Methods for treating obesity, diabetes and liver dysfunction

Abstract

The present disclosure relates to compositions and methods for improving glucose control, increasing lean body mass, reducing fat mass, treating obesity, diabetes, and / or treating liver dysfunction in a subject, more particularly to compositions and uses comprising a GDF-8 inhibitor and a GLP-1 agonist, and to compositions and uses comprising a GDF-8 inhibitor, an activin A inhibitor, and a GLP-1 agonist.

Description

[Technical Field]

[0001] Reference to sequence table XML This application contains a Sequence Listing that has been submitted electronically in XML format. This Sequence Listing XML is incorporated herein by reference. The XML file, created on September 19, 2023, is named 40848_0107WOU1_SL.xml and is 24.7 kilobytes in size.

[0002] CROSS-REFERENCE TO RELATED APPLICATIONS This application is a PCT international patent application filed on September 21, 2023, claiming priority to U.S. Provisional Patent Application No. 63 / 376,582, filed on September 21, 2022, and U.S. Provisional Patent Application No. 63 / 508,458, filed on June 15, 2023, the entire contents of each of which are incorporated herein by reference.

[0003] The present disclosure relates to compositions and methods for improving glucose control, increasing lean body mass, reducing fat mass, treating obesity, diabetes, and / or treating liver dysfunction in a subject, more particularly to compositions comprising a GDF-8 inhibitor and a GLP-1 agonist and uses thereof, and to compositions comprising a GDF-8 inhibitor, an activin A inhibitor, and a GLP-1 agonist and uses thereof. [Background technology]

[0004] Obesity is a global problem affecting over one-third of the world's population. In the United States, the average obesity rate exceeds 20%. The costs of obesity-related diseases are enormous, totaling $190.2 billion, or approximately 21% of annual U.S. healthcare costs. Obesity is an epidemic characterized by chronic low-grade inflammation associated with dysfunctional (increased) fat mass. Obesity is an important underlying risk factor for developing other diseases such as heart disease, stroke, and diabetes. Even modest weight loss (5-10% of initial weight) reduces the risk of developing obesity-related diseases such as heart disease and diabetes.

[0005] Diabetes mellitus is a chronic condition characterized by high blood sugar levels and insulin resistance. If left untreated, high blood sugar levels can lead to long-term complications, including heart disease, stroke, diabetic retinopathy, and lower limb amputation. Treatment of diabetes involves controlling and reducing blood sugar levels, which includes medications such as insulin and metformin, as well as exercise and dietary modifications.

[0006] Growth differentiation factor-8 (GDF8, also known as myostatin) is a secreted ligand that belongs to the transforming growth factor-β (TGF-β) superfamily of growth factors. GDF8 plays a central role in the development and maintenance of skeletal muscle and acts as a negative regulator of muscle mass. While the phenotype of myostatin-null mice demonstrates the importance of GDF8 in controlling muscle size during development, muscle hypertrophy can also be induced in mature muscle through inhibition of GDF8 with neutralizing antibodies, decoy receptors, or other antagonists. Administration of GDF8-neutralizing antibodies has been reported to result in a 10-30% increase in muscle mass. The observed increase in muscle mass is due to an increase in fiber diameter, as opposed to muscle fiber hyperplasia (fiber number). Several studies have also reported improvements in muscle strength or performance, including twitch and tetanic force, commensurate with the increase in size. The use of a cleavage-resistant version of the GDF8 propeptide also results in an increase in muscle size. Antibodies and therapeutic methods against GDF8 are disclosed, for example, in U.S. Patent No. 5,629,999. Anti-GDF8 antibodies are also described, for example, in U.S. Patent No. 5,629,999; ... and U.S. Patent No. 5,629,999; U.S. Patent No. 5,629,999;

[0007] Activin belongs to the transforming growth factor-beta (TGF-β) superfamily and exerts a wide range of biological effects on cell proliferation, differentiation, metabolism, homeostasis, apoptosis, immune response, and tissue repair. Activin A is a disulfide-linked homodimer (two beta A chains) that binds to and activates heteromeric complexes of type I (Act RI-A and Act RI-B) and type II (Act RII-A and Act RII-B) serine-threonine kinase receptors.

[0008] Antibodies to activin A and their uses are disclosed, for example, in US Pat. Nos. 5,629,999, 5,729,945 ... and 5,729,945.

[0009] Compositions and therapeutic methods comprising anti-GDF8 antibodies and anti-activin A antibodies are disclosed, for example, in US Pat. No. 6,223,999.

[0010] One approach used to treat obesity and control blood glucose involves glucagon-like peptide (GLP)-1 receptor agonists, which target the incretin pathway. Glucagon-like peptide (GLP)-1 is a peptide hormone secreted by enteroendocrine cells in the intestinal tract. Upon oral glucose administration, GLP-1 binds to its receptor, resulting in insulin secretion and a decrease in blood glucose levels (incretin effect). However, GLP-1 is rapidly inactivated and degraded by the enzyme dipeptidyl peptidase 4 (DPP4) and has a very short half-life of 1.5 minutes. Therefore, GLP-1 receptor agonists, including long-acting derivatives of GLP-1 and fusion proteins containing GLP-1, have been investigated for diabetes control. GLP-1 analogs, fusion proteins, and GLP-1 receptor agonists are disclosed, for example, in U.S. Patent No. 6,223,669, U.S. Patent No. 6,223,669, U.S. Patent No. 6,224 ... [Prior art documents] [Patent documents]

[0011] [Patent Document 1] US$8,840,894 [Patent Document 2] U.S. Patent No. 6,096,506 [Patent Document 3] U.S. Patent No. 7,320,789 [Patent Document 4] U.S. Patent No. 7,261,893 [Patent Document 5] U.S. Patent No. 7,807,159 [Patent Document 6] U.S. Patent No. 7,888,486 [Patent Document 7] U.S. Patent No. 7,635,760 [Patent Document 8] U.S. Patent No. 7,632,499 [Patent Document 9] U.S. Patent Application Publication No. 2006 / 0263354 [Patent Document 10] U.S. Patent Application Publication No. 2007 / 0178095 [Patent Document 11] U.S. Patent Application Publication No. 2008 / 0299126 [Patent Document 12] U.S. Patent Application Publication No. 2010 / 0166764 [Patent Document 13] U.S. Patent Application Publication No. 2009 / 0148436 [Patent Document 14] International Patent Application Publication No. 2004 / 037861 [Patent Document 15] International Patent Application Publication No. 2007 / 047112 [Patent Document 16] International Patent Application Publication No. 2010 / 070094 [Patent Document 17] US8,309,082 [Patent Document 18] US9,718,881; [Patent Document 19] International Patent Application Publication No. 2008 / 031061 [Patent Document 20] US8,871,209 [Patent Document 21] US7452966 [Patent Document 22] US8389689 [Patent Document 23] US8496149 [Patent Document 24] US8497240 [Patent Document 25] US8557769 [Patent Document 26] US8883447 [Patent Document 27] US8895694 [Patent Document 28] US9409966 [Patent Document 29] US20160194371 [Patent Document 30] US20140024586 [Patent Document 31] US20140073563 [Patent Document 32] US20120148586 [Patent Document 33] US20170114115 [Patent Document 34] US20170112904 [Patent Document 35] US20160361390 [Patent Document 36] US20150313908 [Patent Document 37] US20150259416 [Patent Document 38] WO2017074715 [Patent Document 39] WO2016127887 [Patent Document 40] WO2015021871 [Patent Document 41] WO2014113357 [Patent Document 42] EP3034514 [Patent Document 43] EP2470198 [Patent Document 44] EP2373681 Summary of the Invention [Problem to be solved by the invention]

[0012] Because high fat mass is associated with serious conditions such as congestive heart failure, high blood pressure / hypertension, pulmonary embolism, osteoarthritis, lymphedema, gastroesophageal reflux disease, chronic renal failure, cancer, fatty liver disease, and even depression, there remains a need for therapies that reduce total fat mass and / or android fat mass in a subject. Additionally, there remains a need for agents that treat obesity and diabetes while also treating the associated liver problems. [Means for solving the problem]

[0013] In one aspect, the present disclosure provides a composition comprising a growth differentiation factor-8 (GDF-8) inhibitor and an incretin inhibitor. In another aspect, the present disclosure provides a composition comprising a GDF-8 inhibitor, an activin A inhibitor, and an incretin inhibitor.

[0014] In another aspect, the present disclosure provides a composition comprising a growth differentiation factor-8 (GDF-8) inhibitor and a glucagon-like peptide-1 (GLP-1) agonist. In another aspect, the present disclosure provides a composition comprising a GDF-8 inhibitor, an activin A inhibitor, and a GLP-1 agonist.

[0015] In one embodiment of the composition according to the present disclosure, the GDF-8 inhibitor is a GDF-8-specific binding protein. In another embodiment, the GDF8 inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to GDF-8. In a further embodiment, the anti-GDF8 antibody or antigen-binding fragment thereof comprises a heavy chain complementarity-determining region (HCDR) of the heavy chain variable region (HCVR) comprising SEQ ID NO: 4 and a light chain complementarity-determining region (LCDR) of the light chain variable region (LCVR) comprising SEQ ID NO: 5. In yet a further embodiment, the anti-GDF8 antibody or antigen-binding fragment thereof comprises heavy chain complementarity-determining regions (HCDR1, HCDR2, and HCDR3) comprising the amino acid sequences of SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8, respectively, and three light chain complementarity-determining regions (LCDR1, LCDR2, and LCDR3) comprising the amino acid sequences of SEQ ID NO: 9, TTS, and SEQ ID NO: 11, respectively.

[0016] In one embodiment of a composition according to the present disclosure, the activin A inhibitor is an activin A-specific binding protein. In another embodiment, the activin A inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to activin A. In a further embodiment, the anti-activin A antibody or antigen-binding fragment thereof comprises a heavy chain complementarity-determining region (HCDR) of the heavy chain variable region (HCVR) comprising SEQ ID NO: 12, and a light chain complementarity-determining region (LCDR) of the light chain variable region (LCVR) comprising SEQ ID NO: 13. In yet a further embodiment, the anti-activin A antibody or antigen-binding fragment thereof comprises heavy chain complementarity-determining regions (HCDR1, HCDR2, and HCDR3) comprising the amino acid sequences of SEQ ID NO: 14, SEQ ID NO: 15, and SEQ ID NO: 16, respectively, and three light chain complementarity-determining regions (LCDR1, LCDR2, and LCDR3) comprising the amino acid sequences of SEQ ID NO: 17, GAS, and SEQ ID NO: 19, respectively.

[0017] In one embodiment of the composition according to the present disclosure, the GLP-1 agonist is a GLP-1 receptor agonist.In another embodiment, the GLP-1 agonist is selected from the group consisting of exenatide (long-acting), dulaglutide, liraglutide, tirzepatide, and semaglutide.In a further embodiment, the GLP-1 agonist is a GLP-1 specific binding protein.In yet a further embodiment, the GLP-1 agonist is an antibody or an antigen-binding fragment thereof that specifically binds to GLP-1.

[0018] In one embodiment, a composition according to the present disclosure is for use in improving glucose control, increasing lean body mass, reducing fat mass, treating obesity, treating diabetes, and / or treating liver problems associated with increased fat mass, obesity, and / or diabetes in a subject. In another embodiment, a composition according to the present disclosure is for use in improving glucose control, increasing lean body mass, reducing fat mass, treating obesity, and / or treating diabetes in a subject without worsening liver problems associated with increased fat mass, obesity, and / or diabetes.

[0019] In one aspect, the present disclosure provides a method for improving glucose control, increasing lean body mass, reducing fat mass, reducing total cholesterol, reducing LDL cholesterol, increasing HDL cholesterol, treating obesity, treating diabetes, and / or treating liver problems associated with increased fat mass, obesity and / or diabetes in a subject, the method comprising administering to the subject a composition comprising a GDF8 inhibitor and a GLP-1 agonist.

[0020] In another aspect, the present disclosure provides a method for improving glucose control, increasing lean body mass, reducing fat mass, treating obesity, treating diabetes, and / or treating liver problems associated with increased fat mass, obesity, and / or diabetes in a subject, the method comprising administering to the subject a GDF8 inhibitor, an activin A inhibitor, and a GLP-1 agonist.

[0021] In one embodiment of the method according to the present disclosure, improving glucose control is demonstrated / assessed by reducing glycosylated hemoglobin (HbA1c). In another embodiment of the method according to the present disclosure, the GDF8 inhibitor, the GLP-1 agonist, and, if present, the activin A inhibitor are administered to the subject in a single composition. In yet another embodiment, the GDF8 inhibitor, the GLP-1 agonist, and, if present, the activin A inhibitor are administered to the subject in at least two separate compositions. In yet another embodiment, the GDF8 inhibitor, the GLP-1 agonist, and, if present, the activin A inhibitor are administered to the subject in three separate compositions.

[0022] In one embodiment of the method according to the present disclosure, the GDF-8 inhibitor is a GDF-8-specific binding protein. In another embodiment, the GDF8 inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to GDF-8. In a further embodiment, the anti-GDF8 antibody or antigen-binding fragment thereof comprises a heavy chain complementarity-determining region (HCDR) of the heavy chain variable region (HCVR) comprising SEQ ID NO: 4 and a light chain complementarity-determining region (LCDR) of the light chain variable region (LCVR) comprising SEQ ID NO: 5. In yet a further embodiment, the anti-GDF8 antibody or antigen-binding fragment thereof comprises heavy chain complementarity-determining regions (HCDR1, HCDR2, and HCDR3) comprising SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8, respectively, and three light chain complementarity-determining regions (LCDR1, LCDR2, and LCDR3) comprising SEQ ID NO: 9, TTS, and SEQ ID NO: 11, respectively.

[0023] In one embodiment of the method according to the present disclosure, the activin A inhibitor is an activin A-specific binding protein. In another embodiment, the activin A inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to activin A. In a further embodiment, the anti-activin A antibody or antigen-binding fragment thereof comprises a heavy chain complementarity-determining region (HCDR) of the heavy chain variable region (HCVR) comprising SEQ ID NO: 12, and a light chain complementarity-determining region (LCDR) of the light chain variable region (LCVR) comprising SEQ ID NO: 13. In yet a further embodiment, the anti-activin A antibody or antigen-binding fragment thereof comprises heavy chain complementarity-determining regions (HCDR1, HCDR2, and HCDR3) comprising SEQ ID NO: 14, SEQ ID NO: 15, and SEQ ID NO: 16, respectively, and three light chain complementarity-determining regions (LCDR1, LCDR2, and LCDR3) comprising SEQ ID NO: 17, GAS, and SEQ ID NO: 19, respectively.

[0024] In one embodiment of the method according to the present disclosure, the GLP-1 agonist is a GLP-1 receptor agonist.In another embodiment, the GLP-1 agonist is selected from the group consisting of exenatide (long-acting), dulaglutide, liraglutide, tirzepatide, and semaglutide.In yet another embodiment, the GLP-1 agonist is a GLP-1 specific binding protein.In yet another embodiment, the GLP-1 agonist is an antibody or an antigen-binding fragment thereof that specifically binds to GLP-1.

[0025] In one embodiment of the method according to the present disclosure, Subjects will be assessed at 12 weeks of administration of the one or more inhibitors and the agonist to determine: i) a reduction in fat mass of at least about 35%; ii) an increase in lean mass of at least about 6%; iii) a decrease in fasting glucose of at least about 15%; iv) a reduction in HbA1c of at least about 6%; v) a reduction in LDL of at least about 14%; vi) an increase of at least about 14% in LDL; vii) a reduction of at least about 35% in NEFA; and viii) at least about a 55% decrease in TG The subject exhibits at least one parameter change selected from the group consisting of: In another embodiment, the subject receives, at 12 weeks from administration of the one or more inhibitors and the agonist, i) a reduction in fasting glucose of at least about 25%; iv) a reduction in HbA1c of at least about 25%; v) a reduction in LDL of at least about 50%; vi) an increase of at least about 60% in LDL; vii) at least about a 50% reduction in NEFA; and viii) at least about a 65% reduction in TG The subject exhibits at least one parameter change selected from the group consisting of:

[0026] In certain embodiments of the compositions or methods according to the present disclosure, a single antigen-binding molecule comprises a GDF8-specific binding domain and an activin A-specific binding domain. In one embodiment of this aspect of the disclosure, the antigen-binding molecule is a bispecific antibody comprising a first variable domain that specifically binds GDF8 and a second variable domain that specifically binds activin A.

[0027] In one aspect, the present disclosure provides the use of a GDF-8 inhibitor and a GLP-1 agonist in the manufacture of a medicament for improving glucose control, increasing lean body mass, reducing fat mass, treating obesity, treating diabetes, and / or treating liver problems associated with increased fat mass, obesity, and / or diabetes in a subject. In another aspect, the present disclosure provides the use of a GDF-8 inhibitor, an activin A inhibitor, and a GLP-1 agonist in the manufacture of a medicament for improving glucose control, increasing lean body mass, reducing fat mass, treating obesity, treating diabetes, and / or treating liver problems associated with increased fat mass, obesity, and / or diabetes in a subject.

[0028] Other embodiments of the present disclosure will become apparent from consideration of the detailed description that follows. [Brief explanation of the drawings]

[0029] [Figure 1A] Figure 1A shows changes in body weight (Figure 1A), fat mass (Figure 1B), and lean mass (Figure 1C) measured by MRI over time. Semaglutide via osmotic pump: 7 μg / day. IgG4P (isotype control): 20 mg / kg. REGN1033 (α-GDF8): 10 mg / kg. REGN2477 (α-activin A): 10 mg / kg. [Figure 1B] Same as above. [Figure 1C] Same as above. [Figure 2A]Figures 2A and 2B show the percent change in muscle weight and adipose tissue weight, respectively, compared to control at the end of the study. For final muscle weight (Figure 2A), TA weight (% change vs. control) is shown in the left bar, and quadriceps weight (% change vs. control) is shown in the right bar. For final adipose tissue weight (Figure 2B), subQ fat weight (% change vs. control) is shown in the left bar, and gonadal fat weight (% change vs. control) is shown in the right bar. [Figure 2B] Same as above. [Figure 3] 3A-3C show the percent change in weight of other peripheral organs (FIG. 3A pancreas, FIG. 3B heart, FIG. 3C spleen) compared to controls at the end of the study. [Figure 4] 4A-4C show post-feeding blood glucose and insulin levels over the study period: Fig. 4A: post-feeding blood glucose, Fig. 4B: post-feeding insulin, both over time; Fig. 4C: post-feeding glucagon at day 20. [Figure 5] Figures 5A and 5B show glucose tolerance measurements (day 23) and fasting glucose (days 23 and 26). Figure 5A shows blood glucose over time. Figure 5B shows (left bar) AUC glucose, (middle bar) 4-hour fasting blood glucose, and (right bar) 6-hour fasting glucose. [Figure 6A] Figure 6A shows pancreatic weight (left bar), beta cell mass (middle bar), and alpha cell mass (right bar) determined by IHC for either insulin (beta cells) or glucagon (alpha cells). Representative images of alpha cell staining are shown in Figure 6B. [Figure 6B] Figure 6A shows pancreatic weight (left bar), beta cell mass (middle bar), and alpha cell mass (right bar) determined by IHC for either insulin (beta cells) or glucagon (alpha cells). Representative images of alpha cell staining are shown in Figure 6B. [Figure 7-1]7A-7F show measurements of circulating ALT (FIG. 7A), AST (FIG. 7B), cholesterol (Chol) (FIG. 7C), triglycerides (Trig) (FIG. 7D), and free fatty acids (NEFA) (FIG. 7E) at the end of the study, and triglyceride measurements from liver tissue (FIG. 7F). [Figure 7-2] Continued from Figure 7-1. [Figure 8] (FIG. 8A) H&E staining of liver tissue and (FIG. 8B) histological quantification of lipid droplets. [Figure 9] (FIG. 9A) Liver α-SMA (smooth muscle actin) staining as an indicator of fibrosis, and (FIG. 9B) its quantification. [Figure 10] FIG. 1 shows a timeline of an obese NHP study on the weight, hepatic, and metabolic effects of adding myostatin / activin A blockade to GLP-1R agonism. [Figure 11] Figures 11A and 11B show that adding anti-myostatin treatment to semaglutide results in greater weight loss than semaglutide monotherapy: Figure 11A shows the % change in body weight by group from day 0 (D0); Figure 11B shows the % change in body weight by group from the start of antibody dosing (day 14 (D14)). [Figure 12A] 12A-12C show that adding an anti-myostatin to semaglutide results in greater fat loss than semaglutide monotherapy, and that adding anti-activin A to anti-myostatin + semaglutide also increases lean mass. Total mass (FIG. 12A), total fat mass (FIG. 12B), and total lean mass (FIG. 12C) are shown over time for each treatment group. Numbers on the line graphs correspond to the percentage change from baseline at week 12 (W12). [Figure 12B] Same as above. [Figure 12C] Same as above. [Figure 13A]13A-C show that the triple combination of semaglutide, anti-myostatin treatment, and anti-activin A treatment showed the greatest reduction in HbA1c% after 12 weeks of treatment. Fasting glucose (FIG. 13A), HbA1c (FIG. 13B), and insulin (FIG. 13C) are shown over time for each treatment group. The numbers above each group are the % change from baseline at 12 weeks. [Figure 13B] Same as above. [Figure 13C] Same as above. [Figure 14A] 14A-14E show that the triple combination of semaglutide, anti-myostatin treatment, and anti-activin A treatment demonstrated the greatest reduction in LDL and greatest increase in HDL after 12 weeks of treatment. Total cholesterol (FIG. 14A), LDL (FIG. 14B), HDL (FIG. 14C), NEFA (FIG. 14D), and TG (FIG. 14E) are shown over time for each treatment group. The numbers above each group are the % change from baseline at 12 weeks. [Figure 14B] Same as above. [Figure 14C] Same as above. [Figure 14D] Same as above. [Figure 14E] Same as above. [Figure 15A] 15A-B show AST (FIG. 15A, top), ALT (FIG. 15A, bottom), and AST / ALT ratio (FIG. 15B) for various treatment groups over 12 weeks. [Figure 15B] Same as above. [Figure 15C] Same as above. [Figure 16A] 16A-B show total energy intake (7-day mean values) (FIG. 16A) and cumulative energy intake over time (FIG. 16B) (through 12 weeks) for various treatment groups. [Figure 16B] 16A-B show total energy intake (7-day mean values) (FIG. 16A) and cumulative energy intake over time (FIG. 16B) (through 12 weeks) for various treatment groups. [Figure 17A]17A-B show total water intake (7-day average) (FIG. 17A) and cumulative water intake over time (FIG. 17B) (through 12 weeks) for various treatment groups. [Figure 17B] 17A-B show total water intake (7-day average) (FIG. 17A) and cumulative water intake over time (FIG. 17B) (through 12 weeks) for various treatment groups. [Figure 18] FIG. 1 shows that myostatin (GDF8) and activin A blockade synergistically increase muscle mass in mice. [Figure 19A] 19A-19D show changes over time in thigh muscle volume in postmenopausal women (FIG. 19A), total lean mass in obese non-human primates (FIG. 19B), android fat mass in postmenopausal women (FIG. 19C), and total fat mass in obese non-human primates (FIG. 19D), with each group receiving a different treatment. [Figure 19B] Same as above. [Figure 19C] Same as above. [Figure 19D] Same as above. [Figure 20] 20A-B show raw energy expenditure (FIG. 20A) and energy expenditure per kg of lean mass (FIG. 20B) in obese non-human primates receiving various treatment combinations. [Figure 21A] 21A shows energy expenditure versus total lean mass for the no combination treatment groups (vehicle, semaglutide, trevoglumab + galetusumab) and for the combination treatment groups (semaglutide + trevoglumab, semaglutide + trevoglumab + galetusumab) (top for both sets of treatment groups, bottom left for the combination treatment groups, bottom right for the no combination treatment group); and energy expenditure versus percent change in lean mass from baseline for the no combination treatment groups (vehicle, semaglutide, trevoglumab + galetusumab) and for the combination treatment groups (semaglutide + trevoglumab, semaglutide + trevoglumab + galetusumab) (top for both sets of treatment groups, bottom left for the no combination treatment group, bottom right for the combination treatment group). [Figure 21B] Same as above. [Figure 22-1] 22A and 22B show measurements of HbA1c (FIG. 22A), LDL-C (FIG. 22B), ApoB (FIG. 22C), HbA1c change vs. baseline (FIG. 22D), LDL-C change vs. baseline (FIG. 22E), and ApoB change vs. baseline (FIG. 22F) in the obese non-human primate treatment groups of FIGS. 21A and 21B. [Figure 22-2] Continuation of Figure 22-1. [Figure 22-3] Continuation of Figure 22-2. [Figure 23] FIG. 10 is a schematic diagram illustrating the research design of a non-human primate study of obesity on the body weight, hepatic, and metabolic effects of adding myostatin / activin A blockade to GLP-1R agonism. DETAILED DESCRIPTION OF THE INVENTION

[0030] Before describing the present disclosure, it is to be understood that the present disclosure is not limited to the particular methods and experimental conditions described, as such methods and conditions may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, as the scope of the present disclosure will be limited only by the appended claims.

[0031] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this disclosure belongs. As used herein, the term "about," when used in reference to a specific recited numerical value, means that the value may vary by 1% or less from the recited value. For example, as used herein, the expression "about 100" includes 99 and 101, and all values ​​therebetween (e.g., 99.1, 99.2, 99.3, 99.4, etc.). Ranges can be expressed herein as "about" or "approximately" from one particular value and / or to another particular value. When such a range is expressed, another embodiment includes from one particular value and / or to the other particular value.

[0032] "Comprising" or "containing" or "including" means that at least the named compound, element, particle, or method step is present in a composition or article or method, but does not exclude the presence of other such compounds, materials, particles, or method steps, even if they have the same function as the named one.

[0033] antigen-specific binding proteins The present disclosure relates to a composition comprising an agonist and an antigen-specific binding protein.More specifically, in certain embodiments, the present disclosure provides a composition comprising a GLP-1 agonist and a GDF-8 specific binding protein, and a composition comprising a GLP-1 agonist and a GDF-8 specific binding protein and an activin A specific binding protein.

[0034] As used herein, the phrase "antigen-specific binding protein" refers to a protein comprising at least one domain that specifically binds to a particular antigen. Exemplary categories of antigen-specific binding proteins include antibodies, antigen-binding portions of antibodies, peptides that specifically interact with a particular antigen (e.g., peptibodies), receptor molecules that specifically interact with a particular antigen, and proteins that comprise the ligand-binding portion of a receptor that specifically binds to a particular antigen.

[0035] The present disclosure includes antigen-specific binding proteins that specifically bind to GDF-8, i.e., "GDF-8-specific binding proteins." The term "GDF-8" (also referred to as "growth differentiation factor-8" and "myostatin") refers to the protein (mature protein) having the amino acid sequence of SEQ ID NO: 25 (SEQ ID NO: 1). According to the present disclosure, a GDF-8-specific binding protein specifically binds to GDF-8 but does not bind to other ActRIIB ligands, such as GDF3, BMP2, BMP4, BMP7, BMP9, BMP10, GDF11, activin A, activin B, activin AB, Nodal, etc.

[0036] The present disclosure also includes antigen-specific binding proteins that specifically bind to activin A, i.e., "activin A-specific binding proteins." Activins are homo- and heterodimeric molecules comprising a βA subunit and / or a βB subunit. The βA subunit has the amino acid sequence of SEQ ID NO: 2, and the βB subunit has the amino acid sequence of SEQ ID NO: 3. Activin A is a homodimer of two βA subunits; activin B is a homodimer of two βB subunits; and activin AB is a heterodimer of one βA subunit and one βB subunit. An activin A-specific binding protein can be an antigen-specific binding protein that specifically binds to the βA subunit. Because the βA subunit is found in both activin A and activin AB molecules, an "activin A-specific binding protein" can be an antigen-specific binding protein that specifically binds to activin A and activin AB (through its interaction with the βA subunit). Thus, according to the present disclosure, an activin A-specific binding protein specifically binds to activin A, or to activin A and activin AB, but does not bind to other ActRIIB ligands, such as activin B, GDF3, GDF8, BMP2, BMP4, BMP7, BMP9, BMP10, GDF11, Nodal, etc.

[0037] In the context of the present disclosure, molecules such as ActRIIB-Fc (e.g., "ACE-031"), which contain the ligand-binding portion of the ActRIIB receptor, are not considered "GDF8-specific binding proteins" or "activin A-specific binding proteins" because such molecules bind to multiple ligands in addition to GDF8, activin A, and activin AB.

[0038] In one embodiment, myostatin (GDF8) and activin A inhibition can alternatively be provided by an antibody that binds to ActRIIb-Fc molecule or ActRIIB. When male CB17 SCID mice were treated with anti-activin A antibody, anti-GDF8 antibody, anti-activin A antibody + anti-GDF8 antibody, or ActRIIB.hFc, the increase in TA muscle mass in response to the combination of anti-GDF8 + anti-activin A was significantly greater than the increase in these parameters observed in subjects treated with anti-GDF8 monotherapy or anti-activin A monotherapy. Animals treated with ActRIIB-Fc also showed a significantly greater increase in muscle mass (Figure 18). In another embodiment, the antibody that binds to ActRIIB is bimagrumab.

[0039] The present disclosure includes antigen-specific binding proteins that specifically bind to GLP-1 and / or GLP-1R, i.e., "GLP-1-specific binding proteins." The term "GLP-1R" refers to glucagon-like peptide 1 receptor, including recombinant GLP-1R proteins or fragments thereof. GLP-1R has a sequence of 463 residues (NCBI Accession No. NP_002053, SEQ ID NO: 20). Donnelly, 2011, Br J Pharmacol 166(1):27-41 (2011). Glucagon-like peptide 1 (GLP-1) is a 31-amino acid peptide hormone released from intestinal L cells following nutrient consumption. Binding of GLP-1 to GLP-1R enhances glucose-induced insulin secretion from pancreatic beta cells, increases insulin expression, inhibits beta cell apoptosis, promotes beta cell neogenesis, reduces glucagon secretion, delays gastric emptying, promotes satiety, and enhances peripheral glucose disposal.

[0040] Antigen-binding molecules with two different antigen-specific binding domains The present disclosure also includes antigen-binding molecules comprising two distinct antigen-specific binding domains. In particular, the present disclosure includes antigen-binding molecules comprising a GDF8-specific binding domain and an activin A-specific binding domain. The term "antigen-specific binding domain," as used herein, includes polypeptides comprising or consisting of: (i) an antigen-binding fragment of an antibody molecule, (ii) a peptide (e.g., a peptibody) that specifically interacts with a particular antigen, and / or (iii) a ligand-binding portion of a receptor that specifically binds to a particular antigen. For example, the present disclosure includes bispecific antibodies comprising one arm comprising a first heavy chain variable region / light chain variable region (HCVR / LCVR) pair that specifically binds to GDF8, and another arm comprising a second HCVR / LCVR pair that specifically binds to activin A. Thus, a composition comprising a GDF-8-specific binding protein and an activin A-specific binding protein (and a GLP-1 agonist) may actually comprise a single binding protein comprising both a GDF8-specific binding domain and an activin A-specific binding domain.

[0041] specific binding "Specifically binds" and like terms, as used herein, refers to an antigen-specific binding protein or antigen-specific binding domain that binds to an antigen with a dissociation constant (K) of 500 pM or less. D) and does not bind to other unrelated antigens under normal test conditions. An "unrelated antigen" is a protein, peptide, or polypeptide that has less than 95% amino acid identity with each other. Methods for determining whether two molecules specifically bind to each other are well known in the art and include, for example, equilibrium dialysis, surface plasmon resonance, etc. For example, as used in the context of the present disclosure, an antigen-specific binding protein or antigen-specific binding domain has a K of less than about 500 pM, less than about 400 pM, less than about 300 pM, less than about 200 pM, less than about 100 pM, less than about 90 pM, less than about 80 pM, less than about 70 pM, less than about 60 pM, less than about 50 pM, less than about 40 pM, less than about 30 pM, less than about 20 pM, less than about 10 pM, less than about 5 pM, less than about 4 pM, less than about 2 pM, less than about 1 pM, less than about 0.5 pM, less than about 0.2 pM, less than about 0.1 pM, or less than about 0.05 pM, as measured by surface plasmon resonance assay. D The present invention also includes molecules that bind to specific antigens (e.g., GDF-8 or activin A and / or AB or GLP-1 / GLP-1R) or portions thereof comprising:

[0042] As used herein, an antigen-specific binding protein or antigen-specific binding domain "does not bind" to a particular molecule if, when tested for binding to that molecule in a surface plasmon resonance assay at 25°C, the protein or binding domain exhibits a K of greater than 1000 pM in such assay or its equivalent. D In this case, the binding may be either negative or may not be able to occur at all.

[0043] The term "surface plasmon resonance," as used herein, refers to an optical phenomenon that allows for the analysis of real-time interactions through the detection of changes in protein concentration within a biosensor matrix, for example, using a BIAcore™ system (Biacore Life Sciences division of GE Healthcare, Piscataway, NJ).

[0044] "K D The term "" as used herein refers to the equilibrium dissociation constant of a particular protein-protein interaction (e.g., antibody-antibody interaction). Unless otherwise indicated, the K D The K value is determined by surface plasmon resonance assay at 25°C. D Points to a value.

[0045] Antibodies and antigen-binding fragments of antibodies As indicated above, an antigen-specific binding protein can comprise or consist of an antibody or an antigen-binding fragment of an antibody. Furthermore, in the case of an antigen-binding molecule comprising two different antigen-specific binding domains, one or both of the antigen-specific binding domains can comprise or consist of an antigen-binding fragment of an antibody.

[0046] The term "antibody," as used herein, is intended to refer to an immunoglobulin molecule comprising four polypeptide chains: two heavy (H) chains and two light (L) chains interconnected by disulfide bonds, and multimers thereof (e.g., IgM). Each heavy chain contains a heavy chain variable region (herein referred to as HCVR or V H The heavy chain constant region is made up of three domains: H 1. C H 2 and C H Each light chain comprises a light chain variable region (herein referred to as LCVR or V L The light chain constant region contains one domain (C L 1) V H Area and V L The region can be further subdivided into regions of hypervariability called complementarity determining regions (CDRs), which are separated by more conserved regions called framework regions (FRs). H and V Lis composed of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. In various embodiments of the present disclosure, the FRs of an antibody (or antigen-binding portion thereof) of the present disclosure may be identical to human germline sequences or may be naturally or artificially modified. An amino acid consensus sequence can be defined based on a parallel analysis of two or more CDRs.

[0047] The term "antibody," as used herein, also includes antigen-binding fragments of a complete antibody molecule. The terms "antigen-binding portion" of an antibody, "antigen-binding fragment" of an antibody, and the like, as used herein, include any naturally occurring, enzymatically obtainable, synthetic, or genetically engineered polypeptide or glycoprotein that specifically binds to an antigen to form a complex. Antigen-binding fragments of antibodies can be derived from complete antibody molecules using any suitable standard technique, such as, for example, proteolytic digestion or recombinant genetic engineering techniques involving the manipulation and expression of DNA encoding the variable and, optionally, constant domains of the antibody. Such DNA is known and / or readily available, for example, from commercial sources, DNA libraries (including, for example, phage-antibody libraries), or can be synthesized. The DNA can be sequenced and manipulated chemically or using molecular biology techniques, for example, to place one or more variable and / or constant domains in the appropriate configuration, or to introduce codons, generate cysteine ​​residues, modify, add, or delete amino acids, etc.

[0048] Non-limiting examples of antigen-binding fragments include: (i) Fab fragments; (ii) F(ab')2 fragments; (iii) Fd fragments; (iv) Fv fragments; (v) single-chain Fv (scFv) molecules; (vi) dAb fragments; and (vii) minimal recognition units consisting of amino acid residues that mimic the hypervariable regions (e.g., isolated complementarity-determining regions (CDRs) such as CDR3 peptides) of antibodies or constrained FR3-CDR3-FR4 peptides. Other engineered molecules, such as domain-specific antibodies, single-domain antibodies, domain-deleted antibodies, chimeric antibodies, CDR-grafted antibodies, diabodies, triabodies, tetrabodies, minibodies, nanobodies (e.g., monovalent nanobodies, bivalent nanobodies, etc.), small modular immunopharmaceuticals (SMIPs), and shark variable IgNAR domains, are also encompassed within the term "antigen-binding fragment" as used herein.

[0049] An antigen-binding fragment of an antibody will usually contain at least one variable domain, which may be of any size or amino acid composition and will generally contain at least one CDR adjacent to or in frame with one or more framework sequences. L V associated with the domain H For antigen-binding fragments containing domains, V H Domains and V L The domains can be positioned relative to each other in any suitable arrangement. For example, the variable region can be a dimer, with the V H -V H , V H -V L or V L -V L Alternatively, the antigen-binding fragment of an antibody may contain a dimer of each of the monomeric V H Domain or V L It may contain domains.

[0050] In certain embodiments, an antigen-binding fragment of an antibody can contain at least one variable domain covalently linked to at least one constant domain. Non-limiting exemplary configurations of variable and constant domains that can be found in antigen-binding fragments of antibodies of the present disclosure include: (i) V H -C H 1;(ii)V H -C H 2;(iii)V H -C H 3;(iv)V H -C H 1-C H 2;(v)V H -C H 1-C H 2-C H 3;(vi)V H -C H 2-C H 3;(vii)V H -C L ;(viii)V L -C H 1;(ix)V L -C H 2;(x)V L -C H 3;(xi)V L -C H 1-C H 2;(xii)V L -C H 1-C H 2-C H 3;(xiii)V L -CH2-C H 3; and (xiv) V L -C LIn any configuration of the variable and constant domains, including any of the exemplary configurations listed above, the variable and constant domains may be directly linked to each other or may be linked by a full or partial hinge or linker region. The hinge region can consist of at least two (e.g., 5, 10, 15, 20, 40, 60, or more) amino acids that provide a flexible or semi-flexible linkage between adjacent variable domains and / or adjacent constant domains in a single polypeptide molecule. Furthermore, antigen-binding fragments of antibodies of the present disclosure can be non-covalently associated with each other and / or with one or more monomeric V H Domain or V L The variable domains may comprise homodimers or heterodimers (or other multimers) of any of the above listed variable and constant domain configurations, with the domains non-covalently associated (e.g., by disulfide bonds).

[0051] The molecules of the present disclosure may comprise or consist of human antibodies and / or recombinant human antibodies, or fragments thereof. The term "human antibody," as used herein, includes antibodies having variable and constant regions derived from human germline immunoglobulin sequences. Human antibodies may still contain amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo), e.g., in the CDRs, particularly CDR3. However, the term "human antibody," as used herein, is not intended to include antibodies in which CDR sequences derived from the germline of another mammalian species, such as a mouse, have been grafted onto human framework sequences.

[0052] Molecules of the present disclosure may comprise or consist of recombinant human antibodies or antigen-binding fragments thereof. The term "recombinant human antibody," as used herein, is intended to include all human antibodies that are produced, expressed, generated, or isolated by recombinant means, such as antibodies expressed using recombinant expression vectors transfected into host cells (described further below), antibodies isolated from recombinant combinatorial human antibody libraries (described further below), antibodies isolated from animals (e.g., mice) transgenic for human immunoglobulin genes (see, e.g., Taylor et al. (1992) Nucl. Acids Res. 20:6287-6295), or antibodies produced, expressed, generated, or isolated by any other means involving splicing human immunoglobulin gene sequences into other DNA sequences. Such recombinant human antibodies have variable and constant regions derived from human germline immunoglobulin sequences. In certain embodiments, however, such recombinant human antibodies are subjected to in vitro mutagenesis (or, when animals transgenic for human Ig sequences are used, in vivo somatic mutagenesis), resulting in the V H Area and V L The amino acid sequence of the region is human germline V H Array and V L These sequences are derived from and related to sequences, but which, while not naturally occurring within the human antibody germline repertoire in vivo, may not exist.

[0053] All amino acid abbreviations used in this disclosure are those accepted by the United States Patent and Trademark Office as set forth in 37 C.FR §1.822(B)(J). The amino acid sequence of an antibody or antigen-binding fragment thereof can be numbered using any known numbering scheme, including those described by Kabat et al. ("Kabat" numbering scheme); Al-Lazikani et al., 1997, J. Mol. Biol. 273:927-948 ("Chothia" numbering scheme); MacCallum et al., 1996, J. Mol. Biol. 262:732-745 ("Contact" numbering scheme); Lefranc et al., 2003, Dev. Comp. Immunol. 27:55-77 ("IMGT" numbering scheme); and Honegge and Pluckthun, 2001, J. Mol. Biol. 309:657-70 ("AHo" numbering scheme).

[0054] As used herein, an "agonist" antibody or antigen-binding fragment thereof is an antibody or fragment that increases or enhances at least one biological activity of an antigen, such as GLP-1 and / or GLP-1-R. Such enhancement or enhancement may be mediated by the antibody itself, or, when the antibody is part of an antibody-drug conjugate or antibody-linked drug conjugate, by a payload or linker-payload. For example, an agonist antibody or fragment can induce stimulation of the adenylate cyclase pathway, resulting in increased cyclic AMP synthesis and insulin release, when the cell is a mammalian pancreatic beta cell. Another biological activity of GLP-1R may be cAMP-dependent activation of protein kinase A (PKA) and / or cAMP-regulated guanine nucleotide exchange factor 2 (Epac2). An agonist antibody or fragment may reduce glucose levels or reduce body weight when administered to a subject.

[0055] Anti-GDF8 antibodies and antigen-binding fragments thereof In certain specific embodiments of the present disclosure, GDF-8 inhibitor is GDF-8 specific binding protein, and said protein or GDF8 specific binding domain comprises or consists of anti-GDF8 antibody or its antigen-binding fragment.Anti-GDF8 antibody is described in, for example, U.S. Patent No. 6,096,506; U.S. Patent No. 7,320,789; U.S. Patent No. 7,261,893; U.S. Patent No. 7,807,159; U.S. Patent No. 7,888,486; U.S. Patent No. 7,635,760; U.S. Patent No. 7,632,499; U.S. Patent Application Publication No. 2007 / 0178095; U.S. Patent No. 2010 / 0166764; U.S. Patent No. 2009 / 0148436; and International Patent Application Publication No. WO2010 / 070094. Anti-GDF8 antibodies are also described in U.S. Patent Application No. 13 / 115,170, filed May 25, 2011, and published as US20110293630, including antibodies designated 8D12, H4H1657N2, and H4H1669P. In one embodiment, the anti-GDF8 antibody is REGN1033, also known as H4H1657N2. Any of the anti-GDF8 antibodies, or antigen-binding fragments thereof, described and / or listed in any of the aforementioned patents or publications can be used in the context of the present disclosure, so long as such antibodies and / or antigen-binding fragments "specifically bind" to GDF8, as that term is defined herein.

[0056] In one embodiment, the anti-GDF8 antibody or antigen-binding fragment thereof comprises a heavy chain complementarity determining region (HCDR) of the heavy chain variable region (HCVR) comprising SEQ ID NO: 4, and a light chain complementarity determining region (LCDR) of the light chain variable region (LCVR) comprising SEQ ID NO: 5. In another embodiment, the anti-GDF8 antibody or antigen-binding fragment thereof comprises heavy chain complementarity determining regions (HCDR1, HCDR2, and HCDR3) comprising SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8, respectively, and three light chain complementarity determining regions (LCDR1, LCDR2, and LCDR3) comprising SEQ ID NO: 9, TTS, and SEQ ID NO: 11, respectively.

[0057] Anti-activin A antibodies and antigen-binding fragments thereof In certain specific embodiments of the present disclosure, the activin A inhibitor is an activin A-specific binding protein, and the protein or activin A-specific binding domain comprises or consists of an antibody or antigen-binding fragment thereof that specifically binds to activin A. In certain embodiments, the activin A-specific binding protein specifically binds to the βA subunit. An antigen-specific binding protein that specifically binds to the βA subunit can recognize both activin A (βA / βA homodimer) and activin AB (βA / PB heterodimer). Thus, according to the present disclosure, the activin A-specific binding protein can bind to both activin A and activin AB (but not activin B). Anti-activin A antibodies are described, for example, in U.S. Patent Application Publication No. 2009 / 0234106. In one embodiment, the anti-activin A antibody is REGN2477, also known as H4H10446P2. In another embodiment, the anti-activin A antibody is REGN2376, also known as H4H10430P. Another anti-activin A antibody is designated "MAB3381" and is commercially available from R&D Systems, Inc., Minneapolis, MN. MAB3381 specifically binds to activin A (homodimer) and activin AB (heterodimer). Any of the foregoing anti-activin A antibodies or antigen-binding fragments thereof can be used in the context of the present disclosure, so long as such antibodies and / or antigen-binding fragments "specifically bind" to activin A and / or activin AB as defined herein.

[0058] In one embodiment, the anti-activin A antibody or antigen-binding fragment thereof comprises a heavy chain complementarity determining region (HCDR) of the heavy chain variable region (HCVR) comprising SEQ ID NO: 12, and a light chain complementarity determining region (LCDR) of the light chain variable region (LCVR) comprising SEQ ID NO: 13. In another embodiment, the anti-activin A antibody or antigen-binding fragment thereof comprises heavy chain complementarity determining regions (HCDR1, HCDR2, and HCDR3) comprising SEQ ID NO: 14, SEQ ID NO: 15, and SEQ ID NO: 16, respectively, and three light chain complementarity determining regions (LCDR1, LCDR2, and LCDR3) comprising SEQ ID NO: 17, GAS, and SEQ ID NO: 19, respectively.

[0059] Glucagon-like peptide (GLP)-1 agonist / glucagon-like peptide (GLP)-1 receptor agonist Incretin is a gut-derived hormone that is released in response to nutrient intake and stimulates insulin secretion in conjunction with hyperglycemia. In some embodiments of the compositions and methods of the present disclosure, incretin inhibitors are used in combination with other drugs (e.g., GDF-8 inhibitors, or GDF-8 inhibitors + activin A inhibitors). In further embodiments of the compositions and methods of the present disclosure, GLP-1 agonists or dipeptidyl peptidase IV (DPP-4) inhibitors are used in combination with other drugs (e.g., GDF-8 inhibitors, or GDF-8 inhibitors + activin A inhibitors).

[0060] The term "GLP-1," also known as "glucagon-like peptide 1," refers to a 31-amino acid peptide hormone released from intestinal L-cells following nutrient consumption. GLP-1 binds to the GLP-1 receptor, enhances glucose-induced insulin secretion from pancreatic beta cells, increases insulin expression, inhibits beta cell apoptosis, promotes beta cell neogenesis, reduces glucagon secretion, delays gastric emptying, promotes satiety, and enhances peripheral glucose disposal.

[0061] As used herein, the term "GLP-1 agonist" refers to a compound that promotes, upregulates, or simulates the activity of GLP-1. GLP-1 agonists can activate GLP-1R, and include GLP-1 mimetics, peptide variants, antibodies (including antibodies linked to ligands), and fusion proteins. GLP-1 agonists include GLP-1 receptor agonists (GLP-1 RAs). The GLP-1 agonists described / used herein are GLP-1 receptor agonists. Indeed, for the purposes of this disclosure, the terms "GLP-1 agonist" and "GLP-1R agonist" are used interchangeably. As used herein, the term "GLP-1 receptor agonist" refers to a compound that binds to the GLP-1 receptor. GLP-1 receptor agonists increase glucose-dependent insulin secretion, reduce inappropriate glucagon secretion, delay gastric emptying, and enhance satiety (Trujillo et al., 2021, Ther Adv Endocrinol Metab 12:1-15). GLP-1 agonists can be selected from, for example, small molecule and peptide GLP-1R agonists and allosteric modulators (Graaf et al., 2016, Pharmacol Rev 68:954-1013).

[0062] GLP-1 agonists for use in the present disclosure include peptide agonists currently on the market. In certain embodiments, the GLP-1 agonist mimics the action of glucagon-like peptide 1. Known GLP-1 receptor agonists include albiglutide, exenatide (short-acting and long-acting), efpeglenatide, ITCA650, lixisenatide, liraglutide, dulaglutide, and semaglutide. In certain embodiments, the GLP-1 agonist is selected from the group consisting of exenatide (long-acting), dulaglutide, liraglutide, and semaglutide. In further embodiments of the composition or method according to the present disclosure, the GLP-1 agonist is semaglutide. Semaglutide (sold under the brand name Ozempic, among others) is a glucagon-like peptide-1 receptor agonist that increases insulin production and secretion, thereby enhancing glucose metabolism. In one embodiment, the GLP-1 agonist for use in the methods or compositions according to the present disclosure is a modified peptide drug, such as tirzepatide, that activates both the glucagon-like peptide-1 (GLP-1) receptor and the glucose-dependent insulinotropic polypeptide (GIP) receptor.

[0063] In another embodiment, the GLP-1 agonist / receptor agonist for use in a composition or method according to the present disclosure is an antibody or antigen-binding fragment thereof that specifically binds to GLP-1.

[0064] In certain embodiments, the GLP-1 agonist for use in the compositions and methods according to the present disclosure is an antibody-drug conjugate (ADC) that specifically binds to the glucagon-like peptide 1 receptor (GLP-1R) protein. In further embodiments, the antibody or antigen-binding fragment thereof of the ADC specifically targets the extracellular domain of GLP-1R, where the GLP-1 peptidomimetic functionally activates GLP-1R.

[0065] Antibody-drug conjugates (ATDCs) or antibody-drug conjugates (ADCs) refer to antibodies or antigen-binding fragments thereof linked to a payload (e.g., a GLP-1 peptidomimetic) with or without a linker. An antibody-payload conjugate refers to such an antibody or fragment linked to a payload, while an antibody-linker-payload conjugate refers to an antibody or fragment conjugated to a payload via a linker. The antibody or antigen-binding fragment referred to herein includes embodiments in which the antibody or fragment is conjugated to a payload or linker-payload.

[0066] biological equivalent In certain embodiments, the GDF-8 inhibitors, activin A inhibitors, and / or GLP-1 agonists of the present disclosure encompass proteins having amino acid sequences that vary from the amino acid sequences of the described GDF-8 inhibitors, activin A inhibitors, and / or GLP-1 agonists, but that retain the ability to bind to GDF-8, activin A, and GLP-1, respectively. Such variants contain one or more amino acid additions, deletions, or substitutions when compared to the parent sequence, but exhibit biological activity that is essentially equivalent to the biological activity of the described GDF-8 inhibitors, activin A inhibitors, and / or GLP-1 agonists.

[0067] Two proteins are considered bioequivalent if, for example, they are pharmaceutical equivalents or pharmaceutical substitutes that do not show significant differences in the rate and extent of their absorption when administered at the same molar dose under similar experimental conditions, either in single or multiple doses. Some proteins may be considered equivalents or pharmaceutical substitutes if they are equivalent in their extent of absorption but not in their rate of absorption, and still be considered bioequivalent because such differences in absorption rate are intentional, reflected in labeling, and are not essential, for example, to achieving effective body drug concentrations for chronic use, and are not considered medically significant for the particular pharmaceutical product being studied.

[0068] In one embodiment, two GDF8 inhibitor proteins, two activin A inhibitor proteins, or two GLP-1 agonist proteins are bioequivalent if there are no clinically meaningful differences in their safety, purity, and potency.

[0069] In one embodiment, two GDF8 inhibitor proteins, two activin A inhibitor proteins, or two GLP-1 agonist proteins are bioequivalent if a patient can make one or more switches between the reference product and the biologic product without an expected increased risk of adverse effects or decreased efficacy, including a clinically significant change in immunogenicity, compared to therapy continued without such switches.

[0070] In one embodiment, two GDF8 inhibitor proteins, two activin A inhibitor proteins, or two GLP-1 agonist proteins are bioequivalent if they both act by one or more common mechanisms for one or more conditions of use, so long as such mechanisms are known.

[0071] Bioequivalence can be demonstrated by in vivo and / or in vitro methods. Measurements of bioequivalence include, for example, (a) in vivo tests in humans or other mammals that measure the concentration of the protein or its metabolites as a function of time in blood, plasma, serum, or other biological fluids; (b) in vitro tests that have correlated with and are reasonably predictive of human in vivo bioavailability data; (c) in vivo tests in humans or other mammals that measure the appropriate acute pharmacological effect of the protein (or its target) as a function of time; and (d) in vivo tests in well-controlled clinical trials that establish the safety, efficacy, or bioavailability or bioequivalence of the antigen binding protein.

[0072] Biologically equivalent variants of the GDF8 inhibitor, activin A inhibitor and / or GLP-1 agonist protein of the present disclosure can be constructed, for example, by making various substitutions of residues or sequences or by deleting terminal or internal residues or sequences that are not required for biological activity.For example, cysteine ​​residues that are not essential for biological activity can be deleted or replaced with other amino acids to prevent the formation of unnecessary or incorrect intramolecular disulfide bridges during renaturation.In other contexts, biologically equivalent proteins include variants that contain amino acid changes that modify the glycosylation characteristics of proteins, for example, mutations that eliminate or remove glycosylation.

[0073] Pharmaceutical Compositions and Methods of Administration The present disclosure includes pharmaceutical compositions comprising a GDF8 inhibitor and a GLP-1 agonist. The present disclosure also includes pharmaceutical compositions comprising a GDF8 inhibitor, an activin A inhibitor, and a GLP-1 agonist. The pharmaceutical compositions of the present disclosure are formulated with suitable carriers, excipients, and other agents that provide suitable mobility, delivery, tolerability, etc. Many suitable formulations can be found, for example, in Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, PA. Suitable formulations include, for example, powders, pastes, ointments, jellies, waxes, oils, lipids, lipid (cationic or anionic)-containing vesicles (e.g., LIPOFECTIN™), DNA conjugates, anhydrous absorbent pastes, oil-in-water and water-in-oil emulsions, emulsion carbowax (polyethylene glycol of various molecular weights), semi-solid gels, and carbowax-containing semi-solid mixtures. Further suitable formulations are also described in Powell et al., "Compendium of excipients for parenteral formulations," PDA (1998) J Pharm Sci Technol 52:238-311.

[0074] Various delivery systems, such as liposomal encapsulation, microparticles, microcapsules, recombinant cells capable of expressing mutant viruses, and receptor-mediated endocytosis (see, e.g., Wu et al., 1987, J. Biol. Chem. 262:4429-4432), are known and can be used to administer the pharmaceutical compositions of the present disclosure. Administration methods include, but are not limited to, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, and oral routes. The compositions may be administered by any convenient route, such as by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g., oral, rectal, and intestinal mucosa), or may be administered together with other biologically active agents.

[0075] The pharmaceutical composition of the present disclosure can be delivered subcutaneously or intravenously using a standard needle and syringe. In addition, for subcutaneous delivery, a pen delivery device is easily applied to deliver the pharmaceutical composition of the present disclosure. Such a pen delivery device may be reusable or disposable. Reusable pen delivery devices generally utilize a replaceable cartridge containing the pharmaceutical composition. Once all of the pharmaceutical composition in the cartridge has been administered and the cartridge is empty, the empty cartridge can be easily discarded and replaced with a new cartridge containing the pharmaceutical composition. The pen delivery device can then be reused. In disposable pen delivery devices, there is no replaceable cartridge. Rather, the disposable pen delivery device is pre-filled with the pharmaceutical composition held in a reservoir within the device. Once the reservoir is emptied of the pharmaceutical composition, the entire device is discarded.

[0076] Numerous reusable pen and autoinjector delivery devices are adapted for subcutaneous delivery of the pharmaceutical compositions of the present disclosure. Examples include, but are not limited to, the AUTOPEN™ (Owen Mumford, Inc., Woodstock, UK), the DISETRONIC™ pen (Disetronic Medical Systems, Bergdorf, Switzerland), the HUMALOG MIX 75 / 25™ pen, the HUMALOG™ pen, the HUMALIN 70 / 30™ pen (Eli Lilly and Co., Indianapolis, IN), the NOVOPEN™ I, II, and III (Novo Nordisk, Copenhagen, Denmark), the NOVOPEN JUNIOR™ (Novo Nordisk, Copenhagen, Denmark), the BD™ pen (Becton Dickinson, Franklin Lakes, NJ), the OPTIPEN™, the OPTIPEN PRO™, the OPTIPEN™, to name a few. STARLET™, and OPTICLIK™ (sanofi-aventis, Frankfurt, Germany). Examples of disposable pen delivery devices applicable to subcutaneous delivery of the pharmaceutical compositions of the present disclosure include, but are not limited to, the SOLOSTAR™ Pen (sanofi-aventis), FLEXPEN™ (Novo Nordisk), and KWIKPEN™ (Eli Lilly), SURECLICK™ Autoinjector (Amgen, Thousand Oaks, CA), PENLET™ (Haselmeier, Stuttgart, Germany), EPIPEN (Dey, LP), and HUMIRA™ Pen (Abbott Labs, Abbott Park IL), to name a few.

[0077] In certain circumstances, the pharmaceutical compositions of the present disclosure can be delivered in a controlled release system. In one embodiment, a pump may be used (see Langer, supra; Sefton, 1987, CRC Crit. Ref. Biomed. Eng. 14:201). In another embodiment, a polymeric material may be used; see Medical Applications of Controlled Release, Langer and Wise (eds.), 1974, CRC Press, Boca Raton, Florida. In yet another embodiment, the controlled release system can be placed near the target of the composition, thus requiring only a fraction of the systemic dose (see, e.g., Goodson, 1984, in Medical Applications of Controlled Release, supra, Vol. 2, pp. 115-138). Other controlled release systems are discussed in the review by Langer, 1990, Science 249:1527-1533.

[0078] Injectable preparations include dosage forms for intravenous, subcutaneous, intradermal, and intramuscular injections, infusions, and the like. Such injectable preparations can be produced by known methods. For example, injectable preparations can be produced by dissolving, suspending, or emulsifying the above-mentioned antibody or its salt in a sterile aqueous or oily medium conventionally used for injections. Aqueous media for injections include, for example, physiological saline, isotonic solutions containing glucose and other adjuvants, and the like. These can be used in combination with appropriate solubilizers such as alcohols (e.g., ethanol), polyhydric alcohols (e.g., propylene glycol, polyethylene glycol), nonionic surfactants [e.g., polysorbate 80, HCO-50 (hydrogenated castor oil polyoxyethylene (50 mol) adduct)], and the like. Oily media can be, for example, sesame oil, soybean oil, and the like, which can be used in combination with solubilizers such as benzyl benzoate, benzyl alcohol, and the like. The injectable preparations thus produced are preferably filled into appropriate ampoules.

[0079] Advantageously, the above-mentioned pharmaceutical compositions for oral or parenteral use are prepared into dosage forms with unit doses suitable for the dosage of the active ingredient, such as tablets, pills, capsules, injections (ampoules), suppositories, etc.

[0080] Dosage The amount of active ingredient (for example, GDF8 inhibitor, activin A inhibitor, GLP-1 agonist) that can be administered to a subject is generally a therapeutically effective amount.As used herein, the phrase "therapeutically effective amount" refers to the dose of inhibitor, for example, the dose of antigen-specific binding protein and / or antigen-binding molecule and / or agonist, that causes detectable changes in one or more of the following parameters: lean body mass (increase), fat mass (decrease), body weight (decrease), skeletal muscle mass (increase), plasma ALT and / or AST (decrease), liver triglyceride content / fatty degeneration (decrease), and liver smooth muscle actin (decrease).In a specific embodiment, the therapeutically effective amount of GDF8 inhibitor vs. activin A inhibitor vs. GLP-1 agonist refers to the amount that achieves a clear effect. For example, in one embodiment, a therapeutically effective amount of a GLP-1 agonist is an amount that results in one or more of the following: (a) a reduction in elevated sugar levels to normal levels; and / or (b) a detectable improvement in one or more symptoms or signs of diabetes; and / or (c) an improvement in liver dysfunction associated with NASH and / or treatment of NASH; and / or (d) an improvement in cholesterol, LDL cholesterol, and / or HDL cholesterol.

[0081] The dosage of the active ingredient (eg, GDF8 inhibitor, activin A inhibitor, GLP-1 agonist) may vary depending on the age and size of the subject, the target disease, condition, administration route, and the like.

[0082] In the case of antibodies of the disclosure (e.g., anti-GDF8 antibodies, anti-activin A antibodies, anti-GLP-1 antibodies, anti-GLP-1R antibodies, or bispecific antibodies), a therapeutically effective amount can be from about 0.05 mg to about 600 mg of the respective antibody; e.g., about 0.05 mg, about 0.1 mg, about 1.0 mg, about 1.5 mg, about 2.0 mg, about 10 mg, about 20 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 110 mg, about 120 mg, about 130 mg, about 140 mg, about 150 mg, about 160 mg, about 170 mg, about 180 mg, about 190 mg, about 200 mg, about 210 mg , about 220 mg, about 230 mg, about 240 mg, about 250 mg, about 260 mg, about 270 mg, about 280 mg, about 290 mg, about 300 mg, about 310 mg, about 320 mg, about 330 mg, about 340 mg, about 350 mg, about 360 mg, about 370 mg, about 380 mg, about 390 mg, about 400 mg, about 410 mg, about 420 mg, about 430 mg, about 440 mg, about 450 mg, about 460 mg, about 470 mg, about 480 mg, about 490 mg, about 500 mg, about 510 mg, about 520 mg, about 530 mg, about 540 mg, about 550 mg, about 560 mg, about 570 mg, about 580 mg, about 590 mg, or about 600 mg.

[0083] The amount of an antibody of the disclosure (e.g., an anti-GDF8 antibody, an anti-activin A antibody, an anti-GLP-1 antibody, an anti-GLP-1R antibody, or a bispecific antibody) contained within an individual dose can be expressed in milligrams of antibody per kilogram of patient body weight (i.e., mg / kg). For example, each of the anti-GDF8, anti-activin A, anti-GDF8 / anti-activin A bispecific, and / or anti-GLP-1 antibodies of the disclosure can be administered in a dose ranging from about 0.0001 to about 50 mg / kg of patient body weight (e.g., 0.0001 mg / kg, 0.001 mg / kg, 0.01 mg / kg, 0.1 mg / kg, 0.5 mg / kg, 1.0 mg / kg, 1.5 mg / kg, 2.0 mg / kg, 2.5 mg / kg, 3.0 mg / kg). The compound can be administered to a patient at a dose of 100 mg / kg, 3.5 mg / kg, 4.0 mg / kg, 4.5 mg / kg, 5.0 mg / kg, 5.5 mg / kg, 6.0 mg / kg, 6.5 mg / kg, 7.0 mg / kg, 7.5 mg / kg, 8.0 mg / kg, 8.5 mg / kg, 9.0 mg / kg, 9.5 mg / kg, 10.0 mg / kg, 10.5 mg / kg, 11.0 mg / kg, 11.5 mg / kg, etc.

[0084] In certain embodiments, the GLP-1 agonists of the present disclosure can be administered to a subject (patient) at a dose of about 0.05 mg / mL to about 5 mg / mL (e.g., about: 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.4 mg / mL, 0.5 mg / mL, 0.6 mg / mL, 0.7 mg / mL, 0.8 mg / mL, 0.9 mg / mL, 1.0 mg / mL, 1.1 mg / mL, 1.2 mg / mL, 1.3 mg / mL, 1.4 mg / mL, 1.5 mg / mL, 1.6 mg / mL, 1.7 mg / mL, 2.0 mg / mL, 2.5 mg / mL, 3.0 mg / mL, 3.5 mg / mL, 4.0 mg / mL, 4.5 mg / mL, or 5 mg / mL).

[0085] In certain embodiments, the GLP-1 agonist of the present disclosure can be administered to a subject (patient) at a dose of about 0.000001 mg / kg to about 50 mg / kg of subject's body weight. In further embodiments, the GLP-1 agonist (GLP-1R agonist) of the present disclosure is administered to a subject (patient) at a dose of about 0.00001 mg / kg to about 10 mg / kg of subject's body weight. In yet another embodiment, the GLP-1 agonist (GLP-1R agonist) of the present disclosure is administered to a subject (patient) at a dose of about 1 mg / kg of subject's body weight. In yet another embodiment, the GLP-1 agonist (GLP-1R agonist) of the present disclosure is administered to a subject (patient) at a dose of about 0.1 mg / kg of subject's body weight. In yet another embodiment, the GLP-1 agonist (GLP-1R agonist) of the present disclosure is administered to a subject (patient) at a dose of about 10 μg / kg of subject body weight. In certain embodiments, the GLP-1 receptor agonist of the present disclosure can be administered in one or more doses, including between about 0.01 mg and about 60 mg. In further embodiments, the GLP-1 receptor agonist of the present disclosure can be administered in one or more doses, including between about 0.1 mg and about 6 mg.

[0086] In a specific embodiment, the anti-GDF8 antibody or antigen-binding fragment thereof is administered at a concentration of about 10 mg / kg to about 100 mg / kg. In another embodiment, the anti-GDF8 antibody or antigen-binding fragment thereof is administered at a concentration of about 50 mg / kg. In another specific embodiment, the anti-activin A antibody or antigen-binding fragment thereof is administered at a concentration of about 10 mg / kg to about 100 mg / kg. In another embodiment, the anti-activin A antibody or antigen-binding fragment thereof is administered at a concentration of about 50 mg / kg. In yet another specific embodiment, the GLP-1 agonist is administered at a concentration of about 1 μg / kg to about 100 μg / kg. In yet another specific embodiment, the GLP-1 agonist is administered at a concentration of about 10 μg / kg. In a further embodiment, the GLP-1 agonist is administered at a concentration of about 0.833 mg / mL or about 7 μg / day (infusion). In yet a further embodiment, the anti-GDF8 antibody or antigen-binding fragment thereof is administered at a concentration of about 50 mg / kg, the anti-activin A antibody or antigen-binding fragment thereof is administered at a concentration of about 50 mg / kg, and the GLP-1 agonist is administered at a concentration of about 10 μg / kg.

[0087] The composition of the present disclosure can comprise equal amounts of GDF8 specific binding protein and activin A specific binding protein.Alternatively, the amount of GDF8 specific binding protein in the composition can be less than or greater than the amount of activin A specific binding protein.Alternatively, the composition of the present disclosure can not contain any activin A specific binding protein.Those skilled in the art can use routine experimentation and based on the present disclosure to determine the appropriate amount of each component in the composition of the present disclosure that is required to produce desired therapeutic effect.

[0088] Treatment method As used herein, the terms "treat," "treating," or "treatment" refer to the reduction in severity or amelioration of at least one symptom or sign of a disease or disorder associated with GDF-8, activin A, and / or GLP-1. In some embodiments, the disease or disorder is obesity, diabetes, liver dysfunction, or any other condition associated with hyperglycemia. In one embodiment, the diabetes is type 2 diabetes. The terms can also refer to the inhibition of disease progression or worsening of symptoms. The terms can also refer to a favorable prognosis of a disease, i.e., upon administration of a therapeutic agent such as a composition (or compositions) according to the present disclosure, i.e., a composition comprising a GDF8 inhibitor and a GLP-1 agonist, or a composition comprising a GDF8 inhibitor, an activin A inhibitor, and a GLP-1 agonist, the subject may be free of symptoms or signs, or may have symptoms or signs of reduced intensity. The therapeutic agent can be administered to the subject in a therapeutic dose.

[0089] The terms "prevent," "preventing," or "prevention" each refer to the inhibition of the manifestation of any symptom or sign of a disease or disorder associated with GDF-8, activin A, and / or GLP-1. The terms can also refer to the inhibition of the manifestation of symptoms or signs of a disease or disorder associated with GDF-8, activin A, and / or GLP-1 in a subject at risk of developing such a disease or disorder. In some embodiments, such a disease or disorder is obesity, diabetes, liver dysfunction, or any other condition associated with hyperglycemia. In one embodiment, the diabetes is type 2 diabetes.

[0090] The present disclosure includes compositions and methods for treating conditions or afflictions that can be cured, alleviated, or improved by specifically binding to GDF8 and activin A and exerting agonistic activity against GLP-1, thereby increasing lean body mass and / or reducing fat mass in an individual, or by favorably altering glucose control. For example, the present disclosure includes compositions and methods for improving glucose control, increasing lean body mass, reducing fat mass, treating diabetes, and / or treating obesity in a subject, and / or treating liver problems associated with increased fat mass, obesity, and / or diabetes in a subject, which in certain embodiments comprise administering to the subject a composition comprising a GDF-8 inhibitor and a GLP-1 agonist, and in further embodiments, administering to the subject a composition comprising a GDF-8 inhibitor, an activin A inhibitor, and a GLP-1 agonist. The present disclosure also includes compositions and methods for improving glucose control, increasing lean body mass, reducing fat mass, treating diabetes, and / or treating obesity, and / or treating liver problems associated with increased fat mass, obesity, and / or diabetes in a subject, which methods, in certain embodiments, include administering to a subject a composition comprising a GDF-8 inhibitor and a GLP-1 agonist, and in further embodiments, administering to a subject compositions comprising a GDF-8 inhibitor, an activin A inhibitor, and a GLP-1 agonist, in a single composition or in two or more compositions (e.g., each inhibitor and agonist in a separate composition, or the two inhibitors in one composition and the agonist in another composition).

[0091] Treating liver problems includes lowering signs / symptoms of liver damage (e.g., ALT / AST), lowering liver triglycerides, lowering steatosis, and / or lowering fibrosis (in the liver). In a further embodiment, treating liver problems includes treating liver dysfunction, such as hepatitis (A, B, C, D, and E), fatty liver disease (alcoholic and non-alcoholic), autoimmune diseases (autoimmune hepatitis, primary biliary cirrhosis, primary sclerosing cholangitis), genetic diseases (hemochromatosis, Wilson's disease, alpha-1 antitrypsin deficiency), drug-induced liver disease, cancer (e.g., hepatocellular carcinoma), cirrhosis, and liver failure. Any of the GDF-8 inhibitors (e.g., GDF8-specific binding proteins) and activin A inhibitors (e.g., activin A-specific binding proteins) and GLP-1 agonists (e.g., GLP-1 receptor agonists) disclosed or mentioned herein can be used in the context of such aspects of the disclosure. For example, the therapeutic method of the disclosure includes administering to a subject an anti-GDF8 antibody and a GLP-1 agonist, or an anti-GDF8 antibody, an anti-activin A antibody, and a GLP-1 agonist.

[0092] The present disclosure also includes a method for managing or treating the liver condition or problem associated with increased fat mass, obesity and diabetes by administering a GDF8 inhibitor and a GLP-1 agonist, or a GDF8 inhibitor, an activin A inhibitor and a GLP-1 agonist to a subject in need thereof.In certain embodiments, the management or treatment of liver condition or problem is in the form of reducing plasma ALT and / or AST (markers of liver damage), reducing liver triglyceride content / fatty degeneration, and / or reducing liver active smooth muscle (markers of fibrosis).Therefore, in certain embodiments, administering a composition according to the present disclosure reduces liver damage or its risk in a subject, reduces fatty degeneration or its risk, and / or reduces liver fibrosis or its risk.

[0093] In methods involving administering a GDF-8 inhibitor and a GLP-1 agonist, or a GDF-8 inhibitor, an activin A inhibitor and a GLP-1 agonist to a subject, the GDF-8 inhibitor and activin A inhibitor (if present) and the GLP-1 agonist can be administered to the subject simultaneously or substantially simultaneously, e.g., in a single therapeutic dosage, or in two or more separate dosages administered simultaneously or sequentially, e.g., in separate therapeutic dosages separated in time from each other, or in two or three separate doses administered simultaneously or sequentially.

[0094] The compositions of the present disclosure can be administered to a subject with one or more additional therapeutic agents, including, for example, growth factor inhibitors, immunosuppressants, anti-inflammatory agents, metabolic inhibitors, enzyme inhibitors, cytotoxic / cytostatic agents, and medications for controlling blood glucose levels (e.g., metformin). In some embodiments, the additional therapeutic agent can be selected from the group consisting of insulin or insulin analogs, biguanides (e.g., metformin), thiazolidinediones, sulfonylureas (e.g., clopropamide), glinides (e.g., nateglinide), alpha-glucosidase inhibitors, DPP4 inhibitors (e.g., sitagliptin), pramlintide, bromocriptine, SGLT2 inhibitors (e.g., canagliflozin), antihypertensive drugs, statins, aspirin, dietary modification, exercise, and nutritional supplements. The additional therapeutic agent may be administered before, simultaneously with, or after administration of the GDF-8 inhibitor, activin A inhibitor (if present), and GLP-1 agonist (or a composition comprising same) of the present disclosure.

[0095] Exemplary diseases, disorders, and conditions that can be treated with the compositions of the present disclosure include, but are not limited to, sarcopenia, cachexia (either idiopathic or secondary to other conditions, such as cancer, chronic renal failure, or chronic obstructive pulmonary disease), muscle damage, muscle wasting, and muscle atrophy, such as muscle atrophy or muscle wasting caused by or associated with disuse, immobilization, bed rest, injury, medical or surgical intervention (e.g., hip fracture, total hip replacement, knee replacement, etc.), or the need for mechanical ventilation. The compositions of the present disclosure can also be used to treat, prevent, or ameliorate diseases such as cancer, obesity, diabetes, arthritis, multiple sclerosis, muscular dystrophy, amyotrophic lateral sclerosis, Parkinson's disease, osteoporosis, osteoarthritis, osteopenia, metabolic syndrome (including, but not limited to, diabetes, obesity, nutritional disorders, organ atrophy, chronic obstructive pulmonary disease, and anorexia). The compositions of the present disclosure can also be used to treat, prevent, or ameliorate diseases such as diabetes mellitus, obesity, insulin resistance, hypertension, dyslipidemia, type 2 diabetes, type 1 diabetes, prediabetes, cardiovascular disease, atherosclerosis, congestive heart failure, coronary artery disease, arteriosclerosis, peripheral artery disease, stroke, respiratory dysfunction, kidney disease, fatty liver disease, nonalcoholic steatohepatitis (NASH), and metabolic syndrome. NASH is nonalcoholic fatty liver disease (NAFLD) characterized by fatty degeneration of the liver accompanied by inflammation and hepatocyte ballooning, which can lead to progressive fibrosis, cirrhosis, and hepatocellular carcinoma (Paternostro and Trauner, 2022, J Intern Med 0:1-15).

[0096] The compositions of the present disclosure can also be used to treat, prevent, or ameliorate liver dysfunction, for example, diseases such as hepatitis (A, B, C, D, and E), fatty liver disease (alcoholic and non-alcoholic), autoimmune diseases (autoimmune hepatitis, primary biliary cirrhosis, primary sclerosing cholangitis), genetic diseases (hemochromatosis, Wilson's disease, alpha-1 antitrypsin deficiency), drug-induced liver disease, cancer (e.g., hepatocellular carcinoma), cirrhosis, and liver failure.

[0097] Dosing regimen According to certain embodiments of the present disclosure, multiple doses of a composition of the present disclosure (e.g., a composition comprising a GDF8 inhibitor and a GLP-1 agonist, or a composition comprising a GDF8 inhibitor, an activin A inhibitor, and a GLP-1 agonist) can be administered to a subject over a defined time course. A method according to this aspect of the present disclosure includes sequentially administering multiple doses of a composition of the present disclosure to a subject. As used herein, "sequentially administering" means that each dose of a composition of the present disclosure is administered to a subject at different times, for example, on different days, separated by a predetermined interval (e.g., hours, days, weeks, or months). The present disclosure includes methods comprising sequentially administering to a patient an initial dose of a composition of the present disclosure, followed by one or more secondary doses of a composition of the present disclosure, optionally followed by one or more tertiary doses of a composition of the present disclosure.

[0098] The terms "initial dose," "secondary dose," and "tertiary dose" refer to the time sequence of administration of the compositions of the present disclosure. Thus, the "initial dose" refers to the dose administered at the beginning of a treatment regimen (also referred to as the "baseline dose"); the "secondary dose" refers to the dose administered after the initial dose; and the "tertiary dose" refers to the dose administered after the secondary dose. The initial, secondary, and tertiary doses may all contain the same amount of active ingredient, but will generally differ from each other in terms of administration frequency. However, in certain embodiments, the amount of active ingredient contained in the initial, secondary, and / or tertiary doses will differ from each other during the course of treatment (e.g., adjusted up or down as needed).

[0099] In an exemplary embodiment of the present disclosure, each secondary and / or tertiary dose is administered 1 to 30 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30 or more) days after the immediately preceding dose. The phrase "immediately preceding dose," as used herein, refers to the dose of a composition of the present disclosure that is administered to a patient prior to the administration of the immediately subsequent dose in a multiple administration series, provided there are no intervening doses in the series.

[0100] The method according to this aspect of the disclosure can include administering any number of secondary and / or tertiary doses of the composition of the disclosure to the patient. For example, in certain embodiments, only a single secondary dose is administered to the patient. In other embodiments, two or more (e.g., 2, 3, 4, 5, 6, 7, 8, or more) secondary doses are administered to the patient. Similarly, in certain embodiments, only a single tertiary dose is administered to the patient. In other embodiments, two or more (e.g., 2, 3, 4, 5, 6, 7, 8, or more) tertiary doses are administered to the patient.

[0101] In embodiments involving multiple secondary doses, each secondary dose can be administered with the same frequency as the other secondary doses. For example, each secondary dose can be administered to the patient 1 to 29 days after the immediately preceding dose. Similarly, in embodiments involving multiple tertiary doses, each tertiary dose can be administered with the same frequency as the other tertiary doses. For example, each tertiary dose can be administered to the patient 1 to 60 days after the immediately preceding dose. Alternatively, the frequency with which the secondary and / or tertiary doses are administered to the patient can vary over the course of the treatment regimen. The administration frequency can also be adjusted by the physician during the course of treatment depending on the needs of the individual patient following clinical testing.

[0102] array The sequences referred to herein have the SEQ ID NOs and sequences shown in the informal sequence listing below:

[0103] [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Example]

[0104] The following examples are provided so as to provide those of ordinary skill in the art with a complete disclosure and description of how to make and use the disclosed methods and compositions, and are not intended to limit the scope of what the inventors position as their disclosure. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperatures, etc.), but some experimental error and deviation should be accounted for. Unless otherwise specified, parts are parts by weight, molecular weight is average molecular weight, temperature is in degrees Celsius, and pressure is at or near atmospheric. [Example]

[0105] An in vivo efficacy study to investigate the effects of REGN1033 & REGN2477 alone and in combination with the GLP-1 agonist semaglutide on body composition and glucose metabolism A treatment study was conducted using a diet-induced obese (DIO) mouse model to investigate the effects of GDF8 and activin A inhibition on body composition with or without GLP-1 agonism. DIO mice, fed a high-fat diet for at least 20 weeks, were distributed into four distinct groups balanced by body fat content as determined by MRI and blood glucose. On day 0, all mice were implanted with osmotic pumps (Alzet, Cat#2004), in which the stainless steel flow regulator was replaced with PEEK tubing (DURECT Corporation, Cat#0002496) to allow for MRI measurements during the course of the experiment. Pumps in groups 1 and 2 contained PBS, while pumps used in groups 3 and 4 contained semaglutide (0.833 mg / mL for a 7 μg / day infusion). The following day (day 1), mice were injected with the respective antibodies: Groups 1 and 3: isotype control (at 20 mg / kg); Groups 2 and 4: anti-GDF8 antibody (REGN1033) & anti-activin A antibody (REGN2477) (each at 10 mg / kg). Antibody injections were repeated on days 4 and 7, and then weekly thereafter.

[0106] Body composition using MRI and post-fed blood glucose using a tail-bleed blood glucose meter were measured weekly in the morning. Mice were also bled weekly for insulin measurements. An oral glucose tolerance test (oGTT) was performed on day 23. For the oGTT, mice were fasted for 4 h and baseline glucose was measured. Subsequently, 2 g / kg glucose was administered to mice via oral gavage, and glucose was measured again 30, 60, 90, and 120 min later. On day 26, 6-h fasting blood glucose was measured, and mice were bled for insulin assessment as described above. At the end of the study (days 28 / 29), mice were sacrificed, and tissues (skeletal muscle, fat, pancreas, heart, spleen, and liver) were weighed and collected for further analysis. Mouse plasma was used to measure circulating levels of ALT, AST, nonesterified fatty acids (NEFA), triglycerides (trig), and cholesterol (chol) using a Siemens Advia Chemistry Analyzer. One lobe of the liver was fresh-frozen to allow for triglyceride extraction and quantification using standard protocols. The other lobes of the liver and the pancreas were fixed in 10% formalin, embedded in paraffin, and sectioned for liver histology. H&E and smooth muscle actin (SMA) staining were performed, and slides were subjected to quantification using Halo software. Pancreatic paraffin sections were stained for insulin and glucagon to determine beta- and alpha-cell mass, respectively, using Halo software.

[0107] result Treatment with REGN1033 & REGN2477 in DIO mice resulted in an approximately 10% increase in lean mass and a 15% decrease in fat mass compared to the control group (Group 1). Overall, body weight was unchanged in the isotype groups. Semaglutide treatment reduced body weight (approximately 14% compared to the control group), which was associated with a decrease in fat mass (approximately 18% compared to the control group) and a smaller decrease in lean mass (approximately 3-4% compared to the control group) (Figures 1A-1C). The combination of REGN1033 & REGN2477 and semaglutide (Group 4) appeared to have an additive effect on fat loss (approximately 50% compared to the control group) while maintaining the increase in lean mass observed with the combination of REGN1033 and REGN2477 alone (8% compared to the control group). Due to the increase in lean mass, weight loss in Group 4 was similar to that observed in mice treated with semaglutide alone (Group 3). The greater lean mass mediated by REGN1033 & REGN2477 was the result of increased muscle mass, rather than increased heart or spleen mass, as confirmed by increased skeletal muscle weight (approximately 40% compared to controls) at the end of the study (Figure 2A). In addition, there was a significant reduction in subcutaneous and gonadal fat pads (50% compared to controls) only in Group 4 when REGN1033 & REGN2477 was combined with semaglutide treatment (Figure 2B).

[0108] [Table 2]

[0109] [Table 3]

[0110] [Table 4]

[0111] [Table 5]

[0112] No significant changes in glucose metabolism were observed throughout the study. All semaglutide-treated mice (groups 3 and 4) showed a transient decrease in fed glucose on day 7, but this was no longer observed at later dates (Figure 4A). Fasting glucose was lower in the semaglutide-treated groups at later time points during the study, while no significant changes in glucose tolerance were observed. However, a decrease in circulating insulin levels was observed in all treatment groups compared with the control group, indicating that glucose was controlled using less insulin (Figure 4B). In addition, glucagon levels appeared to be lower in the REGN1033 & REGN2477-treated group (Figure 4C). When REGN1033 & REGN2477 were combined with semaglutide treatment, pancreatic mass was significantly increased in this study (Figure 6A). These effects are thought to be driven by semaglutide and have been previously reported with other GLP-1 agonists or compounds that activate this pathway (PMIDs: 19509017, 22266668). To determine whether the changes in pancreatic mass, insulin, and glucagon were driven by changes in beta or alpha cell mass, pancreatic histology was performed (Figure 6B). There was no significant change in beta cell mass, but a trend toward lower beta cell mass was observed in groups 2 and 4 treated with REGN1033 & REGN2477. Additionally, there was a significant drop in alpha cell mass (approximately 50% compared with the control group) in mice treated with REGN1033 & REGN2477, which could explain the observed decrease in glucagon.

[0113] [Table 6]

[0114] [Table 7]

[0115]

Table 8

[0116]

Table 9

[0117]

Table 10

[0118]

Table 11

[0119]

Table 12

[0120]

Table 13

[0121]

Table 14

[0122]

Table 15

[0123] Because adipose tissue loss is associated with improved liver phenotype, this was also evaluated. Plasma measurements of liver injury markers, ALT and AST, showed clear reductions with all treatments, but this reduction was most pronounced when REGN1033 & REGN2477 were combined with semaglutide treatment (Group 4) (Figures 7A and 7B). In addition, circulating cholesterol was reduced in Group 4 (Figure 7C). Correlating with the improvements in ALT and AST, liver triglyceride content was reduced in all treatment groups (Figure 7F). The combination of REGN1033 & REGN2477 with semaglutide again appeared to have an additive effect in reducing liver triglycerides and ALT. Histological analysis of liver tissue confirmed these results regarding liver triglycerides. In addition, smooth muscle actin staining, a marker of early fibrosis, was reduced in both REGN1033 and REGN2477-treated groups (Figure 9B), indicating that blocking activin A and GDF8 could block liver fibrosis. Neither treatment increased circulating free fatty acids or triglycerides, indicating that fat was being burned and not simply added to the circulation.

[0124] [Table 16]

[0125] [Table 17]

[0126] [Table 18]

[0127] [Table 19]

[0128] [Table 20]

[0129] Overall, these data indicate that the combination of REGN1033 & REGN2477 with semaglutide had an additive effect on fat loss and liver disease improvement (lowering signs of liver damage (ALT / AST), lowering liver triglycerides, reducing steatosis and fibrosis). [Example]

[0130] An efficacy study examining the effect of adding anti-myostatin or a combination of anti-myostatin and anti-activin A to treatment with a GLP-1 agonist An obese NHP study was conducted to examine the weight, liver, and metabolic effects of adding myostatin / activin A blockade to GLP-1R agonism. Figure 10 describes the study and timeline for obese adult male cynomolgus non-human primates (NHPs) with NASH, stratified by weight and % body fat, with n=10 per arm. NHPs have been shown to spontaneously develop obesity, metabolic syndrome, and type 2 diabetes, with metabolic profiles that mirror those of humans (Canter et al., 2018, J Leukoc Biol 104(3):487-497).

[0131] The treatment groups were: · Vehicle; Semaglutide; · Anti-myostatin + anti-activin A; Semaglutide + antimyostatin; and Semaglutide + anti-myostatin + anti-activin A It was decided. REGN1033 is an anti-myostatin antibody, and REGN2477 is an anti-activin A antibody. The GLP-1 agonist used was semaglutide. Semaglutide was administered at 10 μg / kg. The anti-myostatin antibody was administered at 50 mg / kg. The anti-activin A antibody was administered at 50 mg / kg.

[0132] result Adding anti-myostatin treatment to semaglutide resulted in improved weight loss compared with semaglutide monotherapy (Figures 11A and 11B).

[0133] Adding anti-myostatin treatment to semaglutide resulted in improved fat loss compared with semaglutide monotherapy, and adding anti-activin A to anti-myostatin + semaglutide treatment also increased lean mass (Figures 12A-12C). At week 12, adding an anti-myostatin (REGN1033) to semaglutide improved fat loss over semaglutide alone (Figure 12B). Further addition of anti-activin A (REGN2477) also reversed the lean mass loss seen with semaglutide, further increasing lean mass above baseline at week 12 (Figure 12C).

[0134] The triple combination of semaglutide, anti-myostatin antibody and anti-activin A antibody provided the greatest reduction in HbA1c% after 12 weeks of treatment (Figures 13A-13C).

[0135] The triple combination of semaglutide, anti-myostatin antibody and anti-activin A antibody produced the greatest reduction in LDL and the greatest increase in HDL after 12 weeks of treatment (Figures 14A-14E).

[0136] There was an improvement in the AST / ALT ratio at week 12 in all groups (Figures 15A and 15B).

[0137] Food intake was lower following the first dose in all semaglutide groups, but the semaglutide control group recovered to baseline more quickly (Figures 16A and 16B). The anti-myostatin and anti-activin A combination may have slightly increased food intake at the start of dosing, but this change did not magnify over time, as seen by week 12. However, adding an anti-myostatin to semaglutide may help delay the return of the appetite-suppressing effect of semaglutide to baseline, as observed at week 12.

[0138] All groups given semaglutide had lower fluid intake following the first dose, although the triple combination group was less suppressive than the antimyostatin + semaglutide combination (Figure 17A). All semaglutide-treated groups had lower fluid intake from the start of dosing (up to week 12). However, adding antiactivin A to the antimyostatin and semaglutide reduced this thirst suppression.

[0139] Therefore, it was shown that: · Semaglutide and anti-myostatin have synergistic effects on fat loss; · Adding antiactivin A further increases lean mass and further improves fasting glucose, HbA1c, LDL and HDL; The combination of anti-myostatin and anti-activin A alone shows some fat loss and similar lean mass gain to the triple combination but fails to improve circulating lipids and glucose; Food intake was reduced in all semaglutide treatment groups. [Example]

[0140] Myostatin and activin A blockade in non-human primates and humans Postmenopausal women were given 10 mg / kg (anti-myostatin + anti-activin A) IV Q2W. Both were administered for 30 weeks. Thigh muscle volume was measured via MRI and android fat mass was measured via iDXA. Obese non-human primates were administered 50 mg / kg anti-myostatin + 50 mg / kg anti-activin A (QW). Both were administered for 28 weeks. Total lean mass was measured via iDXA and total fat mass was measured via iDXA.

[0141] Myostatin and activin A inhibition resulted in a steady increase in thigh muscle volume (MRI) over the first 10 weeks of treatment in postmenopausal women (Fig. 19A) and a (relatively) steady increase in total lean mass (iDXA) in obese non-human primates (Fig. 19B). Concurrently, myostatin and activin A inhibition resulted in a steady decrease in android fat mass (iDXA) over 30 weeks of treatment in postmenopausal women (Fig. 19C) and a steady decrease in total fat mass (iDXA) over 28 weeks of treatment in obese non-human primates (Fig. 19D).

[0142] Myostatin and activin A inhibition led to significant fat loss in both humans and non-human primates only after significant gains in lean mass, but the effects on fat persisted after lean mass returned to baseline. [Example]

[0143] Adding myostatin and activin A blockade to semaglutide (a GLP-1R agonist) increases energy expenditure in obese non-human primates In this nonhuman primate study of diet-induced obesity, 64 obese male monkeys naive to human immunoglobulin underwent an initial selection screen using a physical examination, metabolic and safety clinical chemistry profiling, hematology, iDEXA (using a GE Lunar iDXA scanner) for body composition, and liver biopsy for scoring of nonalcoholic fatty liver disease plus fibrosis (NAS+). Animals were required to have body fat >25%, fasting glucose ≤350 mg / dL and ≥100 mg / dL, and a liver NAS+ score >4 with a steatosis score >2. From this cohort, 55 monkeys were selected to transition from the high-fat diet on which they had been maintained to a high-fat, high-fructose diet, which is likely to exacerbate any liver disease. The animals were maintained on this diet for 6 weeks, after which they underwent baseline screening for the study with further physical examination for body composition, metabolic and safety clinical chemistry profiling, hematology, and iDEXA (using a GE Lunar iDXA scanner). Food and water intake was also monitored daily. Daily food and water intake over the 6 days was averaged for the baseline readings of these parameters.

[0144] A final cohort of 50 monkeys was selected and balanced across five groups of n=10 each based on the following parameters: body weight, total body fat, and NAS+ score. Secondary parameters considered were fasting glucose, total lean mass, and fasting triglycerides. Baseline characteristics of the five groups are listed in Table 20.

[0145] [Table 21]

[0146] Compounds and Treatment Groups The groupings and dosing for this study are listed below in Table 21, and the design is outlined in Figure 23. There were five groups in this study: vehicle control, GLP-1, α-MSTN, and α-ActA combination, GLP-1 + α-MSTN, and GLP-1 + α-MSTN + α-ActA. All compounds were dosed subcutaneously. GLP-1 was given at 10 μg / kg twice weekly for 20 weeks starting on study day 0, with no dose escalation of GLP-1 due to the low-dose regimen. The group receiving the α-MSTN and α-ActA combination was dosed weekly at 50 mg / kg starting on week 2 through week 12 of the study. All dosing was completed by week 20, after which cohorts were followed for an additional 8-week washout period. The animal numbers listed in Table 21 represent the final counts at the end of the study. There were four deaths that occurred during the course of the study; unless otherwise noted, all data from these monkeys were excluded from the averages.

[0147] [Table 22]

[0148] Liver histology and scoring Liver biopsies were performed twice during the study: once during the animal selection period and once at the end of week 12. Liver tissue samples (approximately 0.5–1.0 cm / sample) were obtained by ultrasound-guided biopsy from animals after an overnight fast under sedation / anesthesia (ketamine, 5–10 mg / kg IM). For all samples, after appropriate location of the right lobe of the liver was identified by ultrasound, the biopsy needle was slowly advanced through the skin, muscle tissue, and liver capsule to reach the liver tissue, and then collected using a 16–18G biopsy needle under direct ultrasound guidance. Biopsies were fixed in 10% neutral-buffered formalin for paraffin embedding. Paraffin-embedded tissue samples were sectioned and simultaneously stained with hematoxylin and eosin (H&E) and Sirius Red. Slides were evaluated for NASH (steatosis, ballooning, inflammation, and fibrosis) by a KBI pathologist.

[0149] double labeled water Total energy expenditure (TEE) was measured using the doubly labeled water method. Subjects were dosed with DLW at 1.51 g / kg of estimated total body water (TBW). The dose consisted of 0.17 g / kg of estimated TBW of 99% 2H2O (Sigma-Aldrich 151882) and 0.3 g / kg of estimated TBW of 97% 2H2O (Sigma-Aldrich 329878) to achieve an initial enrichment relative to body mass. The dose was administered intravenously via a saphenous vein catheter or equivalent, and the syringe was weighed and flushed to ensure all compound was infused. Blood samples were taken at the following time points: 1) pre-dose (baseline), 2) 4-6 hours post-dose (equilibrium), and 3) 1 and 2 weeks post-dose.

[0150] Blood samples (1.5 ml) were collected, transferred to non-liquid sodium heparin anticoagulation tubes, inverted five times, immediately placed on ice, and then centrifuged at 1300 × g for 10 minutes at 4° C. Aliquots were stored at −80° C. and sent to Metabolic Solutions, Inc. for labeling and analysis.

[0151] Briefly, obese non-human primates were administered either i) vehicle, ii) semaglutide, iii) anti-myostatin antibody + anti-activin A antibody, iv) semaglutide + anti-myostatin antibody, or v) semaglutide + anti-myostatin antibody + anti-activin A antibody. Energy expenditure was assessed at the end of the dosing period by DLW performed between weeks 18 and 20 of the study.

[0152] Raw energy expenditure measures showed that primates receiving galetusumab (anti-activin A antibody) + semaglutide + anti-myostatin antibody consumed more energy (Figure 20A). Energy expenditure measures per kg of lean mass showed that the energy gains applied to semaglutide + trevoglumab (anti-myostatin antibody REGN1033) even when normalized per kg of lean mass (Figure 20B).

[0153] Data were graphed to determine whether energy expenditure correlated with lean mass or change in lean mass. The combination treatment groups (semaglutide + trevoglumab, semaglutide + trevoglumab + galetusumab (REGN2477)) consumed more energy than the non-combination treatment groups (vehicle, semaglutide, trevoglumab + galetusumab) (Figure 21A). Furthermore, the combination treatment groups (semaglutide + trevoglumab, semaglutide + trevoglumab + galetusumab) that gained more lean mass also had greater energy expenditure (Figure 21B). Thus, combination-treated primates consumed more energy per gram of lean mass, and their energy expenditure increased in proportion to their gain in lean mass. However, the "boost" in energy expenditure only occurred when both semaglutide and trevoglumab were on-board.

[0154] In addition to semaglutide and anti-myostatin antibodies, activin A blockade also induced further improvements in HbA1c and cholesterol. In obese non-human primate treatment groups, measurements were made of HbA1c (Figure 22A), LDL-C (Figure 22B), ApoB (Figure 22C), change in HbA1c vs. baseline (Figure 22D), change in LDL-C vs. baseline (Figure 22E), and change in ApoB vs. baseline (Figure 22F).

[0155] The combination treatment not only increased lean mass but also improved HbA1c and cholesterol in obese non-human primate subjects.

[0156] The present disclosure is not limited in scope by the specific embodiments described herein. Indeed, various modifications of the present disclosure in addition to those described herein will become apparent to those skilled in the art from the foregoing description and accompanying figures. Such modifications are intended to be encompassed by the appended claims.

Claims

1. growth differentiation factor-8 (GDF-8) inhibitors and glucagon-like peptide-1 (GLP-1) agonists, composition.

2. growth differentiation factor-8 (GDF-8) inhibitors, activin A inhibitors and glucagon-like peptide-1 (GLP-1) agonists, composition.

3. GDF-8 inhibitors are GDF-8 specific binding proteins, The composition according to claim 1 or 2.

4. The GDF-8 inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to GDF-8. The composition according to any one of claims 1 to 3.

5. The anti-GDF8 antibody or antigen-binding fragment thereof comprises a heavy chain complementarity determining region (HCDR) of a heavy chain variable region (HCVR) comprising SEQ ID NO: 4, and a light chain complementarity determining region (LCDR) of a light chain variable region (LCVR) comprising SEQ ID NO:

5. The composition of claim 4.

6. The anti-GDF8 antibody or antigen-binding fragment thereof is heavy chain complementarity determining regions (HCDR1, HCDR2 and HCDR3) comprising SEQ ID NO:6, SEQ ID NO:7 and SEQ ID NO:8, respectively, and three light chain complementarity determining regions (LCDR1, LCDR2 and LCDR3) comprising SEQ ID NO:9, TTS and SEQ ID NO:11, respectively; The composition according to claim 4 or 5.

7. If present, the activin A inhibitor is an activin A specific binding protein. The composition according to any one of claims 2 to 6.

8. When present, the activin A inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to activin A. The composition according to any one of claims 2 to 7.

9. The anti-activin A antibody or antigen-binding fragment thereof comprises a heavy chain complementarity determining region (HCDR) of a heavy chain variable region (HCVR) comprising SEQ ID NO: 12, and a light chain complementarity determining region (LCDR) of a light chain variable region (LCVR) comprising SEQ ID NO:

13. The composition of claim 8.

10. The anti-activin A antibody or antigen-binding fragment thereof comprises heavy chain complementarity determining regions (HCDR1, HCDR2, and HCDR3) comprising SEQ ID NO: 14, SEQ ID NO: 15, and SEQ ID NO: 16, respectively, and three light chain complementarity determining regions (LCDR1, LCDR2, and LCDR3) comprising SEQ ID NO: 17, GAS, and SEQ ID NO: 19, respectively; 10. The composition according to claim 8 or 9.

11. GLP-1 agonists are GLP-1 receptor agonists, The composition according to any one of claims 1 to 10.

12. The GLP-1 agonist is selected from the group consisting of exenatide (long acting), dulaglutide, liraglutide, tirzepatide, and semaglutide; The composition according to any one of claims 1 to 10.

13. The GLP-1 agonist is semaglutide. The composition of claim 12.

14. GLP-1 agonists are GLP-1 specific binding proteins, The composition according to any one of claims 1 to 10.

15. The GLP-1 agonist is an antibody or antigen-binding fragment thereof that specifically binds to GLP-1.

15. The composition of claim 14.

16. For use in improving glucose control, increasing lean body mass, reducing fat mass, treating obesity, treating diabetes, and / or treating liver problems associated with increased fat mass, obesity and / or diabetes in a subject, The composition according to any one of claims 1 to 15.

17. For improving glucose control, increasing lean body mass, reducing fat mass, treating obesity, treating diabetes, and / or treating liver problems associated with increased fat mass, obesity, and / or diabetes in a subject.

1. A method comprising: administering to a subject a GDF8 inhibitor and a GLP-1 agonist; method.

18. For improving glucose control, increasing lean body mass, reducing fat mass, treating obesity, treating diabetes, and / or treating liver problems associated with increased fat mass, obesity, and / or diabetes in a subject.

1. A method comprising: administering to a subject a GDF8 inhibitor, an activin A inhibitor, and a GLP-1 agonist; method.

19. The GDF8 inhibitor and the GLP-1 agonist, and, if present, the activin A inhibitor, are administered to the subject in a single composition; 19. The method of claim 17 or 18.

20. The GDF8 inhibitor and the GLP-1 agonist, and, if present, the activin A inhibitor, are administered to the subject in at least two separate compositions; 19. The method of claim 17 or 18.

21. The method of any one of claims 17 to 20, wherein the GDF-8 inhibitor is a GDF-8 specific binding protein.

22. The GDF-8 inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to GDF-8. The method according to any one of claims 17 to 21.

23. The anti-GDF8 antibody or antigen-binding fragment thereof comprises a heavy chain complementarity determining region (HCDR) of a heavy chain variable region (HCVR) comprising SEQ ID NO: 4 and a light chain complementarity determining region (LCDR) of a light chain variable region (LCVR) comprising SEQ ID NO: 5; 23. The method of claim 22.

24. The anti-GDF8 antibody or antigen-binding fragment thereof is heavy chain complementarity determining regions (HCDR1, HCDR2 and HCDR3) comprising SEQ ID NO:6, SEQ ID NO:7 and SEQ ID NO:8, respectively, and three light chain complementarity determining regions (LCDR1, LCDR2 and LCDR3) comprising SEQ ID NO:9, TTS and SEQ ID NO:11, respectively; 24. The method of claim 22 or 23.

25. If present, the activin A inhibitor is an activin A specific binding protein. The method according to any one of claims 18 to 24.

26. When present, the activin A inhibitor is an antibody or antigen-binding fragment thereof that specifically binds to activin A. The method according to any one of claims 18 to 25.

27. The anti-activin A antibody or antigen-binding fragment thereof comprises a heavy chain complementarity determining region (HCDR) of a heavy chain variable region (HCVR) comprising SEQ ID NO: 12, and a light chain complementarity determining region (LCDR) of a light chain variable region (LCVR) comprising SEQ ID NO:

13.

27. The method of claim 26.

28. The anti-activin A antibody or antigen-binding fragment thereof comprises heavy chain complementarity determining regions (HCDR1, HCDR2, and HCDR3) comprising SEQ ID NO: 14, SEQ ID NO: 15, and SEQ ID NO: 16, respectively, and three light chain complementarity determining regions (LCDR1, LCDR2, and LCDR3) comprising SEQ ID NO: 17, GAS, and SEQ ID NO: 19, respectively; 28. The method of claim 26 or 27.

29. GLP-1 agonists are GLP-1 receptor agonists, The method according to any one of claims 17 to 28.

30. The GLP-1 agonist is selected from the group consisting of exenatide (long acting), dulaglutide, liraglutide, tirzepatide, and semaglutide; The method according to any one of claims 17 to 28.

31. The GLP-1 agonist is semaglutide.

31. The method of claim 30.

32. GLP-1 agonists are GLP-1 specific binding proteins, The method according to any one of claims 17 to 28.

33. The GLP-1 agonist is an antibody or antigen-binding fragment thereof that specifically binds to GLP-1.

33. The method of claim 32.

34. At 12 weeks from administration of said one or more inhibitors and said agonist, the subject: i) at least about a 35% reduction in fat mass; ii) an increase in lean mass of at least about 6%; iii) a reduction in fasting glucose of at least about 15%; iv) a reduction in HbA1c of at least about 6%; v) a reduction in LDL of at least about 14%; vi) an increase in LDL of at least about 14%; vii) at least about a 35% reduction in NEFA; and viii) at least about a 55% decrease in TG exhibiting at least one parameter change selected from the group consisting of: The method according to any one of claims 17 to 33.

35. At 12 weeks from administration of said one or more inhibitors and said agonist, the subject: i) a reduction in fasting glucose of at least about 25%; iv) a reduction in HbA1c of at least about 25%; v) at least about a 50% reduction in LDL; vi) an increase in LDL of at least about 60%; vii) at least about a 50% reduction in NEFA; and viii) at least about a 65% reduction in TG exhibiting at least one parameter change selected from the group consisting of:

35. The method of claim 34.

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