Activin receptor type ii chimeras and methods of use thereof

Abstract

To provide polypeptides that include an extracellular ActRII chimera.SOLUTION: Provided is a polypeptide of the present invention including an extracellular ActRII chimera fused to an Fc domain monomer or moiety. The invention also features pharmaceutical compositions and methods of using the polypeptides to treat diseases and conditions involving weakness and atrophy of muscles, bone damage, low red blood cell levels (e.g., anemia or blood loss), low platelet levels (e.g., thrombocytopenia), low neutrophil levels (e.g., neutropenia), fibrosis, metabolic disorders, and / or pulmonary hypertension.SELECTED DRAWING: None

Description

[Background technology]

[0001] Sequence Listing The instant application contains a Sequence Listing which has been submitted electronically in ASCII format and is hereby incorporated by reference in its entirety.

[0002] Duchenne muscular dystrophy (DMD), facioscapulohumeral muscular dystrophy (FSHD), inclusion body myositis (IBM), and amyotrophic lateral sclerosis (ALS) are examples of muscle diseases involving the weakening and atrophy of muscles and / or motor neurons that control voluntary muscle movement. DMD is caused by mutations in the X-linked dystrophin gene and is characterized by progressive muscle degeneration and weakness in all skeletal muscles. FSHD particularly affects the skeletal muscles of the face, shoulders, upper arms, and lower limbs. IBM is an inflammatory muscle disease that primarily affects the muscles of the thighs and arms that control flexion of the fingers and wrists. ALS is a motor neuron disease characterized by muscle stiffness, muscle spasms, and muscle atrophy throughout the body due to the degeneration of motor neurons. Efforts to improve the treatment and survival of subjects with these devastating muscle diseases have yet to be successful.

[0003] Healthy bone undergoes constant remodeling, involving both bone destruction and bone growth. Bone growth is mediated by osteoblasts, while osteoclasts resorb bone. When these systems are out of balance, either through downregulation of the anabolic program, upregulation of the catabolic system, or a combination of both, pathology occurs, resulting in net bone loss. Therefore, controlling the balance of bone remodeling may be useful for promoting the healing of bone injuries and for treating disorders such as osteoporosis, which are associated with bone loss and demineralization.

[0004] Bone damage can result from a variety of underlying causes, including age- or cancer-related bone loss, genetic conditions, and adverse side effects of drug treatments. The World Health Organization estimates that osteoporosis alone affects 75 million people in the United States, Europe, and Japan and is a significant risk factor for bone damage. Overall bone loss generally represents a condition with few effective treatments. Instead, treatment focuses on immobility, exercise, and dietary modification rather than drugs that directly promote bone growth and increase bone density. Regarding osteoporosis, estrogen, calcitonin, osteocalcin in combination with vitamin K, or high-dose dietary calcium have all been used as therapeutic interventions. Other therapeutic approaches for osteoporosis include bisphosphonates, parathyroid hormone, parathyroid hormone-related protein, calcium receptor agonists, statins, anabolic steroids, lanthanum and strontium salts, and sodium fluoride. However, these treatments are often associated with unwanted side effects.

[0005] Fibrosis is the formation of excess connective tissue in an organ or tissue. Connective tissue formed in response to injury (e.g., injury) or as part of an immune response (e.g., inflammatory reaction) disrupts the structure and function of the organ or tissue in which it is formed, resulting in increased tissue stiffness. Fibrosis can occur in many organs and tissues in the body, including the lungs (e.g., pulmonary fibrosis, cystic fibrosis), liver (e.g., cirrhosis), heart (e.g., endomyocardial fibrosis or fibrosis after myocardial infarction), brain (e.g., glial scar formation), skin (e.g., keloid formation), kidneys (e.g., renal fibrosis), and eyes (e.g., corneal fibrosis), among others. Fibrosis is known to be associated with certain medical treatments (e.g., chemotherapy, radiation therapy, and surgery). Treatment options for patients with fibrosis are limited, and most treatments focus on improving quality of life or temporarily slowing disease progression.

[0006] Anemia is a global health problem with health consequences affecting both morbidity and mortality. In the United States alone, the prevalence of anemia nearly doubled from 2003 to 2012. Symptoms of anemia include fatigue, weakness, shortness of breath, palpitations, and decreased cognitive ability. Children, pregnant women, women of reproductive age, and the elderly are at highest risk of developing anemia. The most common form of anemia is iron deficiency anemia, but anemia can also be caused by chronic disease, blood loss, and red blood cell destruction. While iron deficiency anemia can be treated with iron supplements, many other forms of anemia, such as aplastic anemia, anemia of chronic disease, and hemolytic anemia, may require blood transfusions.

[0007] Thrombocytopenia is a condition characterized by abnormally low levels of platelets (also called thrombocytes) in the blood. It occurs when too few platelets are produced in the bone marrow or when too many are destroyed or accumulated in an enlarged spleen. Patients with thrombocytopenia may experience internal or external bleeding, bleeding under the skin, and / or purpura. Treatment of thrombocytopenia varies depending on its cause and severity and is primarily focused on preventing death or disability from bleeding. Certain types of thrombocytopenia (e.g., immune thrombocytopenia) can be treated with corticosteroids, while others may require splenectomy or platelet transfusions.

[0008] Neutropenia is a condition characterized by an abnormally low number of neutrophils in the blood. Neutrophils typically comprise 45%–75% of all white blood cells in the bloodstream and serve as the primary defense against infection. A reduced neutrophil count can make infections difficult to control and increase the risk of death from infection. In patients with severe neutropenia, infections can rapidly become severe or fatal. While antibiotics are used to treat infections in neutropenic patients, treatment for neutropenia itself is limited and primarily involves stimulating white blood cell production using growth factors such as colony-stimulating factors. Blood transfusions have not been proven effective.

[0009] Myelodysplastic syndromes (MDS) are a group of bone marrow disorders characterized by ineffective hematopoiesis, often accompanied by a dramatic expansion of progenitor cells that fail to mature into functional blood cells. In the United States, there are 60,000 to 170,000 patients with MDS, with 15,000 to 20,000 new cases reported annually. MDS primarily affects older adults, with approximately 75% of patients aged 60 years or older at diagnosis. Median survival ranges from approximately nine years for very low-risk patients to less than one year for high-risk patients. Anemia is the most common consequence of ineffective hematopoiesis due to decreased red blood cell production in MDS patients, affecting 90% of MDS patients. Another consequence is thrombocytopenia. Patients with MDS-associated anemia are commonly treated with red blood cell transfusions and erythropoiesis-stimulating agents (ESAs), although these therapies are not approved. MDS-associated thrombocytopenia is treated with platelet transfusions and platelet-stimulating agents.

[0010] Myelofibrosis is a rare type of bone marrow cancer that disrupts the normal production of blood cells. It can cause extensive scarring in the bone marrow, leading to severe anemia and low platelet counts. Symptoms of myelofibrosis include fatigue, bone pain, easy bruising, easy bleeding, and fever. Patients with advanced or high-risk myelofibrosis may require blood transfusions or bone marrow transplants. Other treatment options include androgen therapy and treatment with thalidomide or related drugs, which have known risks. For patients with intermediate-risk myelofibrosis, treatment is typically directed toward symptom management.

[0011] Pulmonary hypertension (PH) is a serious condition characterized by higher-than-normal pressure in the blood vessels between the lungs and the heart. PH can be classified into five major types, also known as WHO Groups I–V: arterial (PAH), venous (PH secondary to left-sided heart disease), hypoxic (PH caused by lung disease), thromboembolic (PH caused by chronic arterial obstruction, e.g., a blood clot), or other (PH with unknown or multifactorial mechanisms). PAH is characterized by increased pressure in the blood vessels of the lungs, caused by blockage or narrowing of the small blood vessels in the lungs due to scarring. This increases resistance to blood flow through the lungs, causing the right side of the heart to work harder, potentially leading to heart failure, reduced blood oxygenation, and shortened life expectancy. PAH can be idiopathic (e.g., no identifiable cause), hereditary (e.g., familial, often due to genetic mutations), or associated with drug use (e.g., methamphetamine or cocaine use), infection (e.g., HIV infection or schistosomiasis), cirrhosis, congenital heart abnormalities, or connective tissue / autoimmune diseases (e.g., scleroderma or lupus). Treatments for PH include vasodilators, anticoagulants, and supplemental oxygen; however, these treatments manage the symptoms of the disease rather than target the biological mechanisms that cause the disease.

[0012] Excess weight is a growing problem in much of the world; in 2016, approximately 39% of adults aged 18 years and older were found to be overweight, and approximately 13% of the global adult population was found to be obese. Increased visceral and subcutaneous fat contributes to the dysfunction of various organs. Excess weight is a risk factor for a range of complications, including diabetes (e.g., type 1 and type 2 diabetes), cardiovascular disease, and some forms of cancer. Insulin resistance, also associated with obesity, leads to increased insulin production in pancreatic tissue. When pancreatic beta cells are unable to produce enough insulin to meet demand, hyperglycemia develops, leading to the development of type 2 diabetes. Adipocytes, which increase in number with obesity, are thought to play a role in this process. Despite the prevalence of obesity and metabolic disease, few treatment options are available.

[0013] There is a need for new and effective treatments for muscle diseases, bone diseases, anemia, thrombocytopenia, neutropenia, myelofibrosis, myelodysplastic syndromes, fibrosis, PH, and metabolic diseases. Summary of the Invention

[0014] The present invention features polypeptides comprising an extracellular activin receptor type II (ActRII) chimera. In some embodiments, the polypeptides of the present invention comprise an extracellular ActRII chimera fused to the N- or C-terminus of an Fc domain monomer or another moiety. Such moieties can be attached by amino acids or other covalent bonds, which can increase the stability of the polypeptide. Polypeptides comprising an extracellular ActRII chimera fused to an Fc domain monomer can also form dimers (e.g., homodimers or heterodimers) through interaction between two Fc domain monomers. The polypeptides of the present invention can be used to increase lean body mass, muscle mass, and / or muscle strength in subjects with or at risk of developing diseases or conditions involving muscle weakness and atrophy, such as neuromuscular disease, sarcopenia, cachexia, disuse atrophy, treatment-related muscle loss or atrophy, muscle loss or atrophy associated with hypotonia, hypoxia, or burn injury. Polypeptides of the invention can also be used to increase bone mass or bone density in subjects having or at risk of developing a disease or condition involving bone damage, for example, osteoporosis (e.g., primary osteoporosis or secondary osteoporosis), osteopenia, osteopetrosis, fractures, bone loss associated with bone cancer or cancer metastasis, Paget's disease, renal osteodystrophy, treatment-related bone loss, osteogenesis imperfecta, bone loss associated with neuromuscular disease, bone loss due to burns (e.g., bone loss associated with burns), bone loss associated with anorexia, diet-related bone loss, bone loss associated with the treatment of obesity, bone loss associated with reduced gravity, or immobility.Additionally, the polypeptides of the invention can be used to increase red blood cell levels (e.g., increase hemoglobin levels, increase hematocrit, and / or increase red blood cell count), to promote or increase the maturation and / or differentiation of erythroid progenitor cells, to increase the maturation of late stage erythroid progenitor cells, or to mobilize early stage progenitor cells to the erythroid lineage in a subject in need thereof, e.g., a subject having or at risk of developing anemia or blood loss, or to mobilize early stage progenitor cells to the erythroid lineage in a subject in need thereof, e.g., a subject having or at risk of developing thrombocytopenia. Polypeptides of the present invention may be used to increase platelet levels (e.g., increase platelet count), to increase neutrophil levels (e.g., increase neutrophil count) in a subject in need thereof, e.g., a subject having or at risk of developing neutropenia, to prevent or reduce fibrosis in a subject having or at risk of developing fibrosis, or to treat, prevent, or delay the onset or progression of pulmonary hypertension (e.g., arterial, venous, hypoxic, thromboembolic, or other pulmonary hypertension) in a subject having or at risk of developing a metabolic disease, e.g., obesity, type 1 diabetes, or type 2 diabetes. Polypeptides of the present invention may also be used to reduce weight, reduce body fat, increase glucose clearance, increase insulin sensitivity, or reduce fasting insulin levels in a subject having or at risk of developing a metabolic disease, e.g., obesity, type 1 diabetes, or type 2 diabetes.Additionally, the polypeptides of the present invention can also be used to affect myostatin, activin (e.g., activin A and / or activin B), and / or bone morphogenetic protein 9 (BMP9) signaling in subjects having or at risk of developing a disease or condition, including muscle weakness and atrophy, bone damage or bone demineralization, low blood cell levels (e.g., low hemoglobin levels, low hematocrit, and / or low red blood cell count), low platelet levels (e.g., low platelet count), low neutrophil levels (e.g., low neutrophil count), fibrosis, pulmonary hypertension (e.g., arterial, venous, hypoxic, thromboembolic, or other pulmonary hypertension), or metabolic disease (e.g., obesity, type 1 diabetes, or type 2 diabetes).

[0015] Exemplary embodiments of the present invention are described in the following enumerated items. E1. A polypeptide comprising an extracellular ActRII chimera, said chimera comprising: GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRRHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 1), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFC CCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 2), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 3), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGSIEIVKQGCWL DDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 4), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 5), GAILGRAETRECIYYNANWELERTNQSGLER CEGEQX1KRLHCYASWRNSSGTIELVKKGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 6), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIELVKKGCWLDDX2X3CYDRQECVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 7),GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRRHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 8), GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNM CNEKFSYFPEMEVTQPTS (SEQ ID NO: 9), GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 10), GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGSIEIVKQGCWLDDX2X3CYDRT DCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 11), GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 12), GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASW RNSSGTIELVKKGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 13), and GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIELVKKGCWLDDX2X3CYDRQECVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 14), wherein X1 is D or R, X2 is I, F, E, D, Y, S, N, Q, or T, X3 is N or T, X4 is A or E, X5 is T or K, X6 is E or K, X7 is E or D, X8 is N or S, and X9 is Q, E, K, R, D, or N; and optionally, the chimera is truncated from the N-terminus by deletion of 1, 2, 3, 4, 5, 6, 7, 8, or 9 amino acids, wherein the chimera retains the two amino acids before the first cysteine.

[0016] E2. A polypeptide comprising an extracellular ActRII chimera, said chimera comprising: GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRRHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 15), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGN MCNEKFSYFPEMEVTQPTS (SEQ ID NO: 16), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 17), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGSIEIVKQGCWLDDX2X3CYD RTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 18), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 19), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCYA SWRNSSGTIELVKKGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 20), and GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIELVKKGCWLDDX2X3CYDRQECVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 21), wherein X1 is D or R, X2 is I, F, E, D, Y, S, N, Q, or T, X3 is N or T, X4 is A or E, X5 is T or K, X6 is E or K, X7 is E or D, X8 is N or S, and X9 is Q, E, K, R, D, or N; and optionally, the chimera is truncated from the N-terminus by deletion of 1, 2, 3, 4, 5, 6, 7, 8, or 9 amino acids, wherein the chimera retains the two amino acids before the first cysteine.

[0017] E3. The chimera: GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRRHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 1).

[0018] E4. The chimera: GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 2).

[0019] E5. The chimera: The polypeptide described in E1, having a sequence of: GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 3).

[0020] E6. The polypeptide of E1, wherein the chimera has the sequence: GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 4).

[0021] E7. The chimera: The polypeptide described in E1, having the sequence: GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 5).

[0022] E8. The chimera: The polypeptide described in E1, having a sequence of: GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIELVKKGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 6).

[0023] E9. The chimera: The polypeptide described in E1, having a sequence of: GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIELVKKGCWLDDX2X3CYDRQECVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 7).

[0024] E10. The chimera The polypeptide described in E1 having the sequence GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRRHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 8).

[0025] E11. The chimera: The polypeptide described in E1 having the sequence GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 9).

[0026] E12. The chimera The polypeptide described in E1 having the sequence GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 10).

[0027] E13. The chimera The polypeptide described in E1 having the sequence GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 11).

[0028] E14. The chimera: The polypeptide described in E1 having the sequence GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 12).

[0029] E15. The chimera The polypeptide described in E1 having the sequence GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIELVKKGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 13).

[0030] E16. The chimera The polypeptide described in E1 having the sequence GRGEAETRECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIELVKKGCWLDDX2X3CYDRQECVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 14).

[0031] E17. The chimera The polypeptide described in E2 having the sequence: GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRRHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 15).

[0032] E18. The polypeptide of E2, wherein said chimera has the sequence: GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCFATWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 16).

[0033] E19. The polypeptide of E2, wherein said chimera has the sequence: GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWKNISGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 17).

[0034] E20. The chimera The polypeptide described in E2 having a sequence of: GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGSIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 18).

[0035] E21. The chimera: The polypeptide described in E2 having the sequence: GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIEIVKQGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 19).

[0036] E22. The chimera The polypeptide described in E2 having the sequence: GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIELVKKGCWLDDX2X3CYDRTDCVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 20).

[0037] E23. The chimera The polypeptide described in E2 having the sequence: GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX1KRLHCYASWRNSSGTIELVKKGCWLDDX2X3CYDRQECVX4X5X6X7X8PX9VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 21).

[0038] E24. The polypeptide according to any one of E1 to E23, wherein X1 is D. E25. The polypeptide according to any one of E1 to E23, wherein X1 is R. E26. The polypeptide according to any one of E1 to E25, wherein X2 is I.

[0039] E27. The polypeptide according to any one of E1 to E25, wherein X2 is F. E28. The polypeptide according to any one of E1 to E25, wherein X2 is E. E29. The polypeptide according to any one of E1 to E25, wherein X2 is D.

[0040] E30. The polypeptide according to any one of E1 to E25, wherein X2 is Y. E31. The polypeptide according to any one of E1 to E25, wherein X2 is S. E32. The polypeptide according to any one of E1 to E25, wherein X2 is N.

[0041] E33. The polypeptide according to any one of E1 to E25, wherein X2 is Q. E34. The polypeptide according to any one of E1 to E25, wherein X2 is T. E35. The polypeptide according to any one of E1 to E34, wherein X3 is N.

[0042] The polypeptide according to any one of E1 to E34, wherein E36.X3 is T. E37. The polypeptide according to any one of E1 to E36, wherein X4 is A. The polypeptide according to any one of E1 to E36, wherein E38.X4 is E.

[0043] E39. The polypeptide according to any one of E1 to E38, wherein X5 is T. E40. The polypeptide according to any one of E1 to E38, wherein X5 is K. The polypeptide according to any one of E1 to E40, wherein E41.X6 is E.

[0044] The polypeptide according to any one of E1 to E40, wherein E42.X6 is K. The polypeptide according to any one of E1 to E42, wherein E43.X7 is E. The polypeptide according to any one of E1 to E42, wherein E44.X7 is D.

[0045] The polypeptide according to any one of E1 to E44, wherein E45.X8 is N. The polypeptide according to any one of E1 to E44, wherein E46.X8 is S. The polypeptide of any one of E1 to E46, wherein E47.X9 is Q.

[0046] The polypeptide according to any one of E1 to E46, wherein E48.X9 is E. E49. The polypeptide according to any one of E1 to E46, wherein X9 is K. The polypeptide of any one of E1 to E46, wherein E50.X9 is R.

[0047] The polypeptide of any one of E1 to E46, wherein E51.X9 is D. The polypeptide of any one of E1 to E46, wherein E52.X9 is N. E53. The polypeptide according to any one of E1 to E52, wherein X5 is T, X6 is E, X7 is E, and X8 is N.

[0048] E54. The polypeptide according to any one of E1 to E52, wherein X5 is T, X6 is K, X7 is E, and X8 is N. E55. The polypeptide according to any one of E1 to E54, wherein X2 is E and X3 is T.

[0049] E56. The polypeptide according to any one of E1 to E54, wherein X2 is I or F and X3 is N. The polypeptide of E56, wherein X2 is I.

[0050] The polypeptide of E56, wherein E58.X2 is F. E59. The polypeptide of E1 or E2, wherein the chimera has the sequence of any one of SEQ ID NOs: 22-43.

[0051] E60. The polypeptide of E59, wherein said chimera has the sequence of SEQ ID NO: 22. E61. The polypeptide of E59, wherein said chimera has the sequence of SEQ ID NO: 23. E62. The polypeptide of E59, wherein said chimera has the sequence of SEQ ID NO: 24.

[0052] E63. The polypeptide of E59, wherein said chimera has the sequence of SEQ ID NO: 25. E64. The polypeptide of E59, wherein said chimera has the sequence of SEQ ID NO: 40. E65. The polypeptide of E59, wherein said chimera has the sequence of SEQ ID NO: 41.

[0053] E66. The polypeptide of E59, wherein said chimera has the sequence of SEQ ID NO: 28. E67. The polypeptide of E59, wherein said chimera has the sequence of SEQ ID NO: 42. E68. The polypeptide of E59, wherein said chimera has the sequence of SEQ ID NO: 43.

[0054] E69. The polypeptide of E59, wherein said chimera has the sequence of SEQ ID NO: 37. E70. The polypeptide of any one of E1-E69, wherein said chimera is truncated from the N-terminus by the deletion of one amino acid.

[0055] E71. The polypeptide of any one of E1-E69, wherein the chimera is truncated from the N-terminus by the deletion of two amino acids. E72. The polypeptide of any one of E1-E69, wherein the chimera is truncated from the N-terminus by the deletion of three amino acids.

[0056] E73. The polypeptide of any one of E1-E69, wherein the chimera is truncated from the N-terminus by the deletion of four amino acids. E74. The polypeptide of any one of E1-E69, wherein said chimera is N-terminally truncated by the deletion of five amino acids.

[0057] E75. The polypeptide of any one of E1-E69, wherein said chimera is N-terminally truncated by the deletion of six amino acids. E76. The polypeptide of any one of E1-E69, wherein the chimera is N-terminally truncated by the deletion of seven amino acids.

[0058] E77. The polypeptide of any one of E1-E69, wherein the chimera is N-terminally truncated by the deletion of eight amino acids. E78. The polypeptide of any one of E1-E69, wherein said chimera is N-terminally truncated by the deletion of 9 amino acids.

[0059] E79. The polypeptide of any one of E1, E2, and E70-E78, wherein the chimera has the sequence of any one of SEQ ID NOs: 111-183. E80. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO:111.

[0060] E81. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 116. E82. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 117. E83. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 118.

[0061] E84. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 119. E85. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 120. E86. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 121.

[0062] E87. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 122. E88. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 126. E89. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 130.

[0063] E90. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 142. E91. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 143. E92. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 146.

[0064] E93. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 147. E94. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 150. E95. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 151.

[0065] E96. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 152. E97. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 153. E98. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 154.

[0066] E99. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 155. E100. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 156.

[0067] E101. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 158. E102. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 159.

[0068] E103. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 162. E104. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 163.

[0069] E105. The polypeptide of E79, wherein said chimera has the sequence of SEQ ID NO: 164. E106. The polypeptide of any one of E1 to E105, wherein the polypeptide (e.g., the chimera) further comprises a C-terminal extension of one or more amino acids (e.g., 1, 2, 3, 4, 5, 6, or more amino acids from wild-type extracellular ActRIIA or ActRIIB).

[0070] E107. The polypeptide of E106, wherein said C-terminal extension is NP. E108. The polypeptide of E106, wherein said C-terminal extension is NPVTPK (SEQ ID NO: 104).

[0071] E109. The polypeptide of any one of E1 to E108, wherein the polypeptide further comprises an Fc domain monomer fused to the C-terminus of the polypeptide (e.g., the C-terminus of the chimera) via a linker.

[0072] E110. The polypeptide of E109, wherein said Fc domain monomer has the sequence of SEQ ID NO:48. E111. The polypeptide of E109 or E110, wherein said polypeptide forms a dimer.

[0073] E112. The polypeptide of any one of E1 to E108, wherein the polypeptide further comprises a wild-type Fc domain fused to the C-terminus of the polypeptide (e.g., the C-terminus of the chimera) via a linker.

[0074] E113. The polypeptide of E112, wherein said wild-type Fc domain has the sequence of SEQ ID NO: 100 or SEQ ID NO: 264. E114. The polypeptide of E112 or E113, wherein the polypeptide has the sequence of any one of SEQ ID NOs: 107-110 and 184-263.

[0075] E115. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 107. E116. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 108.

[0076] E117. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 109. E118. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 110.

[0077] E119. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 184. E120. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 189.

[0078] E121. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 190. E122. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 191.

[0079] E123. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 192. E124. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 193.

[0080] E125. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 194. E126. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 195.

[0081] E127. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 199. E128. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 200.

[0082] E129. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 213. E130. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 214.

[0083] E131. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 215. E132. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 216.

[0084] E133. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 217. E134. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 218.

[0085] E135. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 221. E136. The polypeptide according to E114, wherein said polypeptide has the sequence of SEQ ID NO: 222.

[0086] E137. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 225. E138. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 226.

[0087] E139. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 229. E140. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 230.

[0088] E141. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 231. E142. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 232.

[0089] E143. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 233. E144. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 234.

[0090] E145. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 235. E146. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 236.

[0091] E147. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 237. E148. The polypeptide of E114, wherein said polypeptide has the sequence of SEQ ID NO: 238.

[0092] E149. The polypeptide of any one of E1 to E108, wherein the polypeptide further comprises an Fc domain comprising an amino acid substitution fused to the C-terminus of the polypeptide (e.g., the C-terminus of the chimera) via a linker.

[0093] E150. The polypeptide of E149, wherein said Fc domain does not dimerize. E151. The polypeptide of any one of E1 to E108, wherein the polypeptide further comprises an albumin binding peptide fused to the C-terminus of the polypeptide (e.g., the C-terminus of the chimera) via a linker.

[0094] E152. The polypeptide of E151, wherein said albumin binding peptide has the sequence of SEQ ID NO: 101. E153. The polypeptide of any one of E1-E108, wherein said polypeptide further comprises a fibronectin domain fused to the C-terminus of said polypeptide (e.g., to the C-terminus of said chimera) via a linker.

[0095] E154. The polypeptide of E153, wherein said fibronectin domain has SEQ ID NO: 102. E155. The polypeptide of any one of E1 to E108, wherein said polypeptide further comprises human serum albumin fused to the C-terminus of said polypeptide (e.g., the C-terminus of said chimera) via a linker.

[0096] E156. The polypeptide of E155, wherein said human serum albumin has SEQ ID NO: 103. E157. The polypeptide according to any one of E109 to E156, wherein said linker is an amino acid spacer.

[0097] E158. The polypeptide of E157, wherein said amino acid spacer is GGG, GGGA (SEQ ID NO:49), GGGG (SEQ ID NO:51), GGGAG (SEQ ID NO:81), GGGAGG (SEQ ID NO:82), or GGGAGGG (SEQ ID NO:83).

[0098] E159. The amino acid spacer is GA, GS, GG, GGA, GGS, GGGS (SEQ ID NO: 50), GGGGA (SEQ ID NO: 52), GGGGS (SEQ ID NO: 53), GGGGG (SEQ ID NO: 54), GGAG (SEQ ID NO: 55), GGSG (SEQ ID NO: 56), AGGG (SEQ ID NO: 57), SGGG (SEQ ID NO: 58), GAGA (SEQ ID NO: 59), GSGS (SEQ ID NO: 60), GAGAGA (SEQ ID NO: 61), GSGSGS (SEQ ID NO: 62), GAGAGAGA (SEQ ID NO: 63), GSGSGSGS (SEQ ID NO: 64). ), GAGAGAGAGA (SEQ ID NO: 65), GSGSGSGSGS (SEQ ID NO: 66), GAGAGAGAGA (SEQ ID NO: 67), GSGSGSGSGSGS (SEQ ID NO: 68), GGAGGA (SEQ ID NO: 69), GGSGGS (SEQ ID NO: 70), GGAGGAGGA (SEQ ID NO: 71), GGSGGSGGS (SEQ ID NO: 72), GGAGGAGGAGGA (SEQ ID NO: 73), and GGSGGSGGSGGS (SEQ ID NO: 74), GGAGGGAG (SEQ ID NO: 75), GGSGGGSG (SEQ ID NO: 76), GGAG GGAGGGAG (SEQ ID NO: 77), and GGSGGGSGGGSG (SEQ ID NO: 78), GGGGAGGGGAGGGGA (SEQ ID NO: 79), GGGGSGGGSGGGGS (SEQ ID NO: 80), AAAL (SEQ ID NO: 84), AAAK (SEQ ID NO: 85), AAAR (SEQ ID NO: 86), EGKSSGSGSESKST (SEQ ID NO: 87), GSAGSAAGSGEF (SEQ ID NO: 88), AEAAAKEAAAKA (SEQ ID NO: 89), KESGSVSSEQLAQFRSLD (SEQ ID NO: 90), GENLYFQSGG ( The polypeptide of E157, which is SEQ ID NO: 91), SACYCELS (SEQ ID NO: 92), RSIAT (SEQ ID NO: 93), RPACKIPNDLKQKVMNH (SEQ ID NO: 94), GGSAGGSGSGSSGGSSGASGTGTAGGTGSGSGTGSG (SEQ ID NO: 95), AAANSSIDLISVPVDSR (SEQ ID NO: 96), GGSGGGSEGGGSEGGGSEGGGSEGGGSEGGGSGGGS (SEQ ID NO: 97), EAAAK (SEQ ID NO: 98), or PAPAP (SEQ ID NO: 99).

[0099] E160. The polypeptide of any one of E1-E159, wherein said polypeptide (eg, an ActRII chimeric-Fc fusion protein) has a serum half-life of at least 7 days.

[0100] E161. The polypeptide of any one of E1 to E160, wherein the polypeptide has increased binding to one or more ActRII ligands (e.g., activin A, activin B, myostatin, and / or GDF-11) compared to wild-type ActRIIA and / or wild-type ActRIIB (e.g., wild-type extracellular ActRIIA and / or ActRIIB).

[0101] E162. The polypeptide of any one of E1 to E161, wherein the polypeptide has reduced binding to bone morphogenetic protein 9 (BMP9, e.g., human BMP9) compared to wild-type ActRIIB (e.g., wild-type extracellular ActRIIB).

[0102] E163. The polypeptide of any one of E1-E162, wherein the polypeptide binds to activin A, activin B, and / or myostatin and has reduced or weak binding to human BMP9 (e.g., compared to wild-type extracellular ActRIIB).

[0103] E164. The polypeptide of any one of E1 to E163, wherein said polypeptide does not substantially bind to human BMP9. E165. The polypeptide has a K D The polypeptide according to any one of E1 to E164, which binds to human activin A at

[0104] E166. The polypeptide has a K D The polypeptide according to any one of E1 to E165, which binds to human activin B at the E167. The polypeptide has a K D The polypeptide according to any one of E1 to E166, which binds to human GDF-11 at

[0105] E168. A nucleic acid molecule encoding a polypeptide according to any one of E1 to E167. E169. A vector comprising a nucleic acid molecule according to E168.

[0106] E170. A host cell expressing a polypeptide according to any one of E1 to E167, said host cell comprising a nucleic acid molecule according to E168 or a vector according to E169, said nucleic acid molecule or vector being expressed in said host cell.

[0107] E171. A method for preparing a polypeptide according to any one of E1 to E167, said method comprising: a) providing a host cell containing a nucleic acid molecule according to E168 or a vector according to E169; and b) expressing said nucleic acid molecule or vector in said host cell under conditions allowing the formation of said polypeptide.

[0108] E172. A pharmaceutical composition comprising a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, or a vector according to E169, and one or more pharmaceutically acceptable carriers or excipients.

[0109] E173. The pharmaceutical composition of E172, wherein said polypeptide, nucleic acid molecule, or vector is in a therapeutically effective amount. E174. A construct comprising two identical polypeptides (e.g., homodimers) each comprising an extracellular ActRII chimera described in any one of E1 to E108 (e.g., an ActRII chimera having a sequence of any one of SEQ ID NOS: 1-43 and 111-183) fused to the N-terminus or C-terminus of an Fc domain monomer (SEQ ID NO: 48). The two Fc domain monomers in the two polypeptides interact to form an Fc domain in the construct.

[0110] E175. A construct comprising two different polypeptides (e.g., a heterodimer), each comprising an extracellular ActRII chimera described in any one of E1 to E108 (e.g., an ActRII chimera having a sequence of any one of SEQ ID NOS: 1-43 and 111-183) fused to the N-terminus or C-terminus of an Fc domain monomer (SEQ ID NO: 48). The two Fc domain monomers in the two polypeptides interact to form an Fc domain in the construct.

[0111] E176. A method for increasing lean body mass in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0112] E177. A method for increasing muscle mass in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0113] E178. A method for increasing muscle strength in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0114] E179. The method of any one of E176-E178, wherein the subject has or is at risk of developing a neuromuscular disease, sarcopenia, cachexia, disuse atrophy, treatment-related muscle loss or atrophy, hypotonia, hypoxia-related muscle loss or atrophy, or burn-related muscle loss or atrophy.

[0115] E180. A method for treating a subject having or at risk of developing a muscle disease, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0116] E181. The method of E180, wherein said muscle disease is a neuromuscular disease, sarcopenia, cachexia, disuse atrophy, treatment-related muscle loss or atrophy, hypotonia, hypoxia-related muscle loss or atrophy, or burn-related muscle loss or atrophy.

[0117] E182. A method for affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing a disease or condition involving muscle weakness and atrophy, the method comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0118] E183. The method of E182, wherein said disease or condition is a neuromuscular disease, sarcopenia, cachexia, disuse atrophy, treatment-related muscle loss or atrophy, hypotonia, hypoxia-related muscle loss or atrophy, or burn-related muscle loss or atrophy.

[0119] E184. A method for treating a subject having or at risk of developing a neuromuscular disease, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0120] E185. The method of any one of E179, E181, and E183, wherein the subject has or is at risk of developing a neuromuscular disease, or the disease or condition is a neuromuscular disease.

[0121] E186. A method for treating a subject having or at risk of developing DMD by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0122] E187. A method for treating a subject having or at risk of developing FSHD by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0123] E188. A method for treating a subject having or at risk of developing IBM by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0124] E189. A method for treating a subject having or at risk of developing ALS by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0125] E190. The method of any one of E179, E181, and E183, wherein said subject has or is at risk of developing sarcopenia, or said disease or condition is sarcopenia.

[0126] E191. A method of treating a subject having or at risk of developing sarcopenia by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0127] E192. The method of any one of E179, E181, and E183, wherein the subject has or is at risk of developing disuse atrophy, or the disease or condition is disuse atrophy.

[0128] E193. A method for treating a subject having or at risk of developing disuse atrophy by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0129] E194. The method of any one of E179, E181, and E183, wherein the subject has or is at risk of developing treatment-related muscle loss or atrophy, or the disease or condition is treatment-related muscle loss or atrophy.

[0130] E195. A method of treating a subject having or at risk of developing treatment-associated muscle loss or atrophy by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0131] E196. The method of any one of E179, E181, E183, E194, or E195, wherein said treatment is glucocorticoid treatment, FGF-21 treatment, GLP-1 treatment, treatment with a therapeutic agent comprising FGF-21 or GLP-1, bariatric surgery (e.g., gastric bypass), cancer treatment (e.g., chemotherapy or radiation), or treatment for obesity or type 2 diabetes.

[0132] E197. The method of any one of E179, E181, and E183, wherein the subject has or is at risk of developing hypotonia, or the disease or condition is hypotonia.

[0133] E198. A method for treating a subject having or at risk of developing hypotonia by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0134] E199. The method of any one of E179, E181, and E183, wherein the subject has or is at risk of developing hypoxia-associated muscle loss or atrophy, or the disease or condition is hypoxia-associated muscle loss or atrophy.

[0135] E200. A method of treating a subject having or at risk of developing hypoxia-associated muscle loss or atrophy by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0136] E201. The method of any one of E179, E181, and E183, wherein the subject has or is at risk of developing burn-related muscle loss or atrophy, or the disease or condition is burn-related muscle loss or atrophy.

[0137] E202. A method of treating a subject having or at risk of developing burn-related muscle loss or atrophy by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0138] E203. The method of any one of E179, E181, and E183, wherein said subject has or is at risk of developing cachexia, or said disease or condition is cachexia.

[0139] E204. A method of treating a subject having or at risk of developing cachexia by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0140] E205. The method of any one of E179, E181, E183, E203, and E204, wherein said cachexia is cancer cachexia, HIV-associated cachexia, cardiac cachexia (e.g., cachexia associated with heart failure), cachexia associated with chronic kidney disease, or pulmonary cachexia (e.g., cachexia associated with COPD).

[0141] E206. The method according to any one of E176 to E205, wherein said method increases muscle mass. E207. The method according to any one of E176 to E206, wherein said method increases lean body mass.

[0142] E208. The method according to any one of E176 to E207, wherein said method increases muscle strength. E209. A method of increasing bone mineral density in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0143] E210. A method of reducing bone resorption in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0144] E211. A method of increasing bone formation in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0145] E212. A method of increasing bone strength in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide according to any one of E1-E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0146] E213. A method of reducing the risk or occurrence of bone fracture in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0147] E214. The method of any one of E209-213, wherein the subject has or is at risk of developing osteoporosis, osteopenia, osteopetrosis, fracture, bone cancer or cancer metastasis-associated bone loss, Paget's disease, renal osteodystrophy, treatment-associated bone loss, osteogenesis imperfecta, neuromuscular disease-associated bone loss, burns-associated bone loss, anorexia-associated bone loss, diet-associated bone loss, obesity treatment-associated bone loss, reduced gravity-associated bone loss, or immobility-associated bone loss.

[0148] E215. A method of treating a subject having or at risk of developing a bone disease by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0149] E216. The method of E215, wherein said bone disease is osteoporosis, osteopenia, osteopetrosis, bone fracture, bone loss associated with bone cancer or cancer metastasis, Paget's disease, renal osteodystrophy, treatment-related bone loss, osteogenesis imperfecta, bone loss associated with neuromuscular disease, burn-related bone loss, anorexia-related bone loss, diet-related bone loss, bone loss associated with treatment of obesity, bone loss associated with reduced gravity, or bone loss associated with immobility.

[0150] E217. A method of affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing a disease or condition involving bone damage, by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0151] E218. The method of E217, wherein said disease or condition is osteoporosis, osteopenia, osteopetrosis, bone fracture, bone loss associated with bone cancer or cancer metastasis, Paget's disease, renal osteodystrophy, treatment-related bone loss, osteogenesis imperfecta, bone loss associated with neuromuscular disease, bone loss due to burns, bone loss associated with anorexia, diet-related bone loss, bone loss associated with treatment of obesity, bone loss associated with reduced gravity, or bone loss associated with immobility.

[0152] E219. The method of any one of E214, E216, or E218, wherein said subject has or is at risk of developing osteoporosis, or said disease or condition is osteoporosis.

[0153] E220. A method of treating a subject having or at risk of developing osteoporosis by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0154] E221. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing osteogenesis imperfecta, or said disease or condition is osteogenesis imperfecta.

[0155] E222. A method of treating a subject having or at risk of developing osteogenesis imperfecta by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0156] E223. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing osteopenia, or said disease or condition is osteopenia.

[0157] E224. A method of treating a subject having or at risk of developing osteopenia by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0158] E225. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing a bone fracture, or said disease or condition is a bone fracture.

[0159] E226. A method for treating a subject having or at risk of developing a fracture by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0160] E227. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing bone loss associated with bone cancer or cancer metastasis, or said disease or condition is bone loss associated with bone cancer or cancer metastasis.

[0161] E228. A method of treating a subject having or at risk of developing bone loss associated with bone cancer or cancer metastasis by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0162] E229. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing Paget's disease, or said disease or condition is Paget's disease.

[0163] E230. A method of treating a subject having or at risk of developing Paget's disease by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0164] E231. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing renal osteodystrophy, or said disease or condition is renal osteodystrophy.

[0165] E232. A method of treating a subject having or at risk of developing renal osteodystrophy by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0166] E233. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing treatment-related bone loss, or said disease or condition is treatment-related bone loss.

[0167] E234. A method of treating a subject having or at risk of developing treatment-associated bone loss by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0168] E235. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing diet-related bone loss, or said disease or condition is diet-related bone loss.

[0169] E236. A method of treating a subject having or at risk of developing diet-related bone loss by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0170] E237. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing bone loss associated with low gravity, or said disease or condition is bone loss associated with low gravity.

[0171] E238. A method of treating a subject having or at risk of developing bone loss associated with hypogravity by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0172] E239. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing immobility-associated bone loss, or said disease or condition is immobility-associated bone loss.

[0173] E240. A method of treating a subject having or at risk of developing immobility-associated bone loss by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0174] E241. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing bone loss associated with a neuromuscular disease, or said disease or condition is bone loss associated with a neuromuscular disease.

[0175] E242. A method of treating a subject having or at risk of developing bone loss associated with a neuromuscular disease by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0176] E243. The neuromuscular disease is muscular dystrophy, amyotrophic lateral sclerosis (ALS), autonomic neuropathy, botulism, Charcot-Marie-Tooth disease (CMT), chronic inflammatory demyelinating polyneuropathy, congenital myasthenic syndrome, congenital myopathy, cramp-fasciculation syndrome, dermatomyositis, diabetic neuropathy, distal myopathy, dystrophinopathy, endocrine myopathy, focal muscular atrophy, glycogen storage disease type II, Guillain-Barré syndrome, hereditary spastic paraplegia, inclusion body myositis (IBM), Isaac syndrome, Kearns-Sayre syndrome, Kennedy disease, Lambert-Eaton myasthenic syndrome, metabolic myopathy, The method of any one of E179, E181, E183 to E185, E214, E216, E218, E241, and E242, wherein the disease is metabolic neuropathy, mitochondrial myopathy, motor neuron disease, multiple sclerosis, myasthenia gravis, myotonic dystrophy, necrotizing myopathy, neuromyotonia, Friedreich's ataxia neuropathy, nutritional neuropathy, peripheral neuropathy, polymyositis, primary lateral sclerosis, Schwartz-Jampel syndrome, small fiber neuropathy, spinal and bulbar muscular atrophy, spinal muscular atrophy (SMA), spinal muscular atrophy with respiratory distress syndrome type 1, stiff person syndrome, toxic neuropathy, or Troyer syndrome.

[0177] E244. The method of E243, wherein said neuromuscular disease is muscular dystrophy. E245. The method of E244, wherein said muscular dystrophy is Duchenne muscular dystrophy (DMD), facioscapulohumeral muscular dystrophy (FSHD), Becker muscular dystrophy (BMD), myotonic dystrophy (DM), congenital muscular dystrophy, limb-girdle muscular dystrophy (LGMD), distal muscular dystrophy (DD), oculopharyngeal muscular dystrophy (OPMD), or Emery-Dreifuss muscular dystrophy (EDMD).

[0178] E246. The method of E245, wherein said muscular dystrophy is DMD. E247. The method of E245, wherein said muscular dystrophy is FSHD. E248. The method of E245, wherein said muscular dystrophy is BMD.

[0179] E249. The method according to E245, wherein said muscular dystrophy is DM. E250. The method of E245, wherein said muscular dystrophy is LGMD. E251. The method of E245, wherein said muscular dystrophy is DD.

[0180] E252. The method according to E245, wherein said muscular dystrophy is OPMD. E253. The method according to E245, wherein said muscular dystrophy is EDMD. E254. The method according to E245, wherein said muscular dystrophy is a congenital muscular dystrophy.

[0181] E255. The method of E254, wherein the congenital muscular dystrophy is congenital muscular dystrophy type 1A (MDC1A), congenital muscular dystrophy type 1C (MDC1C), congenital muscular dystrophy type 1D (MDC1D), congenital muscular dystrophy type 1B (MDC1B), Fukuyama congenital muscular dystrophy (FCMD), muscle-eye-brain disease (MEB), Walker-Warburg syndrome (WWS), spondylotonic muscular dystrophy (RSMD1), Ullrich congenital muscular dystrophy (UCMD), or a muscular dystrophy associated with a mutation in integrin α7, integrin α9, docking protein 7, laminin A / C, SECIS-binding protein 2, or choline kinase β.

[0182] E256. The method of E255, wherein said congenital muscular dystrophy is MDC1A. E257. The method of E255, wherein said congenital muscular dystrophy is MDC1B. E258. The method of E255, wherein said congenital muscular dystrophy is MDC1C.

[0183] E259. The method of E255, wherein said congenital muscular dystrophy is MDC1D. E260. The method of E255, wherein said congenital muscular dystrophy is FCMD. E261. The method of E255, wherein said congenital muscular dystrophy is MEB.

[0184] E262. The method according to E255, wherein said congenital muscular dystrophy is WWS. E263. The method of E255, wherein said congenital muscular dystrophy is RSMD1. E264. The method of E255, wherein said congenital muscular dystrophy is UCMD.

[0185] E265. The method of E243, wherein said neuromuscular disease is CMT. E266. The method of E243, wherein said neuromuscular disease is ALS. E267. The method of E243, wherein said neuromuscular disease is SMA.

[0186] E268. The method of E243, wherein said neuromuscular disease is IBM. E269. The method of E243, wherein said neuromuscular disease is myasthenia gravis. E270. The method of E243, wherein said neuromuscular disease is multiple sclerosis.

[0187] E271. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing burns bone loss, or said disease or condition is burns bone loss.

[0188] E272. A method of treating a subject having or at risk of developing bone loss due to burns by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0189] E273. The method of any one of E214, E216, and E218, wherein said subject has or is at risk of developing anorexia-associated bone loss, or said disease or condition is anorexia-associated bone loss.

[0190] E274. A method of treating a subject having or at risk of developing bone loss associated with anorexia by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0191] E275. The method according to any one of E214, E216, and E218 to E220, wherein said osteoporosis is primary osteoporosis. E276. The method according to E275, wherein said primary osteoporosis is age-related osteoporosis or hormone-related osteoporosis.

[0192] E277. The method according to any one of E214, E216, and E218 to E220, wherein said osteoporosis is secondary osteoporosis. E278. The method according to E277, wherein said secondary osteoporosis is immobility-induced osteoporosis or glucocorticoid-induced osteoporosis.

[0193] E279. The method of any one of E214, E216, E218, E227, and E228, wherein said cancer is multiple myeloma. E280. The method of any one of E214, E216, E218, E233, and E234, wherein the treatment is FGF-21 treatment, GLP-1 treatment, treatment with a therapeutic agent comprising FGF-21 or GLP-1, cancer treatment (e.g., chemotherapy or radiation), bariatric surgery (e.g., gastric bypass), androgen or estrogen deprivation therapy, or treatment of obesity or type 2 diabetes.

[0194] E281. The method of any one of E214, E216, E218, E235, and E236, wherein said diet-related bone loss is rickets. E282. The method of any one of E209-E281, wherein said subject is at risk of fracture.

[0195] E283. The method of any one of E209 to E282, wherein said method increases bone formation in said subject. E284. The method of any one of E209 to E283, wherein said method reduces bone resorption in said subject.

[0196] E285. The method of any one of E209 to E284, wherein said method reduces bone loss in said subject. E286. The method of any one of E209-E285, wherein said method increases osteoblast activity or osteoblast formation.

[0197] E287. The method of any one of E209-E286, wherein said method reduces osteoclast activity or osteoclast formation. E288. The method of any one of E209-E287, wherein said method reduces the risk or incidence of fracture.

[0198] E289. The method of any one of E209-E288, wherein said method increases bone strength. E290. The method of any one of E209 to E289, wherein said bone is cortical bone.

[0199] E291. The method according to any one of E209 to E289, wherein said bone is cancellous bone. E292. A method of reducing or preventing fibrosis in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0200] E293. A method of delaying or inhibiting the progression of fibrosis in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0201] E294. A method for reducing the risk of developing fibrosis in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0202] E295. A method of treating a subject having or at risk of developing fibrosis by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0203] E296. A method for reversing fibrosis in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0204] E297. A method of affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors) in a subject having, or at risk of developing, fibrosis or a disease or condition involving fibrosis, by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0205] E298. The method of any one of E292 to E297, wherein the fibrosis or disease or condition comprising fibrosis is chemotherapy-induced fibrosis, radiation-induced fibrosis, pulmonary fibrosis, liver fibrosis, renal fibrosis (e.g., fibrosis associated with chronic kidney disease), corneal fibrosis, cardiac fibrosis, bone marrow fibrosis, myelofibrosis, mediastinal fibrosis, retroperitoneal fibrosis, arthrofibrosis, osteoarthrofibrosis, tissue fibrosis, tumor stroma, desmoplastic tumor, surgical adhesion, hypertrophic scar, or keloid.

[0206] E299. The method of any one of E292 to E297, wherein said fibrosis or disease or condition comprising fibrosis is fibrosis associated with a wound, a burn, hepatitis B or C infection, fatty liver disease, schistosome infection, kidney disease (e.g., chronic kidney disease), heart disease, macular degeneration, Crohn's disease, retinal or vitreoretinopathy, systemic or localized scleroderma, atherosclerosis, or restenosis.

[0207] E300. The method of any one of E292 to E298, wherein said fibrosis is due to chronic kidney disease. E301. The method of any one of E292 to E298, wherein said fibrosis is caused by myelofibrosis.

[0208] E302. The method of E298, wherein said tissue fibrosis is fibrosis affecting a tissue selected from the group consisting of muscle tissue, skin epidermis, skin dermis, tendon, cartilage, pancreatic tissue, uterine tissue, nervous tissue, testis, ovary, adrenal gland, artery, vein, bone marrow, colon, small intestine, large intestine, biliary tract, and intestine.

[0209] E303. The method according to any one of E292 to E302, wherein said method improves the function of a fibrotic tissue or organ. E304. The method of any one of E292-E303, wherein said method delays, inhibits, or reverses the onset or progression of fibrosis.

[0210] E305. The method of any one of E292-E304, wherein said method reduces one or more symptoms of fibrosis (eg, reduces the frequency or severity of fibrosis). E306. A method of increasing red blood cell levels (e.g., increasing hemoglobin levels, red blood cell count, or hematocrit) in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0211] E307. A method of increasing hemoglobin levels in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0212] E308. A method for increasing red blood cell count in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0213] E309. A method of increasing hematocrit in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0214] E310. A method of promoting or increasing red blood cell production in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0215] E311. A method of maturation and / or differentiation of erythroid progenitor cells (e.g., maturation and / or differentiation of early or late (e.g., terminal) stage erythroid progenitor cells, e.g., early stage erythroid progenitor cells such as colony forming unit-erythroid cells (CFU-E) and burst-forming unit-erythroid cells (BFU-E), into proerythroblasts, reticulocytes, or erythrocytes) in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0216] E312. A method of promoting or increasing proerythroblasts (e.g., proerythroblast numbers or proerythroblast numbers) in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0217] E313. A method of promoting or increasing reticulocytes (e.g., reticulocyte count or reticulocyte number) in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0218] E314. A method of promoting or increasing the mobilization of early stage progenitor cells into the erythroid lineage in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0219] E315. A method of promoting or increasing late erythroid precursor maturation (e.g., final maturation such as maturation of reticulocytes into erythrocytes or maturation of erythroblasts into reticulocytes and / or erythrocytes) in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0220] E316. A method for reducing the accumulation of erythroid progenitor cells in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0221] E317. A method of increasing the number of early stage erythroid progenitor and / or precursor cells (e.g., expanding a population of early stage progenitor and / or precursor cells) in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0222] E318. A method of promoting the progression of erythroid precursor cells and / or progenitor cells through erythropoiesis in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0223] E319. The method of any one of E306-E318, wherein said subject has or is at risk of developing anemia or blood loss. E320. A method of affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing a disease or condition involving low red blood cell levels (e.g., low hemoglobin levels, low red blood cell count, or low hematocrit), the method comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0224] E321. The method of E320, wherein said disease or condition is anemia or blood loss. E322. A method of treating a subject having or at risk of developing anemia by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0225] E323. The anemia or blood loss is due to cancer (e.g., solid tumors such as breast cancer, lung cancer, colon cancer, etc.; tumors of the lymphatic system such as chronic lymphocytic leukemia, non-Hodgkin's lymphoma, Hodgkin's lymphoma, etc.; or tumors of the hematopoietic system such as leukemia or multiple myeloma), cancer treatment (e.g., chemotherapy or radiation), treatment of myelofibrosis (e.g., treatment with a JAK inhibitor such as ruxolitinib or fedratinib), kidney disease or renal failure (e.g., chronic kidney disease or acute kidney disease or renal failure), myelodysplastic syndrome, thalassemia (e.g., α-thalassemia or β-thalassemia), nutritional deficiency (e.g., vitamin deficiency such as B-12 or folate deficiency), adverse reaction to a drug, ineffective hematopoiesis, inflammatory disease, or autoimmune disease. the method of E319, E321, or E322, wherein the disease is associated with an infectious disease, splenomegaly, porphyria, vasculitis, hemolysis, bone marrow deficiency, bone marrow transplantation, myelofibrosis, diabetes, liver disease (e.g., acute liver disease or chronic liver disease), bleeding (e.g., acute hemorrhage or chronic hemorrhage), infection, hemoglobinopathy, drug use, alcohol abuse, elderly, Churg-Strauss syndrome, Felty syndrome, Pearson syndrome, dyskeratosis congenita, graft-versus-host disease, hematopoietic stem cell transplantation, myelofibrosis, pancytopenia, pure red cell aplasia, Schönlein-Henoch purpura, Schwachman syndrome (e.g., Shwachman-Diamond syndrome), transfusion contraindication, surgery, trauma, wound, ulcer, urinary tract bleeding, gastrointestinal bleeding, frequent blood donation, or heavy menstrual bleeding.

[0226] E324. The method according to any one of E319 and E321 to E323, wherein the anemia is caused by chronic kidney disease. E325. The method of any one of E319 and E321-E323, wherein the anemia is associated with a myelodysplastic syndrome (eg, the subject has a myelodysplastic syndrome).

[0227] E326. The method of any one of E319 and E321-E323, wherein the anemia is associated with myelofibrosis (eg, the subject has myelofibrosis). E327. The method of any one of E319 and E321-E323, wherein the anemia is associated with ineffective hematopoiesis (eg, ineffective erythropoiesis), has fibrosis.

[0228] E328. The method according to any one of E319 and E321 to E323, wherein the anemia is aplastic anemia, iron deficiency anemia, vitamin deficiency anemia, anemia of chronic disease (also called anemia of inflammation), anemia associated with bone marrow disease, hemolytic anemia, sickle cell anemia, microcytic anemia, hypochromic anemia, sideroblastic anemia, congenital dyserythroid anemia, Diamond-Blackfan anemia, Fanconi anemia, or refractory anemia with excess blasts.

[0229] E329. The method according to E328, wherein said sideroblastic anemia is acquired sideroblastic anemia or congenital sideroblastic anemia. E330. The method of E329, wherein said sideroblastic anemia is congenital sideroblastic anemia.

[0230] E331. The method of E330, wherein the congenital sideroblastic anemia is associated with a mutation in ALAS2, SLC25A38, FECH, GLRX5, HSPA9, HSCB, SLC25A38, or ABCB7.

[0231] E332. The method of E330, wherein said congenital sideroblastic anemia is associated with a mutation in PUS1, YARS2, LARS2, TRNT1, MT-ATP6, NDUFB11, or SLC19A2, or an mtDNA mutation.

[0232] E333. The method of any one of E306 to E332, wherein the method increases red blood cell production, red blood cell count, hemoglobin level, hematocrit, differentiation and / or maturation of erythroid progenitors (e.g., of early and / or late stage erythroid progenitors), late erythroid progenitor maturation, mobilization of early stage progenitors into the erythroid lineage, proerythroblast count, early stage erythroid progenitor and / or precursor cell count (e.g., increasing the early stage progenitor and / or precursor cell population), progression of erythroid progenitors and / or precursor cells through erythropoiesis, and / or reticulocyte count.

[0233] E334. The method of any one of E306 to E333, wherein said method reduces the accumulation of erythroid progenitor cells. E335. The method of any one of E306 to E334, wherein the subject is identified as having anemia prior to administration of the polypeptide of any one of E1 to E167, the nucleic acid molecule of E168, the vector of E169, the construct of E174 or E175, or the pharmaceutical composition of E172 or E173.

[0234] E336. The method of any one of E306 to E334, wherein the method further comprises identifying the subject as having anemia prior to administration of the polypeptide of any one of E1 to E167, the nucleic acid molecule of E168, the vector of E169, the construct of E174 or E175, or the pharmaceutical composition of E172 or E173.

[0235] E337. The method of any one of E306 to E336, wherein the method further comprises assessing red blood cell, hemoglobin, hematocrit, and / or reticulocyte levels after administration of the polypeptide of any one of E1 to E167, the nucleic acid molecule of E168, the vector of E169, the construct of E174 or E175, or the pharmaceutical composition of E172 or E173.

[0236] E338. A method of increasing platelet levels (e.g., platelet count) in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0237] E339. A method of increasing platelet count in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0238] E340. A method of promoting or increasing platelet production in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0239] E341. A method of promoting or increasing megakaryocyte differentiation and / or maturation (e.g., to produce platelets) in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0240] E342. A method of reducing the accumulation of platelet precursor cells in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0241] E343. The method of any one of E338-E342, wherein said subject has or is at risk of developing thrombocytopenia. E344. A method of affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing a disease or condition involving low platelet levels, said method comprising administering to said subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0242] E345. The method of E344, wherein said disease or condition is thrombocytopenia. E346. A method for treating a subject having or at risk of developing thrombocytopenia by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0243] E347. A method of promoting platelet production by contacting megakaryocytes with a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, or a construct according to E174 or E175, in an amount effective to promote platelet production.

[0244] E348. The method of E347, wherein said contacting is in vitro. E349. A method of treating a subject having or at risk of developing thrombocytopenia by administering to said subject platelets produced by the method of E347 or E348.

[0245] E350. The thrombocytopenia is due to bone marrow deficiency, myelodysplastic syndrome, bone marrow transplantation, myelofibrosis, treatment of myelofibrosis (e.g., treatment with a JAK inhibitor such as ruxolitinib or fedratinib), ineffective hematopoiesis, Gaucher disease, aplastic anemia, Fanconi anemia, Diamond-Blackfan anemia, Shwachman-Diamond syndrome, excessive alcohol consumption, cirrhosis of the liver, cancer (e.g., leukemia or lymphoma), autoimmune disease, viral infection, bacterial infection, splenomegaly, vitamin deficiency, cancer treatment, thrombotic thrombocytopenic purpura, idiopathic thrombocytopenic purpura, or disseminated intravascular coagulation. The method according to any one of E343, E345, E346, and E349, wherein the patient's condition is associated with a syndrome, hemolytic uremic syndrome, paroxysmal nocturnal hemoglobinuria, drug-induced platelet reduction (drug-induced thrombocytopenia, e.g., thrombocytopenia caused by treatment with sulfa-containing antibiotics such as heparin, quinine, vancomycin, rifampin, or trimethoprim, or anticonvulsants such as phenytoin), platelet dilution by transfusion, hematopoietic stem cell transplantation, acquired amegakaryocytic thrombocytopenia, Pearson's syndrome, dyskeratosis congenita, or a contraindication to blood transfusion.

[0246] E351. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with a myelodysplastic syndrome (e.g., said subject has a myelodysplastic syndrome).

[0247] E352. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with myelofibrosis (eg, said subject has myelofibrosis).

[0248] E353. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with bone marrow deficiency. E354. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with bone marrow transplantation.

[0249] E355. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with cancer. E356. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with cancer treatment (e.g., chemotherapy or radiation).

[0250] E357. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with hematopoietic stem cell transplantation. E358. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with an autoimmune disease.

[0251] E359. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with a transfusion contraindication. E360. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with myelofibrosis treatment.

[0252] E361. The method of any one of E343, E345, E346, E349, and E350, wherein said thrombocytopenia is associated with ineffective hematopoiesis. E362. The method of any one of E343, E345, E346, and E349, wherein said thrombocytopenia is familial thrombocytopenia.

[0253] E363. The method according to E362, wherein the familial thrombocytopenia is May-Hegglin anomaly, Sebastian syndrome, Fechtner syndrome, Epstein syndrome, Wiskott-Aldrich syndrome, congenital amegakaryocytic thrombocytopenia, platelet storage pool deficiency, Hermansky-Pudlak syndrome, Bernard-Soulier syndrome, von Willebrand disease type 2B, ANKRD26-associated thrombocytopenia, thrombocytopenia-radial coloboma syndrome, familial platelet disorder associated with myeloid malignancies (FPD / AML), thrombocytopenia associated with a mutation in filamin-A, or thrombocytopenia associated with a mutation in GATA-1.

[0254] E364. The method of any one of E343, E345, E346, and E349, wherein said thrombocytopenia is immune thrombocytopenia. E365. The method according to any one of E338 to E364, wherein said method increases platelet count, platelet production, and / or megakaryocyte differentiation and / or maturation.

[0255] E366. The method according to any one of E338 to E365, wherein said method reduces the accumulation of platelet precursor cells. E367. The method of any one of E338 to E366, wherein said method improves blood clotting, reduces bleeding events (e.g., reduces the incidence of bleeding events), and / or reduces bleeding in the skin of said subject.

[0256] E368. The method of any one of E338 to E347 and E349 to E367, wherein the subject is identified as having thrombocytopenia prior to administration of platelets produced by the polypeptide of any one of E1 to E167, the nucleic acid molecule of E168, the vector of E169, the construct of E174 or E175, or the pharmaceutical composition of E172 or E173, or the method of E347 or E348.

[0257] E369. The method of any one of E338 to E347 and E349 to E367, wherein the method further comprises identifying the subject as having thrombocytopenia prior to administration of platelets produced by the polypeptide of any one of E1 to E167, the nucleic acid molecule of E168, the vector of E169, the construct of E174 or E175, or the pharmaceutical composition of E172 or E173, or the method of E347 or E348.

[0258] E370. The method of any one of E338 to E347 and E349 to E369, wherein the method further comprises assessing platelet levels after administration of platelets produced by the polypeptide of any one of E1 to E167, the nucleic acid molecule of E168, the vector of E169, the construct of E174 or E175, or the pharmaceutical composition of E172 or E173, or the method of E347 or E348.

[0259] E371. A method of increasing neutrophil levels (e.g., neutrophil count) in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0260] E372. A method of increasing neutrophil count in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0261] E373. A method of promoting or increasing neutrophil production in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0262] E374. A method of promoting or increasing the differentiation and / or maturation of progenitor cells (e.g., myeloid progenitor cells, myeloblasts, and / or myeloid cells) into neutrophils in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0263] E375. The method of any one of E371-E374, wherein said subject has or is at risk of developing neutropenia. E376. A method of affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing a disease or condition involving low neutrophil levels, said method comprising administering to said subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0264] E377. The method of E376, wherein said disease or condition is neutropenia. E378. A method of treating a subject having or at risk of developing neutropenia by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0265] E379. The neutropenia is caused by bone marrow deficiency, myelodysplastic syndrome, bone marrow transplantation, myelofibrosis, ineffective hematopoiesis, aplastic anemia, Fanconi anemia, Diamond-Blackfan anemia, Shwachman-Diamond syndrome, paroxysmal nocturnal hemoglobinuria, Pearson syndrome, dyskeratosis congenita, cancer (e.g., leukemia), vitamin deficiency, splenomegaly, autoimmune disease, viral infection, bacterial infection, cancer treatment, drugs (e.g., drugs used to treat hyperthyroidism, such as methimazole and propylthiouracil); vancomycin, The method according to E375, E377, or E378, wherein the treatment is associated with neutrophil reduction, inflammation, hematopoietic stem cell transplantation, or transfusion contraindications due to antibiotics such as penicillin G, trimethoprim, and oxacillin; antiviral drugs such as ganciclovir and valganciclovir; anti-inflammatory drugs for ulcerative colitis or rheumatoid arthritis such as sulfasalazine; drugs used to treat arrhythmias such as quinidine and procainamide; anticonvulsants such as phenytoin and valproate; antipsychotics such as clozapine; or levamisole.

[0266] E380. The method of any one of E375, and E377-E379, wherein said neutropenia is associated with myelodysplastic syndrome (eg, said subject has myelodysplastic syndrome).

[0267] E381. The method of any one of E375, and E377-E379, wherein said neutropenia is associated with myelofibrosis (eg, said subject has myelofibrosis). E382. The method of any one of E375 and E377-E379, wherein said neutropenia is associated with bone marrow deficiency.

[0268] E383. The method of any one of E375 and E377-E379, wherein said neutropenia is associated with bone marrow transplantation. E384. The method according to any one of E375 and E377 to E379, wherein said neutropenia is associated with cancer.

[0269] E385. The method according to any one of E375 and E377-E379, wherein said neutropenia is associated with cancer treatment (eg, chemotherapy or radiation). E386. The method according to any one of E375 and E377-E379, wherein said neutropenia is associated with hematopoietic stem cell transplantation.

[0270] E387. The method according to any one of E375 and E377 to E379, wherein said neutropenia is associated with inflammation. E388. The method according to any one of E375 and E377 to E379, wherein said neutropenia is associated with a transfusion contraindication.

[0271] E389. The method according to any one of E375 and E377-E379, wherein said neutropenia is associated with ineffective hematopoiesis. E390. The method of E375, E377, or E378, wherein said neutropenia is chronic idiopathic neutropenia.

[0272] E391. The method of E375, E377, or E378, wherein said neutropenia is familial neutropenia. E392. The method of E391, wherein said familial neutropenia is cyclic neutropenia, chronic benign neutropenia, or severe congenital neutropenia (e.g., neutropenia associated with a mutation in the ELANE (associated with SCN1), HAX1 (associated with SCN3), G6PC3 (associated with SCN4), GFI1 (associated with SCN2), CSF3R, WAS (associated with X-linked neutropenia / X-linked SCN), CXCR4, VPS45A (associated with SCN5), or JAGN1 gene).

[0273] E393. The method according to any one of E371 to E392, wherein said method increases neutrophil numbers, neutrophil production, and / or differentiation and / or maturation of progenitor cells into neutrophils. E394. The method of any one of E371 to E393, wherein said method reduces the subject's susceptibility to infection.

[0274] E395. The method of any one of E371 to E394, wherein the subject is identified as having neutropenia prior to administration of the polypeptide of any one of E1 to E167, the nucleic acid molecule of E168, the vector of E169, the construct of E174 or E175, or the pharmaceutical composition of E172 or E173.

[0275] E396. The method of any one of E371 to E394, wherein the method further comprises identifying the subject as having neutropenia prior to administration of the polypeptide of any one of E1 to E167, the nucleic acid molecule of E168, the vector of E169, the construct of E174 or E175, or the pharmaceutical composition of E172 or E173.

[0276] E397. The method of any one of E371 to E396, wherein the method further comprises assessing neutrophil levels after administration of the polypeptide of any one of E1 to E167, the nucleic acid molecule of E168, the vector of E169, the construct of E174 or E175, or the pharmaceutical composition of E172 or E173.

[0277] E398. A method for treating a subject having or at risk of developing a myelodysplastic syndrome by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0278] E399. Increase red blood cell count; increase hemoglobin levels; increase hematocrit; increase erythropoiesis; increase or induce maturation and / or differentiation of erythroid progenitors; increase or induce late-stage erythroid precursor maturation; promote or increase mobilization of early-stage progenitors into the erythroid lineage; increase proerythroblasts; increase early-stage erythroid precursor numbers (e.g., increase early-stage precursor populations); promote progression of erythroid precursors through erythropoiesis; increase reticulocytes; increase platelet count; increase platelet production; increase or induce differentiation and / or maturation of megakaryocytes , a method for reducing platelet progenitor cell accumulation, increasing neutrophil count, increasing neutrophil production, increasing or inducing differentiation and / or maturation of progenitor cells into neutrophils, improving blood clotting, reducing bleeding events, reducing skin bleeding, and / or reducing susceptibility to infection in a subject having or at risk of developing a myelodysplastic syndrome, by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0279] E400. The myelodysplastic syndrome is myelodysplastic syndrome with unilineage dysplasia (MDS-SLD), myelodysplastic syndrome with multilineage dysplasia (MDS-MLD), myelodysplastic syndrome with ring sideroblasts (including MDS-RS, unilineage dysplasia (MDS-RS-SLD), and multilineage dysplasia (MDS-RS-MLD)), myelodysplastic syndrome with isolated del chromosomal abnormality (myelodysplastic syndrome with isolated del(5q)), myelodysplastic syndrome with excess blasts The method of any one of E323, E325, E350, E351, E379, E380, E398, and E399, wherein the patient is suffering from myelodysplastic syndrome type 1 with excess blasts (MDS-EB-1), or myelodysplastic syndrome type 2 with excess blasts (MDS-EB-2), myelodysplastic syndrome, unclassifiable (MDS-U), or myelodysplastic syndrome / myeloproliferative neoplasm with ring sideroblasts and thrombocytosis (MDS / MPN-RS-T).

[0280] E401. The method according to any one of E323, E325, E350, E351, E379, E380, and E398 to E400, wherein said myelodysplastic syndrome is MDS-SLD.

[0281] E402. The method according to any one of E323, E325, E350, E351, E379, E380, and E398 to E400, wherein said myelodysplastic syndrome is MDS-MLD.

[0282] E403. The method according to any one of E323, E325, E350, E351, E379, E380, and E398 to E400, wherein said myelodysplastic syndrome is MDS-RS-SLD.

[0283] E404. The method according to any one of E323, E325, E350, E351, E379, E380, and E398 to E400, wherein said myelodysplastic syndrome is MDS-RS-MLD.

[0284] E405. The method according to any one of E323, E325, E350, E351, E379, E380, and E398 to E400, wherein said myelodysplastic syndrome is myelodysplastic syndrome with isolated del(5q).

[0285] E406. The method according to any one of E323, E325, E350, E351, E379, E380, and E398 to E400, wherein the myelodysplastic syndrome is MDS-EB-1.

[0286] E407. The method according to any one of E323, E325, E350, E351, E379, E380, and E398 to E400, wherein the myelodysplastic syndrome is MDS-EB-2.

[0287] E408. The method according to any one of E323, E325, E350, E351, E379, E380, and E398 to E400, wherein said myelodysplastic syndrome is MDS-U.

[0288] E409. The method according to any one of E323, E325, E350, E351, E379, E380, and E398 to E400, wherein the myelodysplastic syndrome is MDS / MPN-RS-T.

[0289] E410. The method of any one of E323, E325, E350, E351, E379, E380, and E398-E409, wherein the myelodysplastic syndrome is ring sideroblast-positive myelodysplastic syndrome (RS-positive MDS, e.g., the subject has ring sideroblasts).

[0290] E411. The method of E410, wherein said RS-positive myelodysplastic syndrome is associated with a mutation in a splicing factor. E412. The method according to E411, wherein the mutation in the splicing factor is a mutation in splicing factor 3b subunit 1 (SF3B1).

[0291] E413. The method of any one of E323, E325, E350, E351, E379, E380, E398-E402, and E405-E409, wherein the myelodysplastic syndrome is non-ring sideroblastic myelodysplastic syndrome (non-RS, e.g., the subject lacks ring sideroblasts).

[0292] E414. The method of any one of E323, E325, E350, E351, E379, E380, and E398-E413, wherein the myelodysplastic syndrome is a very low risk, low risk, or intermediate risk myelodysplastic syndrome (e.g., as determined by the Revised International Prognostic Scoring System).

[0293] E415. The method of E414, wherein said myelodysplastic syndrome is a very low risk myelodysplastic syndrome (eg, as determined by the Revised International Prognostic Scoring System).

[0294] E416. The method of E414, wherein said myelodysplastic syndrome is a low-risk myelodysplastic syndrome (eg, as determined by the Revised International Prognostic Scoring System).

[0295] E417. The method of E414, wherein said myelodysplastic syndrome is an intermediate-risk myelodysplastic syndrome (eg, as determined by the Revised International Prognostic Scoring System).

[0296] E418. The method of any one of E323, E325, E350, E351, E379, E380, and E398-E417, wherein said myelodysplastic syndrome is associated with a defect in terminal maturation.

[0297] E419. The method of any one of E323, E325, E350, E351, E379, E380, and E398-E417, wherein said myelodysplastic syndrome is associated with defects in early stages of hematopoiesis (e.g., progenitor cell commitment or differentiation).

[0298] E420. The method of any one of E323, E325, E350, E351, E379, E380, and E398-E419, wherein said myelodysplastic syndrome is associated with elevated endogenous erythropoietin levels.

[0299] E421. The method of any one of E323, E325, E350, E351, E379, E380, and E398-E420, wherein the myelodysplastic syndrome is associated with a hypocellular bone marrow (e.g., the subject has a hypocellular bone marrow).

[0300] E422. A method of treating a subject having or at risk of developing myelofibrosis by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0301] E423. May increase red blood cell count, increase hemoglobin levels, increase hematocrit, increase erythropoiesis, increase or induce maturation and / or differentiation of erythroid progenitors, increase or induce late-stage erythroid precursor maturation, promote or increase mobilization of early-stage progenitors into the erythroid lineage, increase proerythroblasts, increase early-stage erythroid precursor numbers (e.g., increase early-stage precursor populations), promote progression of erythroid precursors through erythropoiesis, increase reticulocytes, increase platelet count, increase platelet count, increase platelet production, or increase differentiation and / or maturation of megakaryocytes. 10. A method of increasing or inducing platelet precursor cell accumulation, increasing neutrophil counts, increasing neutrophil production, increasing or inducing differentiation and / or maturation of progenitor cells into neutrophils, improving blood clotting, reducing bleeding events, reducing skin bleeding, and / or reducing susceptibility to infection in a subject having or at risk of developing myelofibrosis, by administering to said subject a therapeutically effective amount of a polypeptide set forth in any one of E1 to E167, a nucleic acid molecule set forth in E168, a vector set forth in E169, a construct set forth in E174 or E175, or a pharmaceutical composition set forth in E172 or E173.

[0302] E424. The method according to any one of E306 to E423, wherein said subject does not respond well to treatment with erythropoietin (EPO), is susceptible to adverse effects of EPO, or does not respond well to treatment with an erythrocyte maturation agent.

[0303] E425. The method of any one of E306 to E424, wherein said subject has previously been treated with an erythropoiesis-stimulating agent (ESA). E426. The method of any one of E306 to E424, wherein said subject has not been previously treated with an erythropoiesis-stimulating agent (ESA).

[0304] E427. The method of any one of E306 to E426, wherein said subject has a low transfusion burden. E428. The method described in E427, wherein the subject has received 1 to 3 units of RBCs (1 to 3 RBC transfusions) within 8 weeks prior to initiation of treatment with the polypeptide described in any one of E1 to E167, the nucleic acid molecule described in E168, the vector described in E169, the construct described in E174 or E175, or the pharmaceutical composition described in E172 or E173.

[0305] E429. The method of E427, wherein the subject has received 0 units of RBCs (0 RBC transfusions) within 8 weeks prior to initiation of treatment with the polypeptide of any one of E1 to E167, the nucleic acid molecule of E168, the vector of E169, the construct of E174 or E175, or the pharmaceutical composition of E172 or E173.

[0306] E430. The method of any one of E306 to E426, wherein said subject has a high transfusion burden. E431. The method of any one of E306 to E430, wherein said method reduces the subject's need for blood transfusions (eg, reduces transfusion burden).

[0307] E432. A method for preventing (e.g., preventing the onset of) pulmonary hypertension (PH) in a subject in need thereof by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0308] A method for reducing the risk of developing E433.PH in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0309] A method for delaying or inhibiting the progression of E434.PH in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0310] A method for treating a subject having or at risk of developing E435.PH, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0311] A method for affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing E436.PH, by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0312] A method for reducing vascular remodeling in a subject having or at risk of developing E437.PH, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0313] A method for reducing right ventricular hypertrophy in a subject having or at risk of developing E438.PH, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0314] A method for reducing pulmonary vascular resistance in a subject having or at risk of developing E439.PH, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1 to E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0315] E440. The method according to any one of E432 to E439, wherein said PH is pulmonary arterial hypertension (PAH). E441. The method according to E440, wherein said PAH is idiopathic PAH.

[0316] E442. The method according to E440, wherein said PAH is hereditary PAH. E443. The method of E440, wherein said PAH is associated with HIV infection, schistosomiasis, liver cirrhosis, congenital heart abnormalities, portal hypertension, pulmonary veno-occlusive disease, pulmonary capillary hemangiomatosis, a connective tissue disorder, an autoimmune disorder (e.g., scleroderma or lupus), or drug use or abuse (e.g., cocaine or methamphetamine use).

[0317] E444. The method according to any one of E432 to E439, wherein said PH is venous PH. E445. The method of E444, wherein said venous PH is associated with left ventricular systolic dysfunction, left ventricular diastolic dysfunction, valvular heart disease, congenital cardiomyopathy, or congenital or acquired pulmonary vein stenosis.

[0318] E446. The method according to any one of E432 to E439, wherein said PH is hypoxic PH. E447. The method of E446, wherein said hypoxic PH is associated with chronic obstructive pulmonary disease (e.g., emphysema), interstitial lung disease, sleep-disordered breathing (e.g., sleep apnea), pulmonary disease (e.g., pulmonary fibrosis), impaired alveolar hypoventilation, chronic exposure to high altitude, or developmental abnormalities.

[0319] E448. The method according to any one of E432 to E439, wherein said PH is thromboembolic PH. E449. The method of E448, wherein said thromboembolic PH is associated with chronic thromboembolic pulmonary hypertension, pulmonary embolism, angiosarcoma, arteritis, congenital pulmonary stenosis, or parasitic infection.

[0320] E450. The method according to any one of E432 to E439, wherein said PH is other PH. E451. The method of E450, wherein said other PH is associated with a blood disease (e.g., chronic hemolytic anemia, sickle cell disease), a systemic disease (e.g., sarcoidosis, pulmonary Langerhans cell histiocytosis, lymphangioleiomyomatosis, neurofibromatosis, or vasculitis), a metabolic disorder (e.g., glycogen storage disease, Gaucher disease, or thyroid disease), pulmonary neoplastic thrombotic microangiopathy, fibrosing mediastinitis, chronic renal failure, or segmental pulmonary hypertension.

[0321] E452. The method of any one of E432 to E451, wherein the method reduces the frequency or severity of one or more symptoms of PH (e.g., reduces the severity or frequency of one or more of shortness of breath (dyspnea), fatigue, swelling of the legs, feet, abdomen (ascites), or neck (e.g., edema), or chest pain or pressure, fast pulse or heart palpitations, bluish color of the lips or skin (cyanosis), dizziness, or fainting).

[0322] E453. The method of any one of E432-E452, wherein said method reduces pulmonary vascular remodeling. E454. The method according to any one of E432 to E453, wherein said method reduces vascular remodeling in the heart.

[0323] E455. The method according to any one of E432 to E454, wherein said method reduces right ventricular hypertrophy. E456. The method of any one of E432 to E455, wherein said method reduces pulmonary vascular resistance (eg, reduces pulmonary vascular resistance compared to a pre-treatment measurement).

[0324] E457. The method of any one of E432 to E456, wherein said method improves performance in a 6-minute walk test (eg, improves performance compared to measurements taken before treatment).

[0325] E458. The method according to any one of E432 to E457, wherein said method reduces bone loss. E459. The method according to any one of E432 to E458, wherein said method reduces pulmonary artery muscularization or pulmonary artery wall thickening.

[0326] E460. The method according to any one of E432 to E459, wherein said method reduces right ventricular compensation. E461. A method for reducing body fat in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0327] E462. A method for reducing body weight in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0328] E463. A method of reducing blood glucose in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0329] E464. A method of increasing insulin sensitivity in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0330] E465. The method of any one of E461-E464, wherein said subject has or is at risk of developing a metabolic disease. E466. A method of affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of myostatin, activin A, activin B, and / or BMP9 to their receptors) in a subject having or at risk of developing a metabolic disease by administering to the subject a therapeutically effective amount of a polypeptide described in any one of E1-E167, a nucleic acid molecule described in E168, a vector described in E169, a construct described in E174 or E175, or a pharmaceutical composition described in E172 or E173.

[0331] E467. A method for treating and / or preventing a metabolic disease in a subject in need thereof by administering to said subject a therapeutically effective amount of a polypeptide according to any one of E1 to E167, a nucleic acid molecule according to E168, a vector according to E169, a construct according to E174 or E175, or a pharmaceutical composition according to E172 or E173.

[0332] E468. The method according to any one of E465 to E467, wherein the metabolic disease is an age-related metabolic disease. E469. The method according to any one of E465 to E467, wherein said metabolic disease is a therapeutically relevant metabolic disease.

[0333] E470. The method according to E469, wherein said treatment is treatment with a glucocorticoid (e.g. a corticosteroid such as prednisone), a selective serotonin reuptake inhibitor (SSRI, e.g. paroxetine, mirtazapine, fluoxetine, escitalopram, or sertraline), a serotonin-norepinephrine reuptake inhibitor (SNRI), a tricyclic antidepressant (e.g. amitriptyline), a mood stabilizer (e.g. valproic acid or lithium), an antipsychotic (e.g. olanzapine, chlorpromazine, or clozapine), or a diabetes drug (e.g. insulin, chlorpropamide).

[0334] E471. The method according to any one of E465 to E470, wherein said metabolic disease is selected from the group consisting of obesity, type 1 diabetes, and type 2 diabetes. E472. The method according to E471, wherein said metabolic disease is obesity.

[0335] E473. The method according to E471, wherein said metabolic disease is type 1 diabetes. E474. The method according to E471, wherein said metabolic disease is type 2 diabetes. E475. The method according to any one of E461 to E474, wherein said method reduces the body weight and / or percentage of weight gain of said subject.

[0336] E476. The method according to any one of E461 to E475, wherein said method reduces the amount of body fat and / or percentage of body fat gain in said subject. E477. The method according to any one of E461 to E476, wherein said method does not affect the subject's appetite for food intake.

[0337] E478. The method of any one of E461 to E477, wherein said method reduces steatosis in said subject. E479. The method according to any one of E461 to E478, wherein said method reduces the weight of the epididymal and perirenal fat pads of said subject.

[0338] E480. The method according to any one of E461 to E479, wherein said method reduces the amount of subcutaneous fat, visceral fat, and / or liver fat in said subject. E481. The method according to any one of E461 to E480, wherein said method reduces fasting insulin levels in said subject.

[0339] E482. The method according to any one of E461 to E481, wherein said method reduces blood glucose levels in said subject. E483. The method according to any one of E461 to E482, wherein said method increases insulin sensitivity in said subject.

[0340] E484. The method according to any one of E461 to E483, wherein said method increases the rate of glucose clearance in said subject. E485. The method according to any one of E461 to E484, wherein said method improves the serum lipid profile of said subject.

[0341] E486. The method according to any one of E461 to E485, wherein said method delays, reduces or eliminates the need for insulin therapy. E487. The method according to any one of E461 to E486, wherein said method does not result in a loss of lean body mass.

[0342] E488. The method of any one of E176 to E487, wherein said method reduces or inhibits binding of activin A, activin B, and / or myostatin to their receptors (e.g., their endogenous receptors).

[0343] E489. The method of any one of E176 to E208, E243 to E270, and E488, wherein the polypeptide, nucleic acid, vector, construct, or pharmaceutical composition is administered in an amount sufficient to increase muscle mass and / or strength, increase lean body mass, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or reduce or inhibit binding of activin A, activin B, and / or myostatin to their receptors (e.g., their endogenous receptors).

[0344] E490. The method of any one of E209-E291, and E488, wherein the polypeptide, nucleic acid, vector, construct, or pharmaceutical composition is administered in an amount sufficient to increase bone density, decrease bone resorption, decrease bone loss, decrease the rate of bone resorption, increase bone formation, increase the rate of bone formation, decrease osteoclast activity, increase osteoblast activity, increase bone strength, decrease the risk or incidence of fracture, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or decrease or inhibit binding of activin A, activin B, and / or myostatin to their receptors (e.g., their endogenous receptors).

[0345] E491. The method of any one of E292 to E305, and E488, wherein the polypeptide, nucleic acid, vector, construct, or pharmaceutical composition is administered in an amount sufficient to reduce fibrosis, prevent the occurrence of fibrosis, reduce the risk of developing fibrosis, delay the onset of fibrosis, slow or inhibit the progression of fibrosis, treat fibrosis, alleviate one or more symptoms of fibrosis, improve fibrotic tissue or organ function, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or reduce or inhibit binding of activin A, activin B, and / or myostatin to their receptors (e.g., their endogenous receptors).

[0346] E492. The polypeptide, nucleic acid, vector, construct, or pharmaceutical composition increases red blood cell levels, increases hemoglobin levels, increases red blood cell production, increases red blood cell count, increases hematocrit, reduces transfusion needs, increases maturation and / or differentiation of erythroid progenitors (e.g., early and / or late stage erythroid progenitors), increases maturation of late stage erythroid progenitors, mobilizes early stage progenitors to the erythroid lineage, increases reticulocytes, increases proerythroblast count, increases reticulocyte count, increases erythroid progenitors. The method of any one of E306 to E337, E398 to E431, and E488, wherein the compound is administered in an amount sufficient to reduce cell accumulation, increase the number of early stage erythroid progenitor cells and / or precursor cells, promote the progression of erythroid progenitor cells and / or precursor cells through erythropoiesis, treat anemia, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or reduce or inhibit binding of activin A, activin B and / or myostatin to their receptors (e.g., their endogenous receptors).

[0347] E493. The method of any one of E338 to E370, E98 to E431, and E488, wherein the polypeptide, nucleic acid, vector, construct, or pharmaceutical composition is administered in an amount sufficient to increase platelet levels, increase platelet production, increase platelet count, increase or induce megakaryocyte differentiation and / or maturation, decrease platelet progenitor accumulation, improve blood clotting, decrease bleeding events, decrease cutaneous bleeding, treat thrombocytopenia, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or decrease or inhibit binding of activin A, activin B, and / or myostatin to their receptors (e.g., their endogenous receptors).

[0348] E494. The method of any one of E371 to E431, and E488, wherein the polypeptide, nucleic acid, vector, construct, or pharmaceutical composition is administered in an amount sufficient to increase neutrophil levels, increase neutrophil production, increase neutrophil numbers, increase or induce differentiation and / or maturation of progenitor cells into neutrophils, treat neutropenia, reduce susceptibility to infection, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or reduce or inhibit binding of activin A, activin B, and / or myostatin to their receptors (e.g., their endogenous receptors).

[0349] E495. The method of any one of E432 to E460, and E488, wherein the polypeptide, nucleic acid, vector, construct, or pharmaceutical composition is administered in an amount sufficient to prevent PH, reduce the risk of developing PH, reduce the severity or frequency of one or more symptoms of PH, delay the onset of PH, slow or inhibit the progression of PH, treat PH, reduce pulmonary vascular remodeling, reduce vascular remodeling in the heart, reduce right ventricular hypertrophy, reduce pulmonary vascular resistance, improve performance in a 6-minute walk test, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or reduce or inhibit binding of activin A, activin B, and / or myostatin to their receptors (e.g., their endogenous receptors).

[0350] E496. The polypeptide, nucleic acid, vector, construct, or pharmaceutical composition reduces body fat, reduces subcutaneous fat mass, reduces visceral fat and / or liver fat mass, reduces adiposity, reduces epididymal and perirenal fat pad weight, reduces body fat percentage, reduces body weight, reduces percentage of weight gain, reduces fasting insulin levels, reduces blood glucose levels, increases insulin sensitivity, or enhances myostatin, activin A, activin B, and / or BMP9 signaling in the subject. The method of any one of E461 to E488, wherein the compound is administered in an amount sufficient to affect ATP signaling, reduce adipocyte proliferation, reduce or inhibit binding of activin A, activin B and / or myostatin to their receptors (e.g., their endogenous receptors), reduce LDL, reduce triglycerides, improve serum lipid profile, modulate insulin biosynthesis and / or secretion from beta cells, delay, postpone or reduce the need for insulin, or increase glucose clearance.

[0351] E497. The method according to any one of E176 to E496, wherein said method does not cause vascular complications in said subject. E498. The method of E497, wherein said method does not increase vascular permeability or leakage.

[0352] E499. The method of any one of E176 to E498, wherein said subject is a human. definition To facilitate understanding of the present invention, a number of terms and phrases are defined below. Terms defined herein have meanings commonly understood by one of ordinary skill in the art to which the present invention pertains. Terms such as "a," "an," and "the" are not intended to refer solely to a singular entity, but include general classes of which specific examples can be used for illustration. While terms herein are used to describe particular embodiments of the present invention, their use does not limit the invention except as outlined in the claims.

[0353] As used herein, the term "about" refers to a value that is within 10% above or below the stated value. As used herein, any value provided in a range of values ​​includes both the upper and lower limits, and any value subsumed within those limits.

[0354] As used herein, the terms "extracellular activin receptor type II (ActRII) chimera," "extracellular ActRII chimera," and "ActRII chimera" refer to a peptide comprising the soluble extracellular portion of the single transmembrane receptor ActRIIB and the soluble extracellular portion of the single transmembrane receptor ActRIIA. The ActRII chimera described herein results from joining the N-terminal portion of extracellular ActRIIB to the C-terminal portion of extracellular ActRIIA so that the sequences are contiguous (e.g., the ActRIIA sequence continues where the ActRIIB sequence left off, starting after the amino acid at the corresponding position in ActRIIA). The extracellular ActRII chimera may also include one or more amino acid substitutions in the portion of the chimera corresponding to the sequence of ActRIIB compared to wild-type extracellular ActRIIB (e.g., the bolded portion of the sequence in SEQ ID NO: 46 below), and one or more amino acid substitutions in the portion of the chimera corresponding to the sequence of ActRIIA compared to wild-type extracellular ActRIIA (e.g., the bolded portion of the sequence in SEQ ID NO: 47 below). The extracellular ActRII chimera may also have an N-terminal truncation of 1 to 9 amino acids relative to the extracellular portion of ActRIIB or ActRIIA. The sequences of wild-type human ActRIIB (SEQ ID NO: 46) and wild-type human ActRIIA (SEQ ID NO: 47) are shown below, with the signal peptide in italics and the extracellular portion in bold.

[0355] [ka]

[0356] The extracellular ActRII chimera may have the sequence of any one of SEQ ID NOs: 1 to 43. In certain embodiments, the extracellular ActRII chimera has the sequence of any one of SEQ ID NOs: 22 to 43 (Table 2).

[0357] As used herein, the term "N-terminal truncation" refers to the deletion of 1 to 9 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, or 9 amino acids) from the N-terminus of an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOs: 1-43 (e.g., SEQ ID NOs: 22-43)). N-terminal truncation can remove up to two amino acids before the first cysteine ​​(e.g., the two amino acids (RE or QE) before the first cysteine ​​are retained in the N-terminally truncated ActRII chimera).

[0358] As used herein, the term "linker" refers to a link between two elements, such as peptide or protein domains. The polypeptides described herein may include an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)) fused to a moiety. The moiety can increase the stability or improve the pharmacokinetic properties of the polypeptide. The moiety (e.g., an Fc domain monomer, a wild-type Fc domain, an Fc domain containing amino acid substitutions (e.g., one or more substitutions that reduce dimerization), an albumin-binding peptide, a fibronectin domain, or human serum albumin) can be fused to the polypeptide via a linker. The linker can be a covalent bond or a spacer. The term "bond" refers to a chemical bond, such as an amide bond or disulfide bond, or any type of bond generated by a chemical reaction, such as chemical conjugation. The term "spacer" refers to a moiety (e.g., a polyethylene glycol (PEG) polymer) or amino acid sequence (e.g., a sequence of 1 to 200 amino acids) present between two elements, such as peptide or protein domains, to provide space and / or flexibility between the two elements. An amino acid spacer is part of the primary sequence of a polypeptide (e.g., fused to a peptide spaced apart through the polypeptide backbone). For example, the formation of a disulfide bond between two hinge regions forming an Fc domain is not considered a linker.

[0359] As used herein, the term "Fc domain" refers to a dimer of two Fc domain monomers. An Fc domain comprises at least C H 2 domain and C H The Fc domain monomer has at least 80% sequence identity (e.g., at least 85%, 90%, 95%, 97%, or 100% sequence identity) to a human Fc domain comprising three domains. The Fc domain monomer comprises a second and a third antibody constant domain (C H 2 and C HIn some embodiments, the Fc domain monomer also includes a hinge domain. The Fc domain does not include any portion of an immunoglobulin that can act as an antigen recognition region, such as a variable domain or a complementarity determining region (CDR). In a wild-type Fc domain, two Fc domain monomers are joined by two C H The dimerization occurs through interactions between the three antibody constant domains and one or more disulfide bonds formed between the hinge domains of the two dimerized Fc domain monomers. In some embodiments, the Fc domains may be mutated to lack effector function, typical of a "dead Fc domain." In certain embodiments, each of the Fc domain monomers in the Fc domain may be mutated to lack Cγ receptors to reduce interaction or binding between the Fc domain and the Fcγ receptor. H The Fc domain comprises an amino acid substitution in the Fc domain. In some embodiments, the Fc domain comprises one or more amino acid substitutions that reduce or inhibit Fc domain dimerization. The Fc domain can be any immunoglobulin antibody isotype, including IgG, IgE, IgM, IgA, or IgD. Furthermore, the Fc domain can be an IgG subtype (e.g., IgG1, IgG2a, IgG2b, IgG3, or IgG4). The Fc domain can also be a non-naturally occurring Fc domain, such as a recombinant Fc domain.

[0360] As used herein, the term "albumin-binding peptide" refers to an amino acid sequence of 12 to 16 amino acids that has affinity for and functions to bind to serum albumin. Albumin-binding peptides can be of different origins, e.g., human, mouse, or rat. In some embodiments, the albumin-binding peptide has the sequence DICLPRWGCLW (SEQ ID NO: 101).

[0361] As used herein, the term "endogenous" refers to a molecule (e.g., a polypeptide, nucleic acid, or cofactor) that is naturally found in a particular organism (e.g., a human) or in a particular location within an organism (e.g., an organ, tissue, or cell such as a human cell, e.g., a human red blood cell, platelet, neutrophil, or muscle cell).

[0362] As used herein, the term "fibronectin domain" refers to a high molecular weight glycoprotein or fragment thereof of the extracellular matrix that binds to transmembrane receptor proteins, such as integrins, and extracellular matrix components, such as collagen and fibrin. In some embodiments, the fibronectin domain is a fibronectin type III domain having amino acids 610-702 of the sequence of UniProt ID No. P02751 (SEQ ID NO: 102). In other embodiments, the fibronectin domain is an Adnectin protein.

[0363] As used herein, the term "human serum albumin" refers to the albumin protein present in human plasma. Human serum albumin is the most abundant protein in blood. It accounts for approximately half of serum proteins. In some embodiments, human serum albumin has the sequence of UniProt ID number P02768 (SEQ ID NO: 103).

[0364] As used herein, the term "fused" is used to describe the combination or joining of two or more elements, components, or protein domains, e.g., peptides or polypeptides, by means including chemical conjugation, recombinant means, and chemical bonds, e.g., amide bonds. For example, two single peptides in tandem can be fused to form one continuous protein structure, e.g., a polypeptide, by chemical conjugation, chemical bonds, peptide linkers, or any other covalent bonding means. In some embodiments of the polypeptides described herein, an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOs: 1 to 43 (e.g., SEQ ID NOs: 22 to 43)) may be fused in tandem to the N-terminus or C-terminus of a moiety (e.g., an Fc domain monomer (e.g., the sequence of SEQ ID NO: 48), a wild-type Fc domain (e.g., the sequence of SEQ ID NO: 100, or the sequence of SEQ ID NO: 264), an Fc domain containing amino acid substitutions (e.g., one or more substitutions that reduce dimerization), an albumin-binding peptide (e.g., the sequence of SEQ ID NO: 101), a fibronectin domain (e.g., the sequence of SEQ ID NO: 102), or human serum albumin (e.g., the sequence of SEQ ID NO: 103)) via a linker. For example, an extracellular ActRII chimera is fused to a moiety (e.g., an Fc domain monomer, a wild-type Fc domain, an Fc domain containing amino acid substitutions (e.g., one or more substitutions that reduce dimerization), an albumin-binding peptide, a fibronectin domain, or human serum albumin) by a peptide linker, the N-terminus of the peptide linker is fused to the C-terminus of the extracellular ActRII chimera by a chemical bond, e.g., a peptide bond, and the C-terminus of the peptide linker is fused to the N-terminus of the moiety (e.g., an Fc domain monomer, a wild-type Fc domain, an Fc domain containing amino acid substitutions (e.g., one or more substitutions that reduce dimerization), an albumin-binding peptide, a fibronectin domain, or human serum albumin) by a chemical bond, e.g., a peptide bond.

[0365] As used herein, the terms "bone mineral density (BMD)," "bone density," and "bone mass" refer to a measure of the amount of bone mineral (e.g., calcium) in bone tissue. BMD can be measured by well-established clinical techniques known to those skilled in the art (e.g., by single-energy photon or dual-energy photon or X-ray absorptiometry). The concept of BMD relates to the mass of mineral per volume of bone, but clinically it is measured by proxy according to the optical density per square centimeter of bone surface during imaging. BMD measurements are used in clinical medicine as an indirect indicator of osteoporosis and fracture risk. In some embodiments, BMD test results are provided as a T-score, which represents the subject's BMD compared to the ideal or peak bone density of a healthy 30-year-old adult. A score of 0 indicates a BMD equal to the normal reference value for a healthy young adult. The difference between the subject's measured BMD and the reference value for a healthy young adult is measured in standard deviation units (SD). Thus, a T-score between +1 SD and -1 SD may indicate normal BMD, a T-score between -1 SD and -2.5 SD may indicate low bone mass (e.g., osteopenia), and a T-score below -2.5 SD may indicate osteoporosis or severe osteoporosis. In some embodiments, a polypeptide of the present invention comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)), a nucleic acid encoding such a polypeptide, or a vector comprising such a nucleic acid molecule is administered to a subject in need thereof, and the patient has low bone mass (e.g., a T-score between -1 SD and -2.5 SD). In some embodiments, a polypeptide of the present invention comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOs: 1 to 43 (e.g., SEQ ID NOs: 22 to 43)), a nucleic acid encoding such a polypeptide, or a vector comprising such a nucleic acid molecule is administered to a subject in need thereof, and the patient has osteoporosis (e.g., a T-score of less than -2.5 SD).In some embodiments, administration of a polypeptide of the present invention comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOs: 1-43 (e.g., SEQ ID NOs: 22-43)), a nucleic acid encoding such a polypeptide, or a vector comprising such a nucleic acid molecule treats a subject by increasing BMD. In some embodiments, administration of a polypeptide of the present invention comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOs: 1-43 (e.g., SEQ ID NOs: 22-43)), a nucleic acid encoding such a polypeptide, or a vector comprising such a nucleic acid molecule increases the subject's BMD, resulting in an improvement in the subject's T-score (e.g., resulting in an increase in the subject's T-score of 0.1 or more, 0.2 or more, 0.3 or more, 0.4 or more, 0.5 or more, 1.0 or more, or 2.0 or more).

[0366] As used herein, the term "bone strength" refers to a bone measurement determined by bone quality in addition to bone density. Bone quality is influenced by bone shape, microarchitecture, and tissue constituent properties. Bone strength can be used to assess bone fracture risk.

[0367] As used herein, the term "bone disease" refers to a condition characterized by bone damage (e.g., decreased bone density, decreased bone strength, and / or bone loss). Such diseases or conditions can be caused by an imbalance in osteoblast and / or osteoclast activity (e.g., increased bone resorption or decreased bone formation). Bone diseases include primary osteoporosis, secondary osteoporosis, osteopenia, osteopetrosis, bone loss associated with bone cancer or cancer metastasis (e.g., bone loss associated with multiple myeloma), Paget's disease, renal osteodystrophy, osteogenesis imperfecta, bone loss associated with neuromuscular disease, bone loss due to burns (e.g., bone loss associated with burns), bone loss associated with anorexia, treatment-related bone loss, diet-related bone loss, bone loss associated with obesity treatment, bone loss associated with reduced gravity, and bone loss associated with immobility.

[0368] As used herein, the term "bone loss associated with neuromuscular disease" refers to bone loss that occurs in subjects with neuromuscular disease. Poor bone health is often a significant problem for patients with neuromuscular disease. For example, bone mineral density deficiency and increased incidence of fractures are well-known clinical consequences of diseases such as DMD, ALS, and SMA.

[0369] As used herein, the terms "bone remodeling" or "bone metabolism" refer to the process of maintaining bone strength and ionic homeostasis by replacing individual segments of old bone with packets of newly synthesized proteinaceous matrix. Bone is resorbed by osteoclasts and deposited by osteoblasts in a process called ossification. Osteocyte activity plays a key role in this process. Conditions leading to bone loss can be caused by either increased resorption or decreased ossification. In healthy individuals, bone formation exceeds resorption during childhood. As the aging process occurs, resorption exceeds formation. Additionally, older women, after menopause, typically experience a much higher rate of bone resorption due to estrogen deficiency associated with menopause.

[0370] As used herein, the terms "bone resorption" or "bone catabolic activity" refer to the process by which osteoclasts break down bone tissue to release minerals, resulting in the transfer of minerals (e.g., calcium) from bone tissue to the blood. Increased bone resorption rates are associated with aging, including postmenopausal women. High bone resorption rates, or bone resorption rates that exceed the ossification rate, are associated with bone disorders such as decreased bone mineral density, including osteopenia and osteoporosis, and can lead to bone loss. In some embodiments, a polypeptide of the present invention comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)), a nucleic acid encoding such a polypeptide, or a vector comprising such a nucleic acid molecule is administered to a subject in need thereof to reduce bone resorption (e.g., the amount or rate of bone resorption in the subject) (e.g., reduce bone loss).

[0371] As used herein, the terms "osteogenesis," "ossification," "bone formation," or "bone anabolic activity" refer to the process of new bone tissue formation by osteoblasts. In some embodiments, a polypeptide of the present invention comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)), a nucleic acid encoding such a polypeptide, or a vector comprising such a nucleic acid molecule is administered to a subject in need thereof to increase bone formation (e.g., increase the amount or rate of bone formation) in the subject. A decrease in the rate of bone formation, or a rate where the rate of bone formation exceeds the rate of bone resorption, can result in bone loss.

[0372] As used herein, the terms "increase" and "decrease" refer to modulations that result in a greater or lesser amount, respectively, of the function, expression, or activity of a metric compared to a reference. For example, following administration of a polypeptide of the invention comprising an extracellular ActRII chimera in the manner described herein, the amount of a metric marker described herein (e.g., lean body mass) can be increased or decreased in a subject relative to the amount of the marker before administration. Generally, the metric is measured after administration at a time when administration results in the described effect, e.g., at least 1 week, 1 month, 3 months, or 6 months after initiating a treatment regimen.

[0373] As used herein, the term "fibrosis" refers to the pathological process of excessive formation of fibrous connective tissue. Fibrosis is characterized by the accumulation of fibroblasts and the deposition of collagen in excess of normal deposition in any particular tissue. In response to inflammation or tissue injury, nearby fibroblasts migrate into the wound, proliferate, and produce large amounts of collagenous extracellular matrix. When fibrosis occurs in response to injury, the term "scarring" can be used synonymously. Fibrosis can occur in many tissues of the body, including, for example, the lung, skin, liver, kidney, heart, eye, tendon, cartilage, pancreatic tissue, uterine tissue, nervous tissue, testes, ovaries, adrenal glands, arteries, veins, bone marrow, colon, small and large intestine, biliary tract, and intestine.

[0374] As used herein, the term "pulmonary hypertension" or "PH" refers to a disease characterized by elevated blood pressure between the heart and lungs, and may include elevated blood pressure in the pulmonary arteries (pulmonary arterial hypertension), pulmonary veins, or pulmonary capillaries. Pulmonary hypertension has many symptoms, including shortness of breath (dyspnea), fatigue, swelling of the legs, feet, abdomen (ascites), or neck (e.g., edema), or chest pain or pressure, rapid pulse or palpitations, bluish lip or skin color (cyanosis), dizziness, or fainting. PH also reduces exercise tolerance and can lead to heart failure.

[0375] As used herein, the term "pulmonary arterial hypertension" or "PAH" refers to a form of pulmonary hypertension characterized by narrowing or obstruction of small pulmonary arteries, often caused by scarring, and elevated pulmonary artery blood pressure. PAH is also known as WHO Group I pulmonary hypertension. PAH can be diagnosed based on elevated pulmonary artery blood pressure (mean pulmonary artery pressure >25 mmHg at rest) with normal pulmonary artery capillary wedge pressure. PAH can cause shortness of breath, dizziness, syncope, and other symptoms, all of which are exacerbated by exercise. PAH can become a severe disease with significantly reduced exercise tolerance and heart failure. The two major types of PAH include idiopathic PAH (e.g., PAH without an identified predisposition) and hereditary PAH (e.g., PAH associated with mutations in BMPR2, ALK1, SMAD9, caveolin-1, KCNK3, or EIF2AK4). Mutations in the BMPR2 gene account for 70% of familial PAH cases. Risk factors for developing PAH include a family history of PAH, drug use (e.g., methamphetamine or cocaine use), infection (e.g., HIV infection or schistosomiasis), cirrhosis, congenital heart abnormalities, portal hypertension, veno-occlusive disease, pulmonary capillary hemangiomatosis, or connective tissue / autoimmune diseases (e.g., scleroderma or lupus).

[0376] As used herein, the terms "venous pulmonary hypertension" and "venous PH" refer to forms of pulmonary hypertension secondary to left heart disease. Venous PH is also known as WHO Group II PH. Venous PH can be associated with or caused by left ventricular systolic dysfunction (e.g., left ventricular failure), left ventricular diastolic dysfunction, valvular heart disease (e.g., mitral or aortic valve disease), congenital cardiomyopathy, or congenital or acquired pulmonary vein stenosis.

[0377] As used herein, the terms "hypoxic pulmonary hypertension" and "hypoxic PH" refer to a form of pulmonary hypertension resulting from lung disease or chronic hypoxia. This form of PH is also known as WHO Group III PH. Hypoxic PH can be associated with or caused by chronic obstructive pulmonary disease (e.g., emphysema), interstitial lung disease, sleep-disordered breathing (e.g., sleep apnea), lung disease (e.g., pulmonary fibrosis), alveolar hypoventilation disorders, chronic exposure to high altitude, or developmental abnormalities.

[0378] As used herein, the terms "thromboembolic pulmonary hypertension" and "thromboembolic PH" refer to forms of pulmonary hypertension associated with chronic arterial obstruction (e.g., thrombus). Thromboembolic PH is also known as WHO Group IV PH. Thromboembolic PH can be associated with or caused by chronic thromboembolic pulmonary hypertension or other pulmonary artery obstruction (e.g., pulmonary embolism, angiosarcoma, arteritis, congenital pulmonary stenosis, or parasitic infection).

[0379] As used herein, the terms "other pulmonary hypertension" and "other PH" refer to forms of pulmonary hypertension with unknown or multifactorial mechanisms. This form of PH is classified as WHO Group V PH. Other PH may be associated with or caused by hematological disorders (e.g., chronic hemolytic anemia, sickle cell disease), systemic diseases (e.g., sarcoidosis, pulmonary Langerhans cell histiocytosis, lymphangioleiomyomatosis, neurofibromatosis, or vasculitis), metabolic disorders (e.g., glycogen storage disease, Gaucher disease, or thyroid disease), pulmonary neoplastic thrombotic microangiopathy, fibrosing mediastinitis, chronic renal failure, or segmental pulmonary hypertension.

[0380] As used herein, the terms "increasing red blood cell levels" and "promoting red blood cell formation" refer to clinically observable metrics such as hematocrit, red blood cell count, and hemoglobin measurements, and are intended to be neutral with respect to the mechanism by which such changes occur. As used herein, the term "low red blood cell levels" refers to red blood cell count, hematocrit, and hemoglobin measurements that are below the range of values ​​considered normal for a subject's age and sex.

[0381] As used herein, the terms "erythropoiesis" and "erythropoiesis" refer to the production of red blood cells, such as the process of erythropoiesis in which red blood cells are produced in the bone marrow. As used herein, the terms "increasing platelet levels" and "promoting platelet formation" refer to clinically observable metrics, such as platelet count, and are intended to be neutral with respect to the mechanism by which such changes occur. As used herein, the term "low platelet levels" refers to a platelet count below the range of values ​​considered normal for a subject's age and sex. The terms "platelet formation" and "platelet production" refer to the production of platelets, such as the process by which platelets are produced from megakaryocytes.

[0382] As used herein, the terms "increasing neutrophil levels" and "promoting neutrophilogenesis" refer to clinically observable metrics, such as neutrophil count, and are intended to be neutral with respect to the mechanism by which such changes occur. As used herein, the term "low neutrophil levels" refers to neutrophil counts below the range of values ​​considered normal for a subject's age and sex. The terms "neutrophilogenesis" and "neutrophil production" refer to the generation of neutrophils, such as the process by which neutrophils are produced in the bone marrow.

[0383] As used herein, the term "anemia" refers to a hemoglobin or red blood cell abnormality that results in a reduced level of oxygen in the blood. Anemia can be associated with abnormal production, processing, or performance of red blood cells and / or hemoglobin. The term anemia refers to any decrease in the number of red blood cells and / or hemoglobin levels in the blood compared to normal blood levels.

[0384] As used herein, the term "thrombocytopenia" refers to a condition in which the blood contains a lower-than-normal number of platelets, which may be due to a lack of platelet production, accumulation of platelets in an enlarged spleen, or destruction of platelets. Normal platelet levels range from approximately 150,000 to 450,000 platelets per microliter of blood in humans. A platelet count below 150,000 platelets per microliter is lower than normal. A platelet count below 50,000 platelets per microliter of blood can cause bleeding after relatively minor injury, while a platelet count below 10,000 to 20,000 platelets per microliter of blood can cause severe bleeding without any apparent injury.

[0385] As used herein, the term "immune thrombocytopenia" refers to any type of thrombocytopenia resulting from an autoimmune response against an individual's own platelets. Immune thrombocytopenia includes primary immune thrombocytopenia, in which an autoimmune response is the original cause of a decrease in platelet count, such as idiopathic thrombocytopenic purpura. Immune thrombocytopenia also includes secondary immune thrombocytopenia, in which a decrease in platelet count is associated with one or more other diseases that cause an individual's body to trigger an autoimmune response against their own platelets, such as systemic lupus erythematosus (SLE), antiphospholipid syndrome (APS), Evans syndrome, immune-mediated thyroid disease, leukemia (e.g., chronic lymphocytic leukemia or large granular T-lymphocytic leukemia), or chronic infection (e.g., Helicobacter pylori, human immunodeficiency virus (HIV), or hepatitis C).

[0386] As used herein, the term "neutropenia" refers to an abnormally low number of neutrophils in the blood. The typical lower limit of neutrophil count is approximately 1500 cells per microliter of blood. Below this level, the risk of infection increases. The severity of neutropenia is classified as mild (1000-1500 neutrophils per microliter of blood), moderate (500-1000 neutrophils per microliter of blood), or severe (fewer than 500 neutrophils per microliter of blood). Neutropenia has many causes, but typically falls into two major categories: destruction or depletion of neutrophils faster than the bone marrow can produce new ones, or a decrease in neutrophil production in the bone marrow.

[0387] As used herein, the term "low transfusion burden" refers to the condition of a subject who received fewer than 4 units of red blood cells (RBCs) within 8 weeks prior to treatment with an ActRII chimera described herein (e.g., 3, 2, 1, or 0 units of RBCs within 8 weeks). A subject with a low transfusion burden may be identified as anemic based on measurements of mean hemoglobin concentration. A low transfusion burden and a mean hemoglobin concentration of less than 10.0 g / dL on two measurements taken at least one week apart (e.g., one measurement taken within 1 day prior to treatment and the other taken 7-28 days prior that is not affected by an RBC transfusion within 7 days of the measurement) prior to treatment with an ActRII chimera described herein is defined as anemic. In some embodiments, a subject with a low transfusion burden receives 1-3 units of RBCs (1-3 RBC transfusions) within 8 weeks prior to treatment with an ActRII chimera described herein. In some embodiments, a subject with low transfusion burden does not receive any RBCs (0 RBC transfusions) within 8 weeks prior to treatment with an ActRII chimera described herein.

[0388] As used herein, the term "high transfusion burden" refers to a condition in which a subject requires 4 or more units of RBC (e.g., 4, 5, 6, 7, 8 units, or more) within 8 weeks prior to treatment with the ActRII chimera described herein. A subject with a high transfusion burden can be identified as anemic based on measurements of mean hemoglobin concentration. A subject with a high transfusion burden and a mean hemoglobin concentration of 9.0 g / dL or less is defined as anemic.

[0389] As used herein, the term "ineffective hematopoiesis" refers to a failure to produce fully mature hematopoietic cells (e.g., a failure to produce red blood cells, platelets, and neutrophils). Ineffective hematopoiesis can be due to single or multiple defects, such as abnormal proliferation and / or differentiation of progenitor cells (e.g., an overproduction of progenitor cells that fail to complete differentiation), which can lead to an overproliferation or shortage of progenitor cells.

[0390] As used herein, the terms "erythropoiesis-stimulating agents" and "ESAs" refer to a class of drugs that act on the proliferative phase of erythroid development by expanding the pool of early stage progenitor cells. Examples of erythropoiesis-stimulating agents are epoetin alfa and darbepoetin alfa.

[0391] As used herein, the term "metabolic disease" refers to a disease, disorder, or syndrome related to a subject's metabolism, such as breaking down carbohydrates, proteins, and fats in food to release energy, converting chemicals into other substances, and transporting them into cells for energy utilization and / or storage. Some symptoms of metabolic disease include high serum triglycerides, high low-density cholesterol (LDL), low high-density cholesterol (HDL), and / or high fasting insulin levels, elevated fasting blood glucose, abdominal (central) obesity, and elevated blood pressure. Metabolic disease increases the risk of developing other diseases, such as cardiovascular disease. In the present invention, metabolic diseases include, but are not limited to, obesity, type 1 diabetes, and type 2 diabetes.

[0392] As used herein, the term "treatment-related metabolic disease" refers to a metabolic disease (e.g., obesity, type 1 diabetes, or type 2 diabetes) associated with a drug taken by a subject (e.g., a metabolic disease that develops during drug treatment). The drug may be one that the subject continues to take or one that was taken prior to the onset of the metabolic disease. Drugs associated with the development of obesity include glucocorticoids (e.g., corticosteroids such as prednisone), selective serotonin reuptake inhibitors (SSRIs, e.g., paroxetine, mirtazapine, fluoxetine, escitalopram, sertraline), tricyclic antidepressants (e.g., amitriptyline), mood stabilizers (e.g., valproic acid, lithium), antipsychotics (e.g., olanzapine, chlorpromazine, clozapine), and diabetes medications (e.g., insulin, chlorpropamide). Medications associated with the development of diabetes include glucocorticoids (e.g., corticosteroids, which can cause glucocorticoid-induced diabetes), SSRIs, serotonin-norepinephrine reuptake inhibitors (SNRIs), mood stabilizers (e.g., lithium and valproate), and antipsychotics (e.g., olanzapine and clozapine). In some embodiments, the development of obesity can lead to the development of diabetes.

[0393] As used herein, the term "age-related metabolic disease" refers to metabolic diseases (e.g., obesity, type 1 diabetes, or type 2 diabetes) that develop with age. For example, the risk of diabetes increases with age and is more prevalent among older adults, with approximately 25% of adults over 60 years of age having diabetes. Adults can develop type 2 diabetes or new-onset type 1 diabetes. Obesity rates also increase with age, with the highest obesity rates in the United States among adults aged 40-59 (45% prevalence of obesity). Aging also reduces the body's ability to burn fat, leading to an increase in fat around internal organs.

[0394] As used herein, the term "percentage weight gain" refers to the percentage of weight gained compared to a subject's previous weight. Percentage weight gain can be calculated as follows: 100 x [(post-weight - pre-weight) / (pre-weight)] In the present invention, administration of a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOs: 1 to 43 (e.g., SEQ ID NOs: 22 to 43)), a nucleic acid molecule encoding a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOs: 1 to 43 (e.g., SEQ ID NOs: 22 to 43)), or a vector comprising such a nucleic acid molecule to a subject can reduce the subject's weight gain percentage.

[0395] As used herein, the term "appetite for food intake" refers to a subject's natural desire or need for food. A subject's appetite for food intake can be monitored by measuring the amount of food consumed after administration of a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)). In the present invention, administration of a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)), a nucleic acid molecule encoding a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)), or a vector comprising such a nucleic acid molecule to a subject does not affect the subject's appetite for food intake.

[0396] As used herein, the term "adiposity" refers to fat stored in adipose tissue of a subject. In the present invention, administration of a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1 to 43 (e.g., SEQ ID NOS: 22 to 43)), a nucleic acid molecule encoding a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1 to 43 (e.g., SEQ ID NOS: 22 to 43)), or a vector comprising such a nucleic acid molecule to a subject can reduce obesity in the subject without affecting lean body mass.

[0397] As used herein, the term "epididymal and perirenal fat pads" refers to the adipocytes densely packed around the epididymis and kidney. In the present invention, administration of a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1 to 43 (e.g., SEQ ID NOS: 22 to 43)), a nucleic acid molecule encoding a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1 to 43 (e.g., SEQ ID NOS: 22 to 43)), or a vector comprising such a nucleic acid molecule to a subject can reduce the weight of the subject's epididymal and perirenal fat pads.

[0398] As used herein, the term "fasting insulin" refers to a subject's insulin level during a prolonged period (i.e., 12 to 24 hours) without food. Fasting insulin levels are used to diagnose metabolic diseases. Fasting insulin levels are also used as an indicator of whether a subject is at risk for developing metabolic diseases. Typically, fasting insulin levels in patients with type 1 diabetes are lower than those in healthy subjects. In subjects suffering from insulin resistance (i.e., type 2 diabetes), the subject's fasting insulin level is higher than those in healthy subjects. In the present invention, administration of a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1 to 43 (e.g., SEQ ID NOS: 22 to 43)), a nucleic acid molecule encoding a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1 to 43 (e.g., SEQ ID NOS: 22 to 43)), or a vector comprising such a nucleic acid molecule to a subject can regulate the subject's fasting insulin level.

[0399] As used herein, the term "glucose clearance rate" refers to the rate at which glucose is removed from the blood. Glucose clearance rate can be measured by a glucose tolerance test (GTT). In a GTT, a subject is administered a fixed amount of glucose, and then a blood sample is taken to measure the rate at which glucose is removed from the blood. Glucose clearance rate can be used as a parameter in diagnosing and / or determining the risk of developing metabolic diseases such as obesity, diabetes, and insulin resistance.

[0400] As used herein, the term "serum lipid profile" refers to measuring the distribution of various types of lipids and lipoproteins in a subject's serum. Such measurements can be performed by a series of blood tests. The types of lipids and lipoproteins in a subject's serum include, but are not limited to, cholesterol (e.g., high-density lipoprotein (HDL) and low-density lipoprotein (LDL)), triglycerides, and free fatty acids (FFA). The distribution of various types of lipids and lipoproteins can be used as a parameter in diagnosing and / or determining the risk of developing metabolic diseases such as obesity, diabetes, and insulin resistance. High levels of cholesterol, especially low-density lipoprotein, are generally considered to be a sign or risk factor for developing certain metabolic diseases, or in some severe medical cases, cardiovascular disease. In the present invention, administration of a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOs: 1 to 43 (e.g., SEQ ID NOs: 22 to 43)), a nucleic acid molecule encoding a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOs: 1 to 43 (e.g., SEQ ID NOs: 22 to 43)), or a vector comprising such a nucleic acid molecule to a subject improves the subject's serum lipid profile so that cholesterol (particularly low-density lipoprotein) and triglyceride levels are reduced.

[0401] As used herein, the term "C-terminal extension" refers to the addition of one or more amino acids to the C-terminus of an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)). The C-terminal extension can be one or more amino acids, such as 1 to 6 amino acids (e.g., 1, 2, 3, 4, 5, 6, or more amino acids). The C-terminal extension can include amino acids from the corresponding positions of wild-type ActRIIA or ActRIIB. Exemplary C-terminal extensions are the amino acid sequence NP (2-amino acid C-terminal extension) and the amino acid sequence NPVTPK (SEQ ID NO: 104) (6-amino acid C-terminal extension). Any amino acid sequence that does not interfere with the activity of the polypeptide can be used.

[0402] As used herein, the term "percent identity" refers to the percentage of amino acid (or nucleic acid) residues in a candidate sequence that are identical to the amino acid (or nucleic acid) residues in a reference sequence after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent identity (i.e., gaps can be introduced in one or both of the candidate and reference sequences for optimal alignment, and non-homologous sequences can be ignored for comparison purposes). Alignment for purposes of determining percent identity can be achieved in a variety of ways within the skill of the art, for example, using publicly available computer software such as BLAST, ALIGN, or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms needed to achieve maximum alignment over the entire length of the sequences being compared. In some embodiments, the percent amino acid (or nucleic acid) sequence identity of a given candidate sequence to, with, or relative to a given reference sequence (which can be rephrased as a given candidate sequence having or comprising a particular percent amino acid (or nucleic acid) sequence identity to, with, or relative to a given reference sequence) is calculated as follows: 100 x (fraction A / B) where A is the number of amino acid (or nucleic acid) residues assessed as identical in an alignment of the candidate sequence with the reference sequence, and B is the total number of amino acid (or nucleic acid) residues in the reference sequence. In some embodiments where the length of the candidate sequence is not equal to the length of the reference sequence, the percent amino acid (or nucleic acid) sequence identity of the candidate sequence to the reference sequence is not equal to the percent amino acid (or nucleic acid) sequence identity of the reference sequence to the candidate sequence.

[0403] In certain embodiments, a reference sequence aligned for comparison to a candidate sequence may show that the candidate sequence exhibits 50% to 100% identity over the entire length of the candidate sequence or over a selected portion of consecutive amino acid (or nucleic acid) residues of the candidate sequence. The length of the candidate sequence aligned for comparison purposes may be, for example, at least 30%, e.g., at least 40%, e.g., at least 50%, 60%, 70%, 80%, 90%, or 100% of the length of the reference sequence. When a position in the candidate sequence is occupied by the same amino acid (or nucleic acid) residue as the corresponding position in the reference sequence, the molecules are identical at that position.

[0404] As used herein, the term "serum half-life" refers to the time required for the plasma concentration of a therapeutic protein in a subject to decrease by half in the context of administering the therapeutic protein to the subject.The protein may be redistributed or removed from the bloodstream, or may be degraded, for example, by proteolysis.The serum half-life comparison can be performed by comparing the serum half-life of Fc fusion proteins.

[0405] As used herein, the term "lean body mass" refers to components of body composition, including, for example, lean body mass, body fat, and body fluid. Lean body mass is typically calculated by subtracting the weight of body fat and body fluid from total body weight. Typically, a subject's lean body mass is 60% to 90% of their total body weight. In the present invention, administering to a subject a polypeptide comprising an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1 to 43 (e.g., SEQ ID NOS: 22 to 43), a nucleic acid molecule encoding a polypeptide comprising an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1 to 43 (e.g., SEQ ID NOS: 22 to 43)), or a vector comprising such a nucleic acid molecule increases the subject's lean body mass.

[0406] As used herein, the term "affinity" or "binding affinity" refers to the strength of the binding interaction between two molecules. Generally, binding affinity refers to the strength of the total non-covalent interactions between a molecule and its binding partner, e.g., an extracellular ActRII chimera and BMP9 or activin A. Unless otherwise indicated, binding affinity refers to the intrinsic binding affinity, which reflects a 1:1 interaction between members of a binding pair. The binding affinity between two molecules is usually measured using a dissociation constant (K D ) or affinity constant (K A ) Two molecules that have low binding affinity for each other generally bind slowly and tend to dissociate easily, resulting in a large K D Two molecules that have a high affinity for each other generally bind more easily and tend to remain bound longer, with a small K D The K of two interacting molecules is D can be determined using methods and techniques well known in the art, such as surface plasmon resonance. D is k off / k on It is calculated as the ratio of

[0407] As used herein, the term "muscle mass" refers to the primary component of lean body mass. Muscle mass can be measured experimentally by measuring muscle weight. As used herein, the term "neuromuscular disease" refers to a disease in which problems with the nerves and muscles affect voluntary or involuntary muscle function, typically resulting in muscle weakness. Exemplary neuromuscular diseases include amyotrophic lateral sclerosis (ALS), autonomic neuropathies, botulism, Charcot-Marie-Tooth disease (CMT), chronic inflammatory demyelinating polyneuropathy, congenital myasthenic syndromes, congenital myopathies, cramp-fasciculation syndromes, dermatomyositis, diabetic neuropathy, distal myopathy, dystrophinopathy, endocrine myopathy, focal muscular atrophy, glycogen storage disease type II, Guillain-Barré syndrome, hereditary spastic paraplegia, inclusion body myositis (IBM), Isaac syndrome, Kearns-Sayre syndrome, Kennedy disease, Lambert-Eaton myasthenic syndrome, metabolic myopathies, metabolic neuropathies, mitochondrial myopathies, motor neuron diseases, multiple sclerosis, muscular dystrophies (e.g., Duchesne-Holland syndrome, dystrophies), and muscular dystrophies (e.g., Duchesne-Holland syndrome). These include: DMD, Becker's dystrophy (BMD), myotonic myotonic myopathy (DM), facioscapulohumeral myotonic myopathy (FSHD), limb-girdle myotonic myopathy (LGMD), distal dysplasia (DD), oculopharyngeal myotonic myopathy (OPMD), Emery-Dreyfus' dysplasia (EDMD), and congenital (e.g., MDC1A, MDC1B, MDC1C, FCMD, WWS, RSMD1, MEB, and UCMD)), myasthenia gravis, myotonic dystrophy, necrotizing myopathy, neuromyotonia, Friedreich's ataxia neuropathy, nutritional neuropathy, peripheral neuropathy, polymyositis, primary lateral sclerosis, Schwartz-Jampel syndrome, small fiber neuropathy, spinal and bulbar muscular atrophy, spinal muscular atrophy, spinal muscular atrophy with respiratory distress syndrome type 1, spinocerebellar ataxia, stiff-person syndrome, toxic neuropathy, and Treuer syndrome. Neuromuscular diseases can be inherited in an autosomal dominant or recessive pattern, or by spontaneously occurring mutations.

[0408] As used herein, the phrase "affecting myostatin, activin A, activin B, and / or BMP9 signaling" means altering the binding of myostatin, activin A, activin B, and / or BMP9 to their receptors, e.g., ActRIIA, ActRIIB, and BMPRII (e.g., endogenous receptors). In some embodiments, polypeptides comprising extracellular ActRII chimeras described herein reduce or inhibit the binding of myostatin, activin A, activin B, and / or BMP9 to their receptors, e.g., ActRIIA, ActRIIB, and BMPRII (e.g., endogenous ActRIIA and / or ActRIIB).

[0409] As used herein, the term "vascular complication" refers to any damage to blood vessels, such as a vascular disorder or damage to the blood vessel wall. Damage to the blood vessel wall can cause increased vascular permeability or vascular leakage. The term "vascular permeability or vascular leakage" refers to the ability of the blood vessel wall to allow the flow of small molecules, proteins, and cells into and out of the blood vessel. Increased vascular permeability or vascular leakage can be caused by an increase in gaps between endothelial cells lining the blood vessel wall (e.g., an increase in the size and / or number of gaps) and / or thinning of the blood vessel wall.

[0410] As used herein, the term "polypeptide" refers to a single polymer in which the monomers are amino acid residues covalently linked to one another through amide bonds. Polypeptide is intended to encompass any naturally occurring, recombinant, or synthetically produced amino acid sequence.

[0411] As used herein, the term "homodimer" refers to a molecular construct formed by two identical macromolecules, such as proteins or nucleic acids. Two identical monomers can form a homodimer through covalent or non-covalent bonds. For example, an Fc domain can be a homodimer of two Fc domain monomers if the two Fc domain monomers contain the same sequence. In another example, a polypeptide described herein comprising an extracellular ActRII chimera fused to an Fc domain monomer can form a homodimer through the interaction of the two Fc domain monomers that form the Fc domain in the homodimer.

[0412] As used herein, the term "heterodimer" refers to a molecular construct formed by two different macromolecules, such as proteins or nucleic acids. The two monomers may form a heterodimer through covalent or non-covalent bonds. For example, a polypeptide described herein comprising an extracellular ActRII chimera fused to an Fc domain monomer may form a heterodimer through the interaction of two Fc domain monomers, each fused to a different ActRII chimera, that form an Fc domain in the heterodimer.

[0413] As used herein, the term "host cell" refers to a vehicle containing the necessary cellular components, e.g., organelles, required for expressing a protein from a corresponding nucleic acid. The nucleic acid is typically contained in a nucleic acid vector that can be introduced into the host cell by conventional techniques known in the art (transformation, transfection, electroporation, calcium phosphate precipitation, direct microinjection, etc.). The host cell can be a prokaryotic cell, e.g., a bacterial cell, or a eukaryotic cell, e.g., a mammalian cell (e.g., a CHO cell or HEK293 cell).

[0414] As used herein, the term "therapeutically effective amount" refers to a therapeutically effective amount of a compound or compound that is effective in treating a muscle disorder, a condition involving muscle weakness and atrophy (e.g., a neuromuscular disorder such as muscular dystrophy, IBM, ALS, SMA, CMT, myasthenia gravis, or multiple sclerosis; sarcopenia; or cachexia), a disease or condition involving bone damage (e.g., osteoporosis or a condition involving bone damage, such as primary osteoporosis, secondary osteoporosis, osteopenia, osteopetrosis, fractures, bone loss associated with bone cancer or cancer metastasis, Paget's disease, renal osteodystrophy, treatment-related bone loss, osteogenesis imperfecta, bone loss associated with neuromuscular disorders, bone loss due to burns, bone loss associated with anorexia, diet-related bone loss, bone loss associated with the treatment of obesity, or a condition involving bone damage, such as osteoporosis, secondary osteoporosis, osteopenia, osteopetrosis, bone ... osteoporosis, bone loss associated with bone loss, bone loss associated with bone loss, bone loss associated with bone loss, bone loss associated with bone loss, bone loss associated with bone loss, bone loss associated with bone In treating patients with or at risk of developing a disease such as bone loss, bone mass loss associated with low red blood cell levels (e.g., anemia or blood loss), a disease or condition associated with low platelet levels (e.g., thrombocytopenia), a disease or condition associated with low neutrophil levels (e.g., neutropenia), a disease or condition associated with fibrosis, a metabolic disease, or PH (e.g., PAH, venous PH, hypoxic PH, thromboembolic PH, or other PH), a therapeutically effective amount of a polypeptide, nucleic acid, or vector of the present invention, or a pharmaceutical composition comprising a polypeptide, nucleic acid, or vector of the present invention, is effective to achieve the desired therapeutic effect. In particular, a therapeutically effective amount of a polypeptide, nucleic acid, or vector avoids adverse side effects.

[0415] As used herein, the term "pharmaceutical composition" refers to a medicament or pharmaceutical formulation containing an active ingredient and excipients and diluents to make the active ingredient suitable for the method of administration. The pharmaceutical composition of the present invention contains pharmaceutically acceptable ingredients that are compatible with the polypeptide, nucleic acid, or vector. The pharmaceutical composition may be in the form of a tablet or capsule for oral administration, or in aqueous form for intravenous or subcutaneous administration.

[0416] As used herein, the term "pharmaceutically acceptable carrier or excipient" refers to an excipient or diluent in a pharmaceutical composition. A pharmaceutically acceptable carrier must be compatible with other ingredients of the formulation and not deleterious to the recipient. In the present invention, a pharmaceutically acceptable carrier or excipient must provide adequate pharmaceutical stability for a polypeptide comprising an extracellular ActRII chimera, a nucleic acid molecule(s) encoding the polypeptide, or a vector comprising such a nucleic acid molecule(s). The nature of the carrier or excipient will vary depending on the method of administration. For example, for intravenous administration, an aqueous carrier is generally used. For oral administration, a solid carrier is preferred.

[0417] As used herein, the term "treatment and / or prevention" refers to the use of the methods and compositions of the present invention to treat or prevent a disease, such as a muscle disease (e.g., muscular dystrophy, IBM, SMA, CMT, ALS, myasthenia gravis, or multiple sclerosis; neuromuscular diseases such as sarcopenia; or cachexia), a bone disease (e.g., a disease or condition involving bone damage, e.g., osteoporosis, osteopenia, osteopetrosis, bone fractures, bone loss associated with bone cancer or cancer metastasis, Paget's disease, renal osteodystrophy, treatment-related bone loss, osteogenesis imperfecta, neuromuscular diseases, etc.), or a combination of these. "Bone loss" refers to the treatment and / or prevention of a muscle, bone, low blood cell, low platelet, low neutrophil, metabolic, or fibrotic disease or PH (e.g., bone loss associated with a muscle, bone, low blood cell, low platelet, low neutrophil, metabolic, or fibrotic disease or PH) after the subject has developed the muscle, bone, low blood cell, low platelet, low neutrophil, metabolic, or fibrotic disease or PH and / or has already been diagnosed with the muscle, bone, low blood cell, low platelet, low neutrophil, metabolic, or fibrotic disease or PH. Preventing muscle, bone, low blood count, low platelet count, low neutrophil count, metabolic, or fibrotic disease, or PH refers to steps or treatments taken when a subject is at risk of developing muscle, bone, low blood count, low platelet count, low neutrophil count, metabolic, or fibrotic disease, or PH.The subject may exhibit signs or mild symptoms that are determined by a physician to be indicative of or risk factors for developing a muscle, bone, low blood count, low platelet count, low neutrophil count, metabolic, or fibrotic disease, or PH; may have another disease or condition that is associated with the development of a muscle, bone, low blood count, low platelet count, low neutrophil count, metabolic, or fibrotic disease, or PH; may be undergoing treatment (e.g., surgery, chemotherapy, or radiation) that may cause anemia, thrombocytopenia, neutropenia, fibrosis, obesity, or diabetes, or loss of bone density; or may have a family history or genetic predisposition to developing a muscle, bone, low blood count, low platelet count, low neutrophil count, metabolic, or fibrotic disease, or PH, but may not yet have the disease.

[0418] As used herein, the term "subject" refers to a mammal, for example, preferably a human. Mammals include, but are not limited to, humans and domestic animals and livestock, such as monkeys (e.g., cynomolgus monkeys), mice, rats, dogs, cats, horses, sheep, goats, rabbits, and cows. [Brief explanation of the drawings]

[0419] [Figure 1] 1 is a schematic diagram showing the amino acid sequences of exemplary extracellular ActRII chimeras. The sequence corresponding to wild-type extracellular ActRIIA is shown in bold, and the sequence corresponding to wild-type extracellular ActRIIB is shown in italics. Amino acid substitutions not derived from the ActRIIA or ActRIIB sequences are shown in bold italics. [Figure 2] 1 is a bar graph showing the effect of extracellular ActRII chimeras on body weight. Mice received a single hydrodynamic injection of a plasmid construct encoding the indicated ActRII chimera or vehicle. [Figure 3] 1 is a bar graph showing the effect of extracellular ActRII chimeras on lean body mass. Mice received a single hydrodynamic injection of a plasmid construct encoding the indicated ActRII chimera or vehicle. [Figure 4]1 is a bar graph showing the effect of extracellular ActRII chimeras on body weight. Mice received intraperitoneal injections of the indicated recombinant ActRII chimeras or vehicle control twice weekly for 4 weeks. [Figure 5] 1 is a bar graph showing the effect of extracellular ActRII chimeras on lean body mass. Mice received intraperitoneal injections of the indicated recombinant ActRII chimeras or vehicle control twice weekly for 4 weeks. [Figure 6A] This is a series of bar graphs showing the effects of extracellular ActRII chimeras on hematological parameters. Ten-week-old wild-type male C57Bl / 6 mice received hydrodynamic tail vein injections (HDIs) containing vehicle (n = 7) or vehicle containing expression vectors for either chimera 1 / 2-Fc (n = 9), chimera 1 / 2b-Fc (n = 8), chimera 3-Fc (n = 8), chimera 4-Fc (n = 8), chimera 5-Fc (n = 8), or chimera 2 I65E N66T E81Q-Fc (n = 9) at a dose of 0.75 mg / kg. Seven days after HDI, blood was collected from the submandibular vein of restrained, conscious mice. EDTA was used as an anticoagulant, and blood was analyzed using a Heska HT5 veterinary hematology analyzer. The effect of extracellular ActRII chimeras on red blood cell levels is shown. Data are presented as mean ± SEM. Statistics are presented compared to vehicle treatment using one-way ANOVA with Fisher's LSD post-hoc test. ***P<0.001, ****P<0.0001. [Figure 6B]This is a series of bar graphs showing the effects of extracellular ActRII chimeras on hematological parameters. Ten-week-old wild-type male C57Bl / 6 mice received hydrodynamic tail vein injections (HDIs) containing vehicle (n = 7) or vehicle containing expression vectors for either chimera 1 / 2-Fc (n = 9), chimera 1 / 2b-Fc (n = 8), chimera 3-Fc (n = 8), chimera 4-Fc (n = 8), chimera 5-Fc (n = 8), or chimera 2 I65E N66T E81Q-Fc (n = 9) at a dose of 0.75 mg / kg. Seven days after HDI, blood was collected from the submandibular vein of restrained, conscious mice. EDTA was used as an anticoagulant, and blood was analyzed using a Heska HT5 veterinary hematology analyzer. The effect of extracellular ActRII chimeras on hemoglobin levels is shown. Data are presented as mean ± SEM. Statistics are presented compared to vehicle treatment using one-way ANOVA with Fisher's LSD post-hoc test. ***P<0.001, ****P<0.0001. [Figure 6C] This is a series of bar graphs showing the effects of extracellular ActRII chimeras on hematological parameters. Ten-week-old wild-type male C57Bl / 6 mice received hydrodynamic tail vein injections (HDIs) containing vehicle (n = 7) or vehicle containing expression vectors for either chimera 1 / 2-Fc (n = 9), chimera 1 / 2b-Fc (n = 8), chimera 3-Fc (n = 8), chimera 4-Fc (n = 8), chimera 5-Fc (n = 8), or chimera 2 I65E N66T E81Q-Fc (n = 9) at a dose of 0.75 mg / kg. Seven days after HDI, blood was collected from the submandibular vein of restrained, conscious mice. EDTA was used as an anticoagulant, and blood was analyzed using a Heska HT5 veterinary hematology analyzer. The effect of extracellular ActRII chimeras on hematocrit levels is shown. Data are presented as mean ± SEM. Statistics are presented compared to vehicle treatment using one-way ANOVA with Fisher's LSD post-hoc test. ***P<0.001, ****P<0.0001. [Figure 6D]This is a series of bar graphs showing the effects of extracellular ActRII chimeras on hematological parameters. Ten-week-old wild-type male C57Bl / 6 mice received hydrodynamic tail vein injections (HDIs) containing vehicle (n = 7) or vehicle containing expression vectors for either chimera 1 / 2-Fc (n = 9), chimera 1 / 2b-Fc (n = 8), chimera 3-Fc (n = 8), chimera 4-Fc (n = 8), chimera 5-Fc (n = 8), or chimera 2 I65E N66T E81Q-Fc (n = 9) at a dose of 0.75 mg / kg. Seven days after HDI, blood was collected from the submandibular vein of restrained, conscious mice. EDTA was used as an anticoagulant, and blood was analyzed using a Heska HT5 veterinary hematology analyzer. The effect of extracellular ActRII chimeras on absolute reticulocyte counts is shown. Data are presented as mean ± SEM. Statistics are presented compared to vehicle treatment using one-way ANOVA with Fisher's LSD post-hoc test. ***P<0.001, ****P<0.0001. [Figure 7] This is a bar graph showing the effect of ActRII chimeras on lean body mass. Ten-week-old wild-type male C57Bl / 6 mice received hydrodynamic tail vein injections (HDIs) containing vehicle (n = 7) or vehicle containing expression vectors for either chimera 1 / 2-Fc (n = 9), chimera 1 / 2b-Fc (n = 8), chimera 3-Fc (n = 8), chimera 4-Fc (n = 8), chimera 5-Fc (n = 8), or chimera 2 I65E N66T E81Q-Fc (n = 9) at a dose of 0.75 mg / kg. Prior to administration, mouse body weight and lean body mass were measured using a miniature rodent nuclear magnetic resonance (NMR) analyzer (Bruker, Minispec LF50). The study was terminated after 28 days, and final body weight and lean body mass were measured. The effect of extracellular ActRII chimeras on lean body mass is shown. Data are presented as mean±SEM. Statistics are presented compared to vehicle treatment using one-way ANOVA with Fisher's LSD post-hoc test. *P≦0.05 and ****P<0.0001. [Figure 8A]This is a series of bar graphs showing the effects of extracellular ActRII chimeras on bone mineral density and structure. Ten-week-old wild-type male C57Bl / 6 mice received a hydrodynamic tail vein injection (HDI) containing either vehicle (n = 7) or vehicle containing expression vectors for either chimera 1 / 2-Fc (n = 9), chimera 1 / 2b-Fc (n = 8), chimera 3-Fc (n = 8), chimera 4-Fc (n = 8), chimera 5-Fc (n = 8), or chimera 2 I65E N66T E81Q-Fc (n = 9) at a dose of 0.75 mg / kg. μCT analysis was used to quantify tibial trabecular morphometry and tibial bone mineral density. The effects of extracellular ActRII chimeras on tibial bone mineral density, trabecular volume fraction at the proximal end of the tibia, trabecular spacing within the proximal end of the tibia, and trabecular number at the proximal end of the tibia are shown. Data are presented as mean ± SEM. Statistics are presented compared to vehicle treatment using one-way ANOVA with Fisher's LSD post-hoc test. *P≦0.05, **P<0.01, ***P<0.001, ****P<0.0001. [Figure 8B] This is a series of bar graphs showing the effects of extracellular ActRII chimeras on bone mineral density and structure. Ten-week-old wild-type male C57Bl / 6 mice received a hydrodynamic tail vein injection (HDI) containing either vehicle (n = 7) or vehicle containing expression vectors for either chimera 1 / 2-Fc (n = 9), chimera 1 / 2b-Fc (n = 8), chimera 3-Fc (n = 8), chimera 4-Fc (n = 8), chimera 5-Fc (n = 8), or chimera 2 I65E N66T E81Q-Fc (n = 9) at a dose of 0.75 mg / kg. μCT analysis was used to quantify tibial trabecular morphometry and tibial bone mineral density. The effects of extracellular ActRII chimeras on tibial bone mineral density, trabecular volume fraction at the proximal end of the tibia, trabecular spacing within the proximal end of the tibia, and trabecular number at the proximal end of the tibia are shown. Data are presented as mean ± SEM. Statistics are presented compared to vehicle treatment using one-way ANOVA with Fisher's LSD post-hoc test. *P≦0.05, **P<0.01, ***P<0.001, ****P<0.0001. [Figure 8C]This is a series of bar graphs showing the effects of extracellular ActRII chimeras on bone mineral density and structure. Ten-week-old wild-type male C57Bl / 6 mice received a hydrodynamic tail vein injection (HDI) containing either vehicle (n = 7) or vehicle containing expression vectors for either chimera 1 / 2-Fc (n = 9), chimera 1 / 2b-Fc (n = 8), chimera 3-Fc (n = 8), chimera 4-Fc (n = 8), chimera 5-Fc (n = 8), or chimera 2 I65E N66T E81Q-Fc (n = 9) at a dose of 0.75 mg / kg. μCT analysis was used to quantify tibial trabecular morphometry and tibial bone mineral density. The effects of extracellular ActRII chimeras on tibial bone mineral density, trabecular volume fraction at the proximal end of the tibia, trabecular spacing within the proximal end of the tibia, and trabecular number at the proximal end of the tibia are shown. Data are presented as mean ± SEM. Statistics are presented compared to vehicle treatment using one-way ANOVA with Fisher's LSD post-hoc test. *P≦0.05, **P<0.01, ***P<0.001, ****P<0.0001. [Figure 8D] This is a series of bar graphs showing the effects of extracellular ActRII chimeras on bone mineral density and structure. Ten-week-old wild-type male C57Bl / 6 mice received a hydrodynamic tail vein injection (HDI) containing either vehicle (n = 7) or vehicle containing expression vectors for either chimera 1 / 2-Fc (n = 9), chimera 1 / 2b-Fc (n = 8), chimera 3-Fc (n = 8), chimera 4-Fc (n = 8), chimera 5-Fc (n = 8), or chimera 2 I65E N66T E81Q-Fc (n = 9) at a dose of 0.75 mg / kg. μCT analysis was used to quantify tibial trabecular morphometry and tibial bone mineral density. The effects of extracellular ActRII chimeras on tibial bone mineral density, trabecular volume fraction at the proximal end of the tibia, trabecular spacing within the proximal end of the tibia, and trabecular number at the proximal end of the tibia are shown. Data are presented as mean ± SEM. Statistics are presented compared to vehicle treatment using one-way ANOVA with Fisher's LSD post-hoc test. *P≦0.05, **P<0.01, ***P<0.001, ****P<0.0001. DETAILED DESCRIPTION OF THE INVENTION

[0420] The present invention features polypeptides comprising extracellular activin receptor type II (ActRII) chimeras. In some embodiments, the polypeptides of the invention comprise an extracellular ActRII chimera fused to a moiety (e.g., an Fc domain monomer, a wild-type Fc domain, an Fc domain containing amino acid substitutions (e.g., one or more substitutions that reduce dimerization), an albumin-binding peptide, a fibronectin domain, or human serum albumin). Polypeptides comprising an extracellular ActRII chimera fused to an Fc domain monomer can also form dimers (e.g., homodimers or heterodimers) through interaction between two Fc domain monomers. The ActRII chimeras described herein may have reduced binding to bone morphogenetic protein 9 (BMP9) compared to wild-type extracellular ActRIIB, or may have weaker or no binding affinity for BMP9 compared to the binding affinity for activins (e.g., activin A and / or activin B) and myostatin. The present invention also provides methods for treating diseases and conditions associated with muscle wasting and atrophy by increasing muscle mass, lean body mass, and / or muscle strength, treating or preventing bone damage by increasing bone density, increasing bone formation, or decreasing bone resorption, treating or preventing fibrosis, and improving red blood cell levels (e.g., red blood cell count, hemoglobin level, or hematocrit), red blood cell production, or the maturation and / or differentiation of erythroid progenitor cells (e.g., early or late stages (e.g., maturation and / or differentiation of early (terminal) stage erythroid progenitors into proerythroblasts, reticulocytes, or erythrocytes), late precursor (erythroid precursor) maturation (e.g., terminal maturation such as maturation of reticulocytes into erythrocytes or maturation of erythroblasts into reticulocytes and / or erythrocytes), mobilization of early stage progenitors into the erythroid lineage, by decreasing the accumulation of erythroid progenitors (e.g., by stimulating progenitors to progress to maturation), by increasing the number of early stage erythroid progenitors and / or precursors (e.g., expanding the early stage progenitor and / or precursor population to replenish polychromatic erythroblasts,methods for treating or preventing low red blood cell levels (e.g., anemia or blood loss) by increasing platelet levels (e.g., platelet count, megakaryocyte differentiation and / or maturation, and / or platelet production) or by decreasing platelet precursor cell accumulation (e.g., by stimulating precursor cells to promote maturation); methods for treating or preventing low platelet levels (e.g., thrombocytopenia) by increasing neutrophil levels (e.g., , methods for treating or preventing low neutrophil levels (e.g., neutropenia) by increasing neutrophil numbers (e.g., neutrophil production) or the differentiation and / or maturation of precursor cells (e.g., bone marrow progenitor cells, myeloblasts, or myeloid cells) into neutrophils; methods for treating or preventing pulmonary hypertension (PH) (e.g., PAH, venous PH, hypoxic PH, thromboembolic PH, or other PH); methods for treating or preventing metabolic disease (e.g., obesity, type 1 diabetes, or type 2 diabetes); or methods for affecting myostatin, activin A, activin B, and / or BMP9 signaling in a subject.

[0421] I. Extracellular activin receptor type II chimeras Activin type II receptor is a single transmembrane domain receptor that regulates the signal of ligands of the transforming growth factor beta (TGF-β) superfamily. Ligands of the TGF-β superfamily are involved in many physiological processes, such as muscle growth, vascular growth, cell differentiation, homeostasis, hematopoiesis, and osteogenesis. Examples of ligands of the TGF-β superfamily include activins (e.g., activin A and activin B), inhibins, growth differentiation factors (GDFs) (e.g., GDF8, also known as myostatin, and GDF11), and bone morphogenetic proteins (BMPs) (e.g., BMP9).

[0422] Myostatin and activin are known to be involved in regulating skeletal muscle growth. For example, myostatin-null mice exhibit a significant increase in skeletal muscle mass. Myostatin is also involved in promoting fibrosis. Mice lacking myostatin exhibit reduced muscle fibrosis, and injection of myostatin-coated beads induces muscle fibrosis in mice. Mice overexpressing activin subunits, which result in the production of diffusible activin A, also exhibit fibrosis. Furthermore, activin is abundantly expressed in bone tissue and regulates bone formation by controlling the function of both osteoblasts and osteoclasts. Activin A has been reported to be upregulated in bone diseases and inhibit osteoblast activity. Myostatin is also involved in bone homeostasis through increased bone formation and inhibition of osteoblast activity. The TGF-β signaling pathway also regulates hematopoiesis. Activin-mediated signaling prevents the differentiation of erythroid, platelet, and neutrophil progenitors, maintaining progenitor cells in a quiescent state, while BMP-mediated signaling promotes progenitor differentiation. Homeostasis of this process is essential to ensure the proper replenishment of all cell types, including erythrocytes, leukocytes, and platelets, in the blood. Relatedly, the activin receptor ligand GDF11 has been found to be overexpressed in mouse models of hemolytic anemia and associated with defective erythropoiesis. Elevated activin A levels have also been observed in clinical and experimental pulmonary hypertension. Furthermore, activin is highly expressed in adipose tissue, and increased levels of myostatin and activin receptors have been observed in the subcutaneous and visceral fat of obese mice. Furthermore, myostatin has been shown to be elevated in skeletal muscle and plasma of obese and insulin-resistant women, and both type I and type II activin receptors have been linked to pancreatic function and diabetes. These data suggest that increased signaling through activin receptors, either through increased expression of activin receptor ligands (e.g., activin A, activin B, myostatin) or increased expression of the activin receptors themselves, may contribute to a variety of diseases and conditions, including muscle atrophy or wasting, fibrosis, bone disease, anemia, thrombocytopenia, neutropenia, pulmonary hypertension, and metabolic disease.Thus, methods of reducing or inhibiting activin A, activin B, or myostatin signaling may be used in the treatment of diseases and conditions including muscle atrophy or weakness, fibrosis, bone damage, low red blood cell levels (e.g., anemia), low platelet levels (e.g., thrombocytopenia), low neutrophil levels (e.g., neutropenia), pulmonary hypertension (e.g., PAH, venous PH, hypoxic PH, thromboembolic PH, or other PH), or metabolic disorders (e.g., obesity, type 1 diabetes, or type 2 diabetes).

[0423] There are two types of activin type II receptors: ActRIIA and ActRIIB. Studies have shown that BMP9 binds to ActRIIB with approximately 300-fold higher binding affinity than ActRIIA (see, for example, Townson et al., J. Biol. Chem. 287:27313, 2012). ActRIIA-Fc is known to have a longer half-life than ActRIIB-Fc. The present invention describes an extracellular ActRII chimera constructed by combining portions of extracellular ActRIIA and ActRIIB, with the aim of generating a protein that binds to ActRII ligands (e.g., activin A, activin B, myostatin, and GDF11) and retains the function of wild-type extracellular ActRII protein (e.g., the ability to increase muscle mass and / or lean body mass or red blood cell levels). In some embodiments, the ActRII chimera exhibits reduced BMP9 binding compared to wild-type extracellular ActRIIB and can prevent or reduce the disruption of endogenous BMP9 signaling. In some embodiments, the chimera possesses properties of both ActRIIA (e.g., lower binding affinity for BMP9, ability to increase red blood cell levels, and / or longer serum half-life as an Fc fusion protein) and ActRIIB (e.g., ability to increase muscle mass).In one example, the ActRII chimera has reduced binding affinity for BMP9 compared to wild-type extracellular ActRIIB and is used to promote increased lean body mass, muscle mass, bone mineral density, and / or red blood cell levels (e.g., increased red blood cell production and / or red blood cell mass or volume), loss of body weight and / or body fat, and / or a reduction in muscle disease (e.g., a neuromuscular disease such as muscular dystrophy, IBM, SMA, CMT, ALS, myasthenia gravis, or multiple sclerosis; sarcopenia; or cachexia), bone disease (e.g., a disease or condition associated with bone damage, e.g., resulting in the treatment of osteoporosis, osteopenia, osteopetrosis, fractures, bone loss associated with bone cancer or cancer metastasis, Paget's disease, renal osteodystrophy, treatment-related bone loss, osteogenesis imperfecta, bone loss associated with neuromuscular disease, bone loss due to burns, bone loss associated with anorexia, diet-related bone loss, bone loss associated with the treatment of obesity, bone loss associated with hypogravity, or bone loss associated with immobility), anemia, thrombocytopenia, neutropenia, metabolic disease (e.g., obesity, type 1 diabetes, or type 2 diabetes), fibrosis, or PH.

[0424] The ActRII chimeras exhibit similar or improved binding to activin (e.g., activin A and / or activin B) and / or myostatin compared to wild-type extracellular ActRIIA and / or ActRIIB, and are made to compete with endogenous activin receptors for ligand binding, thereby reducing or inhibiting endogenous activin receptor signaling.As a result, the chimeras can be used to treat disorders in which activin receptor signaling is elevated, such as bone disease, muscle disease, fibrosis, PH, metabolic disease, thrombocytopenia, neutropenia, and / or anemia, and can result in decreased bone resorption or osteoclast activity, increased bone formation or bone density, increased muscle mass, lean body mass, or strength, decreased fibrosis (e.g., decreased fibrosis or slowed or stopped the progression of fibrosis), increased red blood cell levels (e.g., increased hemoglobin levels, hematocrit, or red blood cell count, e.g., increased erythrocyte count, e ... increased red blood cell production and / or red blood cell mass or volume), increased maturation and / or differentiation of erythroid progenitors (e.g., maturation and / or differentiation of early or late (e.g., terminal) stage erythroid progenitors, e.g., maturation and / or differentiation of early stage erythroid progenitors, e.g., colony forming unit-erythroid (CFU-E) and burst forming unit-erythroid (BFU-E) proerythroblasts, reticulocytes, or erythrocytes), mobilization of early stage progenitors into the erythroid lineage, increased maturation of late stage erythroid progenitors Increased maturation (e.g., maturation of reticulocytes into erythrocytes, or final maturation such as maturation of erythroblasts into reticulocytes and / or erythrocytes), decreased accumulation of erythroid progenitors (e.g., by stimulating progenitors to proceed to maturation), increased numbers of early stage erythroid progenitors and / or precursors (e.g., expansion of early stage progenitor and / or precursor cell populations), progression of erythroid progenitors and / or precursors through erythropoiesis (e.g., through the erythropoietic pathway), increased platelet levels (e.g., platelet count, megakaryocyte differentiation and and / or maturation, and / or increased platelet production), decreased accumulation of platelet precursor cells (e.g., by stimulating precursor cells to progress to maturation), increased neutrophil levels (e.g., increased neutrophil count, e.g., increased neutrophil production), increased differentiation and / or maturation of precursor cells (e.g., myeloid precursor cells, myeloblasts, or myeloid cells) into neutrophils, decreased symptoms or progression of PH, or decreased body fat, weight, blood glucose levels, or insulin resistance (e.g., resulting in increased insulin sensitivity).

[0425] The wild-type amino acid sequences of the extracellular portions of human ActRIIA and ActRIIB are shown below. Human ActRIIA, extracellular portion (SEQ ID NO: 44): GAILGRSETQECLFFNANWEKDRTNQTGVEPCYGDKDKRRHCFATWKNISGSIEIVKQGCWLDDINCYDRTDCVEKKDSPEVYFCCCEGNMCNEKFSYFPEMEVTQPTS Human ActRIIB, extracellular portion (SEQ ID NO: 45): GRGEAETRECIYYNANWELERTNQSGLERCEGEQDKRLHCYASWRNSSGTIELVKKGCWLDDFNCYDRQECVATEENPQVYFCCCEGNFCNERFTHLPEAGGPEVTYEPPPTAPT The polypeptides described herein include extracellular ActRII chimeras containing sequences from both the extracellular portion of ActRIIB and the extracellular portion of ActRIIA. The ActRII chimeras described herein result from linking the N-terminal portion of extracellular ActRIIB (SEQ ID NO: 45 shown above) to the C-terminal portion of extracellular ActRIIA (SEQ ID NO: 44 shown above) so that the sequences are contiguous (e.g., the ActRIIA sequence begins after the amino acid at the corresponding position in ActRIIA and continues where the ActRIIB sequence left off). In some embodiments, the N-terminus of the ActRII chimera comprises the six amino acids found at the N-terminus of extracellular ActRIIA linked to the fifth amino acid of extracellular ActRIIB. In some embodiments, the N-terminus of the ActRII chimera begins with the first amino acid located at the N-terminus of extracellular ActRIIB. Thus, in some embodiments, the N-terminal portion of ActRIIB begins with the amino acid (A) at the fifth position of SEQ ID NO: 45, while in other embodiments (e.g., embodiments in which the chimera does not include the six amino acids found at the N-terminus of extracellular ActRIIA), the N-terminal portion of ActRIIB begins with the amino acid (G) at the first position of SEQ ID NO: 45. In some embodiments, the N-terminus of the ActRII chimera comprises the first 10 amino acids found at the N-terminus of extracellular ActRIIA linked to the ninth amino acid of extracellular ActRIIB, in which case the N-terminal portion of ActRIIB begins with the ninth amino acid (E) of SEQ ID NO: 45. The extracellular ActRII chimera may also include one or more amino acid substitutions in the portion of the chimera corresponding to the sequence of ActRIIB compared to the sequence of wild-type extracellular ActRIIB (e.g., SEQ ID NO: 45, above), and one or more amino acid substitutions in the portion of the chimera corresponding to the sequence of ActRIIA compared to the sequence of wild-type extracellular ActRIIA (e.g., SEQ ID NO: 44, above). Amino acid substitutions at nine different positions can be introduced into the extracellular ActRII chimera (Table 1).The extracellular ActRII chimera may have one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, or 9) amino acid substitutions relative to the wild-type sequence (e.g., relative to the sequence of wild-type extracellular ActRIIB (sequence number 45) if the portion of the chimera corresponds to a region of wild-type extracellular ActRIIB, or relative to the sequence of wild-type extracellular ActRIIA (sequence number 44) if the portion of the chimera corresponds to a region of wild-type extracellular ActRIIA). The positions at which amino acid substitutions can be made and the amino acids that can be substituted at these positions are listed in Table 1.

[0426] Amino acid substitutions can alter the activity and / or binding affinity of the extracellular ActRII chimera of the invention. In some embodiments, the extracellular ActRII chimera binds to activin A, activin B, myostatin, and / or GDF11 with sufficient affinity to compete with endogenous activin receptors for binding to one or more of these ligands. In some embodiments, the extracellular ActRII chimera of the invention exhibits reduced, weak, or substantially no binding to BMP9 (e.g., compared to wild-type ActRII). BMP9 binding can be reduced in extracellular ActRII chimeras comprising the amino acid sequence TEEN or TKEN at positions X3, X4, X5, and X6. In some embodiments, a polypeptide of the invention comprising an extracellular ActRII chimera comprising the sequence TEEN at positions X3, X4, X5, and X6 (e.g., any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)) can have the amino acid E at position X4 substituted with the amino acid K. In some embodiments, a polypeptide of the present invention comprising an extracellular ActRII chimera (e.g., any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)) comprising the sequence TKEN at positions X3, X4, X5, and X6 can have the amino acid K at position X4 substituted with the amino acid E. The sequences TEEN and TKEN can be used interchangeably in the extracellular ActRII chimeras of the present invention (e.g., the chimeras of Tables 1 and 2, e.g., SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)). The extracellular ActRII chimera of the present invention can further comprise a C-terminal extension (e.g., additional amino acids at the C-terminus). The C-terminal extension can add one or more additional amino acids (e.g., 1, 2, 3, 4, 5, 6, or more additional amino acids) to the C-terminus of any of the chimeras shown in Tables 1 and 2 (e.g., SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)). The C-terminal extension can correspond to the sequence from the same position in wild-type ActRIIA or ActRIIB. For example, C-terminal extensions that can be included in the extracellular ActRII chimeras of the present invention are the amino acid sequence NP and the amino acid sequence NPVTPK (sequence number 104), which correspond to sequences found at the same positions in wild-type ActRIIA.

[0427] [Table 1-1]

[0428] [Table 1-2]

[0429] [Table 1-3]

[0430] In some embodiments, in the extracellular ActRII chimeras of SEQ ID NOs: 1 to 21 (shown in Table 1), X1 is D, X2 is I, F, or E, X3 is N or T, X4 is A or E, X5 is T or K, X6 is E or K, X7 is E or D, X8 is N or S, and X9 is E or Q. In some embodiments, in the extracellular ActRII chimeras of SEQ ID NOs: 1 to 21, X1 is D, X2 is I or F, X3 is N, X4 is A or E, X5 is T or K, X6 is E or K, X7 is E or D, X8 is N or S, and X9 is E or Q.

[0431] In some embodiments, the polypeptides described herein comprise an extracellular ActRII chimera having the sequence of any one of SEQ ID NOs: 22-43 (Table 2).

[0432] [Table 2-1]

[0433] [Table 2-2]

[0434] [Table 2-3]

[0435] [Table 2-4]

[0436] In some embodiments, the extracellular ActRII chimeras described herein have an N-terminal truncation of 1 to 9 amino acids (e.g., 1, 2, 3, 4, 5, 6, 7, 8, or 9 amino acids). N-terminal truncations can include the removal of 1 to 9 amino acids from the N-terminus of any of the chimeras shown in Tables 1 and 2 (e.g., SEQ ID NOs: 1-43 (e.g., SEQ ID NOs: 22-43)). N-terminal truncations can remove up to two amino acids before the first cysteine ​​(e.g., the two amino acids (RE or QE) before the first cysteine ​​are retained in the N-terminally truncated ActRII chimera). Exemplary ActRII chimeras with N-terminal truncations are presented in Table 3.

[0437] [Table 3-1]

[0438] [Table 3-2]

[0439] [Table 3-3]

[0440] [Table 3-4]

[0441] [Table 3-5]

[0442] [Table 3-6]

[0443] Table 3-7

[0444] Table 3-8

[0445] Table 3-9

[0446] Table 3-10

[0447] Table 3-11

[0448] Table 3-12

[0449] In some embodiments, a polypeptide of the present invention comprising an extracellular ActRII chimera may further comprise a moiety (e.g., an Fc domain monomer, a wild-type Fc domain, an Fc domain comprising amino acid substitutions (e.g., one or more substitutions that reduce dimerization), an albumin-binding peptide, a fibronectin domain, or human serum albumin) that can be fused to the N-terminus or C-terminus (e.g., the C-terminus) of the extracellular ActRII chimera via a linker or other covalent bond. A polypeptide comprising an extracellular ActRII chimera fused to an Fc domain monomer can form a dimer (e.g., a homodimer or heterodimer) through interaction between two Fc domain monomers that combine to form a dimeric Fc domain. Exemplary polypeptides comprising an ActRII chimera, an Fc domain, and a linker are presented in Table 4 below. In some embodiments, the terminal lysine is not found in the Fc domain amino acid sequence.

[0450] [Table 4-1]

[0451] [Table 4-2]

[0452] [Table 4-3]

[0453] [Table 4-4]

[0454] [Table 4-5]

[0455] [Table 4-6]

[0456] Table 4-7

[0457] Table 4-8

[0458] Table 4-9

[0459] Table 4-10

[0460] Table 4-11

[0461] Table 4-12

[0462] Table 4-13

[0463] Table 4-14

[0464] Table 4-15

[0465] Table 4-16

[0466] Table 4-17

[0467] Table 4-18

[0468] Table 4-19

[0469] Table 4-20

[0470] Table 4-21

[0471] Table 4-22

[0472] Table 4-23

[0473] Table 4-24

[0474] Table 4-25

[0475] Table 4-26

[0476] Table 4-27

[0477] Table 4-28

[0478] Table 4-29

[0479] Table 4-30

[0480] Table 4-31

[0481] Table 4-32

[0482] Table 4-33

[0483] Table 4-34

[0484] Table 4-35

[0485] Table 4-36

[0486] Table 4-37

[0487] [Table 4-38]

[0488] [Table 4-39]

[0489] [Table 4-40]

[0490] [Table 4-41]

[0491] [Table 4-42]

[0492] Furthermore, in some embodiments, the polypeptides described herein (e.g., ActRII chimeric-Fc fusion proteins) have a serum half-life in humans of at least 7 days. The polypeptides have a K D In some embodiments, the polypeptide may bind to activin A at positions X3, X4, X5, and X6. ...

[0493] In some embodiments, the polypeptide has a K of about 800 pM or less. D (e.g., a K of about 800, 700, 600, 500, 400, 300, 200, 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 pM or less) D , for example, a K of about 800 pM to about 30 pM D In some embodiments, the polypeptide may bind to human activin A with a K of 800 pM or less. D (e.g., a K of about 800, 700, 600, 500, 400, 300, 200, 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 pM or less) D , for example, a K of about 800 pM to about 5 pM D The polypeptide may also bind to human activin B with a K of about 5 pM or greater. D (e.g., a K of about 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190, 195, or 200 pM or more) D ) and can bind to growth and differentiation factor 11 (GDF-11).

[0494] II. Fc Domain In some embodiments, the polypeptides described herein may comprise an extracellular ActRII chimera fused to an Fc domain monomer or a fragment of an Fc domain of an immunoglobulin to increase the serum half-life of the polypeptide. A polypeptide comprising an extracellular ActRII chimera fused to an Fc domain monomer may form a dimer (e.g., a homodimer or a heterodimer) through the interaction between two Fc domain monomers forming a dimeric Fc domain. As conventionally known in the art, an Fc domain is a protein structure found at the C-terminus of an immunoglobulin. An Fc domain is a C HThe Fc domain comprises two Fc domain monomers that are dimerized by interactions between three antibody constant domains. Wild-type Fc domains form the minimal structure that binds to Fc receptors, e.g., FcγRI, FcγRIIa, FcγRIIb, FcγRIIIa, FcγRIIIb, and FcγRIV. In some embodiments, the Fc domain can be mutated to lack effector function, typical of a "dead" Fc domain. For example, the Fc domain can contain specific amino acid substitutions known to minimize interactions between the Fc domain and Fcγ receptors. In some embodiments, the Fc domain is derived from an IgG1 antibody and contains amino acid substitutions L234A, L235A, and G237A. In some embodiments, the Fc domain is derived from an IgG1 antibody and contains amino acid substitutions D265A, K322A, and N434A. The above-mentioned amino acid positions are defined according to Kabat (Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991)). The Kabat numbering of amino acid residues can be determined for a given antibody by aligning the homology regions of the antibody's sequence with the "standard" Kabat numbering sequence. Furthermore, in some embodiments, the Fc domain does not induce any immune system-related response. For example, the Fc domain in a polypeptide dimer comprising an extracellular ActRII chimera fused to an Fc domain monomer can be modified to reduce the interaction or binding between the Fc domain and the Fcγ receptor. The sequence of an Fc domain monomer that can be fused to an extracellular ActRII chimera is shown below (SEQ ID NO: 48): THTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPVPIEK TISKAKGQPREPQVYTLPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGPFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK In some embodiments, the Fc domain is derived from an IgG1 antibody and includes amino acid substitutions L12A, L13A, and G15A relative to the sequence of SEQ ID NO: 48. In some embodiments, the Fc domain is derived from an IgG1 antibody and includes amino acid substitutions D43A, K100A, and N212A relative to the sequence of SEQ ID NO: 48. In some embodiments, the terminal lysine is absent in an Fc domain monomer having the sequence of SEQ ID NO: 48. In some embodiments, an extracellular ActRII chimera described herein (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOs: 1-43 (e.g., SEQ ID NOs: 22-43)) can be fused to the N- or C-terminus of an Fc domain monomer (e.g., SEQ ID NO: 48) via conventional genetic or chemical means, e.g., chemical conjugation. If necessary, a linker (e.g., a spacer) can be inserted between the extracellular ActRII chimera and the Fc domain monomer. The Fc domain monomer can be fused to the N-terminus or C-terminus (e.g., the C-terminus) of the extracellular ActRII chimera.

[0495] In some embodiments, the polypeptides described herein may comprise an extracellular ActRII chimera fused to an Fc domain. In some embodiments, the Fc domain comprises one or more amino acid substitutions that reduce or inhibit Fc domain dimerization. In some embodiments, the Fc domain comprises a hinge domain. The Fc domain may be any immunoglobulin antibody isotype, IgG, IgE, IgM, IgA, or IgD. Furthermore, the Fc domain may be an IgG subtype (e.g., IgG1, IgG2a, IgG2b, IgG3, or IgG4). The Fc domain may also be a non-naturally occurring Fc domain, such as a recombinant Fc domain.

[0496] Methods for engineering Fc domains with reduced dimerization are known in the art. In some embodiments, one or more amino acids with large side chains (e.g., tyrosine or tryptophan) are replaced with C H 3-C H 3 dimer interface to prevent dimer formation through steric clashes. In other embodiments, one or more amino acids with small side chains (e.g., alanine, valine, or threonine) may be introduced into the C3 dimer interface to remove favorable interactions. H 3-C H 3 can be introduced into the dimer interface. H Methods for introducing amino acids with large or small side chains into the three domains are described, for example, in Ying et al. (J Biol Chem. 287:19399-19408, 2012), U.S. Patent Application Publication No. 2006 / 0074225, U.S. Patent Nos. 8,216,805 and 5,731,168, Ridgway et al. (Protein Eng. 9:617-612, 1996), Atwell et al. (J Mol Biol. 270:26-35, 1997), and Merchant et al. (Nat Biotechnol. 16:677-681, 1998), all of which are incorporated herein by reference in their entireties.

[0497] In yet another embodiment, the C between two Fc domainsH 3-C H 3C that constitutes the interface H One or more amino acid residues within the three domains are substituted with positively charged amino acid residues (e.g., lysine, arginine, or histidine) or negatively charged amino acid residues (e.g., aspartic acid or glutamic acid) to make the interaction electrostatically unfavorable, depending on the particular charged amino acid introduced. H Methods of introducing charged amino acids into the three domains to dislike or prevent dimer formation are described, for example, in Ying et al. (J Biol Chem. 287:19399-19408, 2012), U.S. Patent Application Publication Nos. 2006 / 0074225, 2012 / 0244578, and 2014 / 0024111, all of which are incorporated herein by reference in their entireties.

[0498] In some embodiments of the invention, the Fc domain comprises one or more of the following amino acid substitutions relative to the sequence of human IgG1: T366W, T366Y, T394W, F405W, Y349T, Y349E, Y349V, L351T, L351H, L351N, L352K, P353S, S354D, D356K, D356R, D356S, E357K, E357R, E357Q, S 364A, T366E, L368T, L368Y, L368E, K370E, K370D, K370Q, K392E, K392D, T394N, P395N, P396T, V397T, V397Q, L398T, D399K, D399R, D399N, F405T, F405H, F405R, Y407T, Y407H, Y407I, K409E, K409D, K409T, and K409I. In some embodiments, the terminal lysine is absent from the Fc domain amino acid sequence. In one particular embodiment, the Fc domain comprises the amino acid substitution T366W relative to the sequence of human IgG1. The sequence of the wild-type Fc domain is shown below in SEQ ID NO: 100.

[0499] DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIE KTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK An exemplary sequence of a wild-type Fc domain lacking the terminal lysine is provided below (SEQ ID NO: 264).

[0500] DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIE KTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG III. Albumin-binding peptides In some embodiments, the polypeptides described herein may comprise an extracellular ActRII chimera fused to a serum protein-binding peptide. Binding to the serum protein-binding peptide can improve the pharmacokinetics of protein pharmaceuticals.

[0501] By way of example, albumin-binding peptides that can be used in the methods and compositions described herein are commonly known in the art. In one embodiment, the albumin-binding peptide comprises the sequence DICLPRWGCLW (SEQ ID NO: 101).

[0502] In the present invention, an albumin-binding peptide can be attached to the N-terminus or C-terminus (e.g., the C-terminus) of an extracellular ActRII chimera described herein (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOs: 1 to 43 (e.g., SEQ ID NOs: 22 to 43)) to increase the serum half-life of the extracellular ActRII chimera. In some embodiments, the albumin-binding peptide is attached to the N-terminus or C-terminus of the extracellular ActRII chimera directly or via a linker.

[0503] In some embodiments, the extracellular ActRII chimera described herein (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)) can be fused to the N- or C-terminus of an albumin-binding peptide (e.g., SEQ ID NOS: 101) via conventional genetic or chemical means, e.g., chemical conjugation. If necessary, a linker (e.g., a spacer) can be inserted between the extracellular ActRII chimera and the albumin-binding peptide. Without wishing to be bound by theory, it is expected that the inclusion of an albumin-binding peptide in the extracellular ActRII chimera described herein can prolong the retention of therapeutic proteins through binding to serum albumin.

[0504] IV. Fibronectin Domains In some embodiments, the polypeptides described herein may comprise an extracellular ActRII chimera fused to a fibronectin domain, which can improve the pharmacokinetics of protein pharmaceuticals.

[0505] A fibronectin domain is a high-molecular-weight glycoprotein or fragment thereof of the extracellular matrix that binds to transmembrane receptor proteins such as integrins and extracellular matrix components such as collagen and fibrin. In some embodiments of the present invention, a fibronectin domain can be attached to the N-terminus or C-terminus (e.g., the C-terminus) of an extracellular ActRII chimera described herein (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)) to increase the serum half-life of the extracellular ActRII chimera. The fibronectin domain can be attached to the N-terminus or C-terminus of the extracellular ActRII chimera directly or via a linker.

[0506] By way of example, fibronectin domains that can be used in the methods and compositions described herein are commonly known in the art. In one embodiment, the fibronectin domain is a fibronectin type III domain having amino acids 610-702 of UniProt ID number P02751 (SEQ ID NO: 102 shown below): GPVEVFITETPSQPNSHPIQWNAPQPSHISKYILRWRPKNSVGRWKEATIPGHLNSYTIKGLKPGVVYEGQLISIQQYGHQEVTRFDFTTTST In another embodiment, the fibronectin domain is an adnectin protein.

[0507] In some embodiments, the extracellular ActRII chimera described herein (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)) can be fused to the N- or C-terminus of a fibronectin domain (e.g., SEQ ID NOS: 102) via conventional genetic or chemical means, e.g., chemical conjugation. If necessary, a linker (e.g., a spacer) can be inserted between the extracellular ActRII chimera and the fibronectin domain. Without wishing to be bound by theory, it is expected that the inclusion of a fibronectin domain in the extracellular ActRII chimera described herein can prolong the retention of therapeutic proteins through binding to integrins and extracellular matrix components such as collagen and fibrin.

[0508] V. Serum Albumin In some embodiments, the polypeptides described herein may comprise an extracellular ActRII chimera fused to serum albumin. Binding to serum albumin can improve the pharmacokinetics of protein pharmaceuticals.

[0509] Serum albumin is a globular protein that is the most abundant blood protein in mammals. It is produced in the liver and constitutes approximately half of serum proteins. It is monomeric and soluble in blood. The most important functions of serum albumin include transporting hormones, fatty acids, and other proteins in the body, buffering pH, and maintaining the osmotic pressure necessary for proper distribution of body fluids between blood vessels and body tissues. In a preferred embodiment, the serum albumin is human serum albumin. In some embodiments of the present invention, human serum albumin can be attached to the N- or C-terminus (e.g., the C-terminus) of the extracellular ActRII chimera described herein (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)) to increase the serum half-life of the extracellular ActRII chimera. Human serum albumin can be attached to the N- or C-terminus of the extracellular ActRII chimera directly or via a linker.

[0510] By way of example, human serum albumin that can be used in the methods and compositions described herein is commonly known in the art. In one embodiment, the serum albumin is selected from the group consisting of the UniProt It contains the sequence of ID number P02768 (SEQ ID NO: 103, shown below).

[0511] MKWVTFISLLFLFSSAYSRGVFRRDAHKSEVAHRFKDLGEENFKALVLIAFAQYLQQCPFEDHVKLVNEVTEFAKTCVADESAENCDKSLHTLFGDKLCTVATLRETYGEMADCCAKQEPERNECFLQHKDDNPNLPRLVRPEVDVMCTAFH DNEETFLKKYLYEIARRHPYFYAPELLFFAKRYKAAFTECCQAADKAACLLPKLDELRDEGKASSAKQRLKCASLQKFGERAFKAWAVARLSQRFPKAEFAEVSKLVTDLTKVHTECCHGDLLECADDRADLAKYICENQDSISSKLKECCE KPLLEKSHCIAEVENDEMPADLPSLAADFVESKDVCKNYEAKDVFLGMFLYEYARRHPDYSVVLLLRLAKTYETTLEKCCAAADPHECYAKVFDEFKPLVEEPQNLIKQNCELFEQLGEYKFQNALLVRYTKKVPQVSTPTLVEVSRNLGK VGSKCCKHPEAKRMPCAEDYLSVVLNQLCVLHEKTPVSDRVTKCCTESLVNRRPCFSALEVDETYVPKEFNAETFTFHADICTLSEKERQIKKQTALVELVKHKPKATKEQLKAVMDDFAAFVEKCCKADDKETCFAEEGKKLVAASQAALGL In some embodiments, the extracellular ActRII chimera described herein (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)) can be fused to the N-terminus or C-terminus of human serum albumin (e.g., SEQ ID NOS: 103) via conventional genetic or chemical means, e.g., chemical conjugation. If necessary, a linker (e.g., a spacer) can be inserted between the extracellular ActRII chimera and human serum albumin. Without wishing to be bound by theory, it is expected that the inclusion of human serum albumin in the extracellular ActRII chimera described herein can prolong the retention of therapeutic proteins.

[0512] VI. Linker The polypeptides described herein may include an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)) fused to a moiety by a linker. In some embodiments, the moiety increases the stability of the polypeptide. Exemplary moieties include an Fc domain monomer, a wild-type Fc domain, an Fc domain containing amino acid substitutions (e.g., one or more substitutions that reduce dimerization), an albumin-binding peptide, a fibronectin domain, or human serum albumin. In the present invention, the linker between the moiety (e.g., an Fc domain monomer (e.g., the sequence of SEQ ID NO: 48), a wild-type Fc domain (e.g., the sequence of SEQ ID NO: 100, or the sequence of SEQ ID NO: 264), an Fc domain containing amino acid substitutions (e.g., one or more substitutions that reduce dimerization), an albumin-binding peptide (e.g., the sequence of SEQ ID NO: 101), a fibronectin domain (e.g., the sequence of SEQ ID NO: 102), or human serum albumin (e.g., the sequence of SEQ ID NO: 103)) and the extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOs: 1 to 43 (e.g., SEQ ID NOs: 22 to 43)) can be an amino acid spacer containing 1 to 200 amino acids. Suitable peptide spacers are known in the art and include, for example, peptide linkers containing flexible amino acid residues such as glycine, alanine, and serine. In some embodiments, the spacer may include a GA, GS, GG, GGA, GGS, GGG, GGGA (SEQ ID NO: 49), GGGS (SEQ ID NO: 50), GGGG (SEQ ID NO: 51), GGGGA (SEQ ID NO: 52), GGGGS (SEQ ID NO: 53), GGGGG (SEQ ID NO: 54), GGAG (SEQ ID NO: 55), GGSG (SEQ ID NO: 56), AGGG (SEQ ID NO: 57), or SGGG (SEQ ID NO: 58) motif, e.g., multiple or repeated motifs.In some embodiments, the spacer can include 2 to 12 amino acids containing a GA or GS motif, e.g., GA, GS, GAGA (SEQ ID NO: 59), GSGS (SEQ ID NO: 60), GAGAGA (SEQ ID NO: 61), GSGSGS (SEQ ID NO: 62), GAGAGAGA (SEQ ID NO: 63), GSGSGSGS (SEQ ID NO: 64), GAGAGAGAGA (SEQ ID NO: 65), GSGSGSGSGS (SEQ ID NO: 66), GAGAGAGAGAGA (SEQ ID NO: 67), and GSGSGSGSGSGS (SEQ ID NO: 68). In some embodiments, the spacer can include 3 to 12 amino acids containing a GGA or GGS motif, e.g., GGA, GGS, GGAGGA (SEQ ID NO: 69), GGSGGS (SEQ ID NO: 70), GGAGGAGGA (SEQ ID NO: 71), GGSGGSGGS (SEQ ID NO: 72), GGAGGAGGAGGA (SEQ ID NO: 73), and GGSGGSGGSGGS (SEQ ID NO: 74). In further embodiments, the spacer can comprise 4 to 12 amino acids, including a GGAG (SEQ ID NO: 55), GGSG (SEQ ID NO: 56), e.g., GGAG (SEQ ID NO: 55), GGSG (SEQ ID NO: 56), GGAGGGAG (SEQ ID NO: 75), GGSGGGSG (SEQ ID NO: 76), GGAGGGAGGGAG (SEQ ID NO: 77), and GGSGGGSGGGSG (SEQ ID NO: 78). In some embodiments, the spacer can comprise a GGGGA (SEQ ID NO: 52), or GGGGS (SEQ ID NO: 53), e.g., GGGGAGGGGAGGGGA (SEQ ID NO: 79) and GGGGSGGGGSGGGGGS (SEQ ID NO: 80) motif. In some embodiments of the present invention, the amino acid spacer between the moiety (e.g., an Fc domain monomer, a wild-type Fc domain, an Fc domain containing amino acid substitutions (e.g., one or more substitutions that reduce dimerization), an albumin-binding peptide, a fibronectin domain, or human serum albumin) and the extracellular ActRII chimera (e.g., an extracellular ActRII chimera having any one of the sequences of SEQ ID NOs: 1 to 43 (e.g., SEQ ID NOs: 22 to 43)) may be GGG, GGGA (SEQ ID NO: 49), GGGG (SEQ ID NO: 51), GGGAG (SEQ ID NO: 81), GGGAGG (SEQ ID NO: 82), or GGGAGGG (SEQ ID NO: 83).

[0513] In some embodiments, the spacer can also include amino acids other than glycine, alanine, and serine, such as AAAL (SEQ ID NO: 84), AAAK (SEQ ID NO: 85), AAAR (SEQ ID NO: 86), EGKSSGSGSESKST (SEQ ID NO: 87), GSAGSAAGSGEF (SEQ ID NO: 88), AEAAAKEAAAKA (SEQ ID NO: 89), KESGSVSSEQLAQFRSLD (SEQ ID NO: 90), GENLYFQSGG (SEQ ID NO: 91), SACYCELS (SEQ ID NO: 92), RSIAT (SEQ ID NO: 93), RPACKIPNDLKQKVMNH (SEQ ID NO: 94), GGSAGGSGSGSSGGSSGASGTGTAGGTGSGSGTGSG (SEQ ID NO: 95), AAANSSIDLISVPVDSR (SEQ ID NO: 96), or GGSGGGSEGGGSEGGGSEGGGSEGGGSEGGGSGGGS (SEQ ID NO: 97). In some embodiments, the spacer can include a motif, such as multiple or repeated motifs of EAAAK (SEQ ID NO: 98). In some embodiments, the spacer can be (XP) n wherein X can be any amino acid (e.g., A, K, or E) and n is 1 to 5), and motifs such as multiple or repeated motifs of PAPAP (SEQ ID NO: 99).

[0514] The length of the peptide spacer and the amino acids used can be adjusted depending on the two proteins involved and the degree of flexibility desired in the final protein fusion polypeptide. The length of the spacer can be adjusted to ensure proper protein folding and avoid aggregate formation.

[0515] VII. Vectors, Host Cells, and Protein Production The polypeptides of the present invention can be produced from host cells. A host cell refers to a vehicle containing the cellular components, e.g., organelles, necessary to express the polypeptides and fusion polypeptides described herein from their corresponding nucleic acids. The nucleic acid can be contained in a nucleic acid vector, which can be introduced into a host cell by conventional techniques known in the art (e.g., transformation, transfection, electroporation, calcium phosphate precipitation, direct microinjection, infection, etc.). The choice of nucleic acid vector depends in part on the host cell to be used. Generally, preferred host cells are of either eukaryotic (e.g., mammalian) or prokaryotic (e.g., bacterial) origin.

[0516] Nucleic Acid Vector Constructs and Host Cells Nucleic acid sequences encoding the amino acid sequences of the polypeptides of the present invention can be prepared by various methods known in the art. These methods include, but are not limited to, oligonucleotide-mediated (or site-directed) mutagenesis, PCR mutagenesis, ligation, and overlap extension PCR. Nucleic acid molecules encoding the polypeptides of the present invention can be obtained using standard techniques, such as gene synthesis. Alternatively, nucleic acid molecules encoding wild-type extracellular ActRIIA or extracellular ActRIIB can be mutated to contain specific amino acid substitutions using standard techniques in the art, such as QuikChange® mutagenesis. Nucleic acid molecules can be synthesized using a nucleotide synthesizer or PCR technology.

[0517] The nucleic acid sequence encoding the polypeptide of the present invention can be inserted into a vector capable of replicating and expressing the nucleic acid molecule in a prokaryotic or eukaryotic host cell. Many vectors are available in the art and can be used for the purposes of the present invention. Each vector can contain various components that can be adjusted and optimized for compatibility with a particular host cell. For example, vector components can include, but are not limited to, an origin of replication, a selectable marker gene, a promoter, a ribosome binding site, a signal sequence, a nucleic acid sequence encoding the protein of interest, and a transcription termination sequence.

[0518] In some embodiments, mammalian cells can be used as host cells of the present invention. Examples of mammalian cell types include, but are not limited to, human embryonic kidney (HEK) (e.g., HEK293, HEK293F), Chinese hamster ovary (CHO), HeLa, COS, PC3, Vero, MC3T3, NS0, Sp2 / 0, VERY, BHK, MDCK, W138, BT483, Hs578T, HTB2, BT20, T47D, NS0 (a mouse myeloma cell line that does not endogenously produce any immunoglobulin chains), CRL7O3O, and HsS78Bst cells. In some embodiments, E. coli cells can also be used as hosts of the present invention. Examples of E. coli strains include, but are not limited to, E. coli 294 (ATCC® 31,446), E. coli λ 1776 (ATCC® 31,537), E. coli BL21(DE3) (ATCC® BAA-1025), and E. coli RV308 (ATCC® 31,608). Different host cells have characteristic and specific mechanisms for post-translational processing and modification (e.g., glycosylation) of protein products. An appropriate cell line or host system can be selected to ensure correct modification and processing of the expressed polypeptide. The above-described expression vectors can be introduced into suitable host cells using conventional techniques in the art, such as transformation, transfection, electroporation, calcium phosphate precipitation, and direct microinjection. Once the vector has been introduced into the host cell for protein production, the host cell is cultured in conventional nutrient media modified as appropriate for inducing promoters, selecting transformants, or amplifying the gene encoding the desired sequence. Methods for expressing therapeutic proteins are known in the art and are described, for example, in Paulina Balbas, Argelia Lorence (eds.) Recombinant Gene Expression: Reviews and Protocols (Methods in Molecular Biology), Humana Press; 2nd ed. 2004 and Vladimir Voynov and See Justin A. Caravella (eds.) Therapeutic Proteins: Methods and Protocols (Methods in Molecular Biology) Humana Press; 2nd ed. 2012.

[0519] Protein production, recovery, and purification Host cells used to produce the polypeptides of the present invention can be grown in media known in the art and suitable for culturing the selected host cells. Examples of media suitable for mammalian host cells include Minimum Essential Medium (MEM), Dulbecco's Modified Eagle's Medium (DMEM), Expi293® Expression Medium, DMEM supplemented with fetal bovine serum (FBS), and RPMI-1640. Examples of media suitable for bacterial host cells include Luria Broth (LB) containing necessary supplements, including a selection agent such as ampicillin. Host cells are cultured at an appropriate temperature, such as about 20°C to about 39°C, e.g., 25°C to about 37°C, preferably 37°C, and at a CO2 level, e.g., 5-10%. The pH of the medium is generally about 6.8 to 7.4, e.g., 7.0, depending primarily on the host organism. When an inducible promoter is used in the expression vector of the present invention, protein expression is induced under conditions suitable for promoter activation.

[0520] In some embodiments, depending on the expression vector and host cells used, the expressed protein may be secreted from the host cells (e.g., mammalian host cells) into the cell culture medium. Protein recovery may include filtering the cell culture medium to remove cellular debris. The protein may be further purified. The polypeptides of the invention may be purified by any method known in the art of protein purification, for example, by chromatography (e.g., ion exchange, affinity, and size exclusion column chromatography), centrifugation, absorption gradient solubility, or any other standard technique for purifying proteins. For example, the protein may be isolated and purified by appropriately selecting and combining affinity columns, such as Protein A columns (e.g., POROS Protein A chromatography), and chromatography columns (e.g., POROS HS-50 cation exchange chromatography), filtration, ultrafiltration, salting out, and dialysis procedures.

[0521] In other embodiments, to recover the expressed protein, host cells can be disrupted, for example, by osmotic shock, sonication, or lysis. Once the cells are disrupted, cellular debris can be removed by centrifugation or filtration. In some cases, the polypeptide can be conjugated to a marker sequence, such as a peptide, to facilitate purification. An example of a marker amino acid sequence is a hexahistidine peptide (His tag), which binds to a nickel-functionalized agarose affinity column with micromolar affinity. Other peptide tags useful for purification include, but are not limited to, the hemagglutinin "HA" tag, which corresponds to an epitope derived from the influenza hemagglutinin protein (Wilson et al., Cell 37:767, 1984).

[0522] Alternatively, the polypeptides of the invention can be produced by cells of a subject (e.g., a human) by administering a vector (e.g., a viral vector (e.g., a retroviral vector, an adenoviral vector, a poxvirus vector (e.g., a vaccinia virus vector such as modified vaccinia Ankara (MVA), an adeno-associated virus vector, and an alphavirus vector)) comprising a nucleic acid molecule encoding a polypeptide of the invention, e.g., in the context of gene therapy. The vector promotes expression of the polypeptide once inside the subject's cells (e.g., by transformation, transfection, electroporation, calcium phosphate precipitation, direct microinjection, infection, etc.), which is then secreted from the cells. If treatment of a disease or disorder is the desired outcome, further treatment may not be necessary. If collection of protein is desired, blood can be collected from the subject and the protein can be purified from the blood by methods known in the art.

[0523] VIII. Pharmaceutical Compositions and Preparations The present invention features pharmaceutical compositions comprising a polypeptide described herein (e.g., an extracellular ActRII chimera (e.g., a polypeptide comprising an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)). In some embodiments, the pharmaceutical compositions of the present invention comprise an extracellular ActRII chimera (e.g., any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)) fused to a moiety (e.g., an Fc domain monomer or dimer thereof, a wild-type Fc domain, an Fc domain containing amino acid substitutions (e.g., one or more substitutions that reduce dimerization), an albumin-binding peptide, a fibronectin domain, or human serum albumin). The therapeutic protein includes a polypeptide comprising an extracellular ActRII chimera having one or more sequences of the polypeptide of the present invention. In some embodiments, the pharmaceutical composition of the present invention comprising the polypeptide of the present invention can be used in combination with other agents (e.g., therapeutic biologics and / or small molecules) or compositions in therapy. In addition to a therapeutically effective amount of the polypeptide, the pharmaceutical composition can include one or more pharmaceutically acceptable carriers or excipients, which can be formulated by methods known to those of skill in the art. In some embodiments, the pharmaceutical composition of the present invention comprises a nucleic acid molecule (DNA or RNA, e.g., mRNA) encoding the polypeptide of the present invention, or a vector comprising such a nucleic acid molecule.

[0524] Acceptable carriers and excipients in pharmaceutical compositions are non-toxic to recipients at the dosages and concentrations used. Acceptable carriers and excipients may include buffers such as phosphate, citrate, HEPES, and TAE; antioxidants such as ascorbic acid and methionine; preservatives such as hexamethonium chloride, octadecyldimethylbenzylammonium chloride, resorcinol, and benzalkonium chloride; proteins such as human serum albumin, gelatin, dextran, and immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, histidine, arginine, and lysine; and carbohydrates such as glucose, mannose, sucrose, and sorbitol. The pharmaceutical composition of the present invention can be administered parenterally in the form of an injectable formulation. The injectable pharmaceutical composition can be formulated using a sterile solution or any pharmaceutically acceptable liquid as a vehicle. Pharmaceutically acceptable vehicles include, but are not limited to, sterile water, saline, and cell culture media (e.g., Dulbecco's Modified Eagle's Medium (DMEM), α-Modified Eagle's Medium (α-MEM), F-12 medium). Preparation methods are known in the art. See, for example, Banga (ed.) Therapeutic Peptides and Proteins: Formulation, Processing and Delivery Systems (3rd ed.) Taylor & Francis Group, CRC Press (2015).

[0525] The pharmaceutical compositions of the present invention can be prepared in microcapsules, such as hydroxymethylcellulose or gelatin microcapsules and poly(methyl methacrylate) microcapsules. The pharmaceutical compositions of the present invention can also be formulated in other drug delivery systems, such as liposomes, albumin microspheres, microemulsions, nanoparticles, and nanocapsules. Such approaches are described in Remington: The Science and Practice of Pharmacy 22 ndPharmaceutical compositions to be used for in vivo administration must be sterile, which is readily accomplished by filtration through sterile filtration membranes.

[0526] The pharmaceutical compositions of the present invention can also be prepared as sustained-release formulations. Suitable examples of sustained-release formulations include semipermeable matrices of solid hydrophobic polymers containing the polypeptides of the present invention. Examples of sustained-release matrices include polyesters, hydrogels, polylactides, copolymers of L-glutamic acid and gamma-ethyl-L-glutamate, non-degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers such as LUPRON DEPOT™, and poly-D-(-)-3-hydroxybutyric acid. Some sustained-release formulations enable release of molecules over several months, e.g., 1 to 6 months, while other formulations release the pharmaceutical compositions of the present invention for shorter periods, e.g., days to weeks.

[0527] The pharmaceutical composition may be prepared in a unit dosage form as needed. The amount of the active ingredient, e.g., the polypeptide of the present invention, contained in the pharmaceutical preparation is such that an appropriate dose within a specified range is provided (e.g., a dose within the range of 0.01 to 100 mg per kg of body weight).

[0528] Pharmaceutical compositions for gene therapy can be present in an acceptable diluent or can comprise a slow-release matrix in which the gene delivery vehicle is embedded. When hydrodynamic injection is used as a delivery method, pharmaceutical compositions containing nucleic acid molecules encoding the polypeptides described herein or vectors (e.g., viral vectors) containing nucleic acid molecules are rapidly delivered intravenously in large volumes of liquid. Vectors that can be used as in vivo gene delivery vehicles include, but are not limited to, retroviral vectors, adenoviral vectors, poxvirus vectors (e.g., vaccinia virus vectors such as modified vaccinia Ankara), adeno-associated virus vectors, and alphavirus vectors.

[0529] IX. Route, Dosage, and Administration Pharmaceutical compositions containing the polypeptide of the present invention as a therapeutic protein can be formulated for, for example, intravenous administration, parenteral administration, subcutaneous administration, intramuscular administration, intraarterial administration, intrathecal administration, or intraperitoneal administration. Pharmaceutical compositions can also be formulated or administered orally, nasally, by spray, aerosol, rectum, or vaginal administration. For injectable formulations, various effective pharmaceutical carriers are known in the art. For example, see ASHP Handbook on Injectable Drugs, Toissel, 18th ed. (2014).

[0530] In some embodiments, pharmaceutical compositions containing nucleic acid molecules encoding polypeptides of the present invention or vectors containing such nucleic acid molecules can be administered by gene delivery. Methods of gene delivery are well known to those skilled in the art. Vectors that can be used for in vivo gene delivery and expression include, but are not limited to, retroviral vectors, adenoviral vectors, poxvirus vectors (e.g., vaccinia virus vectors such as modified vaccinia Ankara (MVA)), adeno-associated virus vectors, and alphavirus vectors. In some embodiments, mRNA molecules encoding polypeptides of the present invention can be directly administered to a subject.

[0531] In some embodiments of the present invention, nucleic acid molecules encoding the polypeptides described herein or vectors containing such nucleic acid molecules can be administered using a hydrodynamic injection platform. In hydrodynamic injection, nucleic acid molecules encoding the polypeptides described herein are placed under the control of a strong promoter within an engineered plasmid (e.g., a viral plasmid). The plasmid is often delivered rapidly intravenously in a large volume of fluid. Hydrodynamic injection uses controlled hydrodynamic pressure within the vein to increase cell permeability, and the increased pressure caused by the rapid injection of a large volume of fluid results in extravasation of the fluid and plasmid from the vein. Expression of the nucleic acid molecule is primarily driven by the liver. In mice, hydrodynamic injection is often performed by injecting the plasmid into the tail vein. In certain embodiments, mRNA molecules encoding the polypeptides described herein can be administered using hydrodynamic injection.

[0532] The dosage of the pharmaceutical compositions of the present invention will depend on factors including the route of administration, the disease being treated, and the physical characteristics, such as age, weight, and general health of the subject. The pharmaceutical compositions of the present invention may contain a dosage of a polypeptide of the present invention ranging from 0.01 to 500 mg / kg (e.g., 0.01, 0.1, 0.2, 0.3, 0.4, 0.5, 0.75, 1, 1.25, 1.5, 1.75, 2, 2.25, 2.5, 2.75, 3, 3.25, 3.5, 3.75, 4, 4.25, 4.5, 4.75, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 100, 150, 200, 250, 300, 350, 400, 450, or 500 mg / kg), or in a more specific embodiment, from about 0.1 to about 30 mg / kg, and in an even more specific embodiment, from about 0.3 to about 30 mg / kg. The dosage can be adapted by a physician according to conventional factors such as the extent of the disease and different parameters of the subject.

[0533] Pharmaceutical compositions are administered in a manner compatible with the dosage formulation, and in an amount that is therapeutically effective to improve or remedy the symptoms. Pharmaceutical compositions are administered in a variety of dosage forms, including intravenous, subcutaneous, and oral dosage forms (e.g., ingestible solutions, drug-release capsules). Generally, therapeutic proteins are administered at 0.1-100 mg / kg, e.g., 0.5-50 mg / kg. Pharmaceutical compositions comprising the polypeptides of the present invention can be administered to a subject in need thereof, for example, daily, weekly, every two weeks, every four weeks, monthly, bimonthly, quarterly, semi-annually, once or more times yearly (e.g., 1-10 times or more), or as medically necessary. In some embodiments, pharmaceutical compositions comprising the polypeptides of the present invention can be administered to a subject in need thereof weekly, every two weeks, every four weeks, monthly, bimonthly, or quarterly. Dosages can be provided in single or multiple administration regimens. The interval between administrations can be shortened as the condition improves or lengthened as the patient's health deteriorates.

[0534] X. Treatment method The present invention is based on the discovery that combining the extracellular portions of ActRIIA and ActRIIB results in an ActRII chimera with improved properties (e.g., improved ligand-binding properties) compared to wild-type extracellular ActRIIA and extracellular ActRIIB. The ActRII chimera generated by combining the extracellular portions of ActRIIA and ActRIIB may possess the beneficial properties of both ActRIIB (e.g., the ability to increase muscle mass and strong binding affinity to activins A and B) and ActRIIA (e.g., reduced binding affinity to BMP9 and / or a longer serum half-life as an Fc fusion protein (e.g., compared to ActRIIB-Fc), and / or the ability to increase red blood cell levels). Because the ActRII chimera contains the extracellular portions of ActRIIA and ActRIIB, it is soluble and can compete with endogenous activin receptors by binding to and sequestering ligands (e.g., activins A and B, myostatin, GDF11) without activating intracellular signaling pathways. Therefore, the extracellular ActRII chimeras described herein can be used to treat diseases or conditions in which elevated activin signaling is involved in the pathogenesis (e.g., diseases or conditions in which increased expression of activin receptors or activin receptor ligands has been observed). For example, myostatin has been implicated in promoting fibrosis, inhibiting skeletal muscle growth, and regulating bone homeostasis. Elevated myostatin levels have been observed in the subcutaneous and visceral fat of obese mice and in the plasma of obese and insulin-resistant women. Furthermore, activin A has been reported to be upregulated in bone disease, clinical and experimental pulmonary hypertension, adipose tissue, and the subcutaneous and visceral fat of obese mice, where it has been found to inhibit osteoblast activity and promote fibrosis. Another activin receptor ligand, GDF11, has been overexpressed in mouse models of hemolytic anemia and associated with defective erythropoiesis. Both type I and type II activin receptors have been linked to pancreatic function and diabetes.Without wishing to be bound by theory, therapeutic agents that bind to activin receptor ligands (e.g., GDF11, myostatin, and / or activin) and reduce their binding to or interaction with endogenous activin receptors (e.g., by sequestering the endogenous ligand) may have therapeutic utility in treating or preventing a variety of diseases or conditions, such as muscle diseases, bone diseases, fibrosis, anemia, thrombocytopenia, neutropenia, metabolic diseases (e.g., obesity, type 1 diabetes, or type 2 diabetes), or PH (e.g., PAH, venous PH, hypoxic PH, thromboembolic PH, or other PH).

[0535] The compositions and methods described herein can be used to treat and / or prevent (e.g., prevent the onset of, or treat a subject diagnosed with) a medical condition, e.g., a condition such as a muscle disorder (e.g., skeletal muscle weakness or atrophy), a bone disorder, low red blood cell levels (e.g., low hemoglobin levels or low red blood cell count, e.g., anemia), fibrosis, thrombocytopenia (e.g., low platelet count), neutropenia (e.g., low neutrophil count), a metabolic disorder (e.g., obesity, type 1 diabetes, or type 2 diabetes), or PH (e.g., PAH, venous PH, hypoxic PH, thromboembolic PH, or other PH). In some embodiments, a polypeptide described herein (e.g., an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43), e.g., a polypeptide comprising an effective amount of an ActRII chimera) can be administered to a subject in need thereof to increase muscle mass and strength. In some embodiments, a polypeptide described herein can be administered to increase lean body mass. A polypeptide described herein can increase muscle mass and / or lean mass compared to measurements obtained before treatment. In some embodiments, the subject The subject may have or be at risk of developing a disease or condition that results in muscle weakness or atrophy (e.g., muscle weakness or atrophy associated with a neuromuscular disease, cachexia, sarcopenia, or treatment). In some embodiments, the methods described herein aim to affect myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of activin A, activin B, myostatin, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing a disease or condition involving muscle weakness and atrophy.

[0536] In some embodiments, a polypeptide described herein (e.g., an extracellular ActRII chimera (an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43), e.g., a polypeptide comprising an effective amount of an ActRII chimera) can be administered to a subject in need thereof to increase bone density, increase bone formation, increase bone strength, reduce the risk or occurrence of fracture, or reduce bone resorption. The polypeptide described herein can increase bone density, increase bone formation, or reduce bone strength compared to measurements obtained before treatment. can reduce bone resorption. In some embodiments, the subject may have or be at risk of developing a disease that results in bone damage (e.g., osteoporosis or osteopenia). In some embodiments, the methods described herein are aimed at affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of activin A, activin B, myostatin, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing a disease or condition involving bone damage.

[0537] In some embodiments, the polypeptides described herein (e.g., extracellular ActRII chimeras (e.g., extracellular ActRII chimeras having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43), e.g., polypeptides comprising an effective amount of ActRII chimeras) increase red blood cell levels (e.g., increase hemoglobin levels, increase red blood cell count, increase red blood cell volume, increase red blood cell mass, increase hematocrit, or increase red blood cell formation or production), increase maturation and / or differentiation of red blood cell progenitor cells, or increasing the maturation and / or differentiation of early or late (e.g., terminal) stage progenitor cells, e.g., early stage erythroid progenitor cells, e.g., burst-forming unit-erythroid (BFU-E) and / or colony-forming unit-erythroid (CFU-E), e.g., BFU-E and / or CFU-E, into proerythroblasts, reticulocytes, or erythrocytes, e.g., increasing the number of proerythroblasts and / or reticulocytes; maturation of late stage progenitor cells (erythroid progenitors) (e.g., maturation of reticulocytes into erythrocytes, or maturation of erythroblasts into reticulocytes and / or erythrocytes); administration in a subject in need thereof to increase the number of early stage erythroid precursors and / or progenitors (e.g., to increase the number of early stage erythroid precursors and / or progenitors (e.g., to expand the early stage precursor population to provide a continuous supply of precursors that replenish polychromatic erythroblasts and allow for a continuous supply of mature reticulocytes), promote the progression of erythroid precursors and / or progenitors through erythropoiesis, and / or decrease the accumulation of erythroid precursors (e.g., by stimulating progenitor cells to progress to maturation); The polypeptides described herein can increase red blood cell levels, increase maturation and / or differentiation of erythroid progenitors, increase maturation of late-stage erythroid progenitors, mobilize early-stage progenitors to the erythroid lineage, increase the number of early-stage erythroid progenitors, promote the progression of erythroid progenitors through erythropoiesis, or reduce the accumulation of erythroid progenitors, compared to measurements obtained before treatment. In some embodiments, the subject may have a disease or condition associated with low red blood cell levels (e.g., anemia or blood loss).In some embodiments, the subject may have or be at risk of developing anemia or blood loss (e.g., the subject may have or be at risk of developing anemia resulting from other diseases or conditions, such as myelodysplastic syndrome, myelofibrosis, chronic kidney disease, rheumatoid arthritis, ineffective hematopoiesis, cancer, or inflammatory diseases (e.g., Crohn's disease, SLE, or ulcerative colitis), or resulting from medical treatments, such as chemotherapy, radiation therapy, or surgery). In some embodiments, the methods described herein are aimed at affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of activin A, activin B, myostatin, and / or BMP9 to their endogenous receptors) in subjects having or at risk of developing a disease or condition associated with low red blood cell levels.

[0538] In some embodiments, a polypeptide described herein (e.g., an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43), e.g., a polypeptide comprising an effective amount of an ActRII chimera) can be administered to a subject in need thereof to increase platelet levels (e.g., increase platelet count), promote megakaryocyte differentiation and / or maturation (e.g., to produce platelets), reduce platelet precursor cell accumulation, improve blood clotting, reduce bleeding events, reduce skin bleeding (e.g., petechiae or purpura), and / or promote or increase platelet formation or production. A polypeptide described herein may increase platelet levels, promote megakaryocyte differentiation and / or maturation, reduce platelet progenitor cell accumulation (e.g., by stimulating progenitor cells to progress to maturity), improve blood clotting, reduce bleeding events, reduce bleeding in the skin, and / or promote or increase platelet formation or production compared to measurements obtained before treatment. In some embodiments, the subject may have a disease or condition associated with low platelet levels (e.g., thrombocytopenia). In some embodiments, megakaryocytes can be contacted in vitro with a polypeptide, a nucleic acid encoding a polypeptide, or a vector comprising a nucleic acid described herein to generate platelets for the treatment of thrombocytopenia.In some embodiments, the subject has or may be at risk of developing thrombocytopenia (e.g., the subject has myelodysplastic syndrome, myelofibrosis, myelofibrosis treatment (e.g., treatment with a JAK inhibitor such as ruxolitinib or fedratinib), hematopoietic failure, Gaucher disease, aplastic anemia, Fanconi anemia, Diamond-Blackfan anemia, Shwachman-Diamond syndrome, excessive alcohol use, cirrhosis of the liver, cancer (e.g., leukemia or lymphoma), immune thrombocytopenia, autoimmune disease (e.g., rheumatoid arthritis or lupus (e.g., SLE)), viral infection (e.g., hepatitis C, HIV, chickenpox, mumps, rubella, parvovirus, or Epstein-Barr virus), bacterial infection (e.g., bacteremia), vitamin deficiency (e.g., may have or be at risk for developing thrombocytopenia due to other diseases or conditions, such as thrombocytopenia due to other conditions, such as vitamin B-12 deficiency, folate deficiency, or iron deficiency), cancer treatment (e.g., chemotherapy or radiation therapy), splenomegaly, thrombotic thrombocytopenic purpura, idiopathic thrombocytopenic purpura, disseminated intravascular coagulation, hemolytic uremic syndrome, paroxysmal nocturnal hemoglobinuria, acquired amegakaryocytic thrombocytopenia, Pearson syndrome, dyskeratosis congenita, genetic disorders (e.g., Wiskott-Aldrich or May-Hegglin syndrome), platelet dilution due to blood transfusions, or medications (e.g., sulfa-containing antibiotics such as heparin, quinine, vancomycin, rifampin, trimethoprim, or anticonvulsants such as phenytoin). In some embodiments, the methods described herein are aimed at affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of activin A, activin B, myostatin, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing a disease or condition associated with low platelet levels.

[0539] In some embodiments, a polypeptide described herein (e.g., an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43), e.g., a polypeptide comprising an effective amount of an ActRII chimera) can be administered to a subject in need thereof to increase neutrophil levels (e.g., increase neutrophil count), promote differentiation and / or maturation of progenitor cells (e.g., bone marrow progenitor cells, myeloblasts, or myeloid cells) into neutrophils, and / or promote or increase the formation or production of neutrophils. The polypeptide described herein may increase neutrophil levels, increase or promote the differentiation and / or maturation of progenitor cells into neutrophils, and / or promote or increase the formation or production of neutrophils, compared to measurements obtained before treatment. In some embodiments, the subject may have a disease or condition associated with low neutrophil levels (e.g., neutropenia).In some embodiments, the subject has or may be at risk of developing neutropenia (e.g., the subject has a myelodysplastic syndrome, myelofibrosis, ineffective hematopoiesis, aplastic anemia, Fanconi anemia, Diamond-Blackfan anemia, Shwachman-Diamond syndrome, paroxysmal nocturnal hemoglobinuria, Pearson syndrome, dyskeratosis congenita, cancer (e.g., leukemia), vitamin deficiency (e.g., B-12 deficiency or folate deficiency), splenomegaly, an autoimmune disease (e.g., granulomatosis with polyangiitis, lupus (e.g., SLE), Evans syndrome, Felty syndrome, Crohn's disease, or rheumatoid arthritis), a viral infection (e.g., chickenpox, Epstein-Barr, hepatitis A, hepatitis B, hepatitis C, HIV / AIDS, cytomegalovirus, dengue fever, measles), a bacterial infection (e.g., erythrocyte sedimentation syndrome, leukemia), or a bacterial infection (e.g., leukemia). may have or be at risk for developing neutropenia due to other diseases or conditions, such as infections (e.g., tuberculosis, salmonella infection, or sepsis), cancer treatments (e.g., chemotherapy or radiation therapy), or treatment with other drugs (e.g., drugs used to treat hyperthyroidism such as methimazole and propylthiouracil, antibiotics such as vancomycin, penicillin G, trimethoprim, and oxacillin, antiviral drugs such as ganciclovir and valganciclovir, anti-inflammatory drugs for ulcerative colitis or rheumatoid arthritis such as sulfasalazine, drugs used to treat cardiac arrhythmias such as quinidine and procainamide, anticonvulsants such as phenytoin and valproate, antipsychotics such as clozapine, or levamisole). In some embodiments, the methods described herein are aimed at affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of activin A, activin B, myostatin, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing a disease or condition associated with low neutrophil levels.

[0540] In some embodiments, a polypeptide described herein (e.g., an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43), e.g., a polypeptide comprising an effective amount of an ActRII chimera) can be administered to a subject in need thereof to prevent or reduce fibrosis. In some embodiments, a polypeptide described herein can be administered to slow or halt the progression of fibrosis, reduce the risk of developing fibrosis, or alleviate (e.g., reduce the frequency or severity of) one or more symptoms of fibrosis. A polypeptide described herein can reduce fibrosis or slow the progression of fibrosis, at least as compared to the progression of fibrosis before treatment, or as compared to the progression of fibrosis in an untreated subject. In some embodiments, the subject has or may be at risk of developing fibrosis (e.g., the subject has a condition such as a wound, hepatitis B or hepatitis C, fatty liver disease, kidney disease (e.g., chronic renal disease), cardiac In some embodiments, the subject may have a disease or condition associated with fibrosis, such as liver disease, or atherosclerosis, or may be undergoing treatment associated with the development of fibrosis, such as chemotherapy, radiation, or surgery. In some embodiments, the polypeptides described herein prevent or delay the onset of fibrosis in a subject at risk for developing fibrosis (e.g., a subject being treated with chemotherapy, radiation therapy, or surgery, or a subject with a disease or condition associated with fibrosis, such as a wound, hepatitis B or hepatitis C, fatty liver disease, kidney disease (e.g., chronic kidney disease), heart disease, or atherosclerosis). In some embodiments, the methods described herein are aimed at affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., decreasing or inhibiting binding of activin A, activin B, myostatin, and / or BMP9 to their endogenous receptors) in a subject having or at risk for developing fibrosis or a disease or condition associated with fibrosis.

[0541] In some embodiments, a polypeptide described herein (e.g., an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43), e.g., a polypeptide comprising an effective amount of an ActRII chimera) can be administered to a subject in need thereof to treat PH, alleviate PH (e.g., reduce the severity or frequency of one or more symptoms of PH, such as shortness of breath (dyspnea), fatigue, swelling of the legs, feet, abdomen (ascites), or neck (e.g., edema), or chest pain or pressure, fast pulse or heart palpitations, bluish color of the lips or skin (cyanosis), dizziness, or fainting), prevent PH (e.g., prevent the onset of PH), reduce the risk of onset of PH, or slow or stop the progression of PH. The polypeptide described herein can alleviate the symptoms of PH or slow the progression of PH compared to the symptoms or progression observed before treatment or compared to the symptoms or progression of PH in an untreated subject.In some embodiments, the subject may have or be at risk for developing PH (e.g., the subject may have idiopathic PAH; the subject may have a disease or condition associated with PAH (e.g., a disease or condition that confers an increased risk of developing PAH), such as HIV infection, schistosomiasis, portal hypertension, pulmonary veno-occlusive disease, pulmonary capillary hemangiomatosis, cirrhosis of the liver, congenital heart abnormalities, connective tissue / autoimmune diseases (e.g., scleroderma or lupus), or drug use or abuse (e.g., methamphetamine or cocaine use). The subject may have a family history of PH (e.g., hereditary PAH); the subject may have a disease or condition associated with venous PH (e.g., a disease or condition that confers an increased risk of developing venous PH), such as left ventricular systolic dysfunction, left ventricular diastolic dysfunction, valvular heart disease, congenital cardiomyopathy, or congenital / acquired pulmonary vein stenosis; the subject may have a disease or condition associated with hypoxic PH (e.g., a disease or condition that confers an increased risk of developing hypoxic PH), such as chronic obstructive pulmonary disease. The subject may have a disease or condition associated with thromboembolic PH (e.g., a disease or condition that confers an increased risk of developing thromboembolic PH), such as chronic thromboembolic pulmonary hypertension or pulmonary artery obstruction (e.g., pulmonary embolism, angiosarcoma, arteritis, congenital pulmonary stenosis, or parasitic infection); or the subject may have a disease or condition associated with thromboembolic PH (e.g., a disease or condition that confers an increased risk of developing thromboembolic PH), such as chronic thromboembolic pulmonary hypertension or pulmonary artery obstruction (e.g., pulmonary embolism, angiosarcoma, arteritis, congenital pulmonary stenosis, or parasitic infection). They may have other diseases or conditions associated with PH (e.g., diseases or conditions that pose an increased risk of developing other PH), such as a blood disorder (e.g., chronic hemolytic anemia, sickle cell disease), a systemic disease (e.g., sarcoidosis, pulmonary Langerhans cell histiocytosis, lymphangioleiomyomatosis, neurofibromatosis, or vasculitis), a metabolic disorder (e.g., glycogen storage disease, Gaucher disease, or thyroid disease), pulmonary neoplastic thrombotic microangiopathy, fibrosing mediastinitis, chronic renal failure, or segmental pulmonary hypertension (pulmonary hypertension limited to one or more lobes of the lung).In some embodiments, the polypeptides described herein prevent or delay the onset of PH in a subject at risk of developing PH (e.g., a subject with a family history of PH (e.g., hereditary PAH) or a subject with a disease or disorder that confers an increased risk of developing PAH, venous PH, hypoxic PH, thromboembolic PH, or other PH). In some embodiments, the methods described herein aim to affect myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of activin A, activin B, myostatin, and / or BMP9 to their receptors) in a subject with or at risk of developing PH or a disease or condition related to PH. In some embodiments, the PH is PAH, venous PH, hypoxic PH, thromboembolic PH, or other PH.

[0542] In some embodiments, a polypeptide described herein (e.g., an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43), e.g., a polypeptide comprising an effective amount of an ActRII chimera) can be administered to a subject in need thereof to reduce body fat (e.g., body fat amount or body fat percentage), reduce body weight or weight gain, reduce fasting insulin levels, increase glucose clearance, reduce LDL, reduce triglycerides, improve serum lipid profile, or increase insulin sensitivity (e.g., reduce insulin resistance) compared to measurements obtained before treatment. The therapeutic effect of the present invention can increase clearance, reduce LDL, reduce triglycerides, improve serum lipid profiles, or increase insulin sensitivity (e.g., reduce insulin resistance). In some embodiments, the subject can have a disease or condition related to obesity or diabetes (e.g., type 1 diabetes or type 2 diabetes). In some embodiments, the subject can have or be at risk of developing a metabolic disease (e.g., obesity, type 1 diabetes, or type 2 diabetes), for example, the subject can be overweight, have a family history of obesity, or have other conditions or risk factors associated with an increased risk of developing obesity or diabetes (e.g., older age, or treatment with glucocorticoids, selective serotonin reuptake inhibitors (SSRIs), tricyclic antidepressants, mood stabilizers, antipsychotics, serotonin-norepinephrine reuptake inhibitors (SNRIs)), have a family history of diabetes, or have prediabetes.In some embodiments, the methods described herein are aimed at affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., reducing or inhibiting binding of activin A, activin B, myostatin, and / or BMP9 to their endogenous receptors) in a subject having or at risk of developing a metabolic disease (e.g., obesity, type 1 diabetes, or type 2 diabetes).

[0543] In some embodiments, polypeptides comprising the extracellular ActRII chimeras described herein reduce or inhibit the binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors, e.g., ActRIIA, ActRIIB, and / or BMPRII. The polypeptides described herein may reduce the binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors compared to the binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors in the absence of the polypeptides of the invention.In some embodiments, affecting myostatin, activin A, activin B, and / or BMP9 signaling (e.g., decreasing or inhibiting the binding of myostatin, activin A, activin B, and / or BMP9 to their endogenous receptors, e.g., ActRIIA, ActRIIB, and / or BMPRII) can result in an increase in muscle mass in a subject, an increase in lean body mass in a subject, an increase in bone mineral density or bone formation in a subject, a decrease in bone resorption in a subject, an increase in red blood cell levels in a subject (e.g., hemoglobin level, hematocrit, red blood cell count, red blood cell volume, or red blood cell mass, e.g., promoting or increasing the formation or production of red blood cells), an increase in the maturation and / or differentiation of erythroid progenitors, an increase in the maturation of late erythroid progenitors, mobilization of early progenitors into the erythroid lineage, a decrease in the accumulation of erythroid progenitors, an increase in the number of early stage erythroid progenitors and / or precursors, an increase in the number of erythroid progenitors and / or precursors through erythropoiesis, or an increase in the number of erythroid progenitors and / or precursors through erythropoiesis. or progression of PH, an increase in platelet levels in a subject (e.g., increased platelet count, megakaryocyte differentiation and / or maturation, and / or platelet formation or production), a decrease in platelet precursor accumulation, improved blood clotting, a decrease in bleeding events, a decrease in skin bleeding, an increase in neutrophil levels in a subject (e.g., increased neutrophil count, e.g., increased neutrophil production or increased formation), an increase in differentiation and / or maturation of precursor cells into neutrophils, a decrease in fibrosis or risk of developing fibrosis in a subject, a delay in the onset of fibrosis, a decrease (e.g., slowing or inhibiting) the progression of fibrosis, a decrease in body fat (e.g., body fat amount or body fat percentage), a decrease in body weight or weight gain, a decrease in fasting insulin levels, an increase in glucose clearance, an improvement in serum lipid profile, an increase in insulin sensitivity (e.g., decreased insulin resistance), a reduction in symptoms of PH, a decrease in risk of developing PH, a delay in the onset of PH, and / or a decrease (e.g., slowing or inhibiting) the progression of PH. PH can be PAH, venous PH, hypoxic PH, thromboembolic PH, or other PH.

[0544] In some embodiments, a polypeptide described herein (e.g., an extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43), e.g., a polypeptide comprising an effective amount of an ActRII chimera) increases muscle mass or strength, increases lean body mass, increases bone mineral density, increases bone formation, increases bone strength, reduces the risk or incidence of fractures, reduces bone resorption, increases red blood cell levels, increases maturation and / or differentiation of erythroid progenitors, increases maturation of late-stage erythroid progenitors, mobilizes early-stage progenitors to the erythroid lineage, reduces erythroid progenitor accumulation, increases the number of early-stage erythroid progenitors and / or precursors, promotes the progression of erythroid progenitors and / or precursors through erythropoiesis, increases platelet levels, or increases megakaryocytic differentiation and / or maturation in a subject, compared to measurements obtained before treatment or compared to measurements obtained from an untreated subject with the same disease or condition. the formation or production of platelets, the reduction of platelet precursor cell accumulation, the improvement of blood clotting, the reduction of bleeding events, the reduction of skin bleeding, the increase of neutrophil levels in a subject, the increase of neutrophil production or formation, the increase or promotion of the differentiation and / or maturation of neutrophil precursor cells, the prevention or reduction of fibrosis (reducing fibrosis, preventing or delaying the onset of fibrosis, slowing or halting the progression of fibrosis), the treatment of a metabolic disease, the reduction of body fat (e.g., the amount of body fat or body fat percentage), the reduction of body weight or weight gain, the reduction of fasting insulin levels, the increase of glucose clearance, the improvement of serum lipid profiles, the prevention or treatment of PH (e.g., reducing the symptoms of PH, preventing or delaying the onset of PH, slowing or halting the progression of PH), or the effect of myostatin, activin A, activin B, and / or BMP9 signaling.An extracellular ActRII chimera (e.g., an extracellular ActRII chimera having the sequence of any one of SEQ ID NOs: 1 to 43 (e.g., SEQ ID NOs: 22 to 43), e.g., an effective amount of an ActRII chimera) may increase muscle mass or strength, increase lean body mass, increase bone density, increase bone formation, increase bone strength, or reduce the risk or occurrence of fractures, compared to measurements obtained before treatment or compared to measurements obtained from an untreated subject with the same disease or condition. may decrease blood cell death, decrease bone resorption, increase red blood cell levels, increase maturation and / or differentiation of red blood cell precursors, increase maturation of late stage red blood cell precursors (e.g., final maturation such as maturation of reticulocytes to red blood cells, or maturation of erythroblasts to reticulocytes and / or red blood cells), mobilize early stage precursors to the erythroid lineage, decrease accumulation of red blood cell precursors, increase the number of early stage red blood cell precursors and / or precursors, increase red blood cell proliferation, may promote the progression of erythroid progenitor cells and / or precursor cells through production; may increase platelet levels in a subject; may increase megakaryocyte differentiation and / or maturation; may increase platelet formation or production; may decrease accumulation of platelet precursor cells; may improve blood clotting; may decrease bleeding events; may decrease skin bleeding; may increase neutrophil levels in a subject; may increase or promote differentiation and / or maturation of progenitor cells into neutrophils; In some embodiments, the methods described herein may increase blood glucose levels, prevent or reduce fibrosis, treat metabolic diseases, reduce body fat (e.g., body fat amount or body fat percentage), reduce body weight or weight gain, reduce fasting insulin levels, increase glucose clearance, improve serum lipid profiles, prevent or treat PH, or affect myostatin, activin A, activin B, and / or BMP9 signaling. In some embodiments, the methods described herein do not cause vascular complications in a subject, such as increased vascular permeability or vascular leakage.

[0545] The present invention also includes methods for treating a subject having or at risk of developing a disease or condition involving muscle weakness or atrophy by administering to the subject an effective amount of a polypeptide described herein (e.g., an extracellular ActRII chimera (a polypeptide comprising an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)). In any of the methods described herein, the subject having or at risk of developing a disease or condition involving muscle weakness or atrophy may have a neuromuscular disease (e.g., muscular dystrophy, IBM, SMA, CMT, ALS, myasthenia gravis, or multiple sclerosis), sarcopenia, cachexia (e.g., cancer cachexia, HIV-associated cachexia, cardiac cachexia (e.g., cachexia associated with heart failure), cachexia associated with chronic kidney disease, or pulmonary The patient has or is at risk of developing a disease or condition involving cachexia (e.g., cachexia associated with COPD), disuse atrophy, treatment-related muscle loss or atrophy (e.g., glucocorticoid treatment, FGF-21 treatment, GLP-1 treatment, bariatric surgery, cancer treatment, or treatment for obesity or type 2 diabetes), muscle loss or atrophy associated with hypotonia, hypoxia, or burns. Muscular dystrophies include Duchenne muscular dystrophy (DMD), facioscapulohumeral muscular dystrophy (FSHD), Becker muscular dystrophy (BMD), myotonic dystrophy (DM), congenital muscular dystrophy, limb-girdle muscular dystrophy (LGMD), distal muscular dystrophy (DD), oculopharyngeal muscular dystrophy (OPMD), and Emery-Dreifuss muscular dystrophy (EDMD).There are 33 types of congenital muscular dystrophies, including congenital muscular dystrophy type 1A (MDC1A, associated with mutations in laminin α2), congenital muscular dystrophy type 1C (MDC1C, associated with mutations in FKRP), congenital muscular dystrophy type 1D (MDC1D, associated with mutations in LARGE), congenital muscular dystrophy type 1B (MDC1B), Fukuyama congenital muscular dystrophy (FCMD, associated with mutations in fukutin), muscle-eye-brain disease (MEB, possibly associated with mutations in POMGnT1), Walker-Warburg syndrome (WWS, B3GNT1 (MDDGA type), POMT1 (MDDGA type), In some embodiments, the methods described herein increase muscle mass, e.g., increase muscle mass, lean body mass, and / or muscle strength, e.g., increase muscle mass, lean body mass, and / or muscle strength, e.g., increase muscle mass, lean body mass, and / or muscle strength compared to measurements obtained before treatment or compared to measurements typically observed in untreated subjects with the same disease or condition. In some embodiments, the muscle is skeletal muscle. In some embodiments, the subject is identified as having a disease or condition that results in muscle weakness and atrophy prior to treatment with an ActRII chimera described herein. In some embodiments, the method includes identifying the subject as having a disease or condition that results in muscle weakness or atrophy prior to treatment with an ActRII chimera described herein (e.g., by assessing lean body mass, muscle mass, or muscle strength, or by genetic testing for congenital muscular dystrophy).The method may further include assessing lean body mass, muscle mass, or muscle strength after administration of an ActRII chimera described herein (e.g., 12 hours, 24 hours, 1, 2, 3, 4, 5, 6, or 7 days, 1, 2, 3, 4, 5, 6, 7, or 8 weeks, or 1, 2, 3, 4, 5, or 6 months or more after initiation of treatment).

[0546] The present invention also includes methods for treating a subject having or at risk of developing a bone disease (e.g., bone injury) by administering to the subject an effective amount of a polypeptide described herein (e.g., a polypeptide comprising an extracellular ActRII chimera having the sequence of any one of SEQ ID NOS: 1-43 (e.g., SEQ ID NOS: 22-43)). In any of the methods described herein, the subject having or at risk of developing a bone disease (e.g., bone injury) has or is at risk of developing a disease or condition including primary osteoporosis, secondary osteoporosis, osteopenia, osteopetrosis, osteogenesis imperfecta, bone fracture, bone loss associated with bone cancer or cancer metastasis, Paget's disease, renal osteodystrophy, treatment-related bone loss, bone loss associated with neuromuscular disease, bone loss due to burns, bone loss associated with anorexia, diet-related bone loss, bone loss associated with treatment of obesity, bone loss associated with reduced gravity, or bone loss associated with immobility. In some embodiments, In some embodiments, primary osteoporosis is age-related or hormone-related osteoporosis (e.g., associated with estrogen depletion). In some embodiments, secondary osteoporosis is immobility-induced osteoporosis or glucocorticoid-induced osteoporosis. In some embodiments, the bone cancer is multiple myeloma, or the bone loss associated with cancer metastasis is caused by multiple myeloma. In some embodiments, treatment-related bone loss occurs due to treatment with FGF-21 or GLP-1, treatment with a therapeutic agent containing FGF-21 or GLP-1, treatment of type 2 diabetes and / or obesity, bariatric surgery, androgen or estrogen deprivation therapy, or cancer treatment (e.g., chemotherapy or radiation). In some embodiments, diet-related bone loss is rickets (e.g., vitamin D deficiency). In some embodiments, low-gravity-related bone loss is load-loss-related bone loss.In some embodiments, the methods described herein increase bone density (e.g., increase bone mass), decrease bone resorption (e.g., decrease bone catabolic activity), increase bone formation (e.g., increase bone anabolic activity or increase bone formation), increase osteoblast activity or formation, and / or decrease osteoclast activity or formation, compared to measurements obtained before treat...

Claims

1. 1. A polypeptide comprising an extracellular activin receptor type II (ActRII) chimera, said chimera comprising: GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRRHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 1), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 2), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 3), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGSIEIIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 4), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 5), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 6), GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRQECVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 7), GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRRHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 8), GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 9), GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCYASWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 10), GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGSIEIIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 11), GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 12), GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 13), GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRQECVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 14), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRRHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 15), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 16), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 17), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGSIEIIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 18), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 19), GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 20), and GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRQECVX ４ X ５ X ６ X ７ X ８ PX ９ VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 21) and In the formula, X １ is D or R, and X ２ is I, F, E, D, Y, S, N, Q, or T, and X ３ is N or T, and X ４ is A or E, and X ５ is T or K, and X ６ is E or K, and X ７ is E or D, and X ８ is N or S, and X ９ is Q, E, K, R, D, or N, and optionally said chimera is truncated from the N-terminus by the deletion of 1, 2, 3, 4, 5, 6, 7, 8, or 9 amino acids, wherein said chimera retains the two amino acids before the first cysteine.

2. The chimera GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRRHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 1).

3. The chimera GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 2).

4. The chimera GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 3).

5. The chimera GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGSIEIIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 4).

6. The chimera GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 5).

7. The chimera GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 6).

8. The chimera GAILGRAETRECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRQECVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 7).

9. The chimera GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRRHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 8).

10. The chimera GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 9).

11. The chimera GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCYASWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 10).

12. The chimera GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGSIEIIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 11).

13. The chimera GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 12).

14. The chimera GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 13).

15. The chimera GRGEAETRECIYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRQECVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 14).

16. The chimera GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRRHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 15).

17. The chimera GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCFATWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 16).

18. The chimera GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWKNISGSIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 17).

19. The chimera GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGSIEIIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 18).

20. The chimera GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIEIVKQGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 19).

21. The chimera GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRTDDCVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 20).

22. The chimera GAILGRSETQECIYYNANWELERTNQSGLERCEGEQX １ KRLHCYASWRNSSGTIELVKKGCWLDDX ２ X ３ CYDRQECVX ４ X ５ X ６ X ７ X ８ PX ９ 2. The polypeptide of claim 1 having the sequence VYFCCCEGNMCNEKFSYFPEMEVTQPTS (SEQ ID NO: 21).

23. X １ The polypeptide according to any one of claims 1 to 22, wherein is D.

24. X １ The polypeptide of any one of claims 1 to 22, wherein is R.

25. X ２ The polypeptide according to any one of claims 1 to 24, wherein is I.

26. X ２ The polypeptide according to any one of claims 1 to 24, wherein is F.

27. X ２ The polypeptide according to any one of claims 1 to 24, wherein is E.

28. X ２ The polypeptide according to any one of claims 1 to 24, wherein is D.

29. X ２ The polypeptide of any one of claims 1 to 24, wherein is Y.

30. X ２ The polypeptide of any one of claims 1 to 24, wherein is S.

31. X ２ The polypeptide according to any one of claims 1 to 24, wherein is N.

32. X ２ The polypeptide of any one of claims 1 to 24, wherein is Q.

33. X ２ The polypeptide of any one of claims 1 to 24, wherein is T.

34. X ３ The polypeptide of any one of claims 1 to 33, wherein is N.

35. X ３ The polypeptide of any one of claims 1 to 33, wherein is T.

36. X ４ The polypeptide of any one of claims 1 to 35, wherein is A.

37. X ４ The polypeptide of any one of claims 1 to 35, wherein is E.

38. X ５ The polypeptide of any one of claims 1 to 37, wherein is T.

39. X ５ The polypeptide of any one of claims 1 to 37, wherein is K.

40. X ６ The polypeptide of any one of claims 1 to 39, wherein is E.

41. X ６ The polypeptide of any one of claims 1 to 39, wherein is K.

42. X ７ The polypeptide according to any one of claims 1 to 41, wherein is E.

43. X ７ The polypeptide according to any one of claims 1 to 41, wherein is D.

44. X ８ The polypeptide of any one of claims 1 to 43, wherein is N.

45. X ８ The polypeptide of any one of claims 1 to 43, wherein is S.

46. X ９ The polypeptide of any one of claims 1 to 45, wherein is Q.

47. X ９ The polypeptide of any one of claims 1 to 45, wherein is E.

48. X ９ The polypeptide of any one of claims 1 to 45, wherein is K.

49. X ９ The polypeptide of any one of claims 1 to 45, wherein is R.

50. X ９ The polypeptide of any one of claims 1 to 45, wherein is D.

51. X ９ The polypeptide of any one of claims 1 to 45, wherein is N.

52. X ５ is T and X ６ is E and X ７ is E and X ８ The polypeptide of any one of claims 1 to 51, wherein is N.

53. X ５ is T and X ６ is K and X ７ is E and X ８ The polypeptide of any one of claims 1 to 51, wherein is N.

54. X ２ is E and X ３ The polypeptide of any one of claims 1 to 53, wherein is T.

55. The polypeptide of claim 1, wherein the chimera has the sequence of any one of SEQ ID NOs: 22 to 43.

56. 56. The polypeptide of any one of claims 1 to 55, wherein the chimera is truncated from the N-terminus by the deletion of one amino acid.

57. 56. The polypeptide of any one of claims 1 to 55, wherein the chimera is truncated from the N-terminus by the deletion of two amino acids.

58. 56. The polypeptide of any one of claims 1 to 55, wherein the chimera is truncated from the N-terminus by the deletion of three amino acids.

59. 56. The polypeptide of any one of claims 1 to 55, wherein the chimera is truncated from the N-terminus by the deletion of four amino acids.

60. 56. The polypeptide of any one of claims 1 to 55, wherein the chimera is truncated from the N-terminus by the deletion of five amino acids.

61. 56. The polypeptide of any one of claims 1 to 55, wherein the chimera is truncated from the N-terminus by the deletion of six amino acids.

62. 56. The polypeptide of any one of claims 1 to 55, wherein the chimera is truncated from the N-terminus by the deletion of seven amino acids.

63. 56. The polypeptide of any one of claims 1 to 55, wherein the chimera is truncated from the N-terminus by the deletion of eight amino acids.

64. 56. The polypeptide of any one of claims 1 to 55, wherein the chimera is truncated from the N-terminus by the deletion of 9 amino acids.

65. 65. The polypeptide of any one of claims 1 and 56-64, wherein the chimera has the sequence of any one of SEQ ID NOs: 111-183.

66. 66. The polypeptide of any one of claims 1 to 65, further comprising a C-terminal extension of one or more amino acids.

67. 67. The polypeptide of claim 66, wherein the C-terminal extension is NP.

68. 67. The polypeptide of claim 66, wherein the C-terminal extension is NPVTPK (SEQ ID NO: 104).

69. 69. The polypeptide of any one of claims 1 to 68, further comprising an Fc domain monomer fused to the C-terminus of the polypeptide via a linker.

70. 70. The polypeptide of claim 69, wherein the Fc domain monomer comprises the sequence of SEQ ID NO:

48.

71. 71. The polypeptide of claim 69 or 70, wherein the polypeptide forms a dimer.

72. 69. The polypeptide of any one of claims 1 to 68, further comprising a wild-type Fc domain fused to the C-terminus of the polypeptide via a linker.

73. 73. The polypeptide of claim 72, wherein the wild-type Fc domain comprises the sequence of SEQ ID NO: 100 or SEQ ID NO:

264.

74. 74. The polypeptide of claim 72 or claim 73, wherein the polypeptide comprises the sequence of any one of SEQ ID NOs: 107-110 and 184-263.

75. 69. The polypeptide of any one of claims 1 to 68, further comprising an Fc domain comprising an amino acid substitution fused to the C-terminus of the polypeptide via a linker.

76. 76. The polypeptide of claim 75, wherein the Fc domain does not form a dimer.

77. 69. The polypeptide of any one of claims 1 to 68, further comprising an albumin binding peptide fused to the C-terminus of the polypeptide via a linker.

78. 78. The polypeptide of claim 77, wherein the albumin-binding peptide comprises the sequence of SEQ ID NO:

101.

79. 69. The polypeptide of any one of claims 1 to 68, further comprising a fibronectin domain fused to the C-terminus of the polypeptide via a linker.

80. 80. The polypeptide of claim 79, wherein the fibronectin domain comprises the sequence of SEQ ID NO:

102.

81. 69. The polypeptide of any one of claims 1 to 68, further comprising human serum albumin fused to the C-terminus of the polypeptide via a linker.

82. 82. The polypeptide of claim 81, wherein the human serum albumin comprises the sequence of SEQ ID NO:

103.

83. 83. The polypeptide of any one of claims 69 to 82, wherein the linker is an amino acid spacer.

84. 84. The polypeptide of claim 83, wherein the amino acid spacer is GGG, GGGA (SEQ ID NO: 49), GGGG (SEQ ID NO: 51), GGGAG (SEQ ID NO: 81), GGGAGG (SEQ ID NO: 82), or GGGAGGG (SEQ ID NO: 83).

85. 84. The polypeptide of claim 83, wherein the amino acid spacer is GGS, GGGS (SEQ ID NO: 50), GGGGS (SEQ ID NO: 53), GGSG (SEQ ID NO: 56), or SGGG (SEQ ID NO: 58).

86. 86. The polypeptide of any one of claims 1 to 85, wherein the polypeptide has a serum half-life of at least 7 days.

87. 87. The polypeptide of any one of claims 1 to 86, wherein the polypeptide binds to activin A, activin B, and / or myostatin and has reduced or weak binding to human BMP9.

88. 88. The polypeptide of claim 86 or 87, wherein the polypeptide does not substantially bind to human BMP9.

89. The polypeptide has a K Ｄ 89. The polypeptide of any one of claims 1 to 88, which binds to human activin A at

90. The polypeptide has a K Ｄ 90. The polypeptide of any one of claims 1 to 89, which binds to human activin B at

91. The polypeptide has a K Ｄ 91. The polypeptide of any one of claims 1 to 90, which binds to human GDF-11 at the

92. A nucleic acid molecule encoding the polypeptide of any one of claims 1 to 91.

93. 93. A vector comprising the nucleic acid molecule of claim 92.

94. 92. A host cell expressing a polypeptide according to any one of claims 1 to 91, wherein the host cell comprises a nucleic acid molecule according to claim 92, or a vector according to claim 93, and wherein the nucleic acid molecule or vector is expressed in the host cell.

95. 92. A method for preparing a polypeptide according to any one of claims 1 to 91, said method comprising: a) providing a host cell comprising the nucleic acid molecule of claim 92 or the vector of claim 93; b) expressing said nucleic acid molecule or vector in said host cell under conditions that allow the formation of said polypeptide; The method comprising:

96. 94. A pharmaceutical composition comprising a polypeptide according to any one of claims 1 to 91, a nucleic acid molecule according to claim 92, or a vector according to claim 93, and one or more pharmaceutically acceptable carriers or excipients.

97. 94. A method of increasing lean body mass in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

98. 94. A method of increasing muscle mass in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

99. 99. The method of claim 97 or 98, wherein the subject has or is at risk of developing a neuromuscular disease, sarcopenia, cachexia, disuse atrophy, treatment-related muscle loss or atrophy, hypotonia, hypoxia-related muscle loss or atrophy, or burn-related muscle loss or atrophy.

100. A method for affecting the transmission of myostatin, activin A, activin B, and / or BMP9 signals in a subject having or at risk of developing a disease or condition involving muscle weakness and / or atrophy, the method comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

101. 101. The method of claim 100, wherein the disease or condition is a neuromuscular disease, sarcopenia, cachexia, disuse atrophy, treatment-related muscle loss or atrophy, muscle hypotonia, hypoxia-related muscle loss or atrophy, or burn-related muscle loss or atrophy.

102. 94. A method of treating a subject having or at risk of developing a neuromuscular disease, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

103. The neuromuscular disease is muscular dystrophy, amyotrophic lateral sclerosis (ALS), autonomic neuropathy, botulism, Charcot-Marie-Tooth disease (CMT), chronic inflammatory demyelinating polyneuropathy, congenital myasthenic syndrome, congenital myopathy, cramp-fasciculation syndrome, dermatomyositis, diabetic neuropathy, distal myopathy, dystrophinopathy, endocrine myopathy, focal muscular atrophy, glycogen storage disease type II, Guillain-Barré syndrome, hereditary spastic paraplegia, inclusion body myositis (IBM), Isaac syndrome, Kearns-Sayre syndrome, Kennedy disease, or Lambert-Eaton myasthenia.

103. The method of any one of claims 99, 101, and 102, wherein the condition is selected from the group consisting of neuropathy, metabolic myopathy, metabolic neuropathy, mitochondrial myopathy, motor neuron disease, multiple sclerosis, myasthenia gravis, myotonic dystrophy, necrotizing myopathy, neuromyotonia, Friedreich's ataxia neuropathy, nutritional neuropathy, peripheral neuropathy, polymyositis, primary lateral sclerosis, Schwartz-Jampel syndrome, small fiber neuropathy, spinal and bulbar muscular atrophy, spinal muscular atrophy (SMA), spinal muscular atrophy with respiratory distress syndrome type 1, stiff person syndrome, toxic neuropathy, and Troyer's syndrome.

104. 104. The method of claim 103, wherein the neuromuscular disease is a muscular dystrophy.

105. 105. The method of claim 104, wherein the muscular dystrophy is Duchenne muscular dystrophy (DMD), facioscapulohumeral muscular dystrophy (FSHD), Becker muscular dystrophy (BMD), myotonic dystrophy (DM), congenital muscular dystrophy, limb-girdle muscular dystrophy (LGMD), distal muscular dystrophy (DD), oculopharyngeal muscular dystrophy (OPMD), or Emery-Dreifuss muscular dystrophy (EDMD).

106. 106. The method of claim 105, wherein the muscular dystrophy is DMD.

107. 106. The method of claim 105, wherein the muscular dystrophy is FSHD.

108. 106. The method of claim 105, wherein the muscular dystrophy is a congenital muscular dystrophy.

109. 109. The method of claim 108, wherein the congenital muscular dystrophy is congenital muscular dystrophy type 1A (MDC1A), congenital muscular dystrophy type 1C (MDC1C), congenital muscular dystrophy type 1D (MDC1D), congenital muscular dystrophy type 1B (MDC1B), Fukuyama congenital muscular dystrophy (FCMD), muscle-eye-brain disease (MEB), Walker-Warburg syndrome (WWS), spondylotonic muscular dystrophy (RSMD1), Ullrich congenital muscular dystrophy (UCMD), or a muscular dystrophy associated with a mutation in integrin alpha 7, integrin alpha 9, docking protein 7, laminin A / C, SECIS binding protein 2, or choline kinase beta.

110. 110. The method of claim 109, wherein the congenital muscular dystrophy is MDC1A.

111. 104. The method of claim 103, wherein the neuromuscular disease is CMT.

112. 104. The method of claim 103, wherein the neuromuscular disease is ALS.

113. 104. The method of claim 103, wherein the neuromuscular disease is SMA.

114. 104. The method of claim 103, wherein the neuromuscular disease is IBM.

115. 94. A method of treating a subject having or at risk of developing disuse atrophy, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

116. 94. A method of treating a subject having or at risk of developing treatment-associated muscle loss or atrophy, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

117. 117. The method of any one of claims 99, 101, and 116, wherein the treatment is glucocorticoid treatment, FGF-21 treatment, GLP-1 treatment, bariatric surgery, cancer treatment, or treatment of obesity or type 2 diabetes.

118. 94. A method of treating a subject having or at risk of developing hypotonia, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

119. 96. A method of treating a subject having or at risk of developing hypoxia-associated muscle loss or atrophy, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

120. 94. A method of treating a subject having or at risk of developing burn-related muscle loss or atrophy, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

121. 94. A method for treating a subject having or at risk of developing sarcopenia, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

122. 94. A method of treating a subject having or at risk of developing cachexia, comprising administering to the subject a therapeutically effective amount of a polypeptide according to any one of claims 1 to 91, a nucleic acid molecule according to claim 92, a vector according to claim 93, or a pharmaceutical composition according to claim 96.

123. 123. The method of any one of claims 99, 101, and 122, wherein the cachexia is cancer cachexia, HIV-associated cachexia, cardiac cachexia, cachexia associated with chronic kidney disease, or pulmonary cachexia.

124. 124. The method of any one of claims 97 to 123, wherein the method increases muscle mass.

125. 125. The method of any one of claims 97 to 124, wherein the method increases lean body mass.

126. 94. A method of increasing bone mineral density in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

127. 94. A method of reducing bone resorption in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

128. 94. A method of increasing bone formation in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

129. 94. A method of increasing bone strength in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

130. 94. A method for reducing the risk or incidence of fracture in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

131. 131. The method of any one of claims 126-130, wherein the subject has or is at risk of developing osteoporosis, osteopenia, osteopetrosis, bone fracture, bone cancer or cancer metastasis-associated bone loss, Paget's disease, renal osteodystrophy, treatment-associated bone loss, osteogenesis imperfecta, neuromuscular disease-associated bone loss, burns-associated bone loss, anorexia-associated bone loss, diet-associated bone loss, obesity treatment-associated bone loss, reduced gravity-associated bone loss, or immobility-associated bone loss.

132. A method for affecting the transmission of myostatin, activin A, activin B, and / or BMP9 signals in a subject having or at risk of developing a disease or condition involving bone damage, the method comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

133. 133. The method of claim 132, wherein the disease or condition is osteoporosis, osteopenia, osteopetrosis, bone fracture, bone loss associated with bone cancer or cancer metastasis, Paget's disease, renal osteodystrophy, treatment-related bone loss, osteogenesis imperfecta, bone loss associated with neuromuscular disease, bone loss due to burns, bone loss associated with anorexia, diet-related bone loss, bone loss associated with the treatment of obesity, bone loss associated with reduced gravity, or bone loss associated with immobility.

134. 94. A method of treating a subject having or at risk of developing a bone disease, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

135. 135. The method of claim 134, wherein the bone disease is osteoporosis, osteopenia, osteopetrosis, bone fracture, bone loss associated with bone cancer or cancer metastasis, Paget's disease, renal osteodystrophy, treatment-related bone loss, osteogenesis imperfecta, bone loss associated with neuromuscular disease, bone loss due to burns, bone loss associated with anorexia, diet-related bone loss, bone loss associated with the treatment of obesity, bone loss associated with reduced gravity, or bone loss associated with immobility.

136. 94. A method of treating a subject having or at risk of developing osteoporosis, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 11 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

137. 94. A method of treating a subject having or at risk of developing osteogenesis imperfecta, comprising administering to the subject a therapeutically effective amount of a polypeptide according to any one of claims 1 to 91, a nucleic acid molecule according to claim 92, a vector according to claim 93, or a pharmaceutical composition according to claim 96.

138. 94. A method of treating a subject having or at risk of developing osteopenia, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

139. 94. A method of treating a subject having or at risk of developing bone loss associated with bone cancer or cancer metastasis, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

140. 94. A method of treating a subject having or at risk of developing Paget's disease, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

141. 94. A method of treating a subject having or at risk of developing renal osteodystrophy, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

142. 94. A method of treating a subject having or at risk of developing treatment-associated bone loss, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

143. 94. A method of treating a subject having or at risk of developing diet-related bone loss, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

144. 94. A method of treating a subject having or at risk of developing bone loss associated with low gravity, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

145. 94. A method of treating a subject having or at risk of developing immobility-associated bone loss, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

146. 94. A method of treating a subject having or at risk of developing bone loss associated with a neuromuscular disease, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

147. 94. A method of treating a subject having or at risk of developing bone loss due to burns, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

148. 94. A method of treating a subject having or at risk of developing bone loss associated with anorexia, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

149. 137. The method of any one of claims 131, 133, 135, and 136, wherein the osteoporosis is primary osteoporosis.

150. 150. The method of claim 149, wherein the primary osteoporosis is age-related osteoporosis or hormone-related osteoporosis.

151. 137. The method of any one of claims 131, 133, 135, and 136, wherein the osteoporosis is secondary osteoporosis.

152. 152. The method of claim 151, wherein the secondary osteoporosis is immobility-induced osteoporosis or glucocorticoid-induced osteoporosis.

153. 140. The method of any one of claims 131, 133, 135, and 139, wherein the cancer is multiple myeloma.

154. 143. The method of any one of claims 131, 133, 135, and 142, wherein the treatment is FGF-21 treatment, GLP-1 treatment, cancer treatment, bariatric surgery, androgen or estrogen deprivation therapy, or treatment of obesity or type 2 diabetes.

155. 144. The method of any one of claims 131, 133, 135, and 143, wherein the diet-related bone loss is rickets.

156. 156. The method of any one of claims 126 to 155, wherein the subject is at risk of fracture.

157. 157. The method of any one of claims 126 to 156, wherein the method increases bone formation in the subject.

158. 158. The method of any one of claims 126 to 157, wherein said method reduces bone resorption in said subject.

159. 159. The method of any one of claims 126 to 158, wherein the method increases osteoblast activity or osteoblast formation.

160. 160. The method of any one of claims 126 to 159, wherein said method reduces osteoclast activity or osteoclast formation.

161. 161. The method of any one of claims 126 to 160, wherein the method reduces the risk or incidence of bone fracture.

162. 162. The method of any one of claims 126 to 161, wherein the method increases bone strength.

163. 163. The method of any one of claims 126 to 162, wherein the bone is cortical bone.

164. 163. The method of any one of claims 126 to 162, wherein the bone is cancellous bone.

165. 94. A method of reducing or preventing fibrosis in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide according to any one of claims 1 to 91, a nucleic acid molecule according to claim 92, a vector according to claim 93, or a pharmaceutical composition according to claim 96.

166. 94. A method of treating a subject having or at risk of developing fibrosis, comprising administering to the subject a therapeutically effective amount of a polypeptide according to any one of claims 1 to 91, a nucleic acid molecule according to claim 92, a vector according to claim 93, or a pharmaceutical composition according to claim 96.

167. 94. A method of slowing or inhibiting the progression of fibrosis in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

168. A method for affecting the transmission of myostatin, activin A, activin B, and / or BMP9 signals in a subject having or at risk of developing fibrosis or a disease or condition involving fibrosis, the method comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

169. 169. The method of any one of claims 165 to 168, wherein the fibrosis is chemotherapy-induced fibrosis, radiation-induced fibrosis, pulmonary fibrosis, liver fibrosis, renal fibrosis, corneal fibrosis, cardiac fibrosis, bone marrow fibrosis, myelofibrosis, mediastinal fibrosis, retroperitoneal fibrosis, osteoarthrofibrosis, arthrofibrosis, tissue fibrosis, tumor stroma, desmoplastic tumor, surgical adhesion, hypertrophic scar, or keloid.

170. 170. The method of claim 169, wherein the tissue fibrosis is fibrosis affecting a tissue selected from the group consisting of muscle tissue, skin epidermis, skin dermis, tendon, cartilage, pancreatic tissue, uterine tissue, nervous tissue, testes, ovaries, adrenal glands, arteries, veins, bone marrow, colon, small intestine, large intestine, biliary tract, and intestine.

171. 169. The method of any one of claims 165 to 168, wherein the fibrosis is fibrosis associated with a wound, a burn, hepatitis B or C infection, fatty liver disease, schistosome infection, kidney disease, chronic kidney disease, heart disease, macular degeneration, retinal or vitreoretinopathy, Crohn's disease, systemic or localized scleroderma, atherosclerosis, or restenosis.

172. 172. The method of any one of claims 165 to 171, wherein said method improves the function of a fibrotic tissue or organ.

173. 94. A method of increasing red blood cell levels in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

174. 94. A method of promoting or increasing red blood cell production in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

175. 94. A method for increasing the maturation and / or differentiation of erythroid progenitor cells in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

176. 94. A method of increasing hemoglobin levels in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

177. 94. A method of increasing hematocrit in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

178. 94. A method for increasing reticulocytes in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

179. 94. A method of increasing the maturation of late stage erythroid progenitor cells in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

180. 94. A method for mobilizing early stage progenitor cells to the erythroid lineage in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

181. 94. A method of increasing the number of early stage erythroid progenitor or precursor cells in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

182. 94. A method of promoting the progression of erythroid precursor or progenitor cells through erythropoiesis in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

183. 183. The method of any one of claims 173 to 182, wherein the subject has or is at risk of developing anemia or blood loss.

184. A method for affecting the transmission of myostatin, activin A, activin B, and / or BMP9 signals in a subject having or at risk of developing a disease or condition involving low red blood cell levels or low hemoglobin levels, the method comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

185. 185. The method of claim 184, wherein the disease or condition is anemia or blood loss.

186. 186. The method of claim 183 or 185, wherein the anemia or blood loss is associated with cancer, cancer treatment, myelofibrosis treatment, chronic kidney disease, acute kidney disease or renal failure, chronic kidney disease or renal failure, myelodysplastic syndrome, thalassemia, nutritional deficiency, ineffective hematopoiesis, adverse reaction to a drug, inflammatory or autoimmune disease, splenomegaly, porphyria, vasculitis, hemolysis, bone marrow deficiency, bone marrow transplant, myelofibrosis, diabetes, acute liver disease, chronic liver disease, acute bleeding, chronic bleeding, infection, hemoglobinopathies, drug use, alcohol abuse, Churg-Strauss syndrome, Felty syndrome, Pearson syndrome, dyskeratosis congenita, graft-versus-host disease, hematopoietic stem cell transplant, myelofibrosis, pancytopenia, pure red cell aplasia, Schönlein-Henoch purpura, Schwachman syndrome, elderly, transfusion contraindications, surgery, trauma, wound, ulcer, urinary tract bleeding, gastrointestinal bleeding, frequent blood donations, or heavy menstrual bleeding.

187. 94. A method of treating a subject having or at risk of developing anemia, comprising administering to the subject a therapeutically effective amount of a polypeptide according to any one of claims 1 to 91, a nucleic acid molecule according to claim 92, a vector according to claim 93, or a pharmaceutical composition according to claim 96.

188. 188. The method of claim 187, wherein the anemia is associated with cancer, cancer treatment, myelofibrosis treatment, chronic kidney disease, acute kidney disease or renal failure, chronic kidney disease or renal failure, myelodysplastic syndrome, thalassemia, nutritional deficiency, adverse reaction to a drug, ineffective hematopoiesis, inflammatory or autoimmune disease, splenomegaly, porphyria, vasculitis, hemolysis, bone marrow deficiency, bone marrow transplant, myelofibrosis, diabetes, acute liver disease, chronic liver disease, acute bleeding, chronic bleeding, infection, hemoglobinopathies, drug use, alcohol abuse, Churg-Strauss syndrome, Felty syndrome, Pearson syndrome, dyskeratosis congenita, graft-versus-host disease, hematopoietic stem cell transplant, myelofibrosis, pancytopenia, pure red cell aplasia, Schönlein-Henoch purpura, Schwachman-Diamond syndrome, elderly, transfusion contraindications, surgery, trauma, wound, ulcer, urinary tract bleeding, gastrointestinal bleeding, frequent blood donations, or heavy menstrual bleeding.

189. The method of claim 186 or 188, wherein the anemia is associated with a myelodysplastic syndrome.

190. 189. The method of claim 186 or 188, wherein the anemia is associated with myelofibrosis.

191. 191. The method of any one of claims 183 and 185 to 190, wherein the anemia is aplastic anemia, iron deficiency anemia, vitamin deficiency anemia, anemia of chronic disease, anemia associated with bone marrow disease, hemolytic anemia, sickle cell anemia, microcytic anemia, hypochromic anemia, congenital dyserythroid anemia, sideroblastic anemia, Diamond-Blackfan anemia, Fanconi anemia, or refractory anemia with excess blasts.

192. 192. The method of claim 191, wherein the sideroblastic anemia is acquired sideroblastic anemia or congenital sideroblastic anemia.

193. 193. The method of claim 192, wherein the congenital sideroblastic anemia is associated with a mutation in ALAS2, SLC25A38, FECH, GLRX5, HSPA9, HSCB, SLC25A38, or ABCB7.

194. 193. The method of claim 192, wherein the congenital sideroblastic anemia is associated with a mutation in PUS1, YARS2, LARS2, TRNT1, MT-ATP6, NDUFB11, or SLC19A2, or an mtDNA mutation.

195. 195. The method of any one of claims 173 to 194, wherein the method increases red blood cell production, red blood cell count, hematocrit, hemoglobin levels, erythroid progenitor differentiation and / or maturation, maturation of late stage erythroid progenitors, mobilization of early stage progenitors into the erythroid lineage, proerythroblast count, early stage erythroid precursor or progenitor cell count, progression of erythroid precursor or progenitor cells through erythropoiesis, and / or reticulocyte count.

196. 196. The method of any one of claims 173 to 195, wherein said method reduces the accumulation of erythroid progenitor cells.

197. 94. A method of increasing platelet levels in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

198. 94. A method for promoting or increasing platelet production in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

199. 94. A method for increasing or inducing megakaryocyte differentiation and / or maturation in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a polypeptide according to any one of claims 1 to 91, a nucleic acid molecule according to claim 92, a vector according to claim 93, or a pharmaceutical composition according to claim 96.

200. 200. The method of any one of claims 197 to 199, wherein the subject has or is at risk of developing thrombocytopenia.

201. A method for affecting the transmission of myostatin, activin A, activin B, and / or BMP9 signals in a subject having or at risk of developing a disease or condition involving low platelet levels, the method comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

202. 202. The method of claim 201, wherein the disease or condition is thrombocytopenia.

203. 94. A method of treating a subject having or at risk of developing thrombocytopenia, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

204. 94. A method of promoting platelet production, comprising contacting megakaryocytes with a polypeptide according to any one of claims 1 to 91, a nucleic acid molecule according to claim 92, or a vector according to claim 93, in an amount effective to promote platelet production.

205. 205. The method of claim 204, wherein said contacting is in vitro.

206. 206. A method of treating a subject having or at risk of developing thrombocytopenia, comprising administering to the subject platelets produced by the method of claim 204 or 205.

207. 207. The method of any one of claims 200, 202, 203, and 206, wherein the thrombocytopenia is associated with bone marrow deficiency, myelodysplastic syndrome, bone marrow transplantation, myelofibrosis, treatment of myelofibrosis, ineffective hematopoiesis, Gaucher disease, aplastic anemia, Fanconi anemia, Diamond-Blackfan anemia, Shwachman-Diamond syndrome, excessive alcohol consumption, cirrhosis of the liver, cancer, autoimmune disease, viral infection, bacterial infection, splenomegaly, vitamin deficiency, cancer treatment, thrombotic thrombocytopenic purpura, idiopathic thrombocytopenic purpura, disseminated intravascular coagulation, hemolytic uremic syndrome, paroxysmal nocturnal hemoglobinuria, drug-induced platelet reduction, transfusion-induced platelet dilution, hematopoietic stem cell transplantation, acquired amegakaryocytic thrombocytopenia, Pearson syndrome, dyskeratosis congenita, or a transfusion contraindication.

208. 208. The method of any one of claims 200, 202, 203, 206, and 207, wherein the thrombocytopenia is associated with a myelodysplastic syndrome.

209. 208. The method of any one of claims 200, 202, 203, 206, and 207, wherein the thrombocytopenia is associated with myelofibrosis.

210. 207. The method of any one of claims 200, 202, 203, and 206, wherein the thrombocytopenia is familial thrombocytopenia.

211. The method of claim 210, wherein the familial thrombocytopenia is May-Hegglin anomaly, Sebastian syndrome, Fechtner syndrome, Epstein syndrome, Wiskott-Aldrich syndrome, congenital amegakaryocytic thrombocytopenia, platelet storage pool deficiency, Hermansky-Pudlak syndrome, Bernard-Soulier syndrome, von Willebrand disease type 2B, ANKRD26-associated thrombocytopenia, thrombocytopenia-radial deficiency syndrome, familial platelet disorder associated with myeloid malignancies (FPD / AML), thrombocytopenia associated with mutations in filamin-A, or thrombocytopenia associated with mutations in GATA-1.

212. 207. The method of any one of claims 200, 202, 203, and 206, wherein the thrombocytopenia is immune thrombocytopenia.

213. 213. The method of any one of claims 197 to 212, wherein said method increases platelet count, platelet production, and / or megakaryocyte differentiation and / or maturation.

214. 214. The method of any one of claims 197 to 213, wherein the method reduces the accumulation of platelet precursor cells.

215. 96. A method of increasing neutrophil levels in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

216. 94. A method of promoting or increasing neutrophil production in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

217. 217. The method of claim 215 or 216, wherein the subject has or is at risk of developing neutropenia.

218. A method for affecting the transmission of myostatin, activin A, activin B, and / or BMP9 signals in a subject having or at risk of developing a disease or condition involving low neutrophil levels, the method comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

219. 219. The method of claim 218, wherein the disease or condition is neutropenia.

220. 94. A method of treating a subject having or at risk of developing neutropenia, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

221. The method of any one of claims 217, 219, and 220, wherein the neutropenia is associated with bone marrow deficiency, myelodysplastic syndrome, bone marrow transplantation, myelofibrosis, ineffective hematopoiesis, aplastic anemia, Fanconi anemia, Diamond-Blackfan anemia, Shwachman-Diamond syndrome, paroxysmal nocturnal hemoglobinuria, Pearson syndrome, dyskeratosis congenita, cancer, vitamin deficiency, splenomegaly, autoimmune disease, viral infection, bacterial infection, cancer treatment, drug-induced neutropenia, inflammation, hematopoietic stem cell transplantation, or transfusion contraindication.

222. 222. The method of any one of claims 217 and 219-221, wherein the neutropenia is associated with a myelodysplastic syndrome.

223. 222. The method of any one of claims 217 and 219-221, wherein the neutropenia is associated with myelofibrosis.

224. 221. The method of any one of claims 217, 219, and 220, wherein the neutropenia is associated with chronic idiopathic neutropenia.

225. 221. The method of any one of claims 217, 219, and 220, wherein the neutropenia is familial neutropenia.

226. 226. The method of claim 225, wherein the familial neutropenia is cyclic neutropenia, chronic benign neutropenia, or severe congenital neutropenia.

227. 227. The method of any one of claims 215 to 226, wherein the method increases neutrophil numbers, neutrophil production, and / or differentiation and / or maturation of progenitor cells into neutrophils.

228. 94. A method of treating a subject having or at risk of developing a myelodysplastic syndrome, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

229. The method of any one of claims 186, 188, 189, 207, 208, 221, 222, and 228, wherein the myelodysplastic syndrome is myelodysplastic syndrome with unilineage dysplasia, myelodysplastic syndrome with multilineage dysplasia, myelodysplastic syndrome with ring sideroblasts, myelodysplastic syndrome with isolated del(5q), myelodysplastic syndrome with excess blasts, myelodysplastic syndrome, unclassifiable type, or myelodysplastic syndrome / myeloproliferative neoplasm with ring sideroblasts and thrombocytosis.

230. 229. The method of any one of claims 186, 188, 189, 207, 208, 221, 222, 228, and 229, wherein the myelodysplastic syndrome is very low risk, low risk, or intermediate risk myelodysplastic syndrome.

231. The method of any one of claims 186, 188, 189, 207, 208, 221, 222, and 228-230, wherein the myelodysplastic syndrome is ring sideroblast-positive myelodysplastic syndrome.

232. 230. The method of any one of claims 186, 188, 189, 207, 208, 221, 222, and 228-230, wherein the myelodysplastic syndrome is non-ring sideroblastic myelodysplastic syndrome.

233. 232. The method of any one of claims 186, 188, 189, 207, 208, 221, 222, and 228-232, wherein the myelodysplastic syndrome is associated with a defect in terminal maturation of hematopoietic cells.

234. 232. The method of any one of claims 186, 188, 189, 207, 208, 221, 222, and 228-232, wherein the myelodysplastic syndrome is associated with an early stage defect in hematopoiesis.

235. 235. The method of any one of claims 186, 188, 189, 207, 208, 221, 222, and 228-234, wherein the myelodysplastic syndrome is associated with hypocellular bone marrow.

236. 94. A method of treating a subject having or at risk of developing myelofibrosis, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

237. The method of any one of claims 173 to 236, wherein the subject does not respond well to treatment with erythropoietin (EPO), is susceptible to adverse effects of EPO, or does not respond well to treatment with an erythrocyte maturation agent.

238. The method of any one of claims 173 to 237, wherein the subject has previously been treated with an erythropoiesis-stimulating agent (ESA).

239. The method of any one of claims 173 to 237, wherein the subject has not been previously treated with an ESA.

240. 240. The method of any one of claims 173 to 239, wherein the subject has a low transfusion burden.

241. 240. The method of any one of claims 173 to 239, wherein the subject has a high transfusion burden.

242. 242. The method of any one of claims 173 to 241, wherein said method reduces the subject's need for blood transfusions.

243. 94. A method for preventing pulmonary hypertension (PH) in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

244. 94. A method of slowing or inhibiting the progression of PH in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

245. 94. A method of treating a subject having or at risk of developing PH, comprising administering to the subject a therapeutically effective amount of a polypeptide of any one of claims 1 to 91, a nucleic acid molecule of claim 92, a vector of claim 93, or a pharmaceutical composition of claim 96.

246. A method for affecting the transmission of myostatin, activin A, activin B, and / or BMP9 signals in a subject having or at risk of developing PH, the method comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

247. The method of any one of claims 243 to 246, wherein the PH is pulmonary arterial hypertension (PAH).

248. The method of claim 247, wherein the PAH is idiopathic PAH.

249. The method of claim 247, wherein the PAH is hereditary PAH.

250. The method of claim 247, wherein the PAH is associated with HIV infection, schistosomiasis, cirrhosis, congenital heart abnormalities, portal hypertension, pulmonary veno-occlusive disease, pulmonary capillary hemangiomatosis, connective tissue disorders, autoimmune disorders, or drug use or abuse.

251. The method of any one of claims 243 to 246, wherein the PH is intravenous PH.

252. The method of claim 251, wherein the venous PH is associated with left ventricular systolic dysfunction, left ventricular diastolic dysfunction, valvular heart disease, congenital cardiomyopathy, or congenital or acquired pulmonary vein stenosis.

253. 247. The method of any one of claims 243 to 246, wherein the pH is hypoxic pH.

254. The method of claim 253, wherein the hypoxic PH is associated with chronic obstructive pulmonary disease, interstitial lung disease, sleep-disordered breathing, pulmonary fibrosis, alveolar hypoventilation disorder, chronic exposure to high altitude, or developmental abnormalities.

255. The method of any one of claims 243 to 246, wherein the PH is thromboembolic PH.

256. 256. The method of claim 255, wherein the thromboembolic PH is associated with chronic thromboembolic pulmonary hypertension, pulmonary embolism, angiosarcoma, arteritis, congenital pulmonary stenosis, or parasitic infection.

257. The method of any one of claims 243 to 246, wherein the pH is other pH.

258. The method of claim 257, wherein the other PH is associated with a blood disorder, a systemic disease, a metabolic disorder, pulmonary neoplastic thrombotic microangiopathy, fibrosing mediastinitis, chronic renal failure, or segmental pulmonary hypertension.

259. 259. The method of any one of claims 243-258, wherein the method reduces the frequency or severity of one or more symptoms of PH.

260. 260. The method of any one of claims 243 to 259, wherein the method reduces pulmonary vascular remodeling or vascular remodeling in the subject's heart.

261. 261. The method of any one of claims 243 to 260, wherein said method reduces right ventricular hypertrophy.

262. 262. The method of any one of claims 243 to 261, wherein said method reduces pulmonary vascular resistance.

263. 263. The method of any one of claims 243 to 262, wherein said method improves performance in a 6-minute walk test.

264. 264. The method of any one of claims 243-263, wherein said method reduces bone loss.

265. 265. The method of any one of claims 243 to 264, wherein the method reduces pulmonary artery muscularization or pulmonary artery wall thickening.

266. 266. The method of any one of claims 243 to 265, wherein the method reduces right ventricular decompensation.

267. 94. A method for reducing body fat in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polypeptide according to any one of claims 1 to 91, a nucleic acid molecule according to claim 92, a vector according to claim 93, or a pharmaceutical composition according to claim 96.

268. 94. A method of reducing body weight in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

269. 94. A method of reducing blood glucose in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

270. 94. A method of increasing insulin sensitivity in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of the polypeptide of any one of claims 1 to 91, the nucleic acid molecule of claim 92, the vector of claim 93, or the pharmaceutical composition of claim 96.

271. 271. The method of any one of claims 267 to 270, wherein the subject has or is at risk of developing a metabolic disease.

272. A method for affecting the transmission of myostatin, activin A, activin B, and / or BMP9 signals in a subject having or at risk of developing a metabolic disease, the method comprising administering to the subject a therapeutically effective amount of a polypeptide described in any one of claims 1 to 91, a nucleic acid molecule described in claim 92, a vector described in claim 93, or a pharmaceutical composition described in claim 96.

273. 94. A method for treating and / or preventing a metabolic disease in a subject, the method comprising administering to the subject a therapeutically effective amount of a polypeptide according to any one of claims 11 to 91, a nucleic acid molecule according to claim 92, a vector according to claim 93, or a pharmaceutical composition according to claim 96.

274. The method of any one of claims 271 to 273, wherein the metabolic disease is an age-related metabolic disease.

275. 274. The method of any one of claims 271 to 273, wherein the metabolic disorder is a therapeutically relevant metabolic disorder.

276. 276. The method of claim 275, wherein the treatment is treatment with a glucocorticoid, a selective serotonin reuptake inhibitor, a serotonin-norepinephrine reuptake inhibitor, a tricyclic antidepressant, a mood stabilizer, an antipsychotic, or a diabetic drug.

277. 277. The method of any one of claims 271 to 276, wherein the metabolic disease is obesity, type 1 diabetes, or type 2 diabetes.

278. 278. The method of claim 277, wherein the metabolic disease is obesity.

279. 278. The method of claim 277, wherein the metabolic disease is type 1 diabetes.

280. 278. The method of claim 277, wherein the metabolic disease is type 2 diabetes.

281. 281. The method of any one of claims 267 to 280, wherein said method reduces the body weight and / or percentage of weight gain in said subject.

282. 282. The method of any one of claims 267 to 281, wherein said method reduces the amount of body fat and / or percentage of body fat gain in said subject.

283. 283. The method of any one of claims 267 to 282, wherein the method does not affect the subject's appetite for food intake.

284. 284. The method of any one of claims 267 to 283, wherein said method reduces adiposity in said subject.

285. 285. The method of any one of claims 267 to 284, wherein said method reduces the weight of the epididymal and perirenal fat pads of said subject.

286. 286. The method of any one of claims 267 to 285, wherein the method reduces the amount of subcutaneous fat, visceral fat, and / or liver fat in the subject.

287. 287. The method of any one of claims 267 to 286, wherein said method reduces fasting insulin levels in said subject.

288. 288. The method of any one of claims 267 to 287, wherein the method reduces blood glucose levels in the subject.

289. 289. The method of any one of claims 267-288, wherein said method increases insulin sensitivity in said subject.

290. 290. The method of any one of claims 267 to 289, wherein said method increases the rate of glucose clearance in said subject.

291. 291. The method of any one of claims 267 to 290, wherein said method improves the serum lipid profile of said subject.

292. 292. The method of any one of claims 267 to 291, wherein said method delays, reduces or eliminates the need for insulin therapy.

293. 292. The method of any one of claims 267 to 291, wherein said method does not result in a loss of lean body mass.

294. 294. The method of any one of claims 97 to 293, wherein the method reduces or inhibits binding of activin A, activin B, and / or myostatin to their receptors.

295. The method of any one of claims 97-125 and 294, wherein the polypeptide, nucleic acid, vector, or pharmaceutical composition is administered in an amount sufficient to increase muscle mass and / or strength, increase lean body mass, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or reduce or inhibit binding of activin A, activin B, and / or myostatin to their receptors.

296. The method of any one of claims 126-164 and 294, wherein the polypeptide, nucleic acid, vector, or pharmaceutical composition is administered in an amount sufficient to increase bone density, decrease bone resorption, decrease the rate of bone resorption, increase bone formation, increase the rate of bone formation, decrease osteoclast activity, increase osteoblast activity, increase bone strength, decrease the risk or occurrence of fracture, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or decrease or inhibit binding of activin A, activin B, and / or myostatin to their receptors.

297. The method of any one of claims 165-172 and 294, wherein the polypeptide, nucleic acid, vector or pharmaceutical composition is administered in an amount sufficient to reduce fibrosis, prevent fibrosis, reduce the risk of developing fibrosis, delay the onset of fibrosis, slow or inhibit the progression of fibrosis, treat fibrosis, alleviate one or more symptoms of fibrosis, improve fibrotic tissue or organ function, affect myostatin, activin A, activin B and / or BMP9 signaling in the subject, or reduce or inhibit binding of activin A, activin B and / or myostatin to their receptors.

298. the polypeptide, nucleic acid, vector, or pharmaceutical composition increases red blood cell levels, increases hemoglobin levels, increases red blood cell production, increases red blood cell count, increases hematocrit, reduces transfusion requirements, increases maturation and / or differentiation of erythroid progenitors, increases maturation of late stage erythroid progenitors, mobilizes early stage progenitors to the erythroid lineage, increases proerythroblast counts, increases reticulocyte counts, reduces accumulation of erythroid progenitors, The method of any one of claims 173-196, 228-242, and 294, administered in an amount sufficient to increase the number of early stage erythroid precursor or progenitor cells, promote the progression of erythroid precursor or progenitor cells through erythropoiesis, treat anemia, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or reduce or inhibit binding of activin A, activin B, and / or myostatin to their receptors.

299. The method of any one of claims 197-214, 228-242, and 294, wherein the polypeptide, nucleic acid, vector, or pharmaceutical composition is administered in an amount sufficient to increase platelet levels, increase platelet production, increase platelet count, increase or induce megakaryocyte differentiation and / or maturation, decrease platelet progenitor cell accumulation, treat thrombocytopenia, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or decrease or inhibit binding of activin A, activin B, and / or myostatin to their receptors.

300. The method of any one of claims 215-242 and 294, wherein the polypeptide, nucleic acid, vector, or pharmaceutical composition is administered in an amount sufficient to increase neutrophil levels, increase neutrophil production, increase neutrophil numbers, increase or induce differentiation and / or maturation of progenitor cells into neutrophils, treat neutropenia, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or reduce or inhibit binding of activin A, activin B, and / or myostatin to their receptors.

301. The method of any one of claims 243-266 and 294, wherein the polypeptide, nucleic acid, vector, or pharmaceutical composition is administered in an amount sufficient to prevent PH, reduce the risk of developing PH, reduce the severity or frequency of one or more symptoms of PH, delay the onset of PH, slow or inhibit the progression of PH, treat PH, reduce pulmonary vascular remodeling, reduce vascular remodeling in the heart, reduce right ventricular hypertrophy, reduce pulmonary vascular resistance, improve performance in a 6-minute walk test, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, or reduce or inhibit binding of activin A, activin B, and / or myostatin to their endogenous receptors.

302. 295. The method of any one of claims 267-294, wherein the polypeptide, nucleic acid, vector, or pharmaceutical composition is administered in an amount sufficient to reduce body fat, reduce subcutaneous fat mass, reduce visceral fat and / or liver fat mass, reduce adiposity, reduce epididymal and perirenal fat pad weight, reduce body fat percentage, reduce body weight, reduce percentage of weight gain, reduce fasting insulin levels, reduce blood glucose levels, increase insulin sensitivity, affect myostatin, activin A, activin B, and / or BMP9 signaling in the subject, reduce adipocyte proliferation, reduce or inhibit binding of activin A, activin B, and / or myostatin to their receptors, reduce LDL, reduce triglycerides, improve serum lipid profile, modulate insulin biosynthesis and / or secretion from beta cells, delay, postpone, or reduce the need for insulin, or increase glucose clearance.

303. 303. The method of any one of claims 97 to 302, wherein the method does not cause vascular complications in the subject.

304. 304. The method of claim 303, wherein the method does not increase vascular permeability or leakage.

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