic complications.

[0332] 5. The antibody or antigen-binding fragment thereof for use according to claims 1 to 4, wherein the antibody or antigen-binding fragment thereof is characterized in that it binds to binding sites from position 336 to position 434 of angiopoietin-2 (ANGPT2) (SEQ ID NO: 30).

[0333] 6. The antibody or antigen-binding fragment thereof for use according to claim 5, wherein the length of the binding site is about 2 to about 20 consecutive amino acids, such as about 2 to about 18, about 5 to about 20, or about 10 to about 20, such as about 15 to about 20 consecutive amino acids.

[0334] 7. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is characterized in that it binds to the epitope of human angiopoietin-2 (ANGPT2) SEQ ID NO: 9 or SEQ ID NO: 30.

[0335] 8. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof comprises:

[0336] a) A complementarity-determining region (CDR) of the heavy chain variable region, the CDR comprising: HCDR1, the HCDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, the HCDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, the HCDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0337] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0338] 9. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof comprises:

[0339] a) The complementarity-determining region (CDR) of the heavy chain variable region, wherein the CDR comprises: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0340] b) The complementarity-determining region (CDR) of the light chain variable region, wherein the complementarity-determining region (CDR) comprises: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3; or the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0341] 10. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof comprises a variable heavy chain of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 31 or SEQ ID NO: 33, and a variable light chain of SEQ ID NO: 7, SEQ ID NO: 11, SEQ ID NO: 32, SEQ ID NO: 34 or SEQ ID NO: 38.

[0342] 11. An antibody or an antigen-binding fragment thereof for treating or preventing kidney disease, condition, or injury in a subject, wherein the antibody or antigen-binding fragment thereof comprises:

[0343] a) The complementarity-determining region (CDR) of the heavy chain variable region, wherein the CDR comprises: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; and

[0344] b) The CDR of the light chain variable region, wherein the CDR comprises: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3.

[0345] 12. An antibody or an antigen-binding fragment thereof for treating or preventing fibrotic diseases, conditions, or injuries in a subject, wherein the antibody or antigen-binding fragment thereof comprises:

[0346] a) The complementarity-determining region (CDR) of the heavy chain variable region, wherein the CDR comprises: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; and

[0347] b) The CDR of the light chain variable region, wherein the CDR comprises: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3.

[0348] 13. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof comprises a variable heavy chain of SEQ ID NO: 8 or SEQ ID NO: 10 and a variable light chain of SEQ ID NO: 7 or SEQ ID NO: 11.

[0349] 14. The antibody or antigen-binding fragment thereof for use according to claims 1 to 4, wherein the antibody or antigen-binding fragment thereof is characterized by binding to the epitope of human angiopoietin-2 (ANGPT2) SEQ ID NO: 9.

[0350] 15. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof comprises:

[0351] a) The complementarity-determining region (CDR) of the heavy chain variable region, said CDR comprising: the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0352] b) The complementarity-determining region (CDR) of the light chain variable region, wherein the complementarity-determining region (CDR) comprises the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0353] 16. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof comprises a variable heavy chain of SEQ ID NO: 31 or SEQ ID NO: 33 and a variable light chain of SEQ ID NO: 32 or SEQ ID NO: 34 or SEQ ID NO: 38.

[0354] 17. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is characterized in that it binds to the epitope of human angiopoietin-2 (ANGPT2) SEQ ID NO: 30.

[0355] 18. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is polyclonal or monoclonal.

[0356] 19. An antibody or an antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antigen-binding fragment is scFv or Fab.

[0357] 20. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody is a humanized antibody or a human antibody.

[0358] 21. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody is a human subtype IgG1.

[0359] 22. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein said antibody or antigen-binding fragment thereof:

[0360] a) Reduce or inhibit human angiopoietin-2 (ANGPT2); and / or

[0361] b) Inducing TIE2 activation; and

[0362] c) Inducing TIE2 phosphorylation.

[0363] 23. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is in fibrotic and / or diseased tissue, such as in a diseased kidney:

[0364] a) Reduce or inhibit human angiopoietin-2 (ANGPT2); and / or

[0365] b) Inducing TIE2 activation; and

[0366] c) Inducing TIE2 phosphorylation.

[0367] 24. The antibody or antigen-binding fragment thereof for use as described in claim 23, wherein the induction of TIE2 activation is ANGPT2-dependent.

[0368] 25. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof normalizes pathological blood vessels and prevents the progression of fibrosis.

[0369] 26. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof prevents or mitigates inadequate perfusion associated with fibrotic diseases such as kidney disease, condition, or injury.

[0370] 27. The antibody or antigen-binding fragment thereof for use as described in claim 26, wherein the perfusion is a renal perfusion.

[0371] 28. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof prevents or reduces capillary thinning associated with fibrotic diseases such as kidney disease, condition, or injury.

[0372] 29. The antibody or antigen-binding fragment thereof used according to claim 28, wherein prevention or mitigation of capillary sparsity is evaluated by quantification of endothelial markers such as endothelial mucin or podocyte marker proteins.

[0373] 30. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof prevents or mitigates the reduction of peritubular capillary fenestrations associated with the kidney disease, condition, or injury.

[0374] 31. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein fibrosis, such as renal fibrosis, is prevented or mitigated.

[0375] 32. The antibody or antigen-binding fragment thereof for use according to claim 31, wherein the fibrosis is renal tubular interstitial fibrosis.

[0376] 33. The antibody or antigen-binding fragment thereof for use according to claims 31 to 32, wherein prevention or mitigation of fibrosis is evaluated by quantification of fibrosis markers such as α-smooth muscle actin (aSMA), vimentin, type I collagen (Col1) or platelet-derived growth factor receptor-β (PDGFR-β), optionally wherein said fibrosis is renal fibrosis.

[0377] 34. A pharmaceutical composition comprising an antibody or an antigen-binding fragment thereof according to any one of the preceding claims for the treatment or prevention of kidney disease, wherein the kidney disease, condition, or injury is associated with kidney fibrosis.

[0378] 35. A pharmaceutical composition comprising an antibody or an antigen-binding fragment thereof according to any one of the preceding claims, for the treatment or prevention of fibrotic diseases.

[0379] 36. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is administered prior to the onset of a fibrotic disease, such as kidney disease, condition, or injury.

[0380] 37. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is administered during an attack of a fibrotic disease, such as kidney disease, condition, or injury.

[0381] 38. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is administered after an onset of fibrotic disease, such as kidney disease, condition, or injury.

[0382] 39. The antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the fibrotic disease, such as kidney disease, condition or injury, is induced by, spread by and / or associated with endothelial dysfunction.

[0383] 40. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the kidney disease, condition or injury is induced by, spread by and / or associated with renal capillary sparsity.

[0384] 41. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the kidney disease, condition or injury is induced by, spread by and / or associated with renal tubular atrophy.

[0385] 42. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the kidney disease, condition or injury is induced by, spread by and / or associated with renal tubular injury.

[0386] 43. The antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the fibrotic disease, such as kidney disease, condition or injury, is induced by oxidative stress, spread by oxidative stress, and / or associated with oxidative stress.

[0387] 44. The antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the fibrotic disease, such as kidney disease, condition or injury, is induced by hypoxia, is spread by hypoxia and / or is associated with hypoxia.

[0388] 45. The antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the kidney disease, condition or injury is chronic kidney disease (CKD).

[0389] 46. ​​The antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the fibrotic disease is a chronic fibrotic disease.

[0390] 47. The antibody or antigen-binding fragment thereof for use according to claim 43, wherein the CKD is induced by, spread by, and / or associated with a coagulation disorder.

[0391] 48. The antibody or antigen-binding fragment thereof for use according to claim 41, wherein the chronic fibrotic disease is induced by, spread by, and / or associated with a coagulation disorder.

[0392] 49. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the kidney disease, condition, or injury is selected from the group consisting of: chronic kidney damage, chronic renal failure syndrome, glomerulonephritis, type I diabetes, type II diabetes, ischemic nephropathy, focal segmental glomerulosclerosis, nephrotic syndrome, cardiorenal syndrome, tubulointerstitial disease associated with metabolic disorders, minimal change disease, hypertensive nephropathy, and malignant hypertensive nephropathy. Diseases, IgA nephropathy, IgM nephropathy, mesangial capillary glomerulonephritis, crescentic glomerulonephritis, diffuse intracapillary proliferative glomerulonephritis, mesangial proliferative glomerulonephritis, focal and segmental proliferative glomerulonephritis, systemic vasculitis, pulmonary hemorrhage nephritis syndrome, Dent disease, systemic lupus erythematosus glomerulonephritis syndrome, allergic purpura, systemic sclerosis, scleroderma, nephropathy associated with urinary tract obstruction, tubulointerstitial nephritis, interstitial nephritis, and lithium nephropathy.

[0393] 50. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the renal fibrosis is induced by, spread by, and / or associated with: chronic kidney injury, chronic renal failure syndrome, glomerulonephritis, type I diabetes, type II diabetes, ischemic nephropathy, focal segmental glomerulosclerosis, nephrotic syndrome, cardiorenal syndrome, tubulointerstitial disease associated with metabolic disorders, minimal change disease, hypertensive nephropathy, malignant nephropathy, etc. Hypertensive kidney disease, IgA nephropathy, IgM nephropathy, mesangial capillary glomerulonephritis, crescentic glomerulonephritis, diffuse intracapillary proliferative glomerulonephritis, mesangial proliferative glomerulonephritis, focal and segmental proliferative glomerulonephritis, systemic vasculitis, pulmonary hemorrhage nephritis syndrome, Dent disease, systemic lupus erythematosus glomerulonephritis syndrome, allergic purpura, systemic sclerosis, scleroderma, nephropathy associated with urinary tract obstruction, tubulointerstitial nephritis, interstitial nephritis and / or lithium nephropathy.

[0394] 51. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the kidney disease, condition, or injury is induced by type 2 diabetes, spread by type 2 diabetes, and / or associated with type 2 diabetes.

[0395] 52. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the fibrotic disease is induced by type 2 diabetes, spread by type 2 diabetes, and / or associated with type 2 diabetes.

[0396] 53. The antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the subject has type 2 diabetes.

[0397] 54. A method for treating a kidney disease, condition, or injury, wherein the kidney disease, condition, or injury is associated with kidney fibrosis, the method comprising administering to a subject in need a therapeutically effective amount of an antibody or an antigen-binding fragment thereof according to any one of the preceding claims, or a pharmaceutical composition according to any one of the preceding claims.

[0398] 55. A method for treating fibrosis, the method comprising administering to a subject in need a therapeutically effective amount of an antibody or an antigen-binding fragment thereof according to any one of the preceding claims, or a pharmaceutical composition according to any one of the preceding claims.

[0399] 56. Use of the antibody or antigen-binding fragment thereof according to any one of the preceding claims in the preparation of a medicament for treating fibrotic diseases, such as kidney diseases, conditions or injuries, wherein said kidney diseases, conditions or injuries are associated with kidney fibrosis.

[0400] 57. A method for reducing or inhibiting human angiopoietin-2 (ANGPT2) signaling, and / or inducing TIE2 activation, and / or inducing TIE2 phosphorylation in fibrotic tissue and / or diseased tissue such as diseased kidney, the method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, wherein the antibody or antigen-binding fragment thereof comprises:

[0401] a) A complementarity-determining region (CDR) of the heavy chain variable region, the CDR comprising: HCDR1, the HCDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, the HCDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, the HCDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0402] b) A complementarity-determining region (CDR) of the light chain variable region, the CDR comprising: LCDR1, the LCDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24, or SEQ ID NO: 35; LCDR2, the LCDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25, or SEQ ID NO: 36; and LCDR3, the LCDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0403] The diseased kidney mentioned above is fibrotic.

[0404] 58. A method for reducing or inhibiting human angiopoietin-2 (ANGPT2) signaling, and / or inducing TIE2 activation, and / or inducing TIE2 phosphorylation in fibrotic tissue and / or diseased tissue such as diseased kidney, the method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, wherein the antibody or antigen-binding fragment thereof comprises:

[0405] a) The complementarity-determining region (CDR) of the heavy chain variable region, wherein the CDR comprises: the HCDR1 amino acid sequence of SEQ ID NO: 12, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; and

[0406] b) The CDR of the light chain variable region, wherein the CDR comprises: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3.

[0407] The diseased kidney mentioned above is fibrotic.

[0408] 59. A method for reducing or inhibiting human angiopoietin-2 (ANGPT2) signaling, and / or inducing TIE2 activation, and / or inducing TIE2 phosphorylation in fibrotic tissue and / or diseased tissue such as diseased kidney, the method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, wherein the antibody or antigen-binding fragment thereof comprises:

[0409] a) The complementarity-determining region (CDR) of the heavy chain variable region, said CDR comprising: the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0410] b) The complementarity-determining region (CDR) of the light chain variable region, wherein the CDR comprises the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0411] The diseased kidney mentioned above is fibrotic.

[0412] 60. The method according to items 55 through 57, wherein the induction of TIE2 activation is ANGPT2 dependent.

[0413] 61. A method for normalizing pathological blood vessels and treating and alleviating fibrotic and / or diseased tissue, such as fibrosis in a diseased kidney, and preventing its progression, said method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, said antibody or antigen-binding fragment thereof comprising:

[0414] a) A complementarity-determining region (CDR) of the heavy chain variable region, the CDR comprising: HCDR1, the HCDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, the HCDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, the HCDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0415] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0416] 62. A method for normalizing pathological blood vessels and treating and alleviating fibrotic and / or diseased tissue, such as fibrosis in a diseased kidney, and preventing its progression, said method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, said antibody or antigen-binding fragment thereof comprising:

[0417] a) The complementarity-determining region (CDR) of the heavy chain variable region, wherein the CDR comprises: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; and

[0418] b) The CDR of the light chain variable region, wherein the CDR comprises: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3.

[0419] 63. A method for normalizing pathological blood vessels and treating and alleviating fibrotic and / or diseased tissue, such as fibrosis in a diseased kidney, and preventing its progression, said method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, said antibody or antigen-binding fragment thereof comprising:

[0420] a) The complementarity-determining region (CDR) of the heavy chain variable region, said CDR comprising: the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0421] b) The complementarity-determining region (CDR) of the light chain variable region, wherein the complementarity-determining region (CDR) comprises the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0422] 64. A method for treating and / or alleviating fibrotic tissue and / or diseased tissue, such as fibrosis in a diseased kidney, and / or preventing its progression, said method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, said antibody or antigen-binding fragment thereof comprising:

[0423] a) A complementarity-determining region (CDR) of the heavy chain variable region, the CDR comprising: HCDR1, the HCDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, the HCDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, the HCDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0424] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0425] 65. A method for treating and / or alleviating fibrotic tissue and / or diseased tissue, such as fibrosis in a diseased kidney, and / or preventing its progression, said method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, said antibody or antigen-binding fragment thereof comprising:

[0426] a) The complementarity-determining region (CDR) of the heavy chain variable region, wherein the CDR comprises: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; and

[0427] b) The CDR of the light chain variable region, wherein the CDR comprises: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3.

[0428] 66. A method for treating and / or alleviating fibrotic tissue and / or diseased tissue, such as fibrosis in a diseased kidney, and / or preventing its progression, said method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, said antibody or antigen-binding fragment thereof comprising:

[0429] a) The complementarity-determining region (CDR) of the heavy chain variable region, said CDR comprising: the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0430] b) The complementarity-determining region (CDR) of the light chain variable region, wherein the complementarity-determining region (CDR) comprises the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0431] 67. The antibody or antigen-binding fragment thereof for use according to claims 61 to 66, wherein the fibrosis is tubulointerstitial fibrosis and / or glomerulosclerosis.

[0432] 68. The antibody or antigen-binding fragment thereof used according to items 61 through 67, wherein the prevention or mitigation of fibrosis is evaluated by quantification of fibrosis markers such as α-smooth muscle actin (aSMA), vimentin, type I collagen (Col1), or platelet-derived growth factor receptor-β (PDGFR-β).

[0433] 69. A method for preventing or treating perfusion inadequacy, such as renal inadequacy, in a subject, wherein the renal disease, condition, or injury is associated with renal fibrosis, the method comprising administering an antibody or an antigen-binding fragment thereof to the subject in need, wherein the antibody or antigen-binding fragment thereof comprises:

[0434] a) A complementarity-determining region (CDR) of the heavy chain variable region, the CDR comprising: HCDR1, the HCDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, the HCDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, the HCDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0435] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0436] 70. A method for preventing or treating perfusion inadequacy, such as renal inadequacy, in a subject, wherein the renal disease, condition, or injury is associated with renal fibrosis, the method comprising administering an antibody or an antigen-binding fragment thereof to the subject in need, wherein the antibody or antigen-binding fragment thereof comprises:

[0437] a. The complementarity-determining region (CDR) of the heavy chain variable region, said CDR comprising: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; and

[0438] b. A CDR of the light chain variable region, the CDR comprising: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3.

[0439] 71. A method for preventing or treating perfusion inadequacy, such as renal inadequacy, in a subject, wherein the renal disease, condition, or injury is associated with renal fibrosis, the method comprising administering an antibody or an antigen-binding fragment thereof to the subject in need, wherein the antibody or antigen-binding fragment thereof comprises:

[0440] a) The complementarity-determining region (CDR) of the heavy chain variable region, said CDR comprising: the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0441] b) The complementarity-determining region (CDR) of the light chain variable region, wherein the complementarity-determining region (CDR) comprises the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0442] 72. A method for preventing or reducing capillary thinning associated with diseased and / or fibrotic tissue, such as a subject's kidney disease, condition, or injury, wherein the kidney disease, condition, or injury is associated with kidney fibrosis, the method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need of it, wherein the antibody or antigen-binding fragment thereof comprises:

[0443] a) A complementarity-determining region (CDR) of the heavy chain variable region, the CDR comprising: HCDR1, the HCDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, the HCDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, the HCDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0444] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0445] 73. A method for preventing or reducing capillary thinning associated with diseased and / or fibrotic tissue, such as a subject's kidney disease, condition, or injury, wherein the kidney disease, condition, or injury is associated with kidney fibrosis, the method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need of it, wherein the antibody or antigen-binding fragment thereof comprises:

[0446] a) The complementarity-determining region (CDR) of the heavy chain variable region, wherein the CDR comprises: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; and

[0447] b) The CDR of the light chain variable region, wherein the CDR comprises: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3.

[0448] 74. A method for preventing or reducing capillary thinning associated with diseased and / or fibrotic tissue, such as a subject's kidney disease, condition, or injury, wherein the kidney disease, condition, or injury is associated with kidney fibrosis, the method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, wherein the antibody or antigen-binding fragment thereof comprises:

[0449] a) The complementarity-determining region (CDR) of the heavy chain variable region, said CDR comprising: the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0450] b) The complementarity-determining region (CDR) of the light chain variable region, wherein the complementarity-determining region (CDR) comprises the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

[0451] 75. The method according to any one of claims 72 to 74, wherein the prevention or mitigation of capillary sparsity is evaluated by quantification of endothelial markers such as endothelial mucin or podocyte marker proteins.

[0452] 76. A method for preventing or treating a reduction in peritubular capillary fenestrations associated with kidney disease, condition, or injury in a subject, wherein the kidney disease, condition, or injury is associated with kidney fibrosis, the method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, wherein the antibody or antigen-binding fragment thereof comprises:

[0453] a) A complementarity-determining region (CDR) of the heavy chain variable region, the CDR comprising: HCDR1, the HCDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4 or SEQ ID NO: 27; HCDR2, the HCDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 28 or SEQ ID NO: 37; and HCDR3, the HCDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 6 or SEQ ID NO: 29; and

[0454] b) The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 24 or SEQ ID NO: 35; LCDR2, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 25 or SEQ ID NO: 36; and LCDR3, which comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 3 or SEQ ID NO: 26.

[0455] 77. A method for preventing or treating a reduction in peritubular capillary fenestrations associated with kidney disease, condition, or injury in a subject, wherein the kidney disease, condition, or injury is associated with kidney fibrosis, the method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, wherein the antibody or antigen-binding fragment thereof comprises:

[0456] a) The complementarity-determining region (CDR) of the heavy chain variable region, wherein the CDR comprises: the HCDR1 amino acid sequence of SEQ ID NO: 4, the HCDR2 amino acid sequence of SEQ ID NO: 5, and the HCDR3 amino acid sequence of SEQ ID NO: 6; and

[0457] b) The CDR of the light chain variable region, wherein the CDR comprises: the LCDR1 amino acid sequence of SEQ ID NO: 1, the LCDR2 amino acid sequence of SEQ ID NO: 2, and the LCDR3 amino acid sequence of SEQ ID NO: 3.

[0458] 78. A method for preventing or treating a reduction in peritubular capillary fenestrations associated with kidney disease, condition, or injury in a subject, wherein the kidney disease, condition, or injury is associated with kidney fibrosis, the method comprising administering an antibody or an antigen-binding fragment thereof to a subject in need, wherein the antibody or antigen-binding fragment thereof comprises:

[0459] a) The complementarity-determining region (CDR) of the heavy chain variable region, said CDR comprising: the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 28, and the HCDR3 amino acid sequence of SEQ ID NO: 29; or the HCDR1 amino acid sequence of SEQ ID NO: 27, the HCDR2 amino acid sequence of SEQ ID NO: 37, and the HCDR3 amino acid sequence of SEQ ID NO: 29; and

[0460] b) The complementarity-determining region (CDR) of the light chain variable region, wherein the complementarity-determining region (CDR) comprises the LCDR1 amino acid sequence of SEQ ID NO: 24, the LCDR2 amino acid sequence of SEQ ID NO: 25, and the LCDR3 amino acid sequence of SEQ ID NO: 26; or the LCDR1 amino acid sequence of SEQ ID NO: 35, the LCDR2 amino acid sequence of SEQ ID NO: 36, and the LCDR3 amino acid sequence of SEQ ID NO: 26.

Claims

1. An antibody or an antigen-binding fragment thereof, said antibody or antigen-binding fragment thereof binding to ANGPT2 and via ANGPT2 binding to the TIE2 receptor, for the treatment or prevention of fibrotic diseases.

2. An antibody or an antigen-binding fragment thereof, said antibody or antigen-binding fragment thereof binding to ANGPT2 and via ANGPT2 binding to a TIE2 receptor, for treating or preventing a subject’s kidney disease, condition or injury, said kidney disease, condition or injury being associated with kidney fibrosis.

3. The antibody or antigen-binding fragment thereof for use according to claim 1 or 2, wherein the antibody or antigen-binding fragment thereof is characterized in that it binds to the binding sites from position 336 to position 434 of angiopoietin-2 (ANGPT2) (SEQ ID NO: 30).

4. The antibody or antigen-binding fragment thereof for use according to claim 3, wherein the length of the binding site is about 2 to about 20 consecutive amino acids, such as about 2 to about 18, about 5 to about 20, or about 10 to about 20, such as about 15 to about 20 consecutive amino acids; and / or wherein the antibody or antigen-binding fragment thereof is characterized in that it binds to the epitope of human angiopoietin-2 (ANGPT2) SEQ ID NO: 9 or SEQ ID NO:

30.

5. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof comprises: a complementarity-determining region (CDR) of a heavy chain variable region, the CDR comprising: HCDR1, the HCDR1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 27 or SEQ ID NO: 4; HCDR2, the HCDR2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 37, SEQ ID NO: 28 or SEQ ID NO: 5; and HCDR3, the HCDR3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 29 or SEQ ID NO: 6; and The complementarity-determining region (CDR) of the light chain variable region comprises: LCDR1, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 35, SEQ ID NO: 24, or SEQ ID NO: 1; LCDR2, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 36, SEQ ID NO: 25, or SEQ ID NO: 2; and LCDR3, which contains an amino acid sequence selected from the group consisting of SEQ ID NO: 26 or SEQ ID NO:

3.

6. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof comprises a variable heavy chain of SEQ ID NO: 33, SEQ ID NO: 10, SEQ ID NO: 31 or SEQ ID NO: 8, and a variable light chain of SEQ ID NO: 34, SEQ ID NO: 11, SEQ ID NO: 32, SEQ ID NO: 7 or SEQ ID NO:

38.

7. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody is a human subtype IgG1.

8. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof: a) Reduce or inhibit human angiopoietin-2 (ANGPT2); and / or b) Inducing TIE2 activation; and c) Inducing TIE2 phosphorylation.

9. The antibody or antigen-binding fragment thereof for use according to claim 8, wherein the induction of TIE2 activation is ANGPT2-dependent.

10. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein fibrosis is prevented or mitigated.

11. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the fibrosis is renal tubular interstitial fibrosis.

12. An antibody or antigen-binding fragment thereof for use according to any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is administered before, during, or after the onset of the fibrotic disease or kidney disease, condition, or injury.

13. The antibody or antigen-binding fragment thereof for use according to any one of claims 2 to 12, wherein the kidney disease, condition or injury is induced by renal capillary sparsity, spread by renal capillary sparsity and / or associated with renal capillary sparsity.

14. The antibody or antigen-binding fragment thereof for use according to any one of claims 2 to 13, wherein the kidney disease, condition or injury is induced by, spread by and / or associated with renal tubular injury.

15. The antibody or antigen-binding fragment thereof for use according to any one of claims 2 to 14, wherein the kidney disease, condition, or injury is selected from the group consisting of: chronic kidney disease, chronic kidney damage, chronic renal failure syndrome, glomerulonephritis, type I diabetes, type II diabetes, ischemic nephropathy, focal segmental glomerulosclerosis, nephrotic syndrome, cardiorenal syndrome, tubulointerstitial disease associated with metabolic disorders, minimal change disease, hypertensive nephropathy, malignant hyperplasia... Hypertensive nephropathy, IgA nephropathy, IgM nephropathy, mesangial capillary glomerulonephritis, crescentic glomerulonephritis, diffuse intracapillary proliferative glomerulonephritis, mesangial proliferative glomerulonephritis, focal and segmental proliferative glomerulonephritis, systemic vasculitis, pulmonary hemorrhage nephritis syndrome, Dent disease, systemic lupus erythematosus glomerulonephritis syndrome, allergic purpura, systemic sclerosis, scleroderma, nephropathy associated with urinary tract obstruction, tubulointerstitial nephritis, interstitial nephritis, and lithium nephropathy.

16. A pharmaceutical composition comprising an antibody or an antigen-binding fragment thereof according to any one of the preceding claims for treating or preventing kidney disease, wherein the kidney disease, condition, or injury is associated with kidney fibrosis.

17. A pharmaceutical composition comprising an antibody or an antigen-binding fragment thereof according to any one of the preceding claims, for the treatment or prevention of fibrotic diseases.

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