PSMP antagonists for use in treatment of fibrosis disease in lung, kidney or liver

Abstract

To provide PSMP antagonists for use in treatment of fibrosis disease in lung, kidney or liver.SOLUTION: Disclosed are antagonists of PC3-secreted microprotein (PSMP) and use of the antagonists for treatment of liver, lung or kidney fibrosis, including various diseases or disorders associated with liver, lung or kidney fibrosis, e.g., non-alcoholic fatty liver disease (NAFLD), alcoholic liver disease (ALD), primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC), drug-induced lung injury, acute kidney injury (AKI), chronic kidney disease (CKD), lupus nephritis, IgA nephropathy, and membranous glomerulonephritis. Also disclosed are PSMP antagonists and use thereof for treatment of graft-versus-host disease (GVHD) and systemic lupus erythematosus (SLE).SELECTED DRAWING: None

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Patent Application No. 62 / 797,440, filed January 28, 2019, U.S. Provisional Patent Application No. 62 / 911,511, filed October 7, 2019, and U.S. Provisional Patent Application No. 62 / 913,937, filed October 11, 2019. Each of the foregoing applications is incorporated herein by reference in its entirety. Sequence Listing Reference

[0002] This application has been submitted in ASCII format via EFS-Web and contains a Sequence Listing, which is incorporated herein by reference in its entirety. The ASCII copy, created on December 26, 2019, is entitled "MAB_0110PC_20191226_Seq_Listing_ST25" and is 11,153 bytes in size. [Background technology]

[0003] Liver fibrosis and nonalcoholic fatty liver disease (NAFLD) Fibrosis is the excessive accumulation of extracellular matrix, which often occurs as a wound healing response to repeated or chronic tissue injury and can lead to the destruction of organ structure and loss of function. Fibrosis affects almost every tissue in the body. The mechanism of fibrosis resolution involves the degradation of the fibrous extracellular matrix and the disappearance of fibrogenic myofibroblasts.

[0004] Liver fibrosis, a wound-healing response to chronic liver injury, is characterized by excessive deposition of extracellular matrix (ECM) in the liver, which can be induced by various causes, including hepatitis virus infection, alcohol abuse, nonalcoholic steatohepatitis (NASH), or primary biliary cholangitis, leading to loss of liver function and destruction of liver structure. Recent evidence of resolution of organ fibrosis in humans has been observed in the liver. Patients with liver fibrosis associated with hepatitis B virus (HBV) or HCV infection who received antiviral therapy have shown recovery from fibrosis and histological improvement, even in cases of cirrhosis.

[0005] Nonalcoholic fatty liver disease (NAFLD) has emerged as a major cause of chronic liver disease. NAFLD represents a complex spectrum of liver diseases ranging from benign hepatic steatosis to its more aggressive necroinflammatory manifestation, NASH. NASH is distinct from other liver diseases because it is closely associated with metabolic syndrome comorbidities, such as insulin resistance and type 2 diabetes, as well as cardiovascular complications related to hypertension and dyslipidemia. These comorbidities are already present in simple NAFLD, which is defined by the absence of fibrosis and primary hepatic inflammation. These non-liver-involving diseases are the major comorbidities and cause of mortality from NAFLD and early fibrotic NASH up to stage 2 fibrosis. Liver-related morbidity and mortality increase significantly only beyond stage 1 fibrosis, particularly once cirrhosis appears. Therefore, prevention and treatment must address two largely independent targets: metabolic complications, which are treatable by various medications, and moderate to advanced fibrosis (stages 2–4), for which no approved drugs exist.

[0006] Common experimental rodent models of liver fibrosis include the administration of hepatotoxins (e.g., carbon tetrachloride [CCl4]) to induce acute hepatocellular injury or bile duct ligation (BDL) to induce cholestasis, resulting in pericentral or periportal liver fibrosis, respectively. NASH models are essentially differentiated by their ability to mimic the pathogenesis and / or natural history (obesity-induced diet models) or histopathological examination (nutrient-deficient diet models or chemically induced models). The methionine- and choline-deficient (MCD) diet is one of the most commonly used diets, producing the most severe phenotype of NASH in the shortest time. The MCD diet induces significant fibrosis compared to other dietary animal models. Genetic models (monogenic or polygenic) are also widely used in NASH research.

[0007] The initiation of fibrosis is critically dependent on an inflammatory phase, during which liver-resident macrophages and Kupffer cells are activated and release transforming growth factor-β (TGF-β) and other proinflammatory cytokines that activate hepatic stellate cells (HSCs). HSCs are responsible for the production of most of the extracellular matrix (ECM) and play a central role in liver fibrogenesis. HSCs are quiescent and reside in the space between hepatocytes and the sinusoidal endothelium (the space of Disse) as retinoid-storing cells. Upon liver injury, HSCs, the primary collagen-synthesizing cells in the liver, are activated, proliferate faster, and transdifferentiate into myofibroblast-like cells that exhibit enhanced chemotaxis, survival, and collagen production. HSC activation is driven by multiple mediators, including chemokines, reactive oxygen species, growth factors, matrix stiffness, matricellular proteins, and damage-associated molecular patterns. Alcoholic Liver Disease (ALD)

[0008] Alcoholic liver disease (ALD) affects millions of patients worldwide each year. The progression of ALD is well characterized and is in fact a continuum of liver disease ranging from steatosis, inflammation and necrosis (steatohepatitis), to fibrosis and cirrhosis, and ultimately, in some cases, hepatocellular carcinoma (HCC). Fatty liver (steatosis) is the initial stage of the hepatic response to binge drinking or chronic ethanol consumption. Accumulation of lipid products, such as triglycerides, in hepatocytes leads to lipid superoxidation and oxidative stress, resulting in apoptosis, liver inflammation, and HSC activation.

[0009] Alcohol-induced fatty liver is accompanied by increased hepatic sterol regulatory element-binding protein (SREBP)-1 and decreased hepatic peroxisome proliferator-activated receptor (PPAR)α activity. SREBP1 is primarily expressed in the liver and is a key transcriptional regulator of cholesterol, fatty acid, and triglyceride biosynthesis. PPARα functions as a master regulator of hepatic lipid metabolism.

[0010] Currently, the most widely used model of alcoholic liver disease involves ad libitum feeding of mice with an ethanol-containing Lieber-DeCarli liquid diet for 4–6 weeks. This model can induce hepatic steatosis and minimal fibrosis. Additionally, a chronic and excessive alcohol-feeding model, also known as the National Institute on Alcohol Abuse and Alcoholism (NIAAA) model, has been reported. See, for example, Bertola et al., Nat. Protoc. 8: 627–37, 2013. The NIAAA model mimics the acute exacerbation of chronic alcoholic liver injury in patients with longstanding chronic alcohol use (chronic) and a recent history of excessive alcohol consumption (binge), similar to the drinking patterns of many patients with alcoholic hepatitis. The protocol for the chronic excessive alcohol feeding model is chronic oral feeding (e.g., 10 days) of a Lieber-DeCarli ethanol liquid diet ad libitum plus a single excessive ethanol feeding (e.g., gavage of a single dose of ethanol, 5 g per kg body weight, 31.5% ethanol). Such chronic excessive ethanol feeding synergistically induced significant steatosis, liver injury, and inflammation in mice. See, e.g., Bertola et al., supra. Primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC)

[0011] Hepatic cholestasis is characterized by defects in bile flow. Among cholestatic diseases, primary sclerosing cholangitis and primary biliary cholangitis are relevant causes of chronic liver disease and are associated with significant morbidity and mortality.

[0012] Primary sclerosing cholangitis (PSC) is a rare chronic cholestatic liver disease of uncertain etiology characterized biochemically by cholestasis and histologically and cholangiographically by fibro-obstructive inflammation of the bile ducts. PSC progresses to cirrhosis, end-stage liver disease, and / or hepatobiliary cancer in a clinically significant proportion of patients, but the disease course can be highly variable.

[0013] Primary biliary cholangitis (PBC), formerly known as primary biliary cirrhosis, is a chronic disease in which the small bile ducts in the liver become injured, inflamed, and eventually destroyed. In the absence of bile ducts, bile builds up, causing liver damage.

[0014] Chronic feeding of 3,5-diethoxycarbonyl-1,4-dihydrocollidine, termed DDC, has been proposed as an in vivo model of cholestatic diseases such as primary sclerosing cholangitis (PSC) and primary biliary cholangitis / primary biliary cirrhosis (PBC) due to the formation of intraductal porphyrin plugs. In these diseases, similar to that in DDC, primary injury is directed at cholangiocytes. Chronic feeding of DDC in mice recapitulates key histopathological features of human cholestatic diseases, including (1) remodeling of bile duct compartments resulting in a ductular reaction, (2) pericanalicular fibrosis, and (3) inflammatory infiltrates. pulmonary fibrosis

[0015] Pulmonary fibrosis is associated with diverse etiologies, including scleroderma (systemic sclerosis), sarcoidosis, infections, and exposure to toxic substances or radiation. Idiopathic pulmonary fibrosis (IPF), the most common form of idiopathic interstitial pneumonia, is usually fatal, with a median survival of 2–3 years. In 2014, the FDA granted fast-track approval for the pro-fibrotic signaling inhibitors pirfenidone and nintedanib for the treatment of IPF based on their slowing of decline in lung function as measured by forced vital capacity and reduction in all-cause mortality. However, the efficacy of these drugs in promoting fibrosis resolution in IPF has not been demonstrated. The most common experimental mouse model of pulmonary fibrosis is intratracheal administration of the chemotherapy drug bleomycin, which induces inflammation followed by fibrosis. Various studies have found that the majority of myofibroblasts in fibrotic lungs are derived from pericytes, with the contribution of mesothelial cells through mesothelial-mesenchymal transition (MMT). Graft-versus-host disease (GVHD)

[0016] Graft-versus-host disease (GVHD) is a common complication following human allogeneic hematopoietic stem cell transplantation (allo-HSCT) and is considered a major obstacle to recovery from HSCT. Many research teams have reported that monocytes and macrophages are attracted to and accumulate in the intestinal mucus, liver, and skin during GVHD, exacerbating the disease. See, for example, Zhang et al., J. Leukoc. Biol. 99: 279-87, 2016. Various mouse models of GVHD have been established to help understand the progression of the disease and to identify effective methods for reducing the prevalence of GVHD. See, for example, Schroeder et al., Dis. Model Mech. 4: 318-333, 2011.

[0017] Clinically, GVHD is defined into two subtypes based on the time of symptom onset: acute GVHD and chronic GVHD. Acute GVHD is the leading cause of death after HSCT, and symptoms include skin rash, gastrointestinal damage, and liver damage, usually occurring within 100 days after HSCT. Chronic GVHD symptoms occur 100 days to several years later and may be limited to a single organ or site within the body or may be widespread. Symptoms may affect any of the following: eyes, mouth, nails, skin, scalp and hair, gastrointestinal tract, lungs, liver, muscles and joints, and genitals. GVHD is classified into grade I and grade II or higher GVHD depending on the severity of acute GVHD. Grade I refers to cutaneous GVHD covering 50 percent or less of the body surface area without liver or gastrointestinal tract involvement, and treatment involves the use of local treatments (e.g., topical steroids) and optimization of preventive measures (e.g., cyclosporine levels). Patients with more severe disease are considered to have grade II or higher GVHD and are typically treated with systemic glucocorticoids (e.g., methylprednisolone). Oral beclomethasone is suggested for use in patients with gastrointestinal involvement, but should be discontinued if gastrointestinal infection is present. Zeiser et al., N. Engl. J. Med. 377: 2167-2179, 2017;Zeiser et al., N. Engl. See J. Med. 377: 2565-2579, 2017.

[0018] Treatment strategies for acute GVHD, based on early data from clinical trials, include the use of costimulatory pathway blockade, targeted anti-interleukin-6 monoclonal antibodies, histone deacetylase inhibitors, kinase inhibitors, proteasome inhibitors, the anti-inflammatory protease inhibitor alpha-1-antitrypsin, CTLA-4 antagonism, CCR5 blockade, and adoptive Treg transfer. These and other recent strategies in development must be tested in prospective phase 3 trials before they become standard of care for acute GVHD. Zeiser See, e.g., et al., N. Engl. J. Med. 377: 2167-2179, 2017.

[0019] For decades, little progress has been made in treating chronic GVHD, and no drugs have been approved by the Food and Drug Administration (FDA) for the treatment of glucocorticoid-dependent or glucocorticoid-refractory patients. Recent advances have been made by reducing the burden of alloreactive T cells and antibody-producing B cells (e.g., by posttransplant treatment with cyclophosphamide, depletion of naive T cells, and depletion of B cells with rituximab). New therapeutic approaches are based on a better understanding of the pathogenesis of chronic GVHD, particularly the prominent roles of B cell signaling and persistent immune activation of certain T cell subsets, Treg cell deficiency, and tissue fibrosis. Prospects include targeting fibrosis and stimulating mechanisms (e.g., using pirfenidone and interleukin-17 or RORγt inhibitors), targeting plasma cells (e.g., using immunoproteasome inhibitors and anti-interleukin-6 receptors), and inhibiting chemokine-induced T cell and B cell recruitment to target organs affected by chronic GVHD identified in biomarker studies (e.g., using CXCL9 inhibitors). Although these developments are promising, glucocorticoids still constitute the standard frontline treatment, despite substantial side effects with long-term use. See Zeiser et al., N. Engl. J. Med. 377: 2565-2579, 2017. Acute Kidney Injury (AKI) and Chronic Kidney Disease (CKD)

[0020] Kidney disease is increasingly recognized as a global health problem. See, e.g., Felix et al., Nat. Rev. Nephrology. 15: 263-27, See, e.g., Yin-Wu Bao et al., Zoological Research. 39: 72-86, 2018. Acute kidney injury (AKI) and chronic kidney disease (CKD) are associated with high morbidity and mortality. AKI is considered a rapid and reversible decline in renal function and is associated with accelerated CKD. Compared with patients without a history of AKI or CKD, patients with AKI are more likely to develop new CKD or end-stage renal disease (ESRD). Conversely, CKD also plays an important role in AKI. Patients with CKD may be at high risk for transient declines in renal function consistent with AKI. See, e.g., Yin-Wu Bao et al., Zoological Research. 39: 72-86, 2018. AKI and CKD are closely linked and are therefore considered an integrated clinical syndrome. See, e.g., Chawla et al., N. Engl. See J. Med. 371: 58-66, 2014.

[0021] The mechanisms of AKI and CKD disease development and progression are still not fully understood. For example, John et al., J. Clin. Invest. 124: 2294-2298, 2014. Thus, few strategies are available for slowing the progression of kidney disease, with the majority of treatment options focused on lowering blood pressure and reducing proteinuria. Clearly, additional therapeutic avenues are needed. Generally, see, e.g., Siew et al., Kidney International. 87: 46-61, 2015.

[0022] Rhabdomyolysis is a severe syndrome caused by skeletal muscle injury and the subsequent release of breakdown products from damaged muscle cells into the systemic circulation. Muscle injury often results from strenuous exercise, muscle hypoxia, drug administration, or drug abuse and can lead to life-threatening complications such as AKI. See, e.g., Koshu et al., Nat. Med. 24: 232-238, 2018. Experimental AKI induced by glycerol injection is a well-established model of rhabdomyolysis. It is characterized by severe cortical acute tubular necrosis and inflammatory cell infiltration. Generally, see, e.g., Yanqiu et al., Stem Cell Research & Therapy 5: 80, 2014.

[0023] Renal fibrosis is an important pathological phenomenon in CKD that contributes to the progressive loss of renal function. Renal fibrosis is accompanied by glomerular sclerosis and / or interstitial fibrosis. Ectopic and excessive deposition of extracellular matrix (ECM) proteins in both glomeruli and interstitial regions is a typical feature of renal fibrosis, thereby amplifying the severity of renal injury.

[0024] Obstructive uropathy is a major cause of end-stage renal disease in children and one of the leading reasons for pediatric kidney transplantation. The most common rodent model used to study AKI and CKD is unilateral ureteral obstruction (UUO), and key features of UUO are interstitial inflammation, tubular cell injury / death, and fibrosis as a result of obstructed urine flow. See, e.g., Elena Martinez-Klimova et al., Biomolecules 9: 141, 2019.

[0025] Systemic lupus erythematosus (SLE) is a systemic autoimmune disease that occurs when the body's immune system attacks its own tissues and organs. Inflammation caused by SLE can affect many different body systems. Renal failure is one of the leading causes of death in people with SLE. Lupus nephritis is a common and severe manifestation of systemic lupus erythematosus (SLE) and is an important cause of both acute kidney injury and end-stage renal disease. See, e.g., Davidson et al., Nat. Rev. Rheumatol, 6: 13-20, 2010. Lupus nephritis is initiated by glomerular deposition of immune complexes, which triggers a cascade of inflammatory events, including Fc receptor and complement activation, recruitment of inflammatory cells, and eventual fibrosis. See, e.g., Anne Davidson, Nat. Rev. Rheumatol. 12: 143-153, 2015. Current approaches to managing lupus nephritis rely on high-dose corticosteroids plus broad-spectrum immunosuppressants. See, e.g., Feng et al., Nat. Rev. Nephrology. 13: 483-495, 2017; Samir et al., J. Am. Soc. Nephrol. 27: 2929-2939, 2016. A well-known mouse model of lupus nephritis utilizes inoculation of female B6D2F1 mice with T lymphocytes from male DBA / 2 mice to induce an immunostimulatory GVH response and lupus-like disease in mice exhibiting symptoms similar to those of SLE patients. See, e.g., Via et al., J. Immunol. 139: 1840-1849, 1987.

[0026] IgA nephropathy, identified 50 years ago by Jacques Berger and once thought to be a rare variant of mesangial proliferative glomerular disease, is the most common glomerular disease worldwide and an important cause of chronic kidney disease and end-stage renal failure. The pathogenic mechanisms of IgA nephropathy are only partially understood. Data from clinical and basic research have suggested multiple hypotheses, including recognition of galactose-deficient IgA1 molecules by specific autoantibodies, resulting in the formation of IgA1-IgG immune complexes deposited in the glomerular mesangium, where they induce renal injury. For example, Jennifer et al. See, e.g., Wang et al., Clin. J. Am. Soc. Nephrol. 12: 677-686, 2017. IgA nephropathy is recognized as an immune complex disease. PC3 secreted microprotein

[0027] PSMP, or PC3 secretory microprotein, was first discovered in PC3 cells and benign and malignant prostate tissues (see Valtonen-Andre et al., Biol. Chem. 388: 289-95, 2007). PSMP is also called prostate-associated microseminoprotein (MSMP) (see Frankenberg et al., BMC Evol. Biol. 11: 373-85, 2011). Apart from its expression in PC3 cells and benign and malignant prostate tissues, the tissue expression of MSMP has been reported to be limited to testis as a member of the beta-microseminoprotein family (see the same document). Studies using omics strategies have revealed that PSMP is a novel chemotactic cytokine that acts as a CCR2 ligand to recruit peripheral blood monocytes and lymphocytes (see Pei et al., J. Immunol. 192: 1878-86, 2014). The affinity of PSMP for CCR2 has been found to be comparable to that of CCL2 for CCR2 (see ibid.).

[0028] Another study demonstrated that PSMP is expressed in human colitis tissue and is significantly upregulated in DSS-induced murine colitis (Pei et al., Sci. Rep. 7: 5107, 2017). PSMP plays a crucial role in promoting DSS colitis by chemoattracting Ly6Chi monocytes in a CCR2-dependent manner (ibid.). Anti-PSMP neutralizing antibodies attenuate colitis by reducing macrophage infiltration and inhibiting the expression of IL-6, TNFα, and CCL2 (ibid.). A recent study found that MSMP gene expression was substantially upregulated in tumors resistant to anti-VEGF therapy compared with control tumors. See Mitamura et al., Oncogene 37: 722-731, 2018. Hypoxia induced MSMP secretion from cancer cells. Histone acetylation under hypoxic conditions in cancer cells reduced recruitment of the transcriptional repressor CCCTC-binding factor (CTCF) to the MSMP enhancer region. MSMP siRNA delivered in vivo using DOPC nanoliposomes restored tumor sensitivity to anti-VEGF therapy. In ovarian cancer patients treated with bevacizumab, serum MSMP concentrations were significantly elevated only in non-responders. See Id. [Prior art documents] [Non-patent literature]

[0029] [Non-Patent Document 1] Bertola et al., Nat. Protoc. 8: 627-37, 2013 [Non-patent document 2] Zhang et al., J. Leukoc. Biol. 99: 279-87, 2016 [Non-patent document 3] Schroeder et al., Dis. Model Mech. 4: 318-333, 2011 [Non-patent document 4] Zeiser et al., N. Engl. J. Med. 377: 2167-2179, 2017 [Non-patent document 5] Zeiser et al., N. Engl. J. Med. 377: 2565-2579, 2017 [Non-patent document 6] Felix et al., Nat. Rev. Nephrology. 15: 263-27, 2019 Summary of the Invention [Means for solving the problem]

[0030] In one aspect, the present invention provides a method for treating liver fibrosis. The method generally comprises administering an effective amount of a PC3-secreted microprotein (PSMP) antagonist to a subject with liver fibrosis. In a related aspect, the present invention provides a PSMP antagonist for use in treating liver fibrosis. In another related aspect, the present invention provides the use of a PSMP antagonist in the manufacture of a medicament for treating liver fibrosis. In some embodiments of the above methods, PSMP antagonists, or uses, the liver fibrosis has progressed to cirrhosis. In other, non-mutually exclusive, embodiments, the liver fibrosis is hepatitis B virus (HBV)-induced liver fibrosis, hepatitis C virus (HCV)-induced liver fibrosis, or alcohol-induced liver fibrosis, or the liver fibrosis is associated with non-alcoholic fatty liver disease. In some variations in which the liver fibrosis has progressed to cirrhosis, the cirrhosis is primary biliary cirrhosis. In some variations, where the liver fibrosis is associated with non-alcoholic fatty liver disease, the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH). In some embodiments, the liver fibrosis is associated with a disease or disorder selected from alcoholic liver disease (ALD), alcoholic hepatitis, alcoholic cirrhosis, chronic hepatitis B, chronic hepatitis C, chronic hepatitis D, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), hemochromatosis, cystic fibrosis, Wilson's disease, biliary atresia, alpha 1 antitrypsin deficiency, galactosemia, glycogen storage disease, inherited digestive disorders, Alagille syndrome, autoimmune hepatitis, primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC, formerly known as primary biliary cirrhosis), infection, drug-induced liver injury, and Budd-Chiari syndrome.

[0031] In another aspect, the present invention provides a method of treating pulmonary fibrosis. The method generally comprises administering an effective amount of a PC3-secreted microprotein (PSMP) antagonist to a subject with pulmonary fibrosis. In a related aspect, the present invention provides a PSMP antagonist for use in treating pulmonary fibrosis. In another related aspect, the present invention provides the use of a PSMP antagonist in the manufacture of a medicament for treating pulmonary fibrosis. In some embodiments, the pulmonary fibrosis is associated with a disease or disorder selected from the group consisting of dermatomyositis, polymyositis, mixed connective tissue disease, systemic lupus erythematosus, rheumatoid arthritis, sarcoidosis, scleroderma, pneumonia, chronic radiation pneumonitis, pneumoconiosis, infectious disease, and drug-induced lung injury.

[0032] In another aspect, the present invention provides a method for treating non-alcoholic fatty liver disease. The method generally comprises administering an effective amount of a PC3 secreted microprotein (PSMP) antagonist to a subject with non-alcoholic fatty liver disease. In a related aspect, the present invention provides a PSMP antagonist for use in treating non-alcoholic fatty liver disease. In another related aspect, the present invention provides the use of a PSMP antagonist in the manufacture of a medicament for treating non-alcoholic fatty liver disease. In some embodiments of the above methods, PSMP antagonists, or uses, the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH).

[0033] In another aspect, the present invention provides a method of treating alcoholic liver disease (ALD). The method generally involves administering to a subject with alcoholic liver disease an effective amount of a PC3 secreted microprotein (PSMP) antagonist. In a related aspect, the present invention provides a PSMP antagonist for use in treating alcoholic liver disease. In another related aspect, the present invention provides the use of a PSMP antagonist in the manufacture of a medicament for treating alcoholic liver disease.

[0034] In another aspect, the present invention provides a method for treating primary sclerosing cholangitis (PSC). The method generally comprises administering to a subject with primary sclerosing cholangitis an effective amount of a PC3-secreted microprotein (PSMP) antagonist. In a related aspect, the present invention provides a PSMP antagonist for use in treating primary sclerosing cholangitis. In another related aspect, the present invention provides use of a PSMP antagonist in the manufacture of a medicament for treating primary sclerosing cholangitis.

[0035] In another aspect, the present invention provides a method of treating primary biliary cholangitis (PBC). The method generally comprises administering to a subject with primary biliary cholangitis an effective amount of a PC3-secreted microprotein (PSMP) antagonist. In a related aspect, the present invention provides a PSMP antagonist for use in treating primary biliary cholangitis. In another related aspect, the present invention provides use of a PSMP antagonist in the manufacture of a medicament for treating primary biliary cholangitis.

[0036] In another aspect, the present invention provides methods of treating graft-versus-host disease (GVHD). The methods generally involve administering to a subject with GVHD an effective amount of a PC3-secreted microprotein (PSMP) antagonist. In a related aspect, the present invention provides a PSMP antagonist for use in treating GVHD. In another related aspect, the present invention provides the use of a PSMP antagonist in the manufacture of a medicament for treating GVHD. In some embodiments, the GVHD being treated is acute GVHD (aGVHD). In other embodiments, the GVHD being treated is chronic GVHD (cGVHD).

[0037] In another aspect, the present invention provides a method of treating systemic lupus erythematosus (SLE). The method generally comprises administering to a subject with SLE an effective amount of a PC3-secreted microprotein (PSMP) antagonist. In a related aspect, the present invention provides a PSMP antagonist for use in treating SLE. In another related aspect, the present invention provides the use of a PSMP antagonist in the manufacture of a medicament for treating SLE.

[0038] In another aspect, the present invention provides a method of treating lupus nephritis. The method generally comprises administering to a subject with lupus nephritis an effective amount of a PC3-secreted microprotein (PSMP) antagonist. In a related aspect, the present invention provides a PSMP antagonist for use in treating lupus nephritis. In another related aspect, the present invention provides the use of a PSMP antagonist in the manufacture of a medicament for treating lupus nephritis.

[0039] In another aspect, the present invention provides a method for treating renal fibrosis. The method generally comprises administering to a subject with renal fibrosis an effective amount of a PC3 secreted microprotein (PSMP) antagonist. In a related aspect, the present invention provides a PSMP antagonist for use in treating renal fibrosis. In another related aspect, the present invention provides the use of a PSMP antagonist in the manufacture of a medicament for treating renal fibrosis. In some embodiments of the above-described methods, PSMP antagonists, or uses, the renal fibrosis is associated with a disease or disorder selected from IgA nephropathy, membranoproliferative glomerulonephritis, membranous glomerulonephritis, crescentic glomerulonephritis, diabetic nephropathy, hypertensive nephropathy, lupus nephritis, hepatic nephropathy, polycystic kidney disease, Alport syndrome, Fabry disease, primary hyperoxaluria, cystinosis, focal segmental glomerulosclerosis (FSGS) caused by coenzyme Q10-related gene mutations, complement 3 glomerulonephritis, acute or subacute immune complex glomerulonephritis, renal vasculitis, systemic lupus erythematosus (SLE), recent-onset renal artery stenosis (fibromuscular or vasculitic), diabetic kidney disease, chronic urate nephropathy, toxic nephropathy, bacterial pyelonephritis, viral nephropathy, multiple myeloma, and obstructive nephropathy.

[0040] In yet another aspect, the present invention provides methods for treating acute kidney injury (AKI) or chronic kidney disease (CKD). The methods generally involve administering an effective amount of a PC3-secreted microprotein (PSMP) antagonist to a subject with AKI or CKD. In a related aspect, the present invention provides a PSMP antagonist for use in treating AKI or CKD. In another related aspect, the present invention provides use of a PSMP antagonist in the manufacture of a medicament for treating AKI or CKD. In some embodiments of the above methods, PSMP antagonists, or uses for treating AKI, the AKI is rhabdomyolysis-induced. In some embodiments of the above-described methods, PSMP antagonists, or uses for treating CKD, the CKD is caused by a disease or disorder selected from IgA nephropathy, membranoproliferative glomerulonephritis, membranous glomerulonephritis, crescentic glomerulonephritis, diabetic nephropathy, hypertensive nephropathy, lupus nephritis, hepatic nephropathy, polycystic kidney disease, Alport syndrome, Fabry disease, primary hyperoxaluria, cystinosis, focal segmental glomerulosclerosis (FSGS) caused by coenzyme Q10-related gene mutations, complement 3 glomerulonephritis, acute or subacute immune complex glomerulonephritis, renal vasculitis, systemic lupus erythematosus (SLE), recent-onset renal artery stenosis (fibromuscular or vasculitic), diabetic kidney disease, chronic uric acid nephropathy, toxic nephropathy, bacterial pyelonephritis, viral nephropathy, multiple myeloma, and obstructive nephropathy.

[0041] In certain embodiments of the above-described methods, PSMP antagonists, or uses, the PSMP antagonist is a soluble protein that specifically binds to PSMP. Particularly suitable soluble protein antagonists include neutralizing anti-PSMP antibodies. In more specific variations, the antibody is a humanized antibody, a chimeric antibody, or a human antibody. In other, non-mutually exclusive, embodiments, the antibody is a single-chain antibody and / or a bispecific antibody. Typically, antibodies for use according to the present invention are monoclonal antibodies. In some variations, the antibody comprises an immunoglobulin constant region, such as an immunoglobulin heavy chain constant region (e.g., an immunoglobulin Fc region).

[0042] In some embodiments, a PSMP antagonist that is a soluble protein (e.g., a neutralizing anti-PSMP antibody) competes for binding to PSMP with an antibody comprising (i) a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and (ii) a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5. In some such embodiments in which the PSMP antagonist is an antibody, the antibody has a complementarity determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref is CDR-H3 of SEQ ID NO: 4 (e.g., CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref a set of VH CDRs with zero amino acid substitutions compared to CDR-H1, Abis CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab is CDR-H3 of SEQ ID NO: 4). Each VH CDR can be defined, for example, according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition of a CDR. In a specific variation, each VH CDR is defined according to the Chothia definition of a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ref has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref has the amino acid sequence set forth in residues 99-108 of SEQ ID NO: 4. In some such embodiments, the set of VH CDRs comprises CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and therefore has zero amino acid substitutions relative to CDR-H1 Ab has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab has the amino acid sequence set forth in residues 99-108 of SEQ ID NO: 4. In some variations, the antibody comprises a humanized VH domain derived from a VH domain having the amino acid sequence set forth in SEQ ID NO: 4. In other embodiments, the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO: 4 (e.g., the antibody is a chimeric antibody comprising a VH domain of SEQ ID NO: 4).

[0043] In other, non-mutually exclusive embodiments, in which the PSMP antagonist is an antibody that competes for binding to PSMP with an antibody comprising (i) a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and (ii) a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5, the antibody has a complementarity determining region (CDR) CDR-L1 Ab , CDR-L2 Ab , and CDR-L3Ab and the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref is the CDR-L1 of SEQ ID NO: 5 (e.g., CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref a set of VL CDRs having zero amino acid substitutions compared to CDR-L1; Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab is CDR-L3 of SEQ ID NO: 5). Each VL CDR can be defined, for example, according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition of a CDR. In a specific variation, each VL CDR is defined according to the Chothia definition of a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref has the amino acid sequence set forth in residues 89-97 of SEQ ID NO: 5. In some such embodiments, the set of VL CDRs comprises CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and therefore has zero amino acid substitutions compared to CDR-L1 Ab has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Abhas the amino acid sequence set forth in residues 89-97 of SEQ ID NO: 5. In some variations, the antibody comprises a humanized VL domain derived from a VL domain having the amino acid sequence set forth in SEQ ID NO: 5. In other embodiments, the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO: 5 (e.g., the antibody is a chimeric antibody comprising the VL domain of SEQ ID NO: 5).

[0044] In some embodiments of the above methods, PSMP antagonists, or uses, the treatment is a combination therapy. definition

[0045] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the described methods and compositions pertain. As used herein, the following terms and phrases have the meanings ascribed to them unless specified otherwise.

[0046] The terms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

[0047] As used herein, the term "antagonist" refers to a compound that decreases the activity of another compound in a biological context.

[0048] Within the scope of this invention, a "PSMP antagonist" is a compound that reduces the receptor-mediated biological activity (e.g., chemotactic activity) of PSMP on target cells. Antagonists may exert their effect by competing with PSMP for binding sites on cell surface receptors, by binding to PSMP and preventing PSMP from binding to cell surface receptors, by otherwise interfering with receptor function, by reducing the production of PSMP, or by other means.

[0049] A "polypeptide" is a polymer of amino acid residues joined by peptide bonds, whether produced naturally or synthetically. Polypeptides of fewer than about 30 amino acid residues are commonly referred to as "peptides."

[0050] A "protein" is a macromolecule containing one or more polypeptide chains. Proteins may also contain non-peptide components, such as carbohydrate groups. Carbohydrates and other non-peptide substituents may be added to proteins by the cell that produces them, and this varies depending on the cell type. Proteins are defined herein in terms of their amino acid backbone structure; substituents such as carbohydrate groups are generally not specified, but may nevertheless be present.

[0051] The term "antibody" is used herein to refer to a protein produced by the body in response to the presence of an antigen and that binds to the antigen, as well as antigen-binding fragments and engineered variants thereof. Thus, the terms "antibody" and "antibodies" include polyclonal antibodies, affinity-purified polyclonal antibodies, monoclonal antibodies, and antigen-binding antibody fragments such as F(ab')2 and Fab fragments. Also included are genetically engineered intact antibodies and fragments, such as chimeric antibodies, humanized antibodies, single-chain Fv fragments, single-chain antibodies, diabodies, minibodies, linear antibodies, multivalent or multispecific hybrid antibodies, and the like. Thus, the term "antibody" is used broadly to include any protein that contains an antigen-binding site and is capable of binding to that antigen.

[0052] The term "engineered antibody" refers to an antibody whose amino acid sequence has been altered from that of a native (i.e., naturally occurring) antibody. Due to the relevance of recombinant DNA techniques in the generation of antibodies, there is no need to be limited to the sequence of amino acids found in natural antibodies; antibodies can be redesigned to obtain desired characteristics. The possible variations are numerous and range from changing just one or a few amino acids to completely redesigning, for example, the variable or constant regions. Changes in the constant region are generally made to improve or alter characteristics such as complement fixation, interaction with cells, and other effector functions. Typically, changes in the variable region are made to improve antigen-binding characteristics, improve the stability of the variable region, or reduce the risk of immunogenicity.

[0053] An "antigen-binding site" is a portion of an antibody that is sufficient for the antibody to bind to an antigen. The smallest such region is typically an immunoglobulin variable domain or fragment thereof, or an engineered variant of an immunoglobulin variable domain or fragment thereof. Single domain binding sites can be derived from camelid antibodies (see Muyldermans and Lauwereys, J. Mol. Recog. 12: 131-140, 1999; Nguyen et al., EMBO J. 19: 921-930, 2000) or from VH domains of other species to generate single domain antibodies ("dAbs"; Ward et al., Nature 341: 544-546, 1989; U.S. Patent No. 6,248,516 to Winter et al.) can be generated. In certain variations, the antigen-binding site is a polypeptide region having only two complementarity-determining regions (CDRs) of a naturally occurring or non-naturally occurring (e.g., mutagenized) heavy chain variable domain or light chain variable domain, or a combination thereof (e.g., see Pessi et al., Nature 362: 367-369, 1993; Qiu et al., Nature Biotechnol. 25: 921-929, 2007). More commonly, the antigen-binding site includes both a heavy chain variable domain and a light chain variable domain that bind to a common epitope. Within the scope of the present invention, in addition to an "antigen-binding site," an antibody may further comprise one or more additional components, such as, for example, one or more of a second antigen-binding site (which may bind to the same or a different epitope or the same or a different antigen), a peptide linker, an immunoglobulin constant domain, an immunoglobulin hinge, an amphipathic helix (see, e.g., Pack and Pluckthun, Biochem. 31: 1579-1584, 1992), a non-peptide linker, an oligonucleotide (see, e.g., Chaudri et al., FEBS Letters 450: 23-26, 1999), etc., and may be a monomeric or multimeric protein.In addition to proteins having the structure of an intact, native antibody (i.e., a tetramer consisting of two immunoglobulin heavy chains and two immunoglobulin light chains), examples of molecules that contain antigen-binding sites are generally known in the art and include, for example, Fv fragments, single-chain Fv fragments (scFv), Fab fragments, diabodies, minibodies, Fab-scFv fusions, bispecific (scFv)4-IgG, and bispecific (scFv)2-Fab (e.g., Hu et al., Cancer Res. 56: 3055-3061, 1996; Atwell et al., Molecular Immunology 33: 1301-1312, 1996; Carter and Merchant, Curr. Opin. Biotechnol. 8: 449-454, 1997; Zuo et al., Protein Engineering 13: 361-367, 2000; and Lu et al., J. Immunol. Methods 267: 213-226, 2002.).

[0054] As used herein, the term "immunoglobulin" refers to a protein consisting of one or more polypeptides substantially encoded by immunoglobulin gene(s). One form of immunoglobulin constitutes the basic structural unit of an intact, native antibody. This form is a tetramer, consisting of two identical pairs of immunoglobulin chains, each pair having one light chain and one heavy chain. In each pair, the light and heavy chain variable regions together are responsible for antigen binding, and the constant regions are responsible for antibody effector functions. Immunoglobulins typically function as antibodies in vertebrate organisms. Five classes of immunoglobulin proteins (IgG, IgA, IgM, IgD, and IgE) have been identified in higher vertebrates. IgG constitutes the major class; it is usually the second most abundant protein found in plasma. In humans, IgG consists of four subclasses, designated IgG1, IgG2, IgG3, and IgG4. The heavy chain constant region of the IgG class is identified using the Greek symbol γ. For example, immunoglobulins of the IgG1 subclass contain a γ1 heavy chain constant region. Each immunoglobulin heavy chain has a constant region consisting of constant region protein domains (CH1, hinge, CH2, and CH3; IgG3 also contains a CH4 domain) that are essentially invariant for a given subclass in a species. DNA sequences encoding human and non-human immunoglobulin chains are known in the art (e.g., Ellison et al., DNA 1: 11-18, 1981;Ellison et al., Nucleic Acids Res. 10: 4071-4079, 1982;Kenten et al., Proc. Natl. Acad. Sci. USA 79: 6661-6665, 1982;Seno et al., Nuc. Acids Res. 11: 719-726, 1983;Riechmann et al., Nature 332: 323-327, 1988;Amster et al., Nuc. Acids Res. 8: 2055-2065, 1980;Rusconi and Kohler, Nature 314: 330-334, 1985;Boss et al., Nuc. Acids Res. 12: 3791-3806, 1984;Bothwell et al., Nature 298: 380-382, 1982;van der Loo et al., Immunogenetics 42: 333-341, 1995;Karlin et al., J. Mol. Evol. 22: 195-208, 1985;Kindsvogel et al., (See DNA 1: 335-343, 1982; Breiner et al., Gene 18: 165-174, 1982; Kondo et al., Eur. J. Immunol. 23: 245-249, 1993; and GenBank Accession No. J00228.) For reviews of immunoglobulin structure and function, see Putnam, The Plasma Proteins, Vol V, Academic Press, Inc., 49-140, 1987; and Padlan, Mol. Immunol. 31: 169-217, 1994. The term "immunoglobulin" is used herein in its general sense to refer to an intact antibody, its component chains, or fragments of chains, depending on the context.

[0055] Full-length immunoglobulin "light chains" (approximately 25 Kd or 214 amino acids) are encoded by a variable region gene at the NH2-terminus (encoding approximately 110 amino acids) and by a kappa or lambda constant region gene at the COOH-terminus. Full-length immunoglobulin "heavy chains" (approximately 50 Kd or 446 amino acids) are encoded by a variable region gene (encoding approximately 116 amino acids) and a gamma, mu, alpha, delta, or epsilon constant region gene (encoding approximately 330 amino acids), the latter defining the antibody's isotype as IgG, IgM, IgA, IgD, or IgE, respectively. Within light and heavy chains, the variable and constant regions are joined by a "J" region of approximately 12 or more amino acids, with the heavy chain also including a "D" region of approximately 10 amino acids (see generally, Fundamental Immunology (Paul, ed., Raven Press, NY, 2nd ed. 1989), Ch. 7).

[0056] Immunoglobulin "Fv" fragments contain a heavy chain variable domain (VH) and a light chain variable domain (VL), which are held together by non-covalent interactions. Thus, immunoglobulin Fv fragments contain a single antigen-binding site. The dimeric structure of Fv fragments can be further stabilized by introducing disulfide bonds by mutagenesis (see Almog et al., Proteins 31: 128-138, 1998).

[0057] As used herein, the term "single-chain antibody" refers to an antibody having an antigen-binding site contained within a single polypeptide chain (e.g., both the variable region from the heavy chain and the variable region from the light chain within a single polypeptide chain). The term "single-chain Fv" refers to a single-chain antibody that contains both the variable region from the heavy chain and the variable region from the light chain, but lacks a constant region. Generally, a single-chain Fv further comprises a polypeptide linker between the VH and VL domains, which enables the single-chain Fv to form the desired structure that enables antigen binding. Single-chain antibodies are described, for example, in Pluckthun in The Pharmacology of Monoclonal Antibodies, vol. 113 (Rosenburg and Moore eds., Springer-Verlag, New York, 1994), pp. 269-315 (see also WIPO Publication WO 88 / 01649; U.S. Pat. Nos. 4,946,778 and 5,260,203; Bird et al., Science 242: 423-426, 1988.) Single-chain antibodies can also be bispecific and / or humanized.

[0058] A "Fab fragment" contains one light chain and the CH1 and variable regions of one heavy chain. The heavy chain of a Fab fragment cannot form disulfide bonds with another heavy chain molecule. A "Fab' fragment" contains one light chain and one heavy chain that contains more of the constant region between the CH1 and CH2 domains, and can therefore form interchain disulfide bonds between the two heavy chains to form a "F(ab')2 fragment" containing two light chains and two heavy chains.

[0059] The terms "Fc fragment" and "Fc region," as used herein, are synonymous and refer to the portion of an antibody responsible for binding to cellular antibody receptors and the complement component C1q. Fc stands for "fragment crystalline," a fragment of an antibody that readily forms protein crystals. The distinct protein fragments originally described by proteolytic digestion allow the overall general structure of immunoglobulin proteins to be defined. As originally defined in the literature, the Fc fragment consists of the disulfide-linked heavy chain hinge region and CH2 and CH3 domains. However, more recently, the term has been applied to a single chain consisting of CH3, CH2, and at least a portion of the hinge sufficient to form a disulfide-linked dimer with a second such chain. For a review of immunoglobulin structure and function, see Putnam, The Plasma Proteins, Vol. V (Academic Press, Inc., 1987), pp. 49-140; and Padlan, Mol. See Immunol. 31: 169-217, 1994. As used herein, the term Fc includes variants of the naturally occurring sequence.

[0060] An immunoglobulin light or heavy chain variable region consists of framework regions and three intervening hypervariable regions. The term "hypervariable region," also referred to herein as "complementarity-determining region" ("CDR"), refers to the amino acid residues of an antibody responsible for binding to an antigen. CDRs can be defined according to any of several known analytical methods. Examples of such methods include the Kabat definition, the Chothia definition, the AbM definition, and the contact definition. The Kabat definition is the standard for numbering antibody residues and is typically used to identify CDR regions. See, e.g., Johnson & Wu, Nucleic Acids Res. 28: 214-8, 2000. The Chothia definition is similar to the Kabat definition, but takes into account the location of certain structural loop regions. See, e.g., Chothia et al., J. Mol. Biol. 196: 901-17, 1986; Chothia et al., Nature 342: 877-83, 1989. The AbM definition uses a suite of integrated computer programs produced by the Oxford Molecular Group to model antibody structure. See, e.g., Martin et al., 1989, Proc. Natl. Acad. Sci. USA 86: 9268-9272, 1989; "AbM TMSee, "Ab Initio Protein Structure Prediction Using a Combined Hierarchical Approach," Oxford, UK; Oxford Molecular, Ltd. The AbM definition models the tertiary structure of an antibody from its primary sequence using a combination of knowledge databases and ab initio methods, such as those described by Samudrala et al., "Ab Initio Protein Structure Prediction Using a Combined Hierarchical Approach," in PROTEINS, Structure, Function and Genetics Suppl. 3:194-198, 1999. The contact definition is based on analysis of available complex crystal structures. See, e.g., MacCallum et al., J. Mol. Biol. 5:732-45, 1996. The CDRs L1, L2, and L3 of the VL domain are also referred to herein as CDR-L1, CDR-L2, and CDR-L3, respectively; the CDRs L1, L2, and L3 of the VH domain are also referred to herein as CDR-L1, CDR-L2, and CDR-L3, respectively. H1, H2, and H3 are also referred to herein as CDR-H1, CDR-H2, and CDR-H3, respectively.

[0061] "Framework regions" are variable domain residues other than hypervariable region residues (e.g., other than residues of CDRs defined by the Kabat, Chothia, AbM, or contact definitions of CDRs). The sequences of framework regions of different light or heavy chains are relatively conserved within a species. Thus, a "human framework region" is a framework region that is substantially identical (about 85% or more, usually 90-95% or more) to the framework regions of naturally occurring human immunoglobulins. The framework region of an antibody is the combined framework regions of the constituent light and heavy chains that serve to position and align the CDRs.

[0062] A "chimeric antibody" is an antibody in which the light and heavy chain genes are constructed, typically by genetic engineering, from immunoglobulin variable and constant region genes belonging to different species. For example, the variable segments of genes from a mouse monoclonal antibody can be joined with segments encoding human constant regions (e.g., human gamma 1 or gamma 3 heavy chain genes and human kappa light chain genes). Thus, therapeutic chimeric antibodies are typically hybrid proteins composed of variable or antigen-binding domains from a mouse antibody and constant domains from a human antibody, although other mammalian species can also be used. Specifically, chimeric antibodies are created by recombinant DNA technology in which all or part of the hinge and constant region of the immunoglobulin light chain, heavy chain, or both, are replaced with the corresponding regions from an immunoglobulin light or heavy chain of another animal. In this way, the antigen-binding portion of the parent monoclonal antibody is grafted onto the framework of an antibody from another species. Chimeric antibodies can be "covered" with a human-like surface, if desired, by replacing exposed residues, resulting in a "veneered antibody."

[0063] As used herein, the term "human antibody" refers to an antibody having an amino acid sequence that corresponds to the amino acid sequence of an antibody produced by a human and / or produced using any of the techniques for producing human antibodies known to those of skill in the art or disclosed herein. This definition of a human antibody encompasses antibodies that comprise at least one human heavy chain polypeptide or that comprise at least one human light chain polypeptide. One such example is an antibody that comprises a murine light chain and a human heavy chain polypeptide. Human antibodies can be produced using a variety of techniques known in the art. In one embodiment, human antibodies are selected from a phage library, where the phage library expresses human antibodies. See, e.g., Vaughan et al., Nature Biotechnology 14: 309-314, 1996; Sheets et al., Proc. Natl. Acad. Sci. USA 95: 6157-6162, 1998; Hoogenboom and Winter, J. Mol. Biol. 227: 381, 1991; Marks et al., J. Mol. Biol. 222: 581, 1991. Human antibodies can also be produced by immunizing animals into which human immunoglobulin loci have been transgenically introduced in place of endogenous loci, e.g., mice in which the endogenous immunoglobulin genes have been partially or completely inactivated. This technique is described, for example, in U.S. Patent Nos. 5,545,807; 5,545,806; 5,569,825; 5,625,126; 5,633,425; 5,661,016; and 7,041,870. Alternatively, human antibodies can be prepared by immortalizing human B lymphocytes that produce antibodies directed against target antigens (such B lymphocytes can be recovered from individuals or immunized in vitro). See, for example, Cole et al., "Monoclonal Antibodies and Cancer Therapy", Alan R. Liss, p. 77, 1985; Boerner et al., J. See Immunol. 147: 86-95, 1991; and U.S. Patent No. 5,750,373.

[0064] The term "humanized immunoglobulin" refers to an immunoglobulin comprising a human framework region and one or more CDRs derived from a non-human (e.g., mouse or rat) immunoglobulin. The non-human immunoglobulin providing the CDRs is referred to as the "donor," and the human immunoglobulin providing the framework is referred to as the "acceptor." Constant regions need not be present, but if present, they will be substantially identical to human immunoglobulin constant regions, i.e., at least about 85-90%, preferably about 95% or greater. Thus, to the greatest extent possible, all portions of a humanized immunoglobulin other than the CDRs are substantially identical to corresponding portions of native human immunoglobulin sequences. In some cases, humanized antibodies may retain non-human residues within the human variable region framework domains to enhance corresponding binding characteristics (e.g., if the antibody is humanized, framework mutations may be necessary to preserve binding affinity). A "humanized antibody" is an antibody comprising a humanized light chain immunoglobulin and / or a humanized heavy chain immunoglobulin. For example, a typical chimeric antibody, as defined above, is not encompassed by a humanized antibody, e.g., because the entire variable region of the chimeric antibody is non-human (or, in the case of a veneered antibody, substantially non-human).

[0065] A "bispecific" antibody is an antibody that has two different antigen-binding sites, each with a different specificity. Bispecific antibodies can be produced by a variety of methods, including, for example, chemical conjugation with a cross-linking agent, somatic cell fusion of two hybridoma lines (quadroma), and genetic engineering. Generally, see, for example, Sedykh et al., Drug Design, Development and Therapy 12: 195-208, 2018.

[0066] The term "diabody" refers to a small antibody fragment having two antigen-binding sites, which comprises a heavy chain variable domain (VH) and a light chain variable domain (VL) connected in the same polypeptide chain (VH-VL). By using a linker that is too short to allow pairing between the two domains on the same chain, these domains are forced to pair with complementary domains on another chain, creating two antigen-binding sites. Diabodies are described in more detail, for example, in EP 404,097; WO 93 / 11161; and Hollinger et al., Proc. Natl. Acad. Sci. USA 90: 6444-6448, 1993.

[0067] The term "minibody" as used herein refers to a polypeptide encoding only two complementarity-determining regions (CDRs) of a heavy chain variable domain or a light chain variable domain, or a combination thereof, whether naturally occurring or non-naturally occurring (e.g., mutagenized). Examples of minibodies are described, for example, by Pessi et al., Nature 362: 367-369, 1993; and Qiu et al., Nature Biotechnol. 25: 921-929, 2007.

[0068] The term "linear antibody" refers to the antibodies described in Zapata et al., Protein Eng. 8:1057-1062, 1995. Briefly, these antibodies comprise a pair of tandem Fd segments (VH-CH1-VH-CH1) that form a pair of antigen-binding regions. Linear antibodies can be bispecific or monospecific.

[0069] As used herein, the term "monoclonal antibody" refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the amino acid sequences of the individual antibodies comprising the population are identical except for possible minor naturally occurring mutations. Monoclonal antibodies are highly specific, being directed against a single antigenic site. Furthermore, in contrast to polyclonal antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a single determinant on the antigen. The modifier "monoclonal" indicates the character of the antibody as being obtained from a substantially homogeneous population of antibodies, and is not to be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies for use in accordance with the present invention can be produced by hybridoma methods, such as those first described by Kohler and Milstein (Nature 256:495, 1975), or by recombinant DNA methods, such as those described in U.S. Pat. No. 4,816,567. The monoclonal antibodies can also be isolated from phage libraries generated using the techniques described in McCafferty et al. (Nature 348: 552-554, 1990), for example.

[0070] References herein to "treating" or "treatment of" liver fibrosis, pulmonary fibrosis, or renal fibrosis include treatment for any of the various diseases or disorders associated with liver fibrosis, pulmonary fibrosis, or renal fibrosis. A disease or disorder is "associated" with liver fibrosis, pulmonary fibrosis, or renal fibrosis if the liver fibrosis, pulmonary fibrosis, or renal fibrosis is interpreted by a clinician as being part of the pathology of that disease or disorder (e.g., an indicator of disease progression and / or a cause of loss of liver, lung, or kidney function). "with." In this context, "pathology" includes fibrosis as either a predicted or actual pathological tissue change as part of disease or disorder progression; thus, a subject having a disease or disorder associated with liver fibrosis, pulmonary fibrosis, or renal fibrosis and receiving administration of a PSMP antagonist in accordance with the present disclosure may or may not have fibrosis as an actual physical manifestation of the disease or disorder at the time of treatment. Furthermore, references herein to treating liver fibrosis, pulmonary fibrosis, or renal fibrosis in a subject and characterizing liver fibrosis, pulmonary fibrosis, or renal fibrosis as "associated with" a disease or disorder mean that the subject being treated has or is at risk of developing the particular disease or disorder.

[0071] Unless the context clearly dictates otherwise, a general reference herein to a "disease or disorder" includes reference to liver fibrosis, pulmonary fibrosis, renal fibrosis, graft-versus-host disease (GVHD), or systemic lupus erythematosus (SLE), as well as a specific disease or disorder associated with liver fibrosis, pulmonary fibrosis, or renal fibrosis (e.g., non-alcoholic fatty liver disease (NAFLD), alcoholic liver disease (ALD), primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC), acute kidney injury (AKI), chronic kidney disease (CKD), lupus nephritis, IgA nephropathy, or membranous glomerulonephritis).

[0072] The term "alternative scaffold" refers to a non-antibody protein that can be diversified in one or more regions to generate one or more binding domains that specifically bind to a target molecule (e.g., PSMP). In some embodiments, the binding domain binds to the target molecule with similar specificity and affinity as an antibody. Exemplary alternative scaffolds include those derived from fibronectin (e.g., Adnectins™), β-sandwich (e.g., iMabs), lipocalin (e.g., Anticalins®), EETI-II / AGRP, BPTI / LACI-D1 / ITI-D2 (e.g., Kunitz domain), Protein A (e.g., Affibody®), ankyrin repeat (e.g., DARPins), gamma-B-crystallin / ubiquitin (e.g., Affilins), CTLD3 (e.g., Tetranectin), Fynomer, and Avimer. Additional information regarding alternative scaffolds is provided in Binz et al., Nat. Biotechnol. 23: 1257-1268, 2005; Skerra, Current Opin. in Biotech. 18: 295-304, 2007; and Silacci et al., J. Biol. Chem. 289: 14392-14398, 2014, each of which is incorporated by reference in its entirety. Alternative scaffold proteins engineered to function as PSMP antagonists (e.g., by specifically binding to and neutralizing PSMP) may be referred to herein as "alternative scaffold PSMP antagonists" or "PSMP antagonists based on alternative scaffolds."

[0073] An "inhibitory polynucleotide" is a DNA or RNA molecule that reduces or prevents the expression (transcription or translation) of a second (target) polynucleotide (e.g., a gene or mRNA encoding PSMP). Inhibitory polynucleotides include antisense polynucleotides, ribozymes, and external guide sequences. The term "inhibitory polynucleotide" further includes DNA and RNA molecules that encode the actual inhibitory species, such as DNA molecules that encode ribozymes.

[0074] The terms "treat" and "treatment" are used broadly to refer to therapeutic and prophylactic interventions that favorably alter a pathological condition.

[0075] The term "effective amount" of a composition, in the context of the treatments described herein, refers to an amount sufficient to produce any one or more beneficial or desired results. For prophylactic use, beneficial or desired results include eliminating or reducing the risk, alleviating the severity, or delaying the onset of a disease or disorder, including the biochemical, histological, and / or physical symptoms of the disease or disorder, its complications, and intermediate pathological phenotypes present during the development of the disease or disorder. For therapeutic use, beneficial or desired results include clinical results such as improving one or more symptoms of the disease or disorder or reducing the incidence thereof, reducing the dose of other medications required to treat the disease or disorder, enhancing the effect of another medication, delaying the progression of the disease or disorder, and / or improving organ function in a patient. For example, with respect to the treatment of liver fibrosis, pulmonary fibrosis, or renal fibrosis by administering a PSMP antagonist to a subject as described herein, an amount of such agent sufficient to delay the onset of, reduce the severity of, or reduce the progression of fibrosis in the subject, or sufficient to result in improved liver function, lung function, or renal function in the subject, is considered an "effective amount" of the PSMP antagonist.

[0076] An effective dosage can be administered in one or more administrations. An effective amount of a composition is administered in an "effective regime" according to the method of the present invention. The term "effective regime" refers to a combination of the amount of a composition administered and the frequency of administration appropriate for achieving treatment. For purposes of the present invention, an effective dosage of a composition is an amount sufficient to directly or indirectly achieve prophylactic or therapeutic treatment. As understood in clinical practice, an effective dosage of a composition may or may not be achieved in conjunction with another drug composition. Thus, an "effective amount" can be considered in the context of administering one or more therapeutic agents, and a single agent can be considered to be administered in an effective amount if the desired result can or is achieved in conjunction with one or more other agents.

[0077] The term "patient" or "subject," with respect to treatment of a disease or disorder described herein, includes mammals such as, for example, humans and other primates. The term also includes domestic animals, such as cattle, pigs, sheep, horses, dogs, and cats.

[0078] The term "combination therapy" refers to a therapeutic regimen that involves providing at least two separate treatments to achieve an indicated therapeutic effect. For example, combination therapy involves administering two or more chemically distinct active ingredients or agents, such as a PSMP antagonist (e.g., an anti-PSMP antibody) and another agent, such as another anti-inflammatory or anti-fibrotic agent. The separate treatments that make up the combination therapy can be delivered, for example, as simultaneous, overlapping, or sequential administration regimens. With respect to the administration of two or more chemically distinct agents, it is understood that the active ingredients can be administered as part of the same composition or as separate compositions. When administered as separate compositions, compositions containing different active ingredients can be administered at the same or different times, by the same or different routes, and using the same or different administration regimens, all as required by the particular situation and as determined by the attending physician.

[0079] Whenever embodiments are described herein with the word "comprising," it is understood that otherwise similar embodiments described with the terms "consisting of" and / or "consisting essentially of" are also provided.

[0080] The term "and / or," e.g., "X and / or Y," shall be understood to mean "X and Y" or "X or Y" and shall be deemed to provide explicit support for both meanings or for either meaning.

[0081] When aspects or embodiments of the invention are described in terms of Markush groups or other alternative groupings, the invention includes the entire group listed together, but also each member of the group individually, and all possible subgroups of the main group, as well as the main group in which one or more of the group members are absent. The invention also envisions the explicit exclusion of any one or more of the group members in the claimed invention. [Brief explanation of the drawings]

[0082] [Figure 1] 1A and 1B illustrate the upregulation of PSMP in cirrhotic and nearby non-tumorous liver tissue (see Example 1). Immunohistochemistry was performed as described in Example 6.

[0083] [Figure 2] 2A and 2B illustrate the upregulation of PSMP in human liver fibrotic tissues of different causes (see Example 1). Immunohistochemistry was performed as described in Example 6.

[0084] [Figure 3-1]3A-3D illustrate the upregulation of PSMP in a mouse model of CCl4-induced liver fibrosis (see Example 1). In the mouse CCl4-induced liver fibrosis model, quantitative real-time PCR assay (FIG. 3A), cytometric bead assay (CBA) (FIGS. 3B and 3C), and immunohistochemistry (FIG. 3D) were performed as described in Example 6. [Figure 3-2] Same as above.

[0085] [Figure 4] 4A and 4B illustrate the upregulation of PSMP in a mouse model of liver fibrosis induced by bile duct ligation (see Example 1). The mouse BDL liver fibrosis model, immunohistochemistry, and quantitative real-time PCR assay were performed as described in Example 6.

[0086] [Figure 5-1] 5A-5K illustrate protection from liver fibrosis in PSMP knockout mice using a CCl4-induced fibrosis model (see Example 2). Mouse CCl4-induced liver fibrosis model, immunohistochemistry, immunoblotting analysis, hydroxyproline determination, and quantitative real-time PCR assay were performed as described in Example 6. [Figure 5-2] Same as above. [Figure 5-3] Same as above. [Figure 5-4] Same as above.

[0087] [Figure 6-1] 6A-6K illustrate protection from liver fibrosis in PSMP knockout mice using a BDL-induced fibrosis model (see Example 2). Mouse BDL-induced liver fibrosis model, immunohistochemistry, immunoblotting analysis, hydroxyproline determination, and quantitative real-time PCR assay were performed as described in Example 6. [Figure 6-2] Same as above. [Figure 6-3] Same as above. [Figure 6-4] Same as above.

[0088] [Figure 7-1] 7A-7L illustrate the protective effect of neutralizing anti-PSMP antibodies in a CCl4-induced fibrosis model (see Example 3). The mouse CCl4-induced liver fibrosis model, antibody treatment, immunohistochemistry, immunoblotting analysis, hydroxyproline determination, and quantitative real-time PCR assay were performed as described in Example 6. [Figure 7-2] Same as above. [Figure 7-3] Same as above. [Figure 7-4] Same as above.

[0089] [Figure 8-1] 8A-8K illustrate the therapeutic effect of neutralizing anti-PSMP antibodies in a CCl4-induced fibrosis model (see Example 3). The mouse CCl4-induced liver fibrosis model, antibody treatment, immunohistochemistry, hydroxyproline determination, and quantitative real-time PCR assay were performed as described in Example 6. [Figure 8-2] Same as above. [Figure 8-3] Same as above. [Figure 8-4] Same as above.

[0090] [Figure 9-1] 9A-9L illustrate the dose-dependent therapeutic effect of neutralizing anti-PSMP antibodies in a CCl4-induced fibrosis model (see Example 3). The mouse CCl4-induced liver fibrosis model, antibody treatment, immunohistochemistry, immunoblotting analysis, hydroxyproline determination, and quantitative real-time PCR assay were performed as described in Example 6. [Figure 9-2] Same as above. [Figure 9-3] Same as above. [Figure 9-4] Same as above.

[0091] [Figure 10-1]10A-10L illustrate the use of the AAV8-hPSMP vector to restore PSMP expression and promote liver fibrosis in PSMP knockout mice (see Example 4). The mouse CCl4-induced liver fibrosis model, AAV8 construction and injection, immunohistochemistry, hydroxyproline determination, and quantitative real-time PCR assay were performed as described in Example 6. [Figure 10-2] Same as above. [Figure 10-3] Same as above. [Figure 10-4] Same as above.

[0092] [Figure 11-1] Figures 11A-11N illustrate the effects of PSMP deficiency on hepatic immune cells and pro-inflammatory cytokine production (see Example 5). In PSMP knockout mice treated with CCl4, reduced hepatic macrophage infiltration (determined by flow cytometry; Figures 11A-11E) and reduced pro-inflammatory cytokine production (determined by cytometric bead assay (CBA); Figures 11K-11N) were observed. Other immune cells, such as neutrophils, B cells, and T cells, were investigated using flow cytometry and found to be unaffected (Figures 11F-11J). The mouse CCl4-induced liver fibrosis model, flow cytometry, and CBA were performed as described in Example 6. [Figure 11-2] Same as above. [Figure 11-3] Same as above. [Figure 11-4] Same as above. [Figure 11-5] Same as above.

[0093] [Figure 12-1]Figures 12A-12G illustrate the upregulation of PSMP and the effects of PSMP deficiency during pulmonary fibrosis in a bleomycin-induced mouse model. See Example 7. PSMP protein levels in lungs and BALF were measured by cytokine bead assay (Figures 12A and 12B). Figure 12C shows representative lung immunohistochemical staining for PSMP. Body mass index was measured for 14 days (Figure 12D). Figure 12E shows representative lung histology with Masson's trichrome staining. α-SMA expression was determined by immunohistochemistry (Figure 12F). Figure 12G shows representative immunohistochemical staining for PSMP in normal human lungs and human lungs with airway fibrosis. Scale bar, 100 μm. *, P<0.05; **, p<0.01, n≧6 / group. [Figure 12-2] Same as above. [Figure 12-3] Same as above.

[0094] [Figure 13] Figure 13 illustrates reduced fibrosis in PSMP knockout mice in a mouse model of nonalcoholic steatohepatitis (NASH). See Example 8.

[0095] [Figure 14] 14 is a graph illustrating the therapeutic effect (prolonged survival time) of a neutralizing anti-PSMP antibody in an acute GVHD (aGVHD) mouse model. See Example 9.

[0096] [Figure 15-1]Figures 15A-15K illustrate the therapeutic effect of a neutralizing anti-PSMP antibody in a chronic ethanol-feeding model. See Example 10. Figure 15A shows a schematic diagram of the experimental design for treatment with the PSMP-neutralizing antibody 3D5 or PBS in a chronic ethanol-feeding mouse model. Serum triglycerides and cholesterol were measured (Figures 15B and 15C). Liver histological examination was performed by H&E staining and Oil Red staining (Figures 15D and 15E) and quantification of Oil Red-positive areas (Figure 15F). Liver Ppara and Srebp-1 mRNA levels were measured by RT-qPCR (Figures 15G and 15H). Further liver histological examination was performed by Sirius Red staining and quantification (Figures 15I and 15J). Liver nonparenchymal cells were stained and analyzed by flow cytometry. Infiltrating macrophages (iMΦ, CD45+Ly6G-F4 / 80+CD11bhigh) were quantified by flow cytometry (Figure 15K) (Scale bar, 100 μm / 50 μm. *, p<0.05; **, p<0.01 by one-way ANOVA, n=3–11 / group). [Figure 15-2] Same as above. [Figure 15-3] Same as above. [Figure 15-4] Same as above. [Figure 15-5] Same as above. [Figure 15-6] Same as above.

[0097] [Figure 15-7]Figures 15L-15T illustrate the therapeutic effects of neutralizing anti-PSMP antibodies in a chronic and excessive alcohol-feeding model. See Example 10. Figure 15L shows a schematic diagram of the experimental design for treatment with the PSMP neutralizing antibody 3D5 or PBS in this model. Liver triglycerides and cholesterol were measured (Figures 15M and 15N). Figure 15O shows representative H&E staining of liver tissue from mice treated with PBS and 3D5. Liver nonparenchymal cells were stained and analyzed by flow cytometry. Infiltrating macrophages (iMΦ, CD45+Ly6G-F4 / 80+CD11bhigh) were quantified by flow cytometry analysis (Figure 15P). Figures 15Q and 15R show representative H&E staining (15Q) and Oil Red O staining (15R) of liver tissue from WT and Psmp- / - mice. Serum ALT levels were measured (Figure 15S). Liver Srebp-1 and Ppara mRNA levels were measured by RT-qPCR (Figures 15T and 15U). Scale bar, 50 μm. *, P<0.05; **, p<0.01 by one-way ANOVA, n≧4 / group. [Figure 15-8] Same as above. [Figure 15-9] Same as above. [Figure 15-10] Same as above. [Figure 15-11] Same as above.

[0098] [Figure 16] Figures 16A-16C illustrate the expression of PSMP in mouse liver injury and fibrosis. See Example 11. WT mice (open bars) and Psmp- / - mice (black bars) treated with the DDC diet or APAP were used to perform immunohistochemical staining of livers for PSMP (Figure 16A) and measurement of PSMP mRNA levels by qRT-PCR (Figures 16B and 16C).

[0099] [Figure 17-1]Figures 17A-17I illustrate the protective effect of PSMP deficiency in DDC mice against liver fibrosis. See Example 12. WT and Psmp- / - mice were fed DDC for 4 weeks. Liver histological examination was performed by H&E staining and Sirius Red staining (Figure 17A) and its quantification (Figures 17B and 17C). α-SMA expression was determined by immunohistochemistry and Western blotting (Figures 17A and 17D). Hepatic mRNA levels of fibrogenic genes were measured by qRT-PCR (Figures 17E-17I). [Figure 17-2] Same as above. [Figure 17-3] Same as above.

[0100] [Figure 18-1] Figures 18A-18G illustrate the effect of neutralizing anti-PSMP antibodies on mouse DDC-induced liver fibrosis. See Example 13. Figure 18A shows a schematic diagram of the experimental design for treatment with PSMP-neutralizing antibody 3D5 or mIgG in mice. Liver histological examination was performed by H&E staining and Sirius Red staining (Figure 18B) and its quantification (Figures 18C and 18D). α-SMA expression was determined by immunohistochemistry and Western blotting (Figures 18B and 18E). Hepatic mRNA levels of fibrogenic genes were measured by qRT-PCR (Figures 18F and 18G). [Figure 18-2] Same as above. [Figure 18-3] Same as above.

[0101] [Figure 19-1]Figures 19A-19G illustrate the protective effect of PSMP deficiency in MCD mice against liver fibrosis. See Example 14. WT and Psmp- / - mice were fed MCD for 6 weeks. Figure 19A shows representative immunohistochemical staining of the liver for PSMP in WT mice. Liver histological examination was performed by H&E staining and Sirius Red staining (Figure 19B) and its quantification (Figures 19C and 19D). Liver mRNA levels of fibrogenic genes were measured by qRT-PCR (Figures 19E-19G). [Figure 19-2] Same as above. [Figure 19-3] Same as above.

[0102] [Figure 20-1] Figures 20A-20D illustrate the effect of a neutralizing anti-PSMP antibody on MCD diet-induced liver fibrosis in mice (treatment from week 4 to week 5.5). See Example 15. Figure 20A shows a schematic diagram of the experimental design for treatment with the PSMP neutralizing antibody 3D5 or PBS in mice. Liver histological examination was performed by H&E staining and Sirius Red staining (Figure 20B) and quantification (Figures 20C and 20D). (Scale bar, 100 μm. **, P<0.01; ***, P<0.001 by Student's t-test, n=5 / group). [Figure 20-2] Same as above.

[0103] [Figure 20-3] Figures 20E-20I illustrate the effect of a neutralizing anti-PSMP antibody on MCD diet-induced liver fibrosis in mice (treatment from week 5 to week 8). See Example 15. Figure 20E shows a schematic diagram of the experimental design for treatment with the PSMP neutralizing antibody 3D5 or PBS in mice. Liver histology was performed by Sirius Red and α-SMA immunohistochemical staining (Figure 20F) and its quantification (Figures 20G-20I). (Scale bar, 100 μm. **, P<0.01 by Student's t-test; control, n=3; MCD-PBS, n=6; MCD-3D5, n=5). [Figure 20-4]Same as above.

[0104] [Figure 21-1] Figures 21A and 21B illustrate PSMP expression in different renal disease tissues. See Example 16. Figure 21A shows representative immunohistochemical staining of PSMP in human normal kidney tissues, LN kidney tissues, and IgAN kidney tissues from nearby clear cell carcinoma. Figure 21B shows a statistical summary of PSMP immunostaining in different renal diseases. LN: lupus nephritis, IgAN: IgA nephropathy, MGN: membranous glomerulonephritis. Scale bar, 100 μm. **, p<0.01; ***, p<0.001 by Student's t-test.

[0105] [Figure 21-2] Figures 21C-21E illustrate the effects of PSMP deficiency or neutralization in a mouse model of acute kidney injury (AKI). See Example 16. Figure 21C shows serum creatinine levels 48 hours after glycerol injection in Psmp- / - or WT mice. Figures 21D and 21E show serum creatinine (21D) and BUN (21E) levels 48 hours after glycerol injection in mice injected with the PSMP-neutralizing antibody 3D5 or PBS. *, P<0.05 by Student's t-test, n=4-6 / group.

[0106] [Figure 21-3] Figures 21F-21H illustrate the therapeutic effect of a neutralizing anti-PSMP antibody in a mouse model of chronic kidney disease (CKD). See Example 16. Figure 21F shows a schematic diagram of the experimental design for treatment with the PSMP neutralizing antibody 3D5 or PBS in UUO mice. Figures 21G and 21H show representative kidney histology by Sirius red staining (21G) and its quantification (21H; gray bars—3D5-treated group, black bars—PBS-treated group). Scale bar, 100 μm. *, P<0.05 by Student's t-test, n=4-6 / group. [Figure 21-4] Same as above. [Figure 21-5] Same as above.

[0107] [Figure 22-1] Figures 22A-22D illustrate the therapeutic effect of a neutralizing anti-PSMP antibody in a chronic GVHD (cGVHD) mouse model. See Example 17. Figure 22A shows body weight monitored over time. Figure 22B shows fur shedding in the control group compared to mice treated with anti-PSMP antibody 3D5. Serum markers of liver injury, ALT and AST, were measured (Figures 22C and 22D). *, P<0.05; **, p<0.01 by Student's t-test, n≧6 / group. [Figure 22-2] Same as above.

[0108] [Figure 23-1] Figures 23A-23C illustrate the therapeutic effect of a neutralizing anti-PSMP antibody in a mouse model of lupus nephritis. See Example 18. Renal function markers serum creatinine (Figure 23A) and blood urea nitrogen (BUN) (Figure 23B), serum total IgG levels (Figure 23C), and glomerular autoantibody deposition (Figure 23D) were measured. Scale bar, 100 μm. *, P<0.05; **, p<0.01 by Student's t-test, n≧6 / group. [Figure 23-2] Same as above. DETAILED DESCRIPTION OF THE INVENTION

[0109] The present invention is generally directed to the use of PSMP antagonists, including, for example, neutralizing anti-PSMP antibodies, for treating diseases. Specifically, in some embodiments, the present invention relates to the use of PSMP antagonists for treating liver fibrosis. In other embodiments, the present invention relates to the use of PSMP antagonists for treating pulmonary fibrosis. In other embodiments, the present invention relates to the use of PSMP antagonists for treating renal fibrosis. In still other embodiments, the present invention relates to the use of PSMP antagonists for treating diseases or disorders associated with liver fibrosis, pulmonary fibrosis, or renal fibrosis, such as a disease or disorder selected from non-alcoholic fatty liver disease (e.g., non-alcoholic steatohepatitis (NASH)), alcoholic liver disease (ALD), primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC), drug-induced lung injury, acute kidney injury (AKI), chronic kidney disease (CKD), lupus nephritis, IgA nephropathy, and membranous glomerulonephritis. In yet another aspect, the invention relates to the use of PSMP antagonists to treat graft-versus-host disease (GVHD) or systemic lupus erythematosus.

[0110] The research described herein shows that PSMP signaling is involved in the pathogenesis of liver fibrosis, lung fibrosis and kidney fibrosis, and that blocking PSMP in vivo can improve fibrosis in disease models.Specifically, the present inventors have found that PSMP expression is upregulated in human liver fibrosis and mouse liver fibrosis (see Example 1).Using PSMP knockout mice and liver fibrosis models, the present inventors have also shown that PSMP deficiency significantly attenuates the development of liver fibrosis (see Example 2), that the restoration of liver PSMP expression promotes liver fibrosis in a CCR2-dependent manner (see Example 4), and that neutralizing PSMP signaling by either preventive or therapeutic treatment regimens significantly improves liver fibrosis (see Example 3). Furthermore, the present inventors have shown that PSMP is required for the establishment of fibrosis in a model of nonalcoholic steatohepatitis (NASH) (see Example 8), that PSMP deficiency protects against liver fibrosis in an MCD diet-induced NASH model (see Example 14), and that neutralizing PSMP attenuates liver fibrosis in an MCD diet-induced NASH model (see Example 15). The present inventors have also found that neutralization or deficiency of PSMP attenuates liver steatosis and liver fibrosis in a chronic ethanol-feeding model, and attenuates liver steatosis, liver injury, and inflammation in a chronic and excessive alcohol-feeding model (also referred to as the NIAAA model) (see Example 10). The studies described herein also show that PSMP is upregulated in DDC diet-induced liver fibrosis and acute liver injury models (see Example 11), that PSMP deficiency confers protection from liver fibrosis in the DDC model (see Example 12), and that neutralizing PSMP attenuates DDC-induced liver fibrosis in mice (see Example 13).Furthermore, the studies described herein demonstrate that PSMP is upregulated in a bleomycin-induced model of pulmonary fibrosis, that PSMP deficiency in this model confers protection from pulmonary fibrosis, and that neutralization of PSMP signaling attenuates pulmonary fibrosis in this model (see Example 7).

[0111] The inventors have also found that PSMP expression is upregulated in human renal disease, that PSMP depletion and neutralization attenuates renal injury in a glycerol-induced rhabdomyolysis model, and that PSMP neutralization attenuates renal fibrosis in a UUO-induced model of acute kidney injury (AKI) and chronic kidney disease (see Example 16).

[0112] The inventors have also found that neutralizing PSMP can effectively treat graft-versus-host disease, as shown by prolonged survival in an acute GVHD mouse model (see Example 9), and by a lack of fur shedding and significantly lower levels of liver injury markers in a chronic GVHD model (see Example 17).

[0113] The inventors have also found that neutralizing PSMP can effectively treat lupus nephritis, a common manifestation of systemic lupus erythematosus (SLE), as demonstrated by improved renal function, reduced serum IgG levels, and reduced glomerular autoantibody deposition in a mouse lupus model (see Example 18).

[0114] Examples of PSMP antagonists for use within the scope of the present invention include molecules that bind to PSMP and reduce the activity of PSMP in cells expressing a PSMP receptor, such as CCR2. Particularly suitable PSMP antagonists include neutralizing anti-PSMP antibodies. Suitable PSMP antagonists also include non-antibody soluble proteins that can inhibit the interaction of PSMP with its receptor, including, for example, alternative scaffold proteins and peptide aptamers that specifically bind to and neutralize PSMP. In some embodiments, the PSMP antagonist is a nucleic acid aptamer that specifically binds to and neutralizes PSMP. In other variations, small molecule antagonists can be used that can inhibit the interaction of PSMP with its receptor or otherwise inhibit PSMP-induced intracellular signaling through the PSMP receptor. Inhibitory polynucleotides that target the PSMP gene or mRNA encoding PSMP can also be used.

[0115] The neutralizing activity of candidate PSMP antagonists (e.g., antibodies or other soluble binding proteins) can be evaluated, for example, in a chemotaxis assay using cells expressing PSMP receptors. For example, chemotaxis assays can utilize cells expressing either or both of the isoforms of human CCR2 (CCR2A and / or CCR2B, the amino acid sequences of which are set forth in SEQ ID NO: 2 and SEQ ID NO: 3, respectively). Chemotaxis assays using CCR2-expressing cells are known in the art. See, for example, Pei et al., J. Immunol. 192: 1878-86, 2014. In one exemplary assay, a plasmid (10 μg) expressing CCR2B (SEQ ID NO: 3) is transiently transfected into HEK293 cells by electroporation (120 V for 20 ms) using an electric pulse generator. After 48 hours, the CCR2B-expressing HEK293 cells are used in a chemotaxis assay using a 48-well micro-chemotaxis chamber. Pretreat PSMP (residues 37-139 of SEQ ID NO: 1) (7 ng / ml, 70 ng / ml, or 700 ng / ml) with candidate PSMP antagonists (10 or 50 μg / ml) for 30 minutes. Cells migrating to the bottom of the filter are fixed and analyzed by three-phase ELISA. Stain using the Step Stain Set. Cells are counted in five randomly selected high-power fields (×400) per well. The chemotactic ability of PSMP toward CCR2B-expressing HEK293 cells in the presence of a candidate antagonist is assessed compared to the chemotactic ability in the absence of the candidate antagonist to determine whether the candidate is effective in neutralizing PSMP activity.

[0116] In some embodiments, a PSMP antagonist according to the present invention is a soluble protein (e.g., an antibody) that specifically binds to PSMP (residues 37-139 of SEQ ID NO: 1). A binding protein is considered to specifically bind if it (1) exhibits a threshold level of binding activity and (2) does not significantly cross-react with a control polypeptide molecule. For example, a threshold level of binding is determined when the protein binds to a PSMP polypeptide, peptide, or epitope with an affinity that is at least 10-fold greater than the binding affinity for a control (non-PSMP) polypeptide. Binding proteins (e.g., antibodies) used within the scope of the present invention may have a binding affinity of at least 10-fold greater than the binding affinity for a control (non-PSMP) polypeptide. 6 M -1 or larger, preferably 10 7 M -1 or greater, more preferably 10 8 M -1 or greater, most preferably 10 9 M -1 or greater binding affinity (K a ) is preferably shown.

[0117] Soluble binding proteins (e.g., antibodies) can be evaluated for binding activity using any of a variety of known assays. For example, one assay system uses a commercially available biosensor instrument (BIAcore™, Pharmacia Biosensor, Piscataway, NJ), in which a binding protein (e.g., an anti-PSMP antibody) is immobilized on the surface of a sensor chip, and a test sample containing a soluble target molecule (e.g., PSMP) is passed through the cell. If the immobilized protein has affinity for the target molecule, it will bind to the target, causing a change in the refractive index of the medium, which is detected as a change in the surface plasmon resonance of the gold film. This system allows the association and dissociation rates to be determined, from which binding affinity can be calculated, allowing the stoichiometry of binding to be assessed. The use of this instrument has been disclosed, for example, by Karlsson (J. Immunol. Methods 145: 229-240, 1991) and Cunningham and Wells (J. Mol. Biol. 234: 554-563, 1993). The binding activity of candidate protein molecules can also be determined using a range of other assay systems known in the art. Such systems include Scatchard analysis (see Scatchard, Ann. NY Acad. Sci. 51: 660-672, 1949) and calorimetric assays (see Cunningham et al., Science 253: 545-548, 1991; Cunningham et al., Science 254: 821-825, 1991) to determine binding affinity.

[0118] In certain embodiments, the PSMP antagonist is a soluble protein (e.g., an antibody) that competes for binding to PSMP (residues 37-139 of SEQ ID NO: 1) with an antibody having the same heavy and light chain variable domains (VH and VL) as monoclonal antibody 3D5. The amino acid sequences of the mAb 3D5 VH and VL domains are set forth herein as SEQ ID NO: 4 and SEQ ID NO: 5, respectively. mAb 3D5 has been shown to effectively neutralize PSMP activity. See Pei et al., J. Immunol. 192: 1878-86, 2014; see also Example 3, infra. The ability of a PSMP-binding protein to compete with an antibody having the VH and VL domains of mAb 3D5 for binding to PSMP can be determined by an assay in which a test PSMP-binding protein (e.g., an antibody) or a functional binding fragment thereof prevents or inhibits the specific binding of a reference antibody having the VH and VL domains of mAb 3D5 (i.e., having the VH and VL domains of SEQ ID NO: 4 and SEQ ID NO: 5, respectively) to PSMP. Typically, such an assay involves the use of a purified target protein (e.g., residues 37-139 of SEQ ID NO: 1, or a fragment thereof) bound to a solid surface or cells bearing the target protein, an unlabeled test protein (i.e., the PSMP-binding protein or candidate PSMP-binding protein), and a labeled reference antibody. Competitive inhibition is measured by determining the amount of label bound to the solid surface or cells in the presence of the test protein. Typically, the test protein is present in excess and / or is bound first. Soluble PSMP-binding proteins identified by competitive assays ("competitive PSMP-binding proteins") include proteins that bind to the same epitope as that bound by the reference antibody, proteins that bind to epitopes that overlap with the epitope bound by the reference antibody, and proteins that bind to epitopes that are sufficiently close to the epitope bound by the reference antibody to create steric hindrance.Typically, when a competing PSMP-binding protein (e.g., a competing anti-PSMP antibody) is present in excess, the competing PSMP-binding protein (e.g., a competing anti-PSMP antibody) inhibits specific binding of the reference antibody to the PSMP target protein by at least 30%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, or at least 75%, and in some cases binding is inhibited by at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, or more. Conversely, if the reference antibody binds, it preferably inhibits binding of a subsequently added competing PSMP-binding protein (e.g., a competing anti-PSMP antibody) by at least 30%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, or at least 75%, and in some cases binding is inhibited by at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, or more.

[0119] In certain preferred embodiments, the PSMP antagonist for use according to the present invention is an antibody. The antibody according to the present invention comprises or consists of at least a portion of an intact antibody that retains antigen-binding specificity. Suitable antibodies include, for example, fully human antibodies; humanized antibodies; chimeric antibodies; antibody fragments, such as Fab, Fab', F(ab)2, F(ab')2, and Fv antibody fragments; single-chain antibodies; and antibody heavy or light chain monomers or dimers, or mixtures thereof. A preferred antibody of the present invention is a monoclonal antibody. Antibodies comprising a light chain may comprise a kappa or lambda light chain.

[0120] In certain embodiments, antibodies of the invention comprise intact immunoglobulins of any isotype, including IgA, IgG, IgE, IgD, or IgM (including subtypes thereof). Preferably, intact immunoglobulins according to the invention comprise intact IgG (e.g., intact IgG1, IgG2, IgG3, or IgG4).

[0121] Methods for preparing and isolating polyclonal antibodies, monoclonal antibodies, and their antigen-binding antibody fragments are well known in the art.See, for example, Current Protocols in Immunology (Cooligan et al. eds., John Wiley and Sons, Inc. 2006); Sambrook et al., Molecular Cloning: A Laboratory Manual (Cold Spring Harbor, NY, 2nd ed. 1989); and Monoclonal Hybridoma Antibodies: Techniques and Applications (Hurrell ed., CRC Press, Inc., Boca Raton, FL, 1982).Antigen-binding fragments, including scFv, can be prepared, for example, using phage display libraries according to methods known in the art. Methods for preparing recombinant human polyclonal antibodies are disclosed in Wiberg et al., Biotechnol Bioeng. 94: 396-405, 2006; Meijer et al., J. Mol. Biol. 358: 764-772, 2006; Haurum et al., U.S. Patent Application Publication No. 2002 / 0009453; and Haurum et al., U.S. Patent Application Publication No. 2005 / 0180967. As will be apparent to those skilled in the art, these methods are equally applicable to generating antibodies against PSMP.

[0122] As will be apparent to those skilled in the art, polyclonal antibodies for use within the scope of the present invention can be generated by inoculating any of a variety of warm-blooded animals, such as horses, cows, goats, sheep, dogs, chickens, rabbits, mice, and rats, with an immunogenic polypeptide or polypeptide fragment. The immunogenicity of the immunogenic polypeptide can be increased by using an adjuvant, such as alum (aluminum hydroxide) or Freund's complete or incomplete adjuvant. Polypeptides useful for immunization also include fusion polypeptides, such as a fusion of PSMP with an immunoglobulin polypeptide or a fusion of PSMP with maltose-binding protein. Polypeptide immunogens can be full-length molecules or portions thereof. If the polypeptide moiety is haptenic, it can be advantageously conjugated or linked to a macromolecular carrier, such as keyhole limpet hemocyanin (KLH), bovine serum albumin (BSA), or tetanus toxoid, for immunization.

[0123] Additionally, antibodies can be screened against known polypeptides related to the antibody target (e.g., orthologs, paralogs, or sequence variants of PSMP) to isolate a population of antibodies that are highly specific for binding to the target protein. Such lack of cross-reactivity with related polypeptide molecules can be demonstrated, for example, by standard Western blot analysis (Current Protocols in Molecular Biology (Ausubel et al. eds., Green and Wiley and Sons, NY 1993)) or ELISA (enzyme immunoassay) (Immunoassay, A Practical Guide (Chan ed., Academic Press, Inc. 1987)) is demonstrated by the antibody detecting the PSMP polypeptide but not known related polypeptides. In another example, antibodies raised against a PSMP polypeptide are adsorbed to related polypeptides attached to an insoluble matrix; antibodies highly specific for the PSMP polypeptide pass through the matrix under appropriate buffer conditions. Screening makes it possible to isolate polyclonal and monoclonal antibodies that are non-cross-reactive with known closely related polypeptides (Antibodies: A Laboratory Manual (Harlow and Lane eds., Cold Spring Harbor Laboratory Press, 1988); Current Protocols in Immunology (Cooligan et al. eds., National Institutes of Health, John Wiley and Sons, Inc. 1995). Screening and isolation of specific antibodies is well known in the art. See Fundamental Immunology (Paul ed., Raven Press 1993); Getzoff et al., Adv. in Immunol. 43: 1-98, 1988; Monoclonal Antibodies: Principles and Practice (Goding ed., Academic Press Ltd. 1996); Benjamin et al., Ann. Rev. Immunol. 2: 67-101, 1984.

[0124] Native monoclonal antibodies ("mAbs") can be prepared, for example, by immunizing a subject animal (e.g., a rat or mouse) with purified immunogenic protein or a fragment thereof. In a typical procedure, each animal receives an initial intraperitoneal (IP) injection of purified protein or fragment, typically in combination with an adjuvant (e.g., complete Freund's adjuvant or RIBI adjuvant (available from Sigma-Aldrich, St. Louis, MO)), followed by booster IP injections of purified protein at intervals of, for example, two weeks. Seven to ten days after the administration of the third booster injection, the animals are bled and serum collected. Further boosts can be administered if necessary. Spleen cells and lymph node cells from animals with high titers are harvested and fused with myeloma cells (e.g., murine SP2 / 0 or Ag8 cells) using conventional methods. The fusion mixture is then cultured on a thymocyte feeder layer or with appropriate media supplements (including commercially available supplements such as Hybridoma Fusion and Cloning Supplement; Roche Diagnostics, Indianapolis, IN). Approximately 10 days after fusion, hybridoma pools that produce specific antibodies are identified using standard assays (e.g., ELISA). Positive pools can be further analyzed for their ability to block or reduce the activity of target proteins. Positive pools are cloned by limiting dilution.

[0125] The amino acid sequence of a native antibody can be altered by applying recombinant DNA techniques. Thus, antibodies can be redesigned to obtain desired characteristics. Modified antibodies can, for example, provide improved stability and / or therapeutic efficacy compared to their unmodified forms. The possible variations are numerous and range from changing just one or a few amino acids to completely redesigning, for example, the variable or constant region. Alterations in the constant region are generally made to improve or alter characteristics such as complement binding, membrane interaction, and other effector functions. Various alterations of the constant region to alter characteristics (e.g., effector functions) are known. For example, Morgan et al., Immunology 86: 319-324, 1995; Lund et al., J. Immunol. 157: 4963-9 157: 4963-4969, 1996; Idusogie et al., J. Immunol. et al., J. Immunol. 143: 2595-2601, 1989;Jefferis et al., Immunological Reviews 163: 59-76, 1998;Armour et al., Eur. J. Immunol., 29: 2613-2624, 1999; PCT Application No. PCT / GB99 / 01441; UK Patent Application No. 9809951.8. Typically, changes in the variable region are made to improve antigen binding characteristics, improve the stability of the variable region, or reduce the risk of immunogenicity. Phage display techniques can also be used. See, e.g., Huse et al., Science 246: 1275-1281, 1989; Ladner et al., U.S. Patent No. 5,571,698.

[0126] For therapeutic antibodies intended for human use, it is usually desirable to humanize the non-human regions of the antibody according to known procedures. Methods for producing humanized antibodies are disclosed, for example, in U.S. Patent Nos. 5,530,101; 5,821,337; 5,585,089; 5,693,762; and 6,180,370. Methods for producing humanized antibodies are also disclosed, for example, in U.S. Patent No. 7,732,578. Typically, a humanized anti-PSMP antibody comprises the complementarity-determining regions (CDRs) of a mouse donor immunoglobulin and the heavy and light chain frameworks of a human acceptor immunoglobulin. Often, framework residues in the human framework regions are replaced with corresponding residues from the donor antibody to alter, preferably improve, antigen binding. These framework substitutions are identified by methods well known in the art, for example, by modeling the interactions of CDR and framework residues to identify framework residues important for antigen binding, and comparing sequences to identify unusual framework residues at particular positions (see, e.g., Queen et al., U.S. Patent No. 5,585,089; Riechmann et al., Nature 332: 323, 1988).

[0127] Non-humanized chimeric antibodies can also be used therapeutically (e.g., in immunosuppressed patients). Thus, in some variations, the antibody for use according to the present invention is, inter alia, a chimeric antibody derived from a non-human anti-PSMP antibody. Preferably, the chimeric antibody comprises a variable region derived from a mouse or rat antibody and a constant region derived from a human; therefore, the chimeric antibody has a longer half-life and is less immunogenic when administered to a human subject. Methods for producing chimeric antibodies are known in the art. See, for example, Morrison, Science 229: 1202, 1985; Oi et al., BioTechniques 4: 214, 1986; Gillies et al., J. Immunol. Methods 125: 191-202, 1989; U.S. Patent Nos. 5,807,715; 4,816,567; and 4,816,397.

[0128] The present invention also encompasses the use of fully human antibodies, such as those selected from human antibody expression libraries (e.g., phage display libraries); those produced in non-human animals (e.g., mice) transgenic for human heavy chain loci and human light chain loci and in which the corresponding endogenous immunoglobulin loci have been inactivated; or those prepared by immortalizing human B lymphocytes that produce antibodies against PSMP target antigens.

[0129] Antibodies for use in accordance with the present invention may be, for example, 5 x 10 -4 Under M, 10 -4 Less than M, 5 x 10 -5 Under M, 10 -5 Less than M, 5 x 10 -6 Under M, 10 -6 Less than M, 5 x 10 -7 Under M, 10 -7 Less than M, 5 x 10 -8 Under M, 10 -8 Less than M, 5 x 10 -9 Under M, 10 -9 Less than M, 5 x 10 -10 Under M, 10 -10Less than M, 5 x 10 -11 Under M, 10 -11 Less than M, 5 x 10 -12 Under M, 10 -12 Less than M, 5 x 10 -13 Under M, 10 -13 Less than M, 5 x 10 -14 Under M, 10 -14 Less than M, 5 x 10 -15 Less than M or 10 -15 A dissociation constant (K d ) has a binding affinity containing

[0130] The antibodies of the present invention further include derivatives that have been modified, for example, by covalently attaching any type of molecule to the antibody so as not to interfere with the antibody's binding to its epitope. Suitable modifications include, for example, fucosylation, glycosylation, acetylation, pegylation, phosphorylation, and amidation. Antibodies and their derivatives may themselves be derivatized by known protecting / blocking groups, proteolytic cleavage, linkage to cellular ligands or other proteins, and the like. In some embodiments of the present invention, at least one heavy chain of the antibody is fucosylated. In certain variations, the fucosylation is N-linked. In certain embodiments, at least one heavy chain of the antibody comprises fucosylated N-linked oligosaccharides.

[0131] Antibodies for use according to the present invention include variants that have one or more amino acid substitutions, deletions, or additions compared to a reference antibody, such that one or more biological properties of the reference antibody are retained. In certain embodiments, the antibody is a variant that has one or more amino acid substitutions, deletions, or additions compared to a reference anti-PSMP neutralizing antibody having the VH and / or VL sequences set forth in SEQ ID NO: 4 and SEQ ID NO: 5, respectively, such that the reference antibody's ability to specifically bind to PSMP and neutralize PSMP activity is retained. Those skilled in the art can easily create variants that have one or more amino acid substitutions, deletions, or additions compared to a reference antibody. Techniques for obtaining these variants, including genetic techniques (such as suppression, deletion, mutation), chemical techniques, and enzymatic techniques, are known to those skilled in the art.

[0132] In some embodiments, a neutralizing anti-PSMP antibody for use in accordance with the present invention comprises one or more CDRs of the anti-PSMP mAb 3D5. Thus, in certain variations, the antibody comprises the heavy chain CDRs (at least one of the CDR-H1, CDR-H2, and CDR-H3 regions) of the 3D5 VH domain (SEQ ID NO: 4) and / or the light chain CDRs (at least one of the CDR-L1, CDR-L2, and CDR-L3 regions) of the 3D5 VL domain (SEQ ID NO: 5). In typical embodiments, the antibody has two or three of the CDRs of the 3D5 VH domain (SEQ ID NO: 4) and / or two or three of the CDRs of the 3D5 VL domain (SEQ ID NO: 5). In some variations, where an antibody has at least one CDR of the 3D5 VH domain, the antibody further comprises at least one CDR of the 3D5 VL domain; in some such embodiments, the antibody has all three heavy chain CDRs and all three light chain CDRs of mAb 3D5 (i.e., CDR-H1, CDR-H2, and CDR-H3 of SEQ ID NO: 4 and CDR-L1, CDR-L2, and CDR-L3 of SEQ ID NO: 5). One or more CDRs may be defined, for example, according to the Chothia, Kabat, AbM, or contact definition of a CDR. Under the Chothia definition of CDRs, CDR-H1, CDR-H2, and CDR-H3 of mAb 3D5 correspond to residues 31-35, 50-69, and 99-108, respectively, of SEQ ID NO: 4, and CDR-L1, CDR-L2, and CDR-L3 of mAb 3D5 correspond to residues 24-34, 50-56, and 89-97, respectively, of SEQ ID NO: 5. In particular variations of the above antibodies, the antibodies comprise a VH domain and / or a VL domain having a human immunoglobulin framework region or a variant thereof having at least 85%, at least 90%, or at least 95% amino acid sequence identity to a human immunoglobulin framework region.

[0133] In some embodiments, the anti-PSMP antibody comprises (a) a heavy chain variable domain that is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO:4, and / or (b) a light chain variable domain that is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO:5.

[0134] In some embodiments, the anti-PSMP antibody for use according to the invention comprises a heavy chain CDR, CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and at least one of the CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref wherein the reference CDRs are CDR-H1, CDR-H2, and CDR-H3, respectively, of the 3D5 VH domain (SEQ ID NO: 4). In other embodiments, the anti-PSMP antibody for use in accordance with the invention comprises a light chain CDR, CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and at least one of the CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref wherein the reference CDRs are CDR-L1, CDR-L2, and CDR-L3 of the 3D5 VL domain (SEQ ID NO: 5), respectively. In certain embodiments, the anti-PSMP antibody comprises both the above-described sets of heavy chain and light chain CDRs. Particularly suitable PSMP antibodies include CDRs CDR-H1 and CDR-H2.Ab , CDR-H2 Ab , and CDR-H3 Ab and a heavy chain variable domain comprising CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and a light chain variable domain comprising: Ref , CDR-H2 Ref , and CDR-H3 Ref and the set of light chain CDRs comprises CDR-L1, CDR-L2, CDR-L3, CDR-L4, CDR-L5, CDR-L6, CDR-L7, CDR-L8, CDR-L9, CDR-L10, CDR-L11, CDR-L12, CDR-L13, CDR-L14, CDR-L15, CDR-L16, CDR-L17, CDR-L18, CDR-L19 ... Ref , CDR-L2 Ref , and CDR-L3 Ref and have six or fewer, typically five or fewer, more typically four or fewer, and most typically three or fewer amino acid substitutions compared to (Illegible) . The CDRs may be defined, for example, according to the Chothia, Kabat, AbM, or contact definitions of CDRs. In particular variations of the above antibodies, the VH and VL domains each have a human immunoglobulin framework region or a variant thereof having at least 85%, at least 90%, or at least 95% amino acid sequence identity to a human immunoglobulin framework region.

[0135] Anti-PSMP antibodies for use in accordance with the present invention include affinity-matured embodiments. Affinity-matured antibodies can be produced by procedures known in the art. See, for example, Marks et al., Bio / Technology 10: 779-783, 1992; Barbas et al., Proc Nat. Acad. Sci. USA 91: 3809-3813, 1994; Schier et al., Gene 169: 147-155, 1995; Yelton et al., See J. Immunol. 155: 1994-2004, 1995; Jackson et al., J. Immunol. 154: 3310-9, 1995; Hawkins et al., J. Mol. Biol. 226: 889-896, 1992; and PCT Publication No. WO2004 / 058184. In some embodiments, the anti-PSMP antibody is obtained by affinity maturation of an antibody comprising one or more CDRs of mAb 3D5 (e.g., obtained by affinity maturation of an antibody comprising (i) a VH domain having CDR-H1, CDR-H2, and CDR-H3 of SEQ ID NO: 4 and (ii) a VL domain having CDR-L1, CDR-L2, and CDR-L3 of SEQ ID NO: 5); in some such variations, the CDRs are defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition of a CDR.

[0136] One method for adjusting antibody affinity is called "library scanning mutagenesis." Generally, library scanning mutagenesis is performed as follows: Using art-recognized methods, one or more amino acid positions in at least one CDR (e.g., two, three, four, five, or six CDRs) are replaced with two or more (e.g., three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, or twenty) amino acids. This generates a small library of clones (in some embodiments, one for each amino acid position analyzed), each with a complexity of two or more members (when two or more amino acids are substituted per position). Generally, the library also includes clones containing the native (unsubstituted) amino acid. A small number of clones from each library, e.g., about 20-80 clones (depending on the complexity of the library), are screened for binding affinity to the target polypeptide (or other binding target) to identify candidates with increased binding, the same binding, decreased binding, or no binding. Binding affinity can be determined, for example, using Biacore™ surface plasmon resonance analysis, which detects differences in binding affinity of about 2-fold or greater. Biacore™ is a powerful tool for determining whether the starting antibody already has a relatively high affinity, e.g., a K of about 10 nM or less. D This is particularly useful when the bond is

[0137] In some embodiments of the invention, the antibody comprises an immunoglobulin constant region (e.g., an immunoglobulin heavy chain constant region, such as an Fc region). In some such embodiments, the constant region is a variant of a naturally occurring (e.g., wild-type) constant region, such as, for example, a constant region that has increased or decreased affinity for human Fc gamma receptors, or that has reduced activity (compared to an unmodified antibody) with respect to any one or more of the following: triggering complement-mediated lysis, stimulating antibody-dependent cell-mediated cytotoxicity (ADCC), or activating microglia. Various modifications of the constant region can be used to achieve optimal levels and / or combinations of effector functions. For example, Morgan et al., Immunology 86: 319-324, 1995;Lund et al., J. Immunol. 157: 4963-9 157: 4963-4969, 1996;Idusogie et al., J. Immunol. 164: 4178-4184, 2000;Tao et al., J. See Immunol. 143: 2595-2601, 1989; and Jefferis et al., Immunological Reviews 163: 59-76, 1998. In some embodiments, the constant region is modified as described in Armour et al., Eur. J. Immunol., 29: 2613-2624, 1999; PCT Application No. PCT / GB99 / 01441, and / or UK Patent Application No. 9809951.8. In other embodiments, the constant region is deglycosylated for N-linked glycosylation. In some embodiments, the constant region is deglycosylated for N-linked glycosylation by mutating the glycosylated amino acid residue in the constant region or an adjacent residue that is part of the N-glycosylation recognition sequence. For example, N-glycosylation site N297 can be mutated to A, Q, K, or H. See Tao et al., J. Immunol. 143: 2595-2601, 1989; Jefferis et al., Immunological Reviews 163: 59-76, 1998. The constant region can also be deglycosylated enzymatically for N-linked glycosylation (e.g., by removing carbohydrates with the enzyme PNGase) or by expression in glycosylation-deficient host cells. Other suitable variant constant regions include sequences based on chimeric domains derived from two or more human immunoglobulin heavy chain CH2 domains that retain FcRn and FcγRIIb binding activity and do not induce significant complement-dependent lysis or cell-mediated destruction of the target. See PCT Publication No. WO 99 / 58572.

[0138] In some embodiments, PSMP antagonists for use according to the present invention are non-antibody proteins based on alternative scaffolds (see, e.g., Silverman et al., Nat. Biotechnol. 23: 1556-61, 2005; Zahnd See, e.g., U.S. Patent No. 7,115,396; Binz et al., Nat. Biotechnol. 23: 1257-1268, 2005; Skerra, Current Opin. in Biotech. 18: 295-304, 2007; Silacci et al., J. Biol. Chem. 289: 14392-14398, 2014. In some embodiments, the alternative scaffold antagonist specifically binds to and neutralizes PSMP. In certain variations, the alternative scaffold PSMP antagonist is an Adnectin™, iMab, lipocalin, Kunitz domain, Affibody®, ankyrin repeat, Afflin, Tetranectin, Fynomer, or Avimer protein. PSMP-binding proteins based on alternative scaffolds can be prepared, for example, by screening expression libraries (e.g., phage display, ribosome display) containing one or more randomized or semi-randomized regions within the alternative scaffold framework. See, e.g., Koide et al., J. Mol. Biol. 284: 1141-1151, 1998; Zahnd et al., supra.

[0139] In other embodiments, the PSMP antagonist for use in accordance with the present invention is a peptide aptamer (e.g., a peptide aptamer that specifically binds and neutralizes PSMP). Peptide aptamers generally consist of a variable peptide loop (e.g., about 5-20 amino acids) embedded as a loop within a typically small, stable protein scaffold. Peptide aptamers can be prepared by selecting aptamers from random pools or peptide libraries for binding affinity with a specific target. For example, peptide aptamers can be isolated from random peptide libraries by yeast two-hybrid screening (see, e.g., Xu et al., Proc. Natl. Acad. Sci. USA 94: 12473, 1997) or from phage display libraries. Peptide aptamers are reviewed, for example, in Reverdatto et al., Curr. Top. Med. Chem. 15: 1082-1101, 2015.

[0140] In yet another embodiment, the PSMP antagonist is a nucleic acid aptamer (e.g., a nucleic acid aptamer that specifically binds to and neutralizes PSMP). Generally, nucleic acid aptamers are oligonucleotides, such as ribonucleic acid (RNA) or single-stranded deoxyribonucleic acid (ssDNA), that can bind to targets with high affinity and specificity due to their specific three-dimensional structure. Nucleic acid aptamers that specifically bind to target macromolecules can be easily isolated from libraries of such oligomers by techniques such as SELEX. See, for example, Stoltenburg et al., Biamal. Eng. 24: 381, 2007. Nucleic acid aptamers can be easily isolated from libraries of such oligomers by techniques such as SELEX. See, for example, Ni et al., Curr. Med. Chem. 18: 4206-4214, 2011; and Esposito et al., Discovery Medicine 11: This was reviewed in J. Med. Chem. Soc., 2011, 487-496.

[0141] According to certain aspects of the present invention, PSMP antagonists are used to treat liver fibrosis. In some embodiments, the treated liver fibrosis has progressed to liver cirrhosis. In other, non-mutually exclusive, embodiments, the fibrosis is associated with a disease or disorder known to be characterized by or result in liver fibrosis. For example, the treated liver fibrosis may be associated with chronic alcohol abuse (e.g., alcoholic liver disease (ALD), alcoholic hepatitis, alcoholic cirrhosis), chronic viral hepatitis (e.g., hepatitis B, hepatitis C, or hepatitis D), accumulation of fat in the liver (non-alcoholic fatty liver disease (e.g., non-alcoholic steatohepatitis (NASH)), accumulation of iron in the body (hemochromatosis), cystic fibrosis, accumulation of copper in the liver (Wilson's disease), poorly formed bile ducts (biliary atresia), alpha-1 The condition may be associated with antitrypsin deficiency, a genetic disorder of carbohydrate metabolism (e.g., galactosemia, glycogen storage disease), a genetic disorder of the digestive system (e.g., Alagille syndrome), an autoimmune liver disease (e.g., autoimmune hepatitis, primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC, formerly known as primary biliary cirrhosis), an infectious disease (e.g., syphilis, brucellosis), a drug-induced liver injury (e.g., methotrexate-induced or isoniazid-induced liver injury), or Budd-Chiari syndrome.

[0142] According to another aspect of the present invention, a PSMP antagonist is used to treat lung fibrosis. In some embodiments, the fibrosis is associated with a disease or disorder known to be characterized by or result in lung fibrosis. For example, the treated lung fibrosis may be associated with dermatomyositis, polymyositis, mixed connective tissue disease, systemic lupus erythematosus, rheumatoid arthritis, sarcoidosis, scleroderma, pneumonia, chronic radiation pneumonitis, pneumoconiosis, infectious disease, or drug-induced lung injury. In some embodiments, where the pulmonary fibrosis is associated with pneumoconiosis, the pneumoconiosis is caused or exacerbated by exposure to silica dust, asbestos fibers, cemented carbide dust, coal dust, grain dust, or bird or animal feces. In some embodiments in which the pulmonary fibrosis is associated with drug-induced lung injury, the lung injury is induced by a chemotherapeutic drug (e.g., methotrexate, cyclophosphamide), a cardiac medication (e.g., a drug used to treat arrhythmia, such as amiodarone), an antibiotic (e.g., nitrofurantoin, ethambutol), or an anti-inflammatory drug (e.g., rituximab, sulfasalazine).

[0143] In yet other aspects of the invention, PSMP antagonists are used to treat non-alcoholic fatty liver disease (NAFLD). In some such variations, the NAFLD is non-alcoholic steatohepatitis (NASH).

[0144] In yet another embodiment of the invention, PSMP antagonists are used to treat alcoholic liver disease (ALD).

[0145] In yet another embodiment of the invention, PSMP antagonists are used to treat primary sclerosing cholangitis (PSC).

[0146] In yet another embodiment of the invention, PSMP antagonists are used to treat primary biliary cholangitis (PBC).

[0147] In yet another aspect of the invention, PSMP antagonists are used to treat renal fibrosis, hi some embodiments, the fibrosis is associated with a disease or disorder characterized by or known to result in renal fibrosis. For example, the renal fibrosis to be treated may be associated with IgA nephropathy, membranoproliferative glomerulonephritis, membranous glomerulonephritis, crescentic glomerulonephritis, diabetic nephropathy, hypertensive nephropathy, lupus nephritis, hepatic nephropathy, polycystic kidney disease, Alport syndrome, Fabry disease, primary hyperoxaluria, cystinosis; focal segmental glomerulosclerosis (FSGS) caused by coenzyme Q10-related gene mutations, complement 3 glomerulonephritis, acute or subacute immune complex glomerulonephritis, renal vasculitis, systemic lupus erythematosus (SLE), recent-onset renal artery stenosis (fibromuscular or vasculitic), diabetic kidney disease, chronic uric acid nephropathy, toxic nephropathy, bacterial pyelonephritis, viral nephropathy, multiple myeloma, or obstructive nephropathy.

[0148] In yet another embodiment of the invention, PSMP antagonists are used to treat acute kidney injury (AKI) or chronic kidney disease (CKD). In some embodiments in which a PSMP antagonist is used to treat CKD, the CKD is caused by IgA nephropathy, membranoproliferative glomerulonephritis, membranous glomerulonephritis, crescentic glomerulonephritis, diabetic nephropathy, hypertensive nephropathy, lupus nephritis, hepatic nephropathy, polycystic kidney disease, Alport syndrome, Fabry disease, primary hyperoxaluria, cystinosis; focal segmental glomerulosclerosis (FSGS) caused by coenzyme Q10-related gene mutations, complement 3 glomerulonephritis, acute or subacute immune complex glomerulonephritis, renal vasculitis, systemic lupus erythematosus (SLE), recent-onset renal artery stenosis (fibromuscular or vasculitic), diabetic kidney disease, chronic uric acid nephropathy, toxic nephropathy, bacterial pyelonephritis, viral nephropathy, multiple myeloma, or obstructive nephropathy.

[0149] In yet another aspect of the invention, PSMP antagonists are used to treat graft-versus-host disease (GVHD). In some embodiments, PSMP antagonists are used to treat acute GVHD (aGVHD). In other embodiments, PSMP antagonists are used to treat chronic GVHD (cGVHD).

[0150] In yet another embodiment of the invention, PSMP antagonists are used to treat systemic lupus erythematosus (SLE).

[0151] In yet another embodiment of the invention, PSMP antagonists are used to treat lupus nephritis.

[0152] In yet another embodiment of the invention, PSMP antagonists are used to treat IgA nephropathy.

[0153] In yet another embodiment of the invention, PSMP antagonists are used to treat membranous glomerulonephritis.

[0154] In yet another aspect of the invention, PSMP antagonists are used to treat diseases or disorders associated with liver fibrosis, such as chronic alcohol abuse (e.g., alcoholic liver disease (ALD), alcoholic hepatitis, alcoholic cirrhosis), chronic viral hepatitis (e.g., hepatitis B, hepatitis C, or hepatitis D), accumulation of fat in the liver (non-alcoholic fatty liver disease (e.g., non-alcoholic steatohepatitis (NASH)), accumulation of iron in the body (hemochromatosis), cystic fibrosis, accumulation of copper in the liver (Wilson's disease), Treating conditions such as poorly formed bile ducts (biliary atresia), alpha 1 antitrypsin deficiency, inherited disorders of sugar metabolism (e.g., galactosemia, glycogen storage disease), inherited digestive system disorders (e.g., Alagille syndrome), autoimmune liver disease (e.g., autoimmune hepatitis, primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC, formerly known as primary biliary cirrhosis), infectious diseases (e.g., syphilis, brucellosis), drug-induced liver injury (e.g., methotrexate-induced or isoniazid-induced liver injury), or Budd-Chiari syndrome.

[0155] In yet other aspects of the invention, PSMP antagonists are used to treat diseases or disorders associated with pulmonary fibrosis, such as dermatomyositis, polymyositis, mixed connective tissue disease, systemic lupus erythematosus, rheumatoid arthritis, sarcoidosis, scleroderma, pneumonia, chronic radiation pneumonitis, pneumoconiosis, infectious diseases, or drug-induced lung injury. In some embodiments in which a PSMP antagonist is used to treat pneumoconiosis, the pneumoconiosis is caused or exacerbated by exposure to silica dust, asbestos fibers, cemented carbide dust, coal dust, grain dust, or bird or animal feces. In some embodiments in which a PSMP antagonist is used to treat drug-induced lung injury, the lung injury is induced by a chemotherapeutic drug (e.g., methotrexate, cyclophosphamide, bleomycin), a cardiac medication (e.g., a drug used to treat arrhythmia, such as amiodarone), an antibiotic (e.g., nitrofurantoin, ethambutol), or an anti-inflammatory drug (e.g., rituximab, sulfasalazine).

[0156] In yet other embodiments of the invention, PSMP antagonists are used to treat diseases or disorders associated with renal fibrosis, such as IgA nephropathy, membranoproliferative glomerulonephritis, membranous glomerulonephritis, crescentic glomerulonephritis, diabetic nephropathy, hypertensive nephropathy, lupus nephritis, hepatic nephropathy, polycystic kidney disease, Alport syndrome, Fabry disease, primary hyperoxaluria, cystinosis; focal segmental glomerulosclerosis (FSGS) caused by coenzyme Q10-related gene mutations, complement 3 glomerulonephritis, acute or subacute immune complex glomerulonephritis, renal vasculitis, systemic lupus erythematosus (SLE), recent-onset renal artery stenosis (fibromuscular or vasculitic), diabetic kidney disease, chronic uric acid nephropathy, toxic nephropathy, bacterial pyelonephritis, viral nephropathy, multiple myeloma, or obstructive nephropathy.

[0157] In each of the embodiments of the methods of treatment described herein, the PSMP antagonist is delivered in a manner consistent with conventional methodologies associated with the management of the disease or disorder for which treatment is sought. In accordance with the disclosure herein, an effective amount of the antagonist is administered to a subject in need of such treatment for a time and under conditions sufficient to treat the disease or disorder.

[0158] Subjects to whom the PSMP antagonists described herein are administered include patients at high risk of developing a particular disease or disorder (e.g., liver fibrosis, pulmonary fibrosis, renal fibrosis, or a disease or disorder associated with liver fibrosis or pulmonary fibrosis), as well as patients exhibiting an existing disease or disorder. In certain embodiments, the subject has been diagnosed with the disease or disorder for which treatment is being sought. Furthermore, the subject can be monitored for any changes in the disease or disorder (e.g., an increase or decrease in clinical symptoms of the disease or disorder) during the course of treatment. Also, in some variations, the subject does not suffer from another disease or disorder requiring treatment that results in inhibition of the PSMP signaling pathway.

[0159] In preventive applications, pharmaceutical compositions or medicants are administered to patients susceptible to or otherwise at risk of a particular disease or disorder in an amount sufficient to eliminate or reduce the risk of the disease or disorder or delay its onset. In therapeutic applications, compositions or medicants are administered to patients suspected of or already suffering from such a disease in an amount sufficient to cure or at least partially halt the symptoms of the disease or disorder and its complications. An amount adequate to achieve this is referred to as a therapeutically or pharmaceutically effective dose or amount. In both preventive and therapeutic regimes, drugs are usually administered in several doses until a sufficient response (e.g., inhibition of fibrosis or fibrosis biomarkers) is achieved. Typically, the response is monitored, and repeated doses are administered once the desired response begins to fade.

[0160] To identify patients for treatment according to the methods of the present invention, acceptable screening methods can be used to determine risk factors associated with a particular disease or disorder or to determine the status of a pre-existing disease or disorder identified in the subject. Such methods can include, for example, determining whether a subject has relatives diagnosed with a particular disease. Screening methods can also include conventional workup to determine familial status for particular diseases known to have a heritable component. For example, various liver diseases are known to have certain heritable components. See, for example, Scorza et al., International Journal of Hepatology, Volume 2014, Article ID 713754, page 11 (genetic defects causing early chronic liver involvement); Severson et al., World J. Gastroenterol. 22: 6742-6756, 2016 (genetic factors affecting the development of nonalcoholic fatty liver disease). For this purpose, molecular diagnostic assays (e.g., nucleic acid-based diagnostic assays) can be routinely used to identify individuals with genetic markers associated with the disease of interest. Screening can also be performed as indicated by known patient symptomatology, age factors, associated risk factors, and the like. These methods allow clinicians to routinely select patients in need of the treatment methods described herein. According to these methods, administration of a PSMP antagonist can be carried out as an independent treatment program or as a follow-up, adjunctive, or coordinated treatment regimen to other treatments.

[0161] For administration, the PSMP antagonist is formulated as a pharmaceutical composition. Pharmaceutical compositions containing a PSMP antagonist can be formulated according to known methods for preparing pharmaceutically useful compositions, in which a therapeutic molecule is combined with a pharmaceutically acceptable carrier to form a mixture. A composition is considered a "pharmaceutically acceptable carrier" if, when combined with an active ingredient, it allows the active ingredient to retain its biological activity and its administration is acceptable to a recipient patient. Examples include, but are not limited to, any of the standard pharmaceutical carriers, such as phosphate-buffered saline solution, water, emulsions such as oil / water emulsions, and various types of wetting agents. Preferred diluents for aerosol or parenteral administration are phosphate-buffered saline (PBS) or normal saline (0.9%). Compositions containing such carriers are formulated by well-known conventional methods (see, for example, Gennaro (ed.), Remington's Pharmaceutical Sciences (Mack Publishing Company, 19th ed. 1995); Allen (ed.), Remington: The Science and Practice of Pharmacy (Pharmaceutic Press, 22nd revised ed.)). The formulation may further include one or more excipients, preservatives, solubilizers, buffers, albumin to prevent protein loss on the vial surface, etc. The monospecific antagonists can be formulated individually or provided as a combined formulation.

[0162] The pharmaceutical composition comprising PSMP antagonist is administered to the subject in an effective amount.According to the method of the present invention, the antagonist can be administered to the subject by various administration forms, including, for example, intramuscular, subcutaneous, intravenous, intraatrial, intraarticular, parenteral, intranasal, intrapulmonary, transdermal, intrapleural, intrathecal, and oral administration routes.For prevention and treatment purposes, the antagonist can be administered to the subject by a single bolus delivery, by continuous delivery over a long period of time (for example, continuous transdermal delivery), or by a repeated administration protocol (for example, hourly, daily, or weekly).The exact dosage is determined by the clinician according to accepted standards, taking into account the nature and severity of the condition to be treated, the constitution of the patient, etc.The determination of dosage is within the level of ordinary skill in the art.

[0163] In this context, determining effective dosages is typically guided by animal model studies, follow-up human clinical trials, and determining effective dosages and administration protocols that significantly reduce the incidence or severity of the target disease or disorder in model subjects. The effective dose of the compositions of the present invention varies depending on many different factors, including the means of administration, the target site, the patient's physiological condition, whether the patient is human or animal, other medications administered, whether the treatment is prophylactic or therapeutic, and the specific activity of the composition itself and its ability to elicit the desired response in an individual. Typically, the patient is a human, but for some diseases, the patient may be a non-human mammal. Typically, the dosage regimen is adjusted to provide an optimal therapeutic response, i.e., to optimize safety and efficacy. Therefore, a therapeutically or prophylactically effective amount is also an amount in which the beneficial effects outweigh any undesirable side effects. For administration of PSMP antagonists, the dosage typically ranges from about 0.1 μg to 100 mg per kg of subject body weight, or from 1 μg to about 50 mg, more usually from 10 μg to 5 mg. In more specific embodiments, the effective amount of the agent is between about 1 μg / kg and about 20 mg / kg, between about 10 μg / kg and about 10 mg / kg, or between about 0.1 mg / kg and about 5 mg / kg. Dosages within this range can be achieved by a single administration or multiple administrations, including, for example, multiple daily, weekly, biweekly, or monthly administrations. For example, in certain variations, the regimen consists of an initial administration followed by multiple subsequent administrations at weekly or biweekly intervals. Another regimen consists of an initial administration followed by multiple subsequent administrations at monthly or bimonthly intervals. Alternatively, administration can be on an ad hoc basis, as indicated by monitoring fibrosis and / or clinical symptoms of the disease or disorder.

[0164] Particularly suitable animal models for evaluating the effectiveness of PSMP antagonists for the treatment of liver fibrosis, pulmonary fibrosis, or renal fibrosis are generally known in the art. For example, one liver fibrosis model uses CCL4 injection. See, for example, You et al., Mol. Med. Rep. 12: 5594-5600, 2015. CCL4 toxic liver fibrosis can be induced, for example, in 6-8 week-old male mice by intraperitoneal (ip) injection of CCl4 (e.g., 1.0 ml per kg body weight, dissolved in corn oil at a ratio of 1:9) twice a week for 4 or 6 weeks. Another exemplary liver fibrosis model uses bile duct ligation (BDL) to induce cholestasis in mice. See, for example, Yongping et al., J. Ethnopharmacol. 169: 200-209, 2015. Using this model, cholestasis and accompanying fibrosis develop in mice (e.g., 6-8 week old male mice) over a period of time (e.g., 14 days) after common bile duct ligation.

[0165] One suitable model of pulmonary fibrosis is the bleomycin-induced pulmonary fibrosis model. See, for example, Rangarajan et al., Nat. Med. 8: 1121-1127, 2018; doi: 10.1038 / s41591-018-0087-6. For bleomycin-induced pulmonary fibrosis, mice (e.g., 6- to 8-week-old female C57Bl / 6J mice) are anesthetized, tracheotomized, and injected with bleomycin (e.g., a single injection containing 3.5 U / kg of bleomycin). Pulmonary fibrosis develops within a certain period (e.g., 14 days) after injection.

[0166] A suitable model of kidney fibrosis is the unilateral ureteral obstruction (UUO) model, in which kidney fibrosis is caused by injury to the renal tubules as a result of obstructed urine flow. See, for example, Elena Martinez-Klimova et al., Biomolecules 9: 141, 2019.

[0167] Animal models of diseases or disorders associated with liver fibrosis, pulmonary fibrosis, or kidney fibrosis are also known. An exemplary model of alcoholic liver fibrosis is described, for example, in Ambade et al. (Hepatology, 2018, doi: 10.1002 / hep.30249). Regarding nonalcoholic steatohepatitis (NASH), various dietary, chemical, and genetic models can be used. See, for example, Hansen et al., Drug Discovery Today, 22, 1707, 2017. For example, one NASH model is a combined chemical and dietary challenge model by administering streptozotocin (STZ) together with a high-fat diet (HFD). In the STZ-HFD model, NASH is induced by injection of STZ (e.g., a single subcutaneous injection of 200 μg of STZ into 2-day-old male mice) and ad libitum feeding of an HFD for 8 weeks, typically after 4 weeks of age. Another exemplary NASH model is a nutritionally deficient diet model—the methionine-choline deficiency (MCD) model—commonly used in preclinical NASH research. In the MCD model, NASH is induced by ad libitum feeding of MCD. Typically, hepatic macrovesicular steatosis and inflammatory cell infiltration develop in MCD mice after 1–3 weeks of feeding, with robust perisinusoidal fibrosis occurring by the 5th to 7th week. Regarding cholestatic diseases such as primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC), chronic feeding of 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) is an exemplary model.

[0168] The most common rodent model used to study acute kidney injury (AKI) and chronic kidney disease (CKD) is the unilateral ureteral obstruction (UUO) model of renal fibrosis (Elena Martinez-Klimova et al., supra). Furthermore, a common mouse model of AKI after rhabdomyolysis involves injection of glycerol into the leg muscle. See, e.g., Yanqiu et al., Stem Cell Research & Therapy 5: 80, 2014. Regarding lupus nephritis, the DBA / 2→B6D2F1 model is a well-known model in which DBA / 2 mouse spleen cells are transfused into B6D2F1 mice. See, e.g., Via et al., J Immunol. 139: 1840-1849, 1987.

[0169] Animal models for evaluating the efficacy of PSMP antagonists for treating GVHD are also generally known. See, for example, Schroeder et al., Dis. See Model Mech. 4: 318-333, 2011. In one exemplary acute GVHD (aGVHD) model, BABL / c mice (e.g., 6-8 week old males) serve as recipients, and C57BL / 6 mice serve as donors. Splenic CD3+ T cells and bone marrow cells from the C57BL / 6 mice are isolated after sacrifice of the mice. Prior to cell injection (e.g., 12 hours prior), recipient mice are exposed to radiation (e.g., two doses of 4 Gy each). 60 (Total dose: 8 Gy) CD3+ cells and bone marrow cells, e.g., 5 × 10 6 vs 1×10 7The mixture is mixed at a ratio of 0.01 to 0.01, and then intravenously injected into BABL / c mice. During the aGVHD period, which is typically defined as one month after injection, the weight and survival rate of the mice are monitored every two days. Furthermore, a well-known chronic GVHD (cGVHD) mouse model is the DBA / 2→B6D2F1 model, which is established by transfusing DBA / 2 mouse spleen cells into B6D2F1 mice (also a model of systemic lupus erythematosus (SLE) and lupus nephritis, see, for example, Via et al., supra). Generally, see, for example, Kim et al., J. Immunol. 181: 7380-7389, 2008.

[0170] The dosage of pharmaceutical compositions can be varied by the attending clinician to maintain the desired concentration at the target site. For example, if an intravenous delivery method is selected, the local concentration of drug in the bloodstream at the target tissue can be approximately 1 to 50 nanomoles per liter of composition, sometimes between about 1.0 nanomoles per liter and 10, 15, or 25 nanomoles per liter, depending on the subject's condition and the expected measured response. Higher or lower concentrations can be selected based on the delivery method, for example, transepidermal delivery versus delivery to mucosal surfaces. Dosage should also be adjusted based on the release rate of the administered formulation, for example, nasal spray versus powder, sustained-release oral or injectable particles, transdermal formulations, etc. To achieve the same serum concentration level, for example, sustained-release particles with a 5 nanomolar release (under standard conditions) should be administered at approximately twice the dose as particles with a 10 nanomolar release.

[0171] Pharmaceutical compositions containing PSMP antagonists can be provided in liquid, aerosol, or solid form.Liquid forms are exemplified by injections, aerosols, droplets, topological solutions, and oral suspensions.Exemplary solid forms include capsules, tablets, and controlled-release forms.The latter are exemplified by miniosmotic pumps and implants.For example, Bremer et al., Pharm. Biotechnol. 10: 239, 1997; Ranade, "Implants in Drug Delivery", in Drug Delivery Systems 95-123 (Ranade and Hollinger, eds., CRC Press 1995); Bremer et al., "Protein Delivery with Infusion Pumps," in Protein Delivery: Physical Systems 239-254 (Sanders and Hendren, eds., Plenum Press 1997); Yewey et al., "Delivery of Proteins from a Controlled Release Injectable Implant," in Protein Delivery: Physical Systems 93-117 (Sanders and Hendren, eds., Plenum Press 1997). Other solid forms include creams, pastes, and other topological applications.

[0172] Liposomes provide a means for delivering therapeutic polypeptides to a subject, for example, intravenously, intraperitoneally, intrathecally, intramuscularly, subcutaneously, or by oral, inhalation, or intranasal administration. Liposomes are microscopic vesicles consisting of one or more lipid bilayers and a surrounding aqueous compartment. See generally Bakker-Woudenberg et al., Eur. J. Clin. Microbiol. Infect. See Dis. 12 (Suppl. 1): S61, 1993; Kim, Drugs 46: 618, 1993; Ranade, "Site-Specific Drug Delivery Using Liposomes as Carriers," in Drug Delivery Systems 3-24 (Ranade and Hollinger, eds., CRC Press 1995). Liposomes are similar in composition to cell membranes, resulting in safe administration of liposomes and biodegradability. Depending on the preparation method, liposomes can be unilamellar or multilamellar, and liposomes can vary in size, ranging in diameter from 0.02 μm to over 10 μm. Various drugs can be encapsulated in liposomes: hydrophobic drugs are distributed within the bilayer, and hydrophilic drugs are distributed in the internal aqueous space(s). See, e.g., Machy et al., Liposomes In Cell Biology And Pharmacology (John Libbey 1987); Ostro et al., American J. Hosp. Pharm. 46: 1576, 1989. Furthermore, by varying liposome size, number of bilayers, lipid composition, and liposome charge and surface characteristics, it is possible to control the therapeutic availability of the encapsulated agent.

[0173] Liposomes can adsorb to virtually any cell type and then slowly release the encapsulated drug. Alternatively, absorbed liposomes can be endocytosed by phagocytic cells. Endocytosis is followed by intralysosomal degradation of the liposomal lipids and release of the encapsulated drug (see Scherphof et al., Ann. NY Acad. Sci. 446:368, 1985). After intravenous administration, small liposomes (0.1-1.0 μm) are typically taken up by cells of the reticuloendothelial system, primarily located in the liver and spleen, while liposomes larger than 3.0 μm are deposited in the lungs. This preferential uptake of small liposomes by cells of the reticuloendothelial system has been used to deliver therapeutic agents to the liver.

[0174] The reticuloendothelial system can be circumvented by several methods, including saturating with large doses of liposomal particles or selectively inactivating macrophages by pharmacological means (see Claassen et al., Biochim. Biophys. Acta 802: 428, 1984). Furthermore, the incorporation of glycolipid-derivatized or polyethylene glycol-derivatized phospholipids into the liposome membrane has been shown to significantly reduce uptake by the reticuloendothelial system (see Allen et al., Biochim. Biophys. Acta 1068: 133, 1991; Allen et al., Biochim. Biophys. Acta 1150: 9, 1993).

[0175] Liposomes can also be prepared to target specific cells or organs by varying the phospholipid composition or by inserting receptors or counterreceptors into the liposomes. For example, liposomes prepared with a high content of nonionic surfactants have been used to target the liver. See, for example, Japanese Patent 04-244,018 to Hayakawa et al.; Kato et al., Biol. Pharm. Bull. 16: 960, 1993. These formulations were prepared by mixing soybean phosphatidylcholine, α-tocopherol, and ethoxylated hydrogenated castor oil (HCO-60) in methanol, concentrating the mixture under vacuum, and then reconstituting the mixture with water. Liposome formulations of dipalmitoylphosphatidylcholine (DPPC) with soybean-derived steryl glucoside mixture (SG) and cholesterol (Ch) have also been shown to target the liver. See Shimizu et al., Biol. Pharm. Bull. 20: 881, 1997.

[0176] Alternatively, various targeting counter-receptors such as antibodies, antibody fragments, carbohydrates, vitamins, and transport proteins can be bound to the surface of liposomes.For example, to target the liver, liposomes can be modified with branched galactosyl lipid derivatives to target the asialoglycoprotein (galactose) receptor, which is exclusively expressed on the surface of hepatocytes.See Kato and Sugiyama, Crit. Rev. Ther. Drug Carrier Syst.14:287,1997;Murahashi et al., Biol. Pharm. Bull.20:259,1997.In a more general method for tissue targeting, target cells are pre-labeled with biotinylated antibodies specific to the counter-receptors expressed by target cells.See Harasym et al., Adv. Drug Deliv. Rev.32:99,1998.After free antibody disappears from plasma, liposomes conjugated with streptavidin are administered. In another approach, targeting antibodies are attached directly to liposomes, see Harasym et al., supra.

[0177] Antibodies and other soluble proteins can be encapsulated in liposomes using standard techniques for protein microencapsulation. See, e.g., Anderson et al., Infect. Immun. 31:1099, 1981; Anderson et al., Cancer Res. 50:1853, 1990; Cohen et al., Biochim. Biophys. Acta 1063:95, 1991; Alving et al., "Preparation and Use of Liposomes in Immunological Studies," in Liposome Technology (Vol. III) 317 (Gregoriadis, ed., CRC Press, 2nd ed. 1993); Wassef et al., Meth. Enzymol. 149: 124, 1987. As noted above, therapeutically useful liposomes can contain a variety of components. For example, liposomes can contain lipid derivatives of poly(ethylene glycol). See Allen et al., Biochim. Biophys. Acta 1150: 9, 1993.

[0178] Degradable polymer microspheres have been designed to maintain high systemic levels of therapeutic proteins. Microspheres are prepared from degradable polymers, such as poly(lactide-co-glycolide) (PLG), polyanhydrides, poly(orthoesters), and nonbiodegradable ethylvinyl acetate polymers, and entrap proteins within the polymer. See, e.g., Gombotz and Pettit, Bioconjugate Chem. 6: 332, 1995; Ranade, "Role of Polymers in Drug Delivery," in Drug Delivery. Systems 51-93 (Ranade and Hollinger, eds., CRC Press 1995);Roskos and Maskiewicz, “Degradable Controlled Release Systems Useful for Protein Delivery”, in Protein Delivery: Physical Systems 45-92 (Sanders and Hendren, eds., Plenum Press 1997);Bartus et al., Science 281: 1161, 1998;Putney and Burke, Nature Biotechnology 16: 153, 1998;Putney, Curr. Opin. Chem. Biol. 2: 548, 1998. Polyethylene glycol (PEG) coated nanospheres can also provide carriers for intravenous administration of therapeutic proteins. See, e.g., Gref et al., Pharm. Biotechnol. 10: 167, 1997.

[0179] Other dosage forms can be devised by those skilled in the art, as shown, for example, in Ansel and Popovich, Pharmaceutical Dosage Forms and Drug Delivery Systems (Lea & Febiger, 5th ed. 1990); Gennaro (ed.), Remington's Pharmaceutical Sciences (Mack Publishing Company, 19th ed. 1995); Ranade and Hollinger, Drug Delivery Systems (CRC Press 1996); and Allen (ed.), Remington: The Science and Practice of Pharmacy (Pharmaceutic Press, 22nd revised ed.).

[0180] The pharmaceutical composition can be provided as a kit containing a container containing a PSMP antagonist composition described herein. The pharmaceutical composition can be provided, for example, in the form of a single-dose or multi-dose injectable solution or as a sterile powder to be reconstituted prior to injection. Alternatively, such a kit can include a dry powder disperser, liquid aerosol generator, or nebulizer for administering the pharmaceutical composition. Such a kit can further include written information regarding the indications and use of the pharmaceutical composition. Table 1: Exemplary sequences [Table 1-1] [Table 1-2]

[0181] The present invention is further illustrated by the following non-limiting examples. [Example]

[0182] Example 1 PSMP expression is upregulated in human and murine liver fibrosis To assess whether PSMP expression is associated with liver disease, PSMP levels were investigated by immunohistochemistry in tissue microarrays containing tissues from different liver diseases. PSMP was significantly upregulated in cirrhotic and adjacent non-tumorous liver tissues (Figures 1A and 1B). This finding was confirmed in human liver fibrosis tissues with different causes, including hepatitis B virus (HBV)-induced cirrhosis (n = 14), hepatitis C virus (HCV)-induced cirrhosis (n = 1), primary biliary cirrhosis (n = 5), and alcohol-induced cirrhosis (n = 2). Immunohistochemical analysis revealed that hepatic PSMP expression was significantly elevated in patients with cirrhosis compared with normal human liver tissue (Figures 2A and 2B). Similar to the human data, PSMP expression was also significantly increased in mouse models of CCl4-induced and BDL-induced liver fibrosis compared with livers from control mice (Figures 3A-3D, 4A, and 4B). These data indicate that PSMP is upregulated in cirrhotic liver and raise the hypothesis that activation of PSMP signaling may be involved in the pathogenesis of liver fibrosis. Example 2 Deficiency of PSMP confers protection from liver fibrosis in mice

[0183] To further explore the role of PSMP signaling in liver fibrosis, we next investigated fibrogenesis in PSMP knockout mice subjected to a toxic fibrosis mouse model induced by CCl4 treatment. Mice were repeatedly exposed to CCl4 (twice per week) for 4 weeks, resulting in the upregulation of Psmp signaling. - / - In mice, Ccr2 - / -Compared to mice with equivalent oil controls, Psmp demonstrated a significant attenuation of liver injury and fibrosis as assessed by hematoxylin and eosin (H&E), Sirius Red staining, and hepatic hydroxyproline content (Figures 5A-5D). - / - In mice, Ccr2 - / - Consistent with the Psmp mice, there was a marked decrease in the upregulation of α-SMA, a marker of HSC activation as assessed by immunohistochemistry and immunoblotting (Figures 5A and 5F). - / - There was a clear trend towards a decrease in Ccr2 in the mice, indicating an improvement in liver injury (Figure 5E). - / - Similar to mice, hepatic mRNA expression of prototypic profibrotic genes (Acta2, Col1a1, Tgfb1, Timp1, and Pdgfrb) was significantly increased during CCl4-induced Psmp - / - The phenotype was reduced in mice (Figures 5G-5K).

[0184] To further analyze the role of PSMP in liver fibrosis, we used another well-established mouse model of liver fibrosis, the BDL liver fibrosis model. - / - The Sirius Red-positive area, hydroxyproline content, and α-SMA expression were significantly reduced in the mice (Figures 6A-6D and 6F). Serum ALT levels and Psmp expression were also significantly reduced. - / - Psmp was elevated after BDL in mice (Figure 6E). - / - In mice, we also found significant decreases in the mRNA levels of Acta2, Col1a1, Tgfb1, Timp1, and Pdgfrb (Figures 6G-6K).

[0185] Taken together, these results suggest that activation of PSMP signaling may be involved in the pathogenesis of hepatic fibrosis. Example 3 Neutralization of PSMP signaling attenuates liver fibrosis in mice

[0186] Based on the significant attenuation of liver fibrosis development in mice with PSMP deficiency, we investigated the effect of 3D5, a specific PSMP-neutralizing antibody, on CCl4-induced liver fibrosis. First, to investigate the protective effect of 3D5 on mice with CCl4-induced liver fibrosis for 4 weeks, mice were treated with 3D5 after each CCl4 injection (Figure 7A). H&E staining and Sirius red staining assays demonstrated reduced liver injury and fibrosis after 3D5 treatment compared with the control and mIgG-treated groups (Figures 7B-7D). Accordingly, 3D5 significantly reduced α-SMA expression in fibrotic livers (Figures 7B and 7G). Furthermore, hepatic hydroxyproline content was significantly reduced in the 3D5-treated group compared with the control and mIgG-treated groups (Figure 7E). Serum ALT levels were significantly reduced in the 3D5-treated group compared with the control and mIgG-treated groups, indicating improved liver function (Figure 7F). qRT-PCR showed that 3D5 also significantly reduced the mRNA levels of fibrogenic genes (Acta2, Col1a1, Tgfb1, Timp1, and Pdgfrb) (Figures 7H-7L).

[0187] Furthermore, to evaluate the therapeutic potential of 3D5 in 6-week CCl4-induced liver fibrosis, mice were treated with 3D5 along with CCl4 from week 4 to week 6 (Figure 8A). Mice treated with 3D5 showed significant reductions in Sirius Red-positive area, hydroxyproline content, α-SMA expression, and serum ALT levels compared with control and mIgG-treated groups (Figures 8B-8F). Consistently, significant reductions in Acta2, Col1a1, Tgfb1, Timp1, and Pdgfrb mRNA levels were also observed in 3D5-treated mice (Figures 8G-8K).

[0188] Different doses of 3D5 (1 mg / kg, 5 mg / kg, and 10 mg / mL) were used to treat mice with CCl4-induced liver fibrosis (Figure 9A). H&E staining and Sirius red staining assays showed that mice with liver fibrosis treated with 3D5 had milder liver fibrosis than the control and mIgG-treated groups in a dose-dependent manner (Figures 9B-9E). The expression of α-SMA, serum ALT, and mRNA expression levels of fibrogenic genes (Acta2, Col1a1, Tgfb1, Timp1, and Pdgfrb) in CCl4-treated mice were also reduced in a dose-dependent manner after treatment with 3D5 (Figures 9B and 9F-9L).

[0189] These results suggest that blockade of PSMP signaling significantly reverses the pathogenesis of liver fibrosis in mice. Example 4 AAV8-hPSMP restores hepatic PSMP expression and promotes liver fibrosis in a CCR2-dependent manner

[0190] To further explore the role of PSMP in liver fibrosis, we used AAV8 (a gene vector isolated from rhesus macaques, which is used for transduction of hepatocytes due to its high affinity for hepatocytes) to investigate the role of PSMP in liver fibrosis. - / - We overexpressed human PSMP in mice (Fig. 10A). As expected, PSMPs that received AAV8-hPSMP were significantly increased. - / - In mice, the level of hPSMP expression was strongly elevated compared to mice receiving control AAV8-null (Figure 10E). Next, we used this technique to investigate the consequences of PSMP overexpression under conditions of chronic liver injury. Four weeks after AAV8 injection, mice were treated with CCl4 for an additional four weeks to induce liver fibrosis (Figure 10A). We found a significant enhancement of liver injury and fibrosis elicited by PSMP overexpression compared to the effects observed in mice injected with AAV8-null (Figures 10B-10D, 10F-10L).

[0191] The data further indicate that PSMP overexpression promotes the development of liver fibrosis. Example 5 PSMP deficiency inhibits hepatic macrophage infiltration and pro-inflammatory cytokine production induced by CCL4 challenge

[0192] In mice and humans, macrophage infiltration into the liver during chronic injury has been convincingly linked to the progression of liver inflammation and fibrosis. Therefore, we investigated the composition of immune cells in the liver after treatment with CCl4 by flow cytometry analysis. - / - In mice, hepatic macrophages (iMΦ, CD11b + F4 / 80 int ) infiltration and CD11b + CCR2 + The accumulation of cells was significantly reduced (Figures 11A-11E). In parallel with intrahepatic macrophages, neutrophils (CD11b + Ly6G + ), B cells and T cells (CD3 + , CD4 + , CD8 + We also investigated other immune cells, such as IL-1, IL-1, and found that these were unaffected except for some increase or decrease in T cells in PSMP-knockout mice (Figures 11F-11J).

[0193] Next, we investigated the consequences of reduced hepatic macrophage accumulation due to PSMP deficiency in the liver. An important function of hepatic macrophages during disease progression is the secretion of pro-inflammatory cytokines. - / - In mice, treatment with CCl4 reduced serum concentrations of CCL2, TNF-α, IL-6, and IL-12, consistent with a reduction in the number of intrahepatic macrophages (Figures 11K-11N).

[0194] These data indicate that PSMP may promote fibrosis by affecting macrophage infiltration and production of pro-inflammatory cytokines. Example 6 Materials and methods for research into liver fibrosis

[0195] This example describes materials and methods for the studies on liver fibrosis described in Examples 1-5. Human samples

[0196] All human liver samples were collected at the hospital. Cirrhotic samples were collected from HBV-induced cirrhosis, HCV-induced cirrhosis, primary biliary cirrhosis, and alcohol-induced cirrhosis. Normal liver tissue was collected from patients undergoing liver resection for non-neoplastic diseases, including hepatic adenoma and focal nodular hyperplasia. animal

[0197] Specific pathogen-free C57BL / 6 mice were purchased from the Peking Experimental Animal Center (Beijing, China). PSMP knockout mice were on a C57BL / 6 background. CCR2 knockout mice were on a C57BL / 6 background and obtained from Professor Yu Zhang (Department of Immunology, Peking University Health Science Center). PSMP knockout mice were crossed with CCR2 knockout mice to obtain mice with double knockout (DKO) of PSMP and CCR2. All animal experiments were performed in accordance with the Guidelines for the Care and Use of Laboratory Animals and approved by the Ethics Committee of Peking University Health Science Center. Mouse liver fibrosis model

[0198] For toxic liver fibrosis, 6- to 8-week-old male mice received intraperitoneal (ip) injections of CCl4 (1.0 ml per kg body weight, dissolved in corn oil at a 1:9 ratio) or vehicle (corn oil) twice weekly for 4 or 6 weeks. Mice were sacrificed 2 days after the final CCl4 (Aladdin, Shanghai, China) injection.

[0199] To induce cholestatic liver fibrosis, 6- to 8-week-old male mice underwent common bile duct ligation. Sham-operated mice underwent the same procedure but without ligation. Bile duct-ligated (BDL) mice developed cholestasis and associated fibrosis over a 14-day period. Antibody treatment

[0200] Monoclonal hybridoma cells expressing a neutralizing antibody against PSMP (3D5; Pei et al., J. Immunol. 192: 1878-86, 2014) were injected into the peritoneal cavity of mice to obtain ascites. mAb 3D5 in the ascites was purified using protein G. For protective treatment, mice were injected (ip) with 5 mg / kg of 3D5 twice a week for 4 weeks after each CCl4 injection. For therapeutic treatment, mice were injected (ip) with 5 mg / kg of 3D5 twice a week from week 4 to week 6 after each CCl4 injection. Construction and injection of murine adeno-associated virus 8 (AAV8)

[0201] Vigene AAV8 delivery system for overexpressing the human PSMP gene in mouse liver Constructed by Bioscience (Shangdong, China). Empty companion adenovirus (AAV8-null) served as a control. Vector genome titers were measured by qPCR using vector-specific primers. Mice were injected with 2 × 10 AAV8 vector genomes. 11 100 μl of virus containing 100 μg of the virus was injected into the tail vein. Cytometric Bead Assay (CBA)

[0202] Microspheres (A37304, Life Technologies) were coated with polyclonal rabbit anti-PSMP antibody according to the manufacturer's instructions. The coated beads were blocked with 20% FBS for 30 minutes and then added to the sample for 2 hours at room temperature. Monoclonal anti-PSMP antibody was then added for 1 hour. The mixture was suspended in PE goat anti-mouse antibody (eBiocience) and incubated for 30 minutes at room temperature. Data were acquired by flow cytometry.

[0203] Cytokines IL-6, CCL2, TNF-α, and IL-12p70 were detected using the BD™ Cytometric Bead Array (CBA) Mouse Inflammation Kit (552364, BD Biosciences) according to the manufacturer's instructions. Data were acquired by flow cytometry. Histological and immunohistochemical staining of the liver

[0204] Liver specimens were preserved in 10% formalin and cut into 4 μm-thick sections. Sections were then stained with hematoxylin and eosin (H&E) by standard procedures and Sirius Red stain for 1 hour. The Sirius Red stain was used to grade the degree of fibrosis according to the Ishak scoring system. Hepatic fibrosis was graded according to the Ishak scoring system: Grade 0 - no fibrosis; Grade 1 - fibrous expansion of some portal tracts with or without short fibrous septa; Grade 2 - fibrous expansion of most portal tracts with or without short fibrous septa; Grade 3 - fibrous expansion of most portal tracts with occasional portal-portal (PP) bridging; Grade 4 - fibrous expansion of portal tracts with prominent bridging (portal-portal (PP) and portal-central (PC) veins); Grade 5 - prominent bridging (PP and / or PC) with occasional nodules (incomplete cirrhosis); Grade 6 - probable or definite cirrhosis. The cumulative histological score ranged from 0 (completely normal) to 6 (severe disease with cirrhosis). Immunohistochemistry

[0205] For immunohistochemistry (IHC), liver specimens were fixed in 10% buffered formalin. PSMP IHC staining was performed as described by Pei et al., J. Immunol. 192: 1878-86, 2014. For α-SMA IHC staining, a monoclonal antibody against α-SMA (Abcam, ab7817) was used. Immunoblotting analysis

[0206] Proteins in the lysates were separated by SDS-PAGE electrophoresis and transferred to PVDF membranes (Millipore Corporation, MA, USA). Blots were incubated with a rabbit polyclonal antibody against α-SMA and visualized using an enhanced chemiluminescence (ECL) system. Densitometry measurements of each band were analyzed using Quantity-One Protein Analysis Software (Bio-Rad Laboratories, USA) and normalized to GAPDH (β-actin) as an internal control. Quantitative real-time PCR assay

[0207] Tissues or cells were homogenized in Trizol (Invitrogen). Reverse transcription (RT) reactions were performed using the MasterMix system (ABM Inc.). Real-time PCR was performed using SYBR Green PCR mix (ABM Inc.). All gene expressions were normalized to Gapdh. Determination of liver hydroxyproline

[0208] Liver samples obtained immediately after mouse sacrifice were subjected to a modified acid hydrolysis protocol to determine their hydroxyproline levels. The liver hydroxyproline content is expressed as mg of hydroxyproline per gram of liver. Serum ALT

[0209] Liver function in mice with liver injury was monitored based on serum levels of AST and ALT, enzyme activities expressed as units per liter (U / L). Flow cytometry

[0210] Liver mononuclear cells were obtained, blocked in 5% fetal bovine serum for 20 min, and then stained with CD45-FITC, CD11b-PE, CCR2-APC, Ly6C-PerCP Cy5.5, Ly6G-PerCP Cy5.5, F4 / 80-APC, and CD11c-APC (BioLegend) for 30 min at 4° C. Data were acquired on a FACS caliber (BD Biosciences) and analyzed using FlowJo 7.6 (TreeStar, Ashland, OR, USA). statistical analysis

[0211] All data are expressed as mean ± standard error of the mean. Statistical differences between two groups were analyzed by Student's t-test using GraphPad Prism 6.0 (GraphPad Software, San Diego, CA, USA). Statistically significant differences between two groups were * 0.01 <p<0.05、 ** 0.001 <p<0.01、および *** Represented by p<0.001. Example 7 PSMP expression during pulmonary fibrosis and the effects of PSMP deficiency or neutralization

[0212] The effects of PSMP expression and PSMP deficiency during pulmonary fibrosis were analyzed using the bleomycin model of fibrosis. In a mouse model, neutralization of PSMP signaling attenuates bleomycin-induced pulmonary fibrosis. method

[0213] Six- to eight-week-old female C57Bl / 6J (WT) mice were used in this study. After intraperitoneal anesthesia, the animals underwent tracheotomy. A single injection containing 3.5 U / kg of bleomycin (BLM) (Nippon Kayaku Co., Ltd.) diluted in saline, or a single injection containing saline alone (NS) (control group), was instilled into the trachea. Fourteen days after bleomycin administration, the animals were euthanized, bronchoalveolar lavage was performed, and lung samples were collected for biochemical and histological analysis. Total lung collagen content was measured using Masson's trichrome staining (Masson's The PSMP-neutralizing antibody 3D5 was injected (ip) once a week for 3 weeks after a single BLM injection, followed by 10 mg / kg of PBS.

[0214] Student's t-test was used for comparison between two groups. A one-tailed methodology was used for all tests. A P value of less than 0.05 was considered significant. Replicates consisted of at least three independent experiments. result

[0215] In the lung tissue of animals instilled with bleomycin, upregulation of PSMP was observed (Figs. 12A and 12C), which was consistent with the phenomenon observed in bronchoalveolar lavage (Fig. 12B). Compared to WT animals, PSMP - / - Mice showed reduced weight loss (Fig. 12D). Consistent with this, the increase in lung collagen content observed in bleomycin-instilled WT animals was due to the increased risk of PSMP - / - The expression of α-SMA, a marker of fibroblast activation, was significantly reduced in 3D5-treated mice compared with PBS-treated mice (Fig. 12E). Expression of α-SMA, a marker of fibroblast activation, was significantly reduced in 3D5-treated mice compared with PBS-treated mice (Fig. 12F). Immunohistochemical analysis revealed significant PSMP staining in human lungs with airway fibrosis, but not in normal tissue (Fig. 12G). Example 8 Deficiency of PSMP confers protection from liver fibrosis in a nonalcoholic steatohepatitis (NASH) model method

[0216] Two-day-old infant mice were subcutaneously administered 200 μg of streptozotocin (STZ, Sigma, MO, USA). When mice reached 4 weeks of age, they were started on a high-fat diet (HFD, consisting of 60 kcal% fat) and continued throughout the experimental period up to 12 weeks. Fibrosis was assessed by Sirius Red staining on paraffin sections. result

[0217] PSMP - / - A significant reduction in fibrosis was observed in the liver tissue of mice (FIG. 13). Example 9 Treatment of acute GVHD with anti-PSMP neutralizing antibodies method

[0218] In this study, 6- to 8-week-old male BABL / c mice served as recipients, and C57BL / 6 mice served as donors. After the mice were sacrificed, splenic CD3+ T cells and bone marrow cells from the C57BL / 6 mice were isolated. 12 hours before injecting the cells into BABL / c mice, the recipient mice were incubated with Co. 60 The CD3+ cells and bone marrow cells were exposed to two doses of radiation (4 Gy each, total 8 Gy). 6 pcs and 1×10 7 The mixture was mixed at a ratio of 0.01 to 0.01 and injected intravenously into BABL / c mice. During the one-month period following injection (acute GVHD (aGVHD) period), the weight and survival rate of the mice in each of the two groups (mAb 3D5-treated group and IgG control group) were monitored every two days.

[0219] 3D5, a neutralizing antibody to PSMP, was injected intraperitoneally at a dose of 5 mg / kg weekly after cell injection, and mouse IgG was used as a negative control treatment. result

[0220] Anti-PSMP neutralizing antibodies extended the survival time of aGVHD mice. Mice that lost more than 30% of their body weight were defined as non-survivable and were sacrificed. As shown in Figure 14, aGVHD mice died approximately 19 days after injection, while mice in the 3D5-treated group died approximately 24 days after injection. The survival times of aGVHD mice in the two groups of mice treated with IgG or 3D5 were summarized, and significant differences were calculated. The survival times of aGVHD mice showed significant differences between the IgG-treated group and the 3D5-treated group. Example 10 Neutralization or deficiency of PSMP signaling attenuates hepatic steatosis and fibrosis in a chronic ethanol feeding model method Mouse model

[0221] The chronic ethanol feeding model uses commercially available control and ethanol Lieber-DeCarli diets that are isocaloric and have the same composition with respect to fat (35% of calories) and protein (18% of calories). The carbohydrate content is 47% of total calories (dextrin-maltose) in the control diet and 11% of total calories in the ethanol diet, with up to 36% of carbohydrate calories replaced by ethanol.

[0222] For the chronic ethanol feeding model using the ethanol Lieber-DeCarli diet, 10-week-old female mice (approximately 22 g body weight) were acclimated to a liquid diet by feeding a control diet for 5 days. The percentage of calories derived from ethanol was gradually increased over a 1-week period, and then maintained on a 5% ethanol diet for 8 weeks. Mice were pair-fed throughout the experiment.

[0223] For the chronic and excessive alcohol feeding model, 8-week-old female mice (approximately 20 g body weight) were acclimated to a liquid diet by feeding a control diet for 5 days. The percentage of calories derived from ethanol was gradually increased over a 1-week period, and then mice were maintained on a 5% ethanol diet for 10 days. On day 11, mice were gavaged with a single dose of ethanol (5 g per kg body weight, 31.5% ethanol). Gavage was always performed early in the morning. After gavage, mice were maintained on the ethanol diet and kept in cages on a warming blanket with circulating water. After gavage, mice were sluggish but conscious and returned to normal behavior within 4–6 hours. Mice were always euthanized 9 hours after gavage.

[0224] For antibody treatment in the chronic ethanol-feeding model, mice were injected (ip) with 10 mg / kg of the PSMP-neutralizing antibody 3D5 or PBS once a week for 4 weeks (at 1-week midpoints starting at week 5 and continuing through week 8 of alcohol feeding). For antibody treatment in the chronic + binge alcohol-feeding model, mice were injected (ip) with 20 mg / kg of 3D5 or PBS 1 hour before gavage. Serum lipid content

[0225] Serum triglyceride or cholesterol content was determined using a triglyceride or cholesterol assay kit (Zhong Sheng, Beijing, China). Histopathological examination

[0226] Hematoxylin and eosin (H&E) staining and Sirius red staining were performed to examine liver morphology and assess liver fibrosis. Liver fibrosis was quantified by Sirius red staining. Liver specimens were fixed in 10% neutral buffered formalin, embedded in paraffin, and cut into 4 μm sections. To analyze intrahepatic fat accumulation, 10 μm sections were cut from frozen samples and stained with Oil Red O (Solarbio Science & Technology, Co., Ltd. Beijing, China) for 10 minutes. These slides were rinsed with water, counterstained with Mayer's hematoxylin, and analyzed by light microscopy. The positive areas were calculated using at least a 10x (×200) field of view. Immunohistochemical staining of the liver

[0227] For immunohistochemical staining, sections were blocked with 10% goat serum solution for 30 minutes at room temperature, then incubated with a primary antibody against α-SMA (Abcam, ab32575) overnight at 4°C. Sections were incubated with a secondary antibody (ZSGB-Bio) and 3,3-diaminobenzidine (DAB), followed by brief counterstaining with H&E stain and mounting with neutral gum. Biochemical assays

[0228] Serum levels of alanine aminotransferase (ALT) were measured using standard enzymatic procedures according to the manufacturer's instructions (Nanjing Jiancheng Bioengineering Institute, Nanjing, China). Serum and liver triglyceride or cholesterol contents were determined using triglyceride or cholesterol assay kits (Zhong Sheng, Beijing, China). Reverse transcription and quantitative real-time polymerase chain reaction (RT-qPCR)

[0229] Tissues or cells were homogenized in TRIzol (Life Technologies, 15596018). Reverse transcription was performed using Hifair II 1st Strand cDNA Synthesis SuperMix (Yeasen Biotech, 11120ES60). Quantitative real-time polymerase chain reaction was performed using Hieff qPCR SYBR Green Master mix (Yeasen Biotech, 11202ES03). All gene expression levels were normalized to Gapdh. result

[0230] Analysis of serum lipid content confirmed that after ethanol feeding, 3D5-treated mice had lower triglyceride and cholesterol levels than PBS-treated mice (Figures 15B and 15C). Liver hematoxylin and eosin (H&E) and oil red O staining revealed that ethanol-fed mice treated with 3D5 had lower degrees of steatosis and fibrosis than PBS-treated mice (Figures 15D–15F). Liver expression levels of Ppara and Srebp-1 mRNA were measured by RT-qPCR. Consistent with these findings, ethanol-fed mice treated with 3D5 downregulated Srebp-1 expression and upregulated Ppara expression compared with PBS (Figures 15G and 15H). Liver fibrosis was further determined by Sirius red staining. As illustrated in Figures 15I and 15J, mice in the PBS-treated group had greater Sirius red staining than mice treated with 3D5. Furthermore, the composition of immune cells in the liver was investigated by flow cytometry analysis in a chronic and excessive alcohol feeding model. In mice treated with 3D5, the infiltrating liver macrophages (iMΦ, F4 / 80 + CD11b high ) accumulation was reduced (Fig. 15K).

[0231] In the chronic and excessive alcohol feeding model (Figure 15L), analysis of liver lipid content confirmed that after ethanol feeding, 3D5-treated mice had lower levels of triglycerides and cholesterol than PBS-treated mice (Figures 15M and 15N). Liver hematoxylin and eosin (H&E) revealed that ethanol-fed mice treated with 3D5 had a lower degree of steatosis than ethanol-fed mice treated with PBS (Figure 15O). The composition of immune cells in the liver was also investigated by flow cytometry analysis. In 3D5-treated mice, infiltrating liver macrophages (iMΦ, F4 / 80 + CD11b high ) accumulation was reduced (Figure 15P).

[0232] In the chronic + excessive alcohol feeding model, Psmps were detected as assessed by H&E and Oil Red O staining. - / - Mice showed significantly attenuated liver injury and steatosis compared to WT mice (Figures 15Q and 15R). Serum levels of alanine aminotransferase (ALT) were significantly elevated in Psmp mice. - / - It was shown that Psmp expression was reduced in mice, indicating an improvement in liver injury (Figure 15S). - / - Srebp-1 expression was downregulated and Ppara expression was upregulated in mice compared with WT mice (Figures 15T and 15U). Example 11 PSMP expression is upregulated in mouse liver injury and fibrosis method

[0233] To induce acute liver injury, 6- to 8-week-old male WT mice and Psmp mice were cultured. - / - Mice were challenged with a single ip injection of acetaminophen (APAP, 300 mg / kg body weight, dissolved in warm PBS (55°C) (Sigma-Aldrich) (n=5 / group). Control mice received a similar volume of vehicle. Mice were euthanized 24 hours after injection.

[0234] For the 5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet, 8-week-old male WT mice were fed a diet supplemented with 0.1% DDC (Sigma-Aldrich, St. Louis, MO) for 4 weeks (n = 5 / group). Control mice received standard mouse chow. For antibody treatment, mice were injected (ip) with 5 mg / kg of PSMP-neutralizing antibody 3D5 or mIgG twice weekly from week 3 to week 4. result

[0235] In mouse models of liver fibrosis induced by the DDC diet or acute liver injury caused by acetaminophen (APAP) treatment, PSMP expression was significantly increased compared to livers from control mice (Figures 16A-16C). Example 12 PSMP deficiency protects against liver fibrosis in DDC mice

[0236] To analyze the role of PSMP in liver fibrosis due to other causes, the DDC diet model (see Example 11, Methods) was used. The DDC diet model reproduces the clinical features of human biliary fibrosis. After 4 weeks of DDC feeding, Psmp - / - Mice also showed a significant reduction in Sirius Red-positive areas and α-SMA expression (Figures 17A-17D). - / - Significant reductions in the mRNA levels of Acta2, Col1a1, Tgfb1, Timp1, and Pdgfr in mice were also observed (Figures 17E-17I). Example 13 Neutralization of PSMP signaling attenuates DDC-induced liver fibrosis in mice

[0237] To evaluate the therapeutic potential of 3D5 against DDC diet-induced liver fibrosis (see Example 11, Methods), mice were treated with 3D5 from week 3 to week 4 (Figure 18A). H&E staining and Sirius red staining assays showed reduced liver injury and fibrosis after 3D5 treatment compared with the mIgG-treated group (Figures 18B-18D). Accordingly, 3D5 significantly reduced α-SMA expression in fibrotic livers (Figures 18B and 18E). Quantitative real-time PCR (qRT-PCR) assays showed that 3D5 also significantly reduced the mRNA levels of fibrogenic genes (Acta2, Col1a1) (Figures 18F and 18G). Example 14 PSMP deficiency confers protection from liver fibrosis in MCD mice method

[0238] On a methionine-choline deficient (MCD) diet, 8-week-old male WT mice and Psmp - / - Mice were fed an MCD diet (Research Diets, A02082002B) for 6 weeks (n=5 / group). Control mice received standard mouse chow. result

[0239] We established a mouse model of nonalcoholic steatohepatitis (NASH) induced by a methionine-choline-deficient (MCD) diet. After 6 weeks of MCD feeding, PSMP expression was significantly increased in wild-type mice (Fig. 19A). - / - Psmp mice showed significantly reduced regional staining with Sirius Red compared to WT mice (Figures 19B-19D). - / - A significant decrease in the mRNA expression levels of Acta2, Col1a1, and Tgfb1 in the liver of mice was also detected (Figures 19E-19G). Example 15 Treatment with a PSMP-neutralizing antibody ameliorates liver fibrosis in mice in an MCD-induced NASH model method

[0240] For antibody treatment, 8-week-old male mice were fed a methionine- and choline-deficient (MCD) diet for 5.5 weeks. Mice were injected (ip) once a week with 10 mg / kg of the PSMP-neutralizing antibody 3D5 or PBS from week 4 to week 5.5. For other antibody treatment, 8-week-old male mice were fed a MCD diet for 8 weeks. Mice were injected (ip) once a week with 10 mg / kg of the PSMP-neutralizing antibody 3D5 or PBS from week 5 to week 8. result

[0241] To evaluate the therapeutic potential of the PSMP-neutralizing antibody 3D5 on liver fibrosis induced by the MCD diet, mice were treated with 3D5 from week 4 to week 5.5 (Figure 20A). H&E and Sirius Red staining assays and their quantification showed reduced liver injury and fibrosis after 3D5 treatment compared with the PBS-treated group (Figures 20B-20D). Further 3D5 treatment was performed in MCD mice from week 5 to week 8 (Figure 20E). Sirius Red and α-SMA immunohistochemical staining and their quantification showed reduced liver fibrosis after 3D5 treatment compared with the PBS-treated group (Figures 20F-20I). Example 16 Deficiency or neutralization of PSMP ameliorates acute kidney injury (AKI) and chronic kidney disease (CKD) in mice method Mouse model

[0242] In the UUO group, the left ureter of 6-week-old male mice (approximately 20 g) was exposed through a left flank incision in the abdomen and ligated twice with 4-0 silk suture. For treatment, mice were injected (ip) with 5 mg / kg of 3D5 or PBS on days 1 and 4 after UUO. Mice were sacrificed 5 days later.

[0243] In the glycerol-induced rhabdomyolysis group, mice were intramuscularly injected with 7.5 ml / kg of 50% glycerol diluted in PBS into the thigh muscle. For treatment, mice were injected (ip) with 5 mg / kg of 3D5 or PBS after the glycerol injection. Mice were sacrificed 48 hours later.

[0244] The left kidney was then removed, sectioned transversely, and fixed in 4% paraformaldehyde for histopathological examination or snap-frozen in liquid nitrogen for other experiments. Biochemical assays

[0245] Determination of serum creatinine and blood urea nitrogen levels is of great value in helping to confirm renal function in clinical settings. Serum levels of creatinine and blood urea nitrogen (BUN) were measured using assay kits according to the manufacturer's instructions (Nanjing Jiancheng Bioengineering Institute, Nanjing, China). Histopathological examination and immunohistochemical analysis

[0246] Renal tissues fixed in paraformaldehyde were embedded in paraffin and sectioned at 4 μm. Sections were dewaxed in xylene, rehydrated through decreasing ethanol concentrations, and stained with Sirius Red. For immunohistochemical staining, sections were blocked with 10% goat serum solution for 30 minutes at room temperature and then incubated overnight at 4°C with a primary antibody against α-SMA (Abcam, ab32575). Sections were incubated with a secondary antibody (ZSGB-Bio) and 3,3-diaminobenzidine (DAB), followed by brief counterstaining with H&E stain and mounting with neutral gum. Positive areas were calculated using at least a 10x (×200) field of view. result PSMP expression is upregulated in human renal disease

[0247] To determine whether PSMP expression is associated with renal disease, we first investigated PSMP levels by immunohistochemistry. Human normal kidney tissue from nearby clear cell carcinoma showed negative or weak expression of PSMP. PSMP was upregulated in renal tubules from patients with lupus nephritis, IgA nephritis, and membranous glomerulonephritis (Figures 21A and 21B). Deficiency and neutralization of PSMP attenuates glycerol-induced rhabdomyolysis

[0248] Psmp in glycerol-induced rhabdomyolysis - / - Mice showed significantly attenuated renal injury compared to WT mice, as assessed by serum creatinine (Figure 21C). Concurrently, we investigated the therapeutic effect of the PSMP-neutralizing antibody 3D5 on mice with glycerol-induced rhabdomyolysis, as determined using serum creatinine and BUN levels (Figures 21D and 21E). Neutralization of PSMP signaling attenuates renal fibrosis in a UUO model

[0249] To investigate the therapeutic effect of the PSMP-neutralizing antibody 3D5 on mice with UUO-induced renal fibrosis, mice were treated with 3D5 on days 1 and 4 after UUO (FIG. 21F). Sirius red staining assay showed reduced renal fibrosis after treatment with 3D5 compared with the PBS-treated group (FIGS. 21G and 21H; gray bars—3D5-treated group, black bars—PBS-treated group). Example 17 Treatment of chronic GVHD with anti-PSMP neutralizing antibodies

[0250] Chronic GVHD is a syndrome with diverse clinical features that manifests as an autoimmune-like syndrome. cGVHD organ damage in mice can involve the skin, gastrointestinal tract, liver, lungs, salivary glands, or oral mucosa. Generally, see, for example, Schroeder et al., Dis. See model Mech. 4: 318-333, 2011. method

[0251] In this study, a chronic GVHD (cGVHD) model was established using 7 × 10 spleen cells from 6-week-old male DBA2 mice. 7 The cells were established by injecting 6-week-old female B6D2F1 mice without irradiation. The anti-PSMP neutralizing antibody 3D5 was injected intraperitoneally at a dose of 2.5 mg / kg weekly after cell injection, and mouse IgG was used as a negative control treatment. Mice were monitored for weight and appearance, and markers of liver function were detected 80 days after splenocyte injection. result

[0252] The anti-PSMP neutralizing antibody 3D5 reversed the progression of cGVHD in mice. As shown in Figure 22A, there was no difference in body weight between the control and 3D5-treated groups. On day 40 after splenocyte injection, fur loss was observed in the eyelid area of ​​mice in the control group, but not in the neutralizing antibody 3D5 group. As shown in Figure 22B, fur loss gradually became more severe from day 40 to day 80, but no fur loss was observed in the neutralizing antibody 3D5 group. On day 80 after splenocyte injection, peripheral blood was collected from the two groups to detect markers of liver function. ALT and AST, markers of liver injury, were significantly higher in serum samples from the control group than in the 3D5-treated group (Figures 22C and 22D). Example 18 Treatment of lupus with anti-PSMP neutralizing antibodies method

[0253] The male DBA / 2 → female B6D2F1 graft-versus-host disease model is also known as a murine lupus nephritis model. In this study, the murine lupus model was established by injecting 7 × 10 spleen cells from 6-week-old male DBA / 2 mice. 7The cells were then injected into 6-week-old female B6D2F1 mice. Mouse IgG or the anti-PSMP neutralizing antibody 3D5 was injected intraperitoneally at a dose of 2.5 mg / kg weekly after cell injection. Markers of renal function were detected 80 days after splenocyte injection. Blood was collected from the tail vein of the mice, and serum samples were obtained for ELISA to detect total IgG in the serum. Total IgG was detected using a Sigma Mouse IgG ELISA kit (catalog no. 11333151001) according to the manufacturer's protocol. To detect autoantibody deposition within the glomeruli, kidneys were embedded and snap-frozen in liquid nitrogen. Sections (8 μm) were fixed in acetone and stained with FITC-conjugated anti-mouse IgG (BD Biosciences). Fluorescence was examined by confocal microscopy. result

[0254] Since PSMP expression was found to be upregulated in renal tissue from patients with lupus nephritis (see Example 16 and Figures 21A and 21B), we investigated the effect of the PSMP-neutralizing antibody 3D5 on a mouse lupus model. Eighty days after splenocyte injection, the renal function of mice treated with 3D5 was examined and compared with that of the control group. As indicated by serum creatinine and blood urea nitrogen (BUN) levels, renal function was significantly improved in the PSMP-neutralizing antibody 3D5 group compared with the control group (Figures 23A and 23B). Renal injury in lupus may be related to autoantibodies deposited in renal tissue. This study showed that the PSMP-neutralizing antibody 3D5 could reduce serum total IgG levels and autoantibody deposition in glomeruli compared with the control group (Figures 23C and 23D), indicating that the neutralizing antibody against PSMP 3D5 has the potential for renal protection during lupus. Embodiment

[0255] Embodiment 1. A method for treating liver fibrosis, comprising administering to a subject having liver fibrosis an effective amount of a PC3-secreted microprotein (PSMP) antagonist.

[0256] Embodiment 2. The method of embodiment 1, wherein the PSMP antagonist is a soluble protein that specifically binds to PSMP.

[0257] Embodiment 3. The method of embodiment 2, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO:4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO:5.

[0258] Embodiment 4 The method of embodiment 2, wherein the soluble protein is an antibody.

[0259] Embodiment 5. The method of embodiment 4, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO:4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO:5.

[0260] Embodiment 6. The antibody has a complementarity determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref is the CDR-H3 of SEQ ID NO:4.

[0261] Embodiment 7. CDR-H1 Ab is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab7. The method of embodiment 6, wherein is CDR-H3 of SEQ ID NO: 4.

[0262] Embodiment 8. The method of embodiment 6, wherein each VH CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0263] Embodiment 9. The method of embodiment 7, wherein each VH CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0264] Embodiment 10. Each VH CDR is defined according to the Chothia definition for a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ref has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref 9. The method of embodiment 8, wherein the nucleotide sequence of SEQ ID NO: 4 is a sequence of amino acids 99-108 of SEQ ID NO: 4.

[0265] Embodiment 11. CDR-H1 Ab has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab 11. The method of embodiment 10, wherein said VLKL has the amino acid sequence set forth in residues 99-108 of SEQ ID NO:4.

[0266] Embodiment 12. The antibody has a complementarity determining region (CDR) CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Refand CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref 12. The method of any one of embodiments 5 to 11, wherein is CDR-L1 of SEQ ID NO: 5.

[0267] Embodiment 13. CDR-L1 Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab is the CDR-L3 of SEQ ID NO:5.

[0268] Embodiment 14. The method of embodiment 12, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0269] Embodiment 15. The method of embodiment 13, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0270] Embodiment 16. Each VL CDR is defined according to the Chothia definition for a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref 15. The method of embodiment 14, wherein said VLKL has the amino acid sequence set forth in residues 89-97 of SEQ ID NO:5.

[0271] Embodiment 17. CDR-L1 Ab has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ab17. The method of embodiment 16, wherein the VLVI has the amino acid sequence set forth in residues 89-97 of SEQ ID NO:5.

[0272] Embodiment 18 The method of embodiment 11, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4.

[0273] Embodiment 19. The method of embodiment 17, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5.

[0274] Embodiment 20 The method of any one of embodiments 4 to 17, wherein the antibody is a humanized or chimeric antibody.

[0275] Embodiment 21 The method of embodiment 4 or 5, wherein the antibody is a human antibody.

[0276] Embodiment 22 The method of any one of embodiments 4 to 19, wherein the antibody is a single-chain antibody.

[0277] Embodiment 23 The method of any one of embodiments 4 to 19, wherein the antibody is a bispecific antibody.

[0278] Embodiment 24 The method of any one of embodiments 4 to 19, wherein the antibody further comprises an immunoglobulin constant region.

[0279] Embodiment 25 The method of any one of embodiments 1 to 24, wherein the liver fibrosis has progressed to cirrhosis.

[0280] Embodiment 26 The method of any one of embodiments 1 to 25, wherein the liver fibrosis is hepatitis B virus (HBV)-induced liver fibrosis, hepatitis C virus (HCV)-induced liver fibrosis, or alcohol-induced liver fibrosis.

[0281] Embodiment 27. The method of embodiment 25, wherein the cirrhosis is primary biliary cirrhosis.

[0282] Embodiment 28 The method of any one of embodiments 1 to 25, wherein the liver fibrosis is associated with non-alcoholic fatty liver disease.

[0283] Embodiment 29. The method of embodiment 28, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH).

[0284] Embodiment 30 The method of any one of embodiments 1 to 29, wherein the treatment of liver fibrosis is a combination therapy.

[0285] Embodiment 31. A PC3 secreted microprotein (PSMP) antagonist for use in the treatment of liver fibrosis.

[0286] Embodiment 32. Use of a PC3 secreted microprotein (PSMP) antagonist in the manufacture of a medicament for treating liver fibrosis.

[0287] Embodiment 33. The PSMP antagonist of embodiment 31 or the use of embodiment 32, wherein the PSMP antagonist is a soluble protein that specifically binds to PSMP.

[0288] Embodiment 34. The PSMP antagonist or use of embodiment 33, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0289] Embodiment 35 The PSMP antagonist or use of embodiment 33, wherein the soluble protein is an antibody.

[0290] Embodiment 36. The PSMP antagonist or use of embodiment 35, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0291] Embodiment 37. The antibody has a complementarity determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref is the CDR-H3 of SEQ ID NO:4.

[0292] Embodiment 38. CDR-H1 Ab is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab is the CDR-H3 of SEQ ID NO:4.

[0293] Embodiment 39. The PSMP antagonist or use of embodiment 37, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition of a CDR.

[0294] Embodiment 40. The PSMP antagonist or use of embodiment 38, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition of a CDR.

[0295] Embodiment 41. Each VH CDR is defined according to the Chothia definition for a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Refhas the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref 40. The PSMP antagonist or use of embodiment 39, wherein the PSMP antagonist or use has the amino acid sequence shown in residues 99 to 108 of SEQ ID NO:4.

[0296] Embodiment 42. CDR-H1 Ab has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab 42. The PSMP antagonist or use of embodiment 41, wherein said PSMP antagonist or use has the amino acid sequence shown in residues 99 to 108 of SEQ ID NO:4.

[0297] Embodiment 43. The antibody has a complementarity determining region (CDR) CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref is the CDR-L1 of SEQ ID NO:5.

[0298] Embodiment 44. CDR-L1 Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab is the CDR-L3 of SEQ ID NO:5.

[0299] Embodiment 45. The PSMP antagonist or use of embodiment 43, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0300] Embodiment 46. The PSMP antagonist or use of embodiment 44, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0301] Embodiment 47. Each VL CDR is defined according to the Chothia definition for a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref 46. ​​The PSMP antagonist or use of embodiment 45, wherein said PSMP antagonist or use has the amino acid sequence set forth in residues 89-97 of SEQ ID NO:5.

[0302] Embodiment 48. CDR-L1 Ab has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ab 48. The PSMP antagonist or use of embodiment 47, wherein said PSMP antagonist or use has the amino acid sequence shown in residues 89 to 97 of SEQ ID NO:5.

[0303] Embodiment 49. The PSMP antagonist or use of embodiment 42, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4.

[0304] Embodiment 50. The PSMP antagonist or use of embodiment 48, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5.

[0305] Embodiment 51 The PSMP antagonist or use of any one of embodiments 35 to 48, wherein the antibody is a humanized or chimeric antibody.

[0306] Embodiment 52 The PSMP antagonist or use of embodiment 35 or 36, wherein the antibody is a human antibody.

[0307] Embodiment 53 The PSMP antagonist or use of any one of embodiments 35 to 50, wherein the antibody is a single-chain antibody.

[0308] Embodiment 54 The PSMP antagonist or use of any one of embodiments 35 to 50, wherein the antibody is a bispecific antibody.

[0309] Embodiment 55 The PSMP antagonist or use of any one of embodiments 35 to 50, wherein the antibody further comprises an immunoglobulin constant region.

[0310] Embodiment 56 The PSMP antagonist or use of any one of embodiments 31 to 55, wherein the liver fibrosis has progressed to cirrhosis.

[0311] Embodiment 57. The PSMP antagonist or use of any one of embodiments 31 to 56, wherein the liver fibrosis is hepatitis B virus (HBV)-induced liver fibrosis, hepatitis C virus (HCV)-induced liver fibrosis, or alcohol-induced liver fibrosis.

[0312] Embodiment 58. The PSMP antagonist or use of embodiment 56, wherein the cirrhosis is primary biliary cirrhosis.

[0313] Embodiment 39 The PSMP antagonist or use of any one of embodiments 31 to 56, wherein the liver fibrosis is associated with non-alcoholic fatty liver disease.

[0314] Embodiment 60. The PSMP antagonist or use of embodiment 59, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH).

[0315] Embodiment 61 The PSMP antagonist or use of any one of embodiments 31 to 60, wherein the treatment of liver fibrosis is a combination therapy.

[0316] Embodiment 62. A method for treating pulmonary fibrosis, comprising administering to a subject having pulmonary fibrosis an effective amount of a PC3 secreted microprotein (PSMP) antagonist.

[0317] Embodiment 63 The method of embodiment 62, wherein the PSMP antagonist is a soluble protein that specifically binds to PSMP.

[0318] Embodiment 64. The method of embodiment 63, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0319] Embodiment 65. The method of embodiment 63, wherein the soluble protein is an antibody.

[0320] Embodiment 66. The method of embodiment 65, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0321] Embodiment 67. The antibody has a complementarity determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref, and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref 67. The method of embodiment 66, wherein is CDR-H3 of SEQ ID NO: 4.

[0322] Embodiment 68. CDR-H1 Ab is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab 68. The method of embodiment 67, wherein is CDR-H3 of SEQ ID NO: 4.

[0323] Embodiment 69. The method of embodiment 67, wherein each VH CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0324] Embodiment 70. The method of embodiment 68, wherein each VH CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0325] Embodiment 71. Each VH CDR is defined according to the Chothia definition for a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ref has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref 70. The method of embodiment 69, wherein said mAb has the amino acid sequence set forth in residues 99-108 of SEQ ID NO:4.

[0326] Embodiment 72. CDR-H1 Ab has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab72. The method of embodiment 71, wherein said mAb has the amino acid sequence set forth in residues 99-108 of SEQ ID NO:4.

[0327] Embodiment 73. The antibody has a complementarity determining region (CDR) CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref 73. The method of any one of embodiments 66 to 72, wherein is CDR-L1 of SEQ ID NO: 5.

[0328] Embodiment 74. CDR-L1 Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab is the CDR-L3 of SEQ ID NO: 5.

[0329] Embodiment 75. The method of embodiment 73, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0330] Embodiment 76. The method of embodiment 74, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0331] Embodiment 77. Each VL CDR is defined according to the Chothia definition for a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref 76. The method of embodiment 75, wherein said mAb has the amino acid sequence set forth in residues 89-97 of SEQ ID NO:5.

[0332] Embodiment 78. CDR-L1 Ab has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ab 78. The method of embodiment 77, wherein said mAb has the amino acid sequence set forth in residues 89-97 of SEQ ID NO:5.

[0333] Embodiment 79. The method of embodiment 72, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4.

[0334] Embodiment 80. The method of embodiment 78, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5.

[0335] Embodiment 81 The method of any one of embodiments 65 to 78, wherein the antibody is a humanized or chimeric antibody.

[0336] Embodiment 82. The method of embodiment 65 or 66, wherein the antibody is a human antibody.

[0337] Embodiment 83 The method of any one of embodiments 65 to 80, wherein the antibody is a single-chain antibody.

[0338] Embodiment 84. The method of any one of embodiments 65 to 80, wherein the antibody is a bispecific antibody.

[0339] Embodiment 85. The method of any one of embodiments 65 to 80, wherein the antibody further comprises an immunoglobulin constant region.

[0340] Embodiment 86 The method of any one of embodiments 65 to 84, wherein the treatment of pulmonary fibrosis is a combination therapy.

[0341] Embodiment 87. A PC3 secreted microprotein (PSMP) antagonist for use in the treatment of pulmonary fibrosis.

[0342] Embodiment 88. Use of a PC3 secreted microprotein (PSMP) antagonist in the manufacture of a medicament for treating pulmonary fibrosis.

[0343] Embodiment 89. The PSMP antagonist of embodiment 87 or the use of embodiment 88, wherein the PSMP antagonist is a soluble protein that specifically binds to PSMP.

[0344] Embodiment 90. The PSMP antagonist or use of embodiment 89, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0345] Embodiment 91 The PSMP antagonist or use of embodiment 89, wherein the soluble protein is an antibody.

[0346] Embodiment 92. The PSMP antagonist or use of embodiment 91, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0347] Embodiment 93. The antibody has a complementarity determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2Ref , and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref is the CDR-H3 of SEQ ID NO:4.

[0348] Embodiment 94. CDR-H1 Ab is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab is the CDR-H3 of SEQ ID NO:4.

[0349] Embodiment 95. The PSMP antagonist or use of embodiment 93, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition of a CDR.

[0350] Embodiment 96. The PSMP antagonist or use of embodiment 94, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition for a CDR.

[0351] Embodiment 97. Each VH CDR is defined according to the Chothia definition for a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ref has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref 96. The PSMP antagonist or use of embodiment 95, wherein said PSMP antagonist or use has the amino acid sequence shown in residues 99 to 108 of SEQ ID NO:4.

[0352] Embodiment 98. CDR-H1 Abhas the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab 98. The PSMP antagonist or use of embodiment 97, wherein said PSMP antagonist or use has the amino acid sequence shown in residues 99 to 108 of SEQ ID NO:4.

[0353] Embodiment 99. The antibody has a complementarity determining region (CDR) CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref The PSMP antagonist or use of any one of embodiments 92 to 98, wherein is CDR-L1 of SEQ ID NO:5.

[0354] Embodiment 100. CDR-L1 Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab is the CDR-L3 of SEQ ID NO:5.

[0355] Embodiment 101. The PSMP antagonist or use of embodiment 99, wherein each VL CDR is defined according to the Chothia, Kabat, AbM, or contact definition for a CDR.

[0356] Embodiment 102. The PSMP antagonist or use of embodiment 100, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0357] Embodiment 103. Each VL CDR is defined according to the Chothia definition for a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref 102. The PSMP antagonist or use of embodiment 101, wherein said PSMP antagonist or use has the amino acid sequence shown in residues 89-97 of SEQ ID NO:5.

[0358] Embodiment 104. CDR-L1 Ab has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ab 104. The PSMP antagonist or use of embodiment 103, wherein said PSMP antagonist or use has the amino acid sequence set forth in residues 89 to 97 of SEQ ID NO:5.

[0359] Embodiment 105. The PSMP antagonist or use of embodiment 98, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4.

[0360] Embodiment 106 The PSMP antagonist or use of embodiment 104, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5.

[0361] Embodiment 107. The PSMP antagonist or use of any one of embodiments 91 to 104, wherein the antibody is a humanized or chimeric antibody.

[0362] Embodiment 108. The PSMP antagonist or use of embodiment 91 or 92, wherein the antibody is a human antibody.

[0363] Embodiment 109. The PSMP antagonist or use of any one of embodiments 91 to 106, wherein the antibody is a single-chain antibody.

[0364] Embodiment 110. The PSMP antagonist or use of any one of embodiments 91 to 106, wherein the antibody is a bispecific antibody.

[0365] Embodiment 111. The PSMP antagonist or use of any one of embodiments 91 to 106, wherein the antibody further comprises an immunoglobulin constant region.

[0366] Embodiment 112. The PSMP antagonist or use of any one of embodiments 87 to 111, wherein the treatment of pulmonary fibrosis is a combination therapy.

[0367] Embodiment 113. A method for treating a disease selected from the group consisting of non-alcoholic fatty liver disease (NAFLD), alcoholic liver disease (ALD), primary sclerosing cholangitis (PSC), and primary biliary cholangitis (PBC), comprising administering to a subject having NAFLD, ALD, PSC, or PBC an effective amount of a PC3-secreted microprotein (PSMP) antagonist.

[0368] Embodiment 114. The method of embodiment 113, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH).

[0369] Embodiment 115 The method of embodiment 133 or 114, wherein the PSMP antagonist is a soluble protein that specifically binds to PSMP.

[0370] Embodiment 116. The method of embodiment 115, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0371] Embodiment 117. The method of embodiment 115, wherein the soluble protein is an antibody.

[0372] Embodiment 118. The method of embodiment 117, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0373] Embodiment 119. The antibody has a complementarity determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref is the CDR-H3 of SEQ ID NO: 4.

[0374] Embodiment 120. CDR-H1 Ab is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab is the CDR-H3 of SEQ ID NO: 4.

[0375] Embodiment 121. The method of embodiment 119, wherein each VH CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0376] Embodiment 122. The method of embodiment 120, wherein each VH CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0377] Embodiment 123. Each VH CDR is defined according to the Chothia definition for a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ref has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref 122. The method of embodiment 121, wherein said mAb has the amino acid sequence set forth in residues 99-108 of SEQ ID NO:4.

[0378] Embodiment 124. CDR-H1 Ab has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab 124. The method of embodiment 123, wherein said mAb has the amino acid sequence set forth in residues 99 to 108 of SEQ ID NO:4.

[0379] Embodiment 125. The antibody has a complementarity-determining region (CDR) CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Refis CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref The method of any one of embodiments 118 to 124, wherein is CDR-L1 of SEQ ID NO: 5.

[0380] Embodiment 126. CDR-L1 Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab is the CDR-L3 of SEQ ID NO: 5.

[0381] Embodiment 127. The method of embodiment 125, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0382] Embodiment 128. The method of embodiment 126, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0383] Embodiment 129. Each VL CDR is defined according to the Chothia definition for a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref 128. The method of embodiment 127, wherein said mAb has the amino acid sequence set forth in residues 89-97 of SEQ ID NO:5.

[0384] Embodiment 130. CDR-L1 Ab has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ab 130. The method of embodiment 129, wherein said mAb has the amino acid sequence set forth in residues 89-97 of SEQ ID NO:5.

[0385] Embodiment 131. The method of embodiment 124, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4.

[0386] Embodiment 132. The method of embodiment 130, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5.

[0387] Embodiment 133. The method of any one of embodiments 117 to 130, wherein the antibody is a humanized or chimeric antibody.

[0388] Embodiment 134. The method of embodiment 117 or 118, wherein the antibody is a human antibody.

[0389] Embodiment 135. The method of any one of embodiments 117 to 132, wherein the antibody is a single-chain antibody.

[0390] Embodiment 136. The method of any one of embodiments 117 to 132, wherein the antibody is a bispecific antibody.

[0391] Embodiment 137. The method of any one of embodiments 117 to 132, wherein the antibody further comprises an immunoglobulin constant region.

[0392] Embodiment 138. The method of any one of embodiments 116 to 136, wherein the treatment of non-alcoholic fatty liver disease is a combination therapy.

[0393] Embodiment 139. A PC3-secreted microprotein (PSMP) antagonist for use in the treatment of a disease selected from the group consisting of non-alcoholic fatty liver disease (NAFLD), alcoholic liver disease (ALD), primary sclerosing cholangitis (PSC), and primary biliary cholangitis (PBC).

[0394] Embodiment 140. Use of a PC3-secreted microprotein (PSMP) antagonist in the manufacture of a medicament for treating a disease selected from the group consisting of non-alcoholic fatty liver disease (NAFLD), alcoholic liver disease (ALD), primary sclerosing cholangitis (PSC), and primary biliary cholangitis (PBC).

[0395] Embodiment 141. The PSMP antagonist of embodiment 139 or the use of embodiment 140, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH).

[0396] Embodiment 142. The PSMP antagonist or use of any one of embodiments 139 to 141, wherein the PSMP antagonist is a soluble protein that specifically binds to PSMP.

[0397] Embodiment 143. The PSMP antagonist or use of embodiment 142, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0398] Embodiment 144. The PSMP antagonist or use of embodiment 142, wherein the soluble protein is an antibody.

[0399] Embodiment 145. The PSMP antagonist or use of embodiment 144, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0400] Embodiment 146. The antibody has a complementarity-determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Aband wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref is the CDR-H3 of SEQ ID NO:4.

[0401] Embodiment 147. CDR-H1 Ab is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab is the CDR-H3 of SEQ ID NO:4.

[0402] Embodiment 148. The PSMP antagonist or use of embodiment 146, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition of a CDR.

[0403] Embodiment 149. The PSMP antagonist or use of embodiment 147, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition of a CDR.

[0404] Embodiment 150. Each VH CDR is defined according to the Chothia definition for a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ref has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref 149. The PSMP antagonist or use of embodiment 148, wherein said PSMP antagonist or use has the amino acid sequence set forth in residues 99 to 108 of SEQ ID NO:4.

[0405] Embodiment 151. CDR-H1 Ab has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab 151. The PSMP antagonist or use of embodiment 150, wherein said PSMP antagonist or use has the amino acid sequence set forth in residues 99 to 108 of SEQ ID NO:4.

[0406] Embodiment 152. The antibody has a complementarity-determining region (CDR) CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref is CDR-L1 of SEQ ID NO:5.

[0407] Embodiment 153. CDR-L1 Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab is the CDR-L3 of SEQ ID NO:5.

[0408] Embodiment 154. The PSMP antagonist or use of embodiment 152, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0409] Embodiment 155. The PSMP antagonist or use of embodiment 153, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0410] Embodiment 156. Each VL CDR is defined according to the Chothia definition for a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref 155. The PSMP antagonist or use of embodiment 154, wherein said PSMP antagonist or use has the amino acid sequence set forth in residues 89 to 97 of SEQ ID NO:5.

[0411] Embodiment 157. CDR-L1 Ab has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ab 157. The PSMP antagonist or use of embodiment 156, wherein said PSMP antagonist or use has the amino acid sequence set forth in residues 89 to 97 of SEQ ID NO:5.

[0412] Embodiment 158. The PSMP antagonist or use of embodiment 151, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4.

[0413] Embodiment 159. The PSMP antagonist or use of embodiment 157, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5.

[0414] Embodiment 160. The PSMP antagonist or use of any one of embodiments 144 to 157, wherein the antibody is a humanized or chimeric antibody.

[0415] Embodiment 161. The PSMP antagonist or use of embodiment 144 or 145, wherein the antibody is a human antibody.

[0416] Embodiment 162. The PSMP antagonist or use of any one of embodiments 144 to 159, wherein the antibody is a single-chain antibody.

[0417] Embodiment 163. The PSMP antagonist or use of any one of embodiments 144 to 159, wherein the antibody is a bispecific antibody.

[0418] Embodiment 164. The PSMP antagonist or use of any one of embodiments 144 to 159, wherein the antibody further comprises an immunoglobulin constant region.

[0419] Embodiment 165. The PSMP antagonist or use according to any one of embodiments 139 to 164, wherein the treatment of non-alcoholic fatty liver disease is a combination therapy.

[0420] Embodiment 166. The method of any one of embodiments 1 to 25, wherein the liver fibrosis is associated with a disease or disorder selected from the group consisting of alcoholic liver disease (ALD), alcoholic hepatitis, alcoholic cirrhosis, chronic hepatitis B, chronic hepatitis C, chronic hepatitis D, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), hemochromatosis, cystic fibrosis, Wilson's disease, biliary atresia, alpha 1-antitrypsin deficiency, galactosemia, glycogen storage disease, inherited digestive disorders, Alagille syndrome, autoimmune hepatitis, primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC), infection, drug-induced liver injury, and Budd-Chiari syndrome.

[0421] Embodiment 167. The PSMP antagonist or use of any one of embodiments 31 to 56, wherein the liver fibrosis is associated with a disease or disorder selected from the group consisting of alcoholic liver disease (ALD), alcoholic hepatitis, alcoholic cirrhosis, chronic hepatitis B, chronic hepatitis C, chronic hepatitis D, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), hemochromatosis, cystic fibrosis, Wilson's disease, biliary atresia, alpha 1-antitrypsin deficiency, galactosemia, glycogen storage disease, inherited digestive disorders, Alagille syndrome, autoimmune hepatitis, primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC), infection, drug-induced liver injury, and Budd-Chiari syndrome.

[0422] Embodiment 168. The method of any one of embodiments 62 to 86, wherein the pulmonary fibrosis is associated with a disease or disorder selected from the group consisting of dermatomyositis, polymyositis, mixed connective tissue disease, systemic lupus erythematosus, rheumatoid arthritis, sarcoidosis, scleroderma, pneumonia, chronic radiation pneumonitis, pneumoconiosis, infectious diseases, and drug-induced lung injury.

[0423] Embodiment 169. The PSMP antagonist or use of any one of embodiments 87 to 112, wherein the pulmonary fibrosis is associated with a disease or disorder selected from the group consisting of dermatomyositis, polymyositis, mixed connective tissue disease, systemic lupus erythematosus, rheumatoid arthritis, sarcoidosis, scleroderma, pneumonia, chronic radiation pneumonitis, pneumoconiosis, infectious diseases, and drug-induced lung injury.

[0424] Embodiment 170. A method for treating a disease or disorder selected from the group consisting of graft-versus-host disease (GVHD), systemic lupus erythematosus (SLE), and lupus nephritis, comprising administering to a subject with GVHD or SLE an effective amount of a PC3-secreted microprotein (PSMP) antagonist.

[0425] Embodiment 171 The method of embodiment 170, wherein the PSMP antagonist is a soluble protein that specifically binds to PSMP.

[0426] Embodiment 172. The method of embodiment 171, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0427] Embodiment 173. The method of embodiment 171, wherein the soluble protein is an antibody.

[0428] Embodiment 174. The method of embodiment 173, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0429] Embodiment 175. The antibody has a complementarity-determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref is the CDR-H3 of SEQ ID NO: 4.

[0430] Embodiment 176. CDR-H1 Ab is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab is the CDR-H3 of SEQ ID NO: 4.

[0431] Embodiment 177. The method of embodiment 175, wherein each VH CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0432] Embodiment 178. The method of embodiment 176, wherein each VH CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0433] Embodiment 179. Each VH CDR is defined according to the Chothia definition for a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ref has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref 178. The method of embodiment 177, wherein said mAb has the amino acid sequence set forth in residues 99 to 108 of SEQ ID NO:4.

[0434] Embodiment 180. CDR-H1 Ab has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab 180. The method of embodiment 179, wherein said mAb has the amino acid sequence set forth in residues 99 to 108 of SEQ ID NO:4.

[0435] Embodiment 181. The antibody has a complementarity-determining region (CDR) CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Refis CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref 181. The method of any one of embodiments 174 to 180, wherein is CDR-L1 of SEQ ID NO: 5.

[0436] Embodiment 182. CDR-L1 Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab is the CDR-L3 of SEQ ID NO: 5.

[0437] Embodiment 183. The method of embodiment 181, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0438] Embodiment 184. The method of embodiment 182, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0439] Embodiment 185. Each VL CDR is defined according to the Chothia definition for a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref 184. The method of embodiment 183, wherein said VLVI has the amino acid sequence set forth in residues 89-97 of SEQ ID NO:5.

[0440] Embodiment 186. CDR-L1 Ab has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ab 186. The method of embodiment 185, wherein said mAb has the amino acid sequence set forth in residues 89-97 of SEQ ID NO:5.

[0441] Embodiment 187. The method of embodiment 180, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4.

[0442] Embodiment 188. The method of embodiment 186, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5.

[0443] Embodiment 189. The method of any one of embodiments 173 to 181, wherein the antibody is a humanized antibody or a chimeric antibody.

[0444] Embodiment 190. The method of embodiment 173 or 174, wherein the antibody is a human antibody.

[0445] Embodiment 191. The method of any one of embodiments 173 to 188, wherein the antibody is a single-chain antibody.

[0446] Embodiment 192. The method of any one of embodiments 173 to 188, wherein the antibody is a bispecific antibody.

[0447] Embodiment 193. The method of any one of embodiments 173 to 188, wherein the antibody further comprises an immunoglobulin constant region.

[0448] Embodiment 194. The method of any one of embodiments 173 to 192, wherein the treatment of the disease is a combination therapy.

[0449] Embodiment 195. A PC3 secreted microprotein (PSMP) antagonist for use in the treatment of a disease or disorder selected from the group consisting of graft-versus-host disease (GVHD), systemic lupus erythematosus (SLE), and lupus nephritis.

[0450] Embodiment 196. Use of a PC3 secreted microprotein (PSMP) antagonist in the manufacture of a medicament for treating a disease or disorder selected from the group consisting of graft-versus-host disease (GVHD), systemic lupus erythematosus (SLE), and lupus nephritis.

[0451] Embodiment 197. The PSMP antagonist of embodiment 195 or the use of embodiment 196, wherein the PSMP antagonist is a soluble protein that specifically binds to PSMP.

[0452] Embodiment 198. The PSMP antagonist or use of embodiment 197, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0453] Embodiment 199. The PSMP antagonist or use of embodiment 197, wherein the soluble protein is an antibody.

[0454] Embodiment 200. The PSMP antagonist or use of embodiment 199, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0455] Embodiment 201. The antibody has a complementarity determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref is the CDR-H3 of SEQ ID NO:4.

[0456] Embodiment 202. CDR-H1Ab is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab is the CDR-H3 of SEQ ID NO:4.

[0457] Embodiment 203. The PSMP antagonist or use of embodiment 201, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition of a CDR.

[0458] Embodiment 204. The PSMP antagonist or use of embodiment 202, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition of a CDR.

[0459] Embodiment 205. Each VH CDR is defined according to the Chothia definition for a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ref has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref 204. The PSMP antagonist or use of embodiment 203, wherein said PSMP antagonist or use has the amino acid sequence shown in residues 99 to 108 of SEQ ID NO:4.

[0460] Embodiment 206. CDR-H1 Ab has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab 206. The PSMP antagonist or use of embodiment 205, wherein said PSMP antagonist or use has the amino acid sequence shown in residues 99 to 108 of SEQ ID NO:4.

[0461] Embodiment 207. The antibody has a complementarity determining region (CDR) CDR-L1 Ab , CDR-L2Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref The PSMP antagonist or use of any one of embodiments 200 to 206, wherein is CDR-L1 of SEQ ID NO:5.

[0462] Embodiment 208. CDR-L1 Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab is the CDR-L3 of SEQ ID NO:5.

[0463] Embodiment 209. The PSMP antagonist or use of embodiment 207, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0464] Embodiment 210. The PSMP antagonist or use of embodiment 208, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0465] Embodiment 211. Each VL CDR is defined according to the Chothia definition for a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref200. The PSMP antagonist or use of embodiment 209, wherein said PSMP antagonist or use has the amino acid sequence set forth in residues 89 to 97 of SEQ ID NO:5.

[0466] Embodiment 212. CDR-L1 Ab has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ab 212. The PSMP antagonist or use of embodiment 211, wherein said PSMP antagonist or use has the amino acid sequence shown in residues 89-97 of SEQ ID NO:5.

[0467] Embodiment 213. The PSMP antagonist or use of embodiment 206, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4.

[0468] Embodiment 214. The PSMP antagonist or use of embodiment 212, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5.

[0469] Embodiment 215. The PSMP antagonist or use of any one of embodiments 199 to 212, wherein the antibody is a humanized or chimeric antibody.

[0470] Embodiment 216. The method of embodiment 199 or 200, wherein the antibody is a human antibody.

[0471] Embodiment 217. The PSMP antagonist or use of any one of embodiments 199 to 214, wherein the antibody is a single-chain antibody.

[0472] Embodiment 218. The PSMP antagonist or use of any one of embodiments 199 to 214, wherein the antibody is a bispecific antibody.

[0473] Embodiment 219. The PSMP antagonist or use of any one of embodiments 199 to 214, wherein the antibody further comprises an immunoglobulin constant region.

[0474] Embodiment 220. The PSMP antagonist or use according to any one of embodiments 195 to 219, wherein the treatment of the disease is a combination therapy.

[0475] Embodiment 221. A method for treating renal fibrosis, comprising administering to a subject having renal fibrosis an effective amount of a PC3-secreted microprotein (PSMP) antagonist.

[0476] Embodiment 222. A method for treating acute kidney injury (AKI) or chronic kidney disease, comprising administering to a subject having AKI or CKD an effective amount of a PC3 secreted microprotein (PSMP) antagonist.

[0477] Embodiment 223 The method of embodiment 221 or 222, wherein the PSMP antagonist is a soluble protein that specifically binds to PSMP.

[0478] Embodiment 224. The method of embodiment 223, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0479] Embodiment 225. The method of embodiment 223, wherein the soluble protein is an antibody.

[0480] Embodiment 226. The method of embodiment 225, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0481] Embodiment 227. The antibody has a complementarity-determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Aband wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref 227. The method of embodiment 226, wherein is CDR-H3 of SEQ ID NO: 4.

[0482] Embodiment 228. CDR-H1 Ab is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab 228. The method of embodiment 227, wherein is CDR-H3 of SEQ ID NO: 4.

[0483] Embodiment 229. The method of embodiment 227, wherein each VH CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0484] Embodiment 230. The method of embodiment 228, wherein each VH CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0485] Embodiment 231. Each VH CDR is defined according to the Chothia definition for a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ref has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref 229. The method of embodiment 229, wherein said mAb has the amino acid sequence set forth in residues 99 to 108 of SEQ ID NO:4.

[0486] Embodiment 232. CDR-H1 Abhas the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab 232. The method of embodiment 231, wherein said mAb has the amino acid sequence set forth in residues 99-108 of SEQ ID NO:4.

[0487] Embodiment 233. The antibody has a complementarity-determining region (CDR) CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref The method of any one of embodiments 226 to 232, wherein is CDR-L1 of SEQ ID NO: 5.

[0488] Embodiment 234. CDR-L1 Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab 234. The method of embodiment 233, wherein is the CDR-L3 of SEQ ID NO: 5.

[0489] Embodiment 235. The method of embodiment 233, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0490] Embodiment 236. The method of embodiment 234, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0491] Embodiment 237. Each VL CDR is defined according to the Chothia definition for a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref 236. The method of embodiment 235, wherein said mAb has the amino acid sequence set forth in residues 89-97 of SEQ ID NO:5.

[0492] Embodiment 238. CDR-L1 Ab has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ab 238. The method of embodiment 237, wherein said mAb has the amino acid sequence set forth in residues 89 to 97 of SEQ ID NO:5.

[0493] Embodiment 239. The method of embodiment 232, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4.

[0494] Embodiment 240. The method of embodiment 238, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5.

[0495] Embodiment 241. The method of any one of embodiments 225 to 233, wherein the antibody is a humanized or chimeric antibody.

[0496] Embodiment 242. The method of embodiment 225 or 226, wherein the antibody is a human antibody.

[0497] Embodiment 243. The method of any one of embodiments 225 to 240, wherein the antibody is a single-chain antibody.

[0498] Embodiment 244. The method of any one of embodiments 225 to 240, wherein the antibody is a bispecific antibody.

[0499] Embodiment 245. The method of any one of embodiments 225 to 240, wherein the antibody further comprises an immunoglobulin constant region.

[0500] Embodiment 246. The method of any one of embodiments 225 to 244, wherein the treatment of the disease is a combination therapy.

[0501] Embodiment 247. The method of any one of embodiments 221 and 223 to 246, wherein the renal fibrosis is associated with a disease or disorder selected from the group consisting of IgA nephropathy, membranoproliferative glomerulonephritis, membranous glomerulonephritis, crescentic glomerulonephritis, diabetic nephropathy, hypertensive nephropathy, lupus nephritis, hepatic nephropathy, polycystic kidney disease, Alport syndrome, Fabry disease, primary hyperoxaluria, cystinosis, focal segmental glomerulosclerosis (FSGS) caused by coenzyme Q10-related gene mutations, complement 3 glomerulonephritis, acute or subacute immune complex glomerulonephritis, renal vasculitis, systemic lupus erythematosus (SLE), recent-onset renal artery stenosis (fibromuscular or vasculitic), diabetic kidney disease, chronic uric acid nephropathy, toxic nephropathy, bacterial pyelonephritis, viral nephropathy, multiple myeloma, and obstructive nephropathy.

[0502] Embodiment 248. The method of any one of embodiments 222 to 246, wherein the AKI is rhabdomyolysis-induced.

[0503] Embodiment 249. The method of any one of embodiments 222 to 246, wherein the CKD is caused by a disease or disorder selected from the group consisting of IgA nephropathy, membranoproliferative glomerulonephritis, membranous glomerulonephritis, crescentic glomerulonephritis, diabetic nephropathy, hypertensive nephropathy, lupus nephritis, hepatic nephropathy, polycystic kidney disease, Alport syndrome, Fabry disease, primary hyperoxaluria, cystinosis, focal segmental glomerulosclerosis (FSGS) caused by coenzyme Q10-related gene mutations, complement 3 glomerulonephritis, acute or subacute immune complex glomerulonephritis, renal vasculitis, systemic lupus erythematosus (SLE), recent-onset renal artery stenosis (fibromuscular or vasculitic), diabetic kidney disease, chronic uric acid nephropathy, toxic nephropathy, bacterial pyelonephritis, viral nephropathy, multiple myeloma, and obstructive nephropathy.

[0504] Embodiment 250. A PC3 secreted microprotein (PSMP) antagonist for use in the treatment of renal fibrosis.

[0505] Embodiment 251. A PC3 secreted microprotein (PSMP) antagonist for use in the treatment of acute kidney injury (AKI) or chronic kidney disease (CKD).

[0506] Embodiment 252. Use of a PC3 secreted microprotein (PSMP) antagonist in the manufacture of a medicament for treating renal fibrosis.

[0507] Embodiment 253. Use of a PC3 secreted microprotein (PSMP) antagonist in the manufacture of a medicament for treating acute kidney injury (AKI) and chronic kidney disease (CKD).

[0508] Embodiment 254. The PSMP antagonist of embodiment 250 or 251 or the use of embodiment 252 or 253, wherein the PSMP antagonist is a soluble protein that specifically binds to PSMP.

[0509] Embodiment 255. The PSMP antagonist or use of embodiment 254, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0510] Embodiment 256. The PSMP antagonist or use of embodiment 254, wherein the soluble protein is an antibody.

[0511] Embodiment 257. The PSMP antagonist or use of embodiment 256, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5.

[0512] Embodiment 258. The antibody has a complementarity-determining region (CDR) CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and CDR-H1 has three or fewer amino acid substitutions compared to Ref is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ref is the CDR-H3 of SEQ ID NO:4.

[0513] Embodiment 259. CDR-H1 Ab is CDR-H1 of SEQ ID NO: 4, and CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, and CDR-H3 Ab is the CDR-H3 of SEQ ID NO:4.

[0514] Embodiment 260. The PSMP antagonist or use of embodiment 258, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition of a CDR.

[0515] Embodiment 261. The PSMP antagonist or use of embodiment 259, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition of a CDR.

[0516] Embodiment 262. Each VH CDR is defined according to the Chothia definition for a CDR, and thus CDR-H1 Ref has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ref has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ref 261. The PSMP antagonist or use of embodiment 260, wherein said PSMP antagonist or use has the amino acid sequence set forth in residues 99 to 108 of SEQ ID NO:4.

[0517] Embodiment 263. CDR-H1 Ab has the amino acid sequence shown in residues 31 to 35 of SEQ ID NO: 4, and CDR-H2 Ab has the amino acid sequence shown in residues 50 to 69 of SEQ ID NO: 4, and CDR-H3 Ab 263. The PSMP antagonist or use of embodiment 262, wherein said PSMP antagonist or use has the amino acid sequence set forth in residues 99 to 108 of SEQ ID NO:4.

[0518] Embodiment 264. The antibody has a complementarity-determining region (CDR) CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Refand CDR-L1 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ref is CDR-L1 of SEQ ID NO: 5, and CDR-L3 Ref The PSMP antagonist or use of any one of embodiments 257 to 263, wherein is CDR-L1 of SEQ ID NO:5.

[0519] Embodiment 265. CDR-L1 Ab is CDR-L1 of SEQ ID NO: 5, and CDR-L2 Ab is CDR-L2 of SEQ ID NO: 5, and CDR-L3 Ab is the CDR-L3 of SEQ ID NO:5.

[0520] Embodiment 266. The PSMP antagonist or use of embodiment 264, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0521] Embodiment 267. The PSMP antagonist or use of embodiment 265, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR.

[0522] Embodiment 268. Each VL CDR is defined according to the Chothia definition for a CDR, and thus CDR-L1 Ref has the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ref has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ref 267. The PSMP antagonist or use of embodiment 266, wherein said PSMP antagonist or use has the amino acid sequence shown in residues 89 to 97 of SEQ ID NO:5.

[0523] Embodiment 269. CDR-L1 Abhas the amino acid sequence shown in residues 24 to 34 of SEQ ID NO: 5, and CDR-L2 Ab has the amino acid sequence shown in residues 50 to 56 of SEQ ID NO: 5, and CDR-L3 Ab 269. The PSMP antagonist or use of embodiment 268, wherein said PSMP antagonist or use has the amino acid sequence set forth in residues 89 to 97 of SEQ ID NO:5.

[0524] Embodiment 270. The PSMP antagonist or use of embodiment 263, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4.

[0525] Embodiment 271. The PSMP antagonist or use of embodiment 269, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5.

[0526] Embodiment 272. The PSMP antagonist or use of any one of embodiments 256 to 269, wherein the antibody is a humanized or chimeric antibody.

[0527] Embodiment 273. The PSMP antagonist or use of embodiment 256 or 257, wherein the antibody is a human antibody.

[0528] Embodiment 274. The PSMP antagonist or use of any one of embodiments 256 to 271, wherein the antibody is a single-chain antibody.

[0529] Embodiment 275. The PSMP antagonist or use of any one of embodiments 256 to 271, wherein the antibody is a bispecific antibody.

[0530] Embodiment 276. The PSMP antagonist or use of any one of embodiments 256 to 271, wherein the antibody further comprises an immunoglobulin constant region.

[0531] Embodiment 277. The PSMP antagonist or use according to any one of embodiments 250 to 276, wherein the treatment of the disease is a combination therapy.

[0532] Embodiment 278. The renal fibrosis is IgA nephropathy, membranoproliferative glomerulonephritis, membranous glomerulonephritis, crescentic glomerulonephritis, diabetic nephropathy, hypertensive nephropathy, lupus nephritis, hepatic nephropathy, polycystic kidney disease, Alport syndrome, Fabry disease, primary hyperoxaluria, cystinosis, focal segmental glomerulosclerosis (FSGS) caused by coenzyme Q10-related gene mutations, complement 3 glomerulonephritis, acute or subacute immune complex glomerulonephritis. The PSMP antagonist or use of any one of embodiments 250, 252 and 254 to 277, which is associated with a disease or disorder selected from the group consisting of glomerulonephritis, renal vasculitis, systemic lupus erythematosus (SLE), recent-onset renal artery stenosis (fibromuscular or vasculitic), diabetic kidney disease, chronic uric acid nephropathy, toxic nephropathy, bacterial pyelonephritis, viral nephropathy, multiple myeloma, and obstructive nephropathy.

[0533] Embodiment 279. The PSMP antagonist or use of any one of embodiments 251 and 253 to 277, wherein the AKI is rhabdomyolysis-induced.

[0534] Embodiment 280. CKD is selected from the group consisting of IgA nephropathy, membranoproliferative glomerulonephritis, membranous glomerulonephritis, crescentic glomerulonephritis, diabetic nephropathy, hypertensive nephropathy, lupus nephritis, hepatic nephropathy, polycystic kidney disease, Alport syndrome, Fabry disease, primary hyperoxaluria, cystinosis, focal segmental glomerulosclerosis (FSGS) caused by coenzyme Q10-related gene mutations, complement 3 glomerulonephritis, acute or subacute immune complex glomerulonephritis. The PSMP antagonist or use of any one of embodiments 251 and 253 to 277, wherein the renal artery stenosis is caused by a disease or disorder selected from the group consisting of uric acid nephritis, renal vasculitis, systemic lupus erythematosus (SLE), recent-onset renal artery stenosis (fibromuscular or vasculitic), diabetic kidney disease, chronic uric acid nephropathy, toxic nephropathy, bacterial pyelonephritis, viral nephropathy, multiple myeloma, and obstructive nephropathy.

[0535] From the foregoing, it will be understood that, although specific embodiments of the invention have been described herein for purposes of illustration, various modifications may be made without departing from the spirit and scope of the invention. Accordingly, the invention is not limited except as by the appended claims. All publications, patents, and patent applications cited herein are hereby incorporated by reference in their entirety for all purposes. The present invention provides, for example, the following items. (Item 1) A PC3 secreted microprotein (PSMP) antagonist for use in the treatment of liver fibrosis, pulmonary fibrosis, or renal fibrosis. (Item 2) Item 3. The PSMP antagonist according to Item 1, wherein the PSMP antagonist is for use in treating liver fibrosis, and optionally the liver fibrosis has progressed to cirrhosis. The liver fibrosis is hepatitis B virus (HBV)-induced liver fibrosis, hepatitis C virus (HCV)-induced liver fibrosis, or alcohol-induced liver fibrosis; or 3. The method of claim 2, wherein the liver fibrosis is associated with a disease selected from the group consisting of non-alcoholic fatty liver disease (NAFLD), alcoholic liver disease (ALD), primary sclerosing cholangitis (PSC), and primary biliary cholangitis (PBC), and optionally the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH). (Item 4) 2. The PSMP antagonist of item 1, wherein the PSMP antagonist is for use in treating pulmonary fibrosis, and optionally the pulmonary fibrosis is associated with drug-induced lung injury. (Item 5) the PSMP antagonist is for use in treating renal fibrosis; Optionally, the renal fibrosis is associated with a disease or disorder selected from the group consisting of lupus nephritis, IgA nephropathy, and membranous glomerulonephritis. (Item 6) A PC3-secreted microprotein (PSMP) antagonist for use in the treatment of a disease selected from the group consisting of non-alcoholic fatty liver disease (NAFLD), alcoholic liver disease (ALD), primary sclerosing cholangitis (PSC), and primary biliary cholangitis (PBC). (Item 7) 7. The PSMP antagonist according to item 6, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH). (Item 8) 1. A PC3 secreted microprotein (PSMP) antagonist for use in the treatment of acute kidney injury (AKI) or chronic kidney disease (CKD), comprising: Optionally, the AKI is rhabdomyolysis-induced; Optionally, the CKD is caused by a disease or disorder selected from the group consisting of lupus nephritis, IgA nephropathy, and membranous glomerulonephritis. PSMP antagonist. (Item 9) A PC3 secreted microprotein (PSMP) antagonist for use in the treatment of a disease or disorder selected from the group consisting of graft-versus-host disease (GVHD), systemic lupus erythematosus (SLE), and lupus nephritis. (Item 10) 10. The PSMP antagonist of any one of items 1 to 9, which is a soluble protein that specifically binds to PSMP. (Item 11) The PSMP antagonist of item 10, wherein the soluble protein competes for binding to PSMP with an antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5. (Item 12) 11. The PSMP antagonist of claim 10, wherein the soluble protein is an antibody. (Item 13) 13. The PSMP antagonist of Item 12, wherein the antibody competes for binding to PSMP with a second antibody comprising a heavy chain variable domain (VH) having the amino acid sequence set forth in SEQ ID NO: 4 and a light chain variable domain (VL) having the amino acid sequence set forth in SEQ ID NO: 5. (Item 14) The antibody has a complementarity determining region (CDR), CDR-H1 Ab , CDR-H2 Ab , and CDR-H3 Ab and wherein the set of VH CDRs is a reference set of CDRs, CDR-H1 Ref , CDR-H2 Ref , and CDR-H3 Ref and having three or fewer amino acid substitutions compared to CDR-H1 Ref is CDR-H1 of SEQ ID NO: 4, CDR-H2 Ref is CDR-H2 of SEQ ID NO: 4, CDR-H3 Ref is the CDR-H3 of SEQ ID NO: 4, Item 14. The PSMP antagonist according to item 13. (Item 15) CDR-H1 Ab is CDR-H1 of SEQ ID NO: 4, CDR-H2 Ab is CDR-H2 of SEQ ID NO: 4, CDR-H3 Ab is the CDR-H3 of SEQ ID NO: 4, Item 15. The PSMP antagonist according to item 14. (Item 16) 15. The PSMP antagonist of item 14, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition for a CDR. (Item 17) 16. The PSMP antagonist of item 15, wherein each VH CDR is defined according to the Chothia, Kabat, AbM, or contact definition for a CDR. (Item 18) Each VH CDR is defined according to the Chothia definition for a CDR, and therefore: CDR-H1 Ref has the amino acid sequence set forth in residues 31-35 of SEQ ID NO:4, CDR-H2 Ref has the amino acid sequence set forth in residues 50 to 69 of SEQ ID NO:4, CDR-H3 Ref has the amino acid sequence set forth in residues 99 to 108 of SEQ ID NO:4, Item 17. The PSMP antagonist according to item 16. (Item 19) CDR-H1 Ab has the amino acid sequence set forth in residues 31-35 of SEQ ID NO:4, CDR-H2 Ab has the amino acid sequence set forth in residues 50 to 69 of SEQ ID NO:4, CDR-H3 Ab has the amino acid sequence set forth in residues 99 to 108 of SEQ ID NO:4, Item 19. The PSMP antagonist according to item 18. (Item 20) The antibody has a complementarity determining region (CDR), CDR-L1 Ab , CDR-L2 Ab , and CDR-L3 Ab and wherein the set of VL CDRs is a reference set of CDRs, CDR-L1 Ref , CDR-L2 Ref , and CDR-L3 Ref and having three or fewer amino acid substitutions compared to CDR-L1 Ref is the CDR-L1 of SEQ ID NO: 5, CDR-L2 Ref is the CDR-L1 of SEQ ID NO: 5, CDR-L3 Ref is the CDR-L1 of SEQ ID NO: 5, 20. The PSMP antagonist of any one of items 14 to 19. (Item 21) CDR-L1 Abis the CDR-L1 of SEQ ID NO: 5, CDR-L2 Ab is the CDR-L2 of SEQ ID NO: 5, CDR-L3 Ab is the CDR-L3 of SEQ ID NO: 5, 21. The PSMP antagonist according to item 20. (Item 22) 21. The PSMP antagonist of item 20, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR. (Item 23) 22. The PSMP antagonist of item 21, wherein each VL CDR is defined according to the Chothia definition, the Kabat definition, the AbM definition, or the contact definition for a CDR. (Item 24) Each VL CDR is defined according to the Chothia definition for a CDR, and therefore: CDR-L1 Ref has the amino acid sequence set forth in residues 24 to 34 of SEQ ID NO:5, CDR-L2 Ref has the amino acid sequence set forth in residues 50-56 of SEQ ID NO:5, CDR-L3 Ref has the amino acid sequence set forth in residues 89 to 97 of SEQ ID NO:5; 23. The PSMP antagonist according to item 22. (Item 25) CDR-L1 Ab has the amino acid sequence set forth in residues 24 to 34 of SEQ ID NO:5, CDR-L2 Ab has the amino acid sequence set forth in residues 50-56 of SEQ ID NO:5, CDR-L3 Ab 25. The PSMP antagonist of item 24, wherein the PSMP antagonist has the amino acid sequence shown in residues 89 to 97 of SEQ ID NO:5. (Item 26) 20. The PSMP antagonist of item 19, wherein the antibody comprises a VH domain having the amino acid sequence set forth in SEQ ID NO:4. (Item 27) 26. The PSMP antagonist of item 25, wherein the antibody comprises a VL domain having the amino acid sequence set forth in SEQ ID NO:5. (Item 28) 26. The PSMP antagonist of any one of items 12 to 25, wherein the antibody is a humanized or chimeric antibody. (Item 29) The PSMP antagonist according to Item 12 or 13, wherein the antibody is a human antibody. 28. The PSMP antagonist of any one of items 12 to 27, wherein the antibody is a single chain antibody.

Claims

[Claim 1] The invention as set forth in the drawings.

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