Myostatin-selective inhibitors for treating metabolic disorders

Abstract

Disclosed herein are therapeutic uses of myostatin-selective inhibitors, such as myostatin-selective inhibitors, for treating metabolic disorders, such as obesity. Data presented herein support the notion that selectively targeting myostatin without inhibiting Activin A can achieve equivalent or enhanced efficacy when used in conjunction with a GLP-1 receptor agonist in improving body composition, as compared to broadly antagonizing the ActRII pathway in conjunction with a GLP-1 receptor agonist. Administration of a myostatin-selective inhibitor can prolong the effect of semaglutide on fat mass loss. Moreover, administration of a myostatin-selective inhibitor can elicit positive effects on bone in all parameters evaluated, indicating that diet- or inflammation-induced bone atrophy may be prevented by selective inhibition of the myostatin pathway.

Description

[0001] Myostatin-Selective Inhibitors for Treating Metabolic Disorders

[0002] CROSS REFERENCE TO RELATED APPLICATIONS

[0003] [1] This application claims priority to and the benefit of U.S. Provisional Application No. 63 / 650,380, filed May 21 , 2024; U.S. Provisional Application No. 63 / 650,719, filed May 22, 2024; U.S. Provisional Application No. 63 / 654,722, filed May 31 , 2024; U.S. Provisional Application No. 63 / 662,567, filed June 21 , 2024; U.S. Provisional Application No. 63 / 664,634, filed June 26, 2024; U.S. Provisional Application No. 63 / 741 ,609, filed January 3, 2025; U.S. Provisional Application No. 63 / 783,714, filed April 4, 2025; and U.S. Provisional Application No. 63 / 785,322, filed April 8, 2025. The content of each of these applications is incorporated herein by reference in its entirety.

[0004] FIELD OF THE INVENTION

[0005] [2] This invention relates to therapeutic use of myostatin inhibitors for cardiometabolic indications, such as obesity, diabetes and liver disorders.

[0006] BACKGROUND

[0007] [3] Among metabolic diseases, obesity is a chronic condition that affects more than one in three adults and about 17 percent of children and adolescents in the United States alone. More than one in three adults is overweight. Being overweight or obese increases the risk of type 2 diabetes, heart disease, stroke, fatty liver disease, kidney disease, and other health issues.

[0008] [4] Weight loss efforts often face challenges. For example, extreme dieting such as ultra-low caloric restriction, is impractical, if not unfeasible to sustain for most patients. For example, when a person with obesity syndrome goes on a diet, 10% weight loss can result in about 20-30% decrease in overall energy expenditure, accompanied by reduced basal metabolic rate, reduced leptin concentrations, and reduced lean mass. It is well known that weight loss from dietary / caloric restriction causes not only adipose (fat) tissue mass reduction, but also results in loss of skeletal muscle mass. Coupled with the body’s natural propensity towards maintaining homeostasis by adapting to lower energy supply, reduced skeletal muscle causes a decrease in metabolic rate, lowering energy expenditure. In fact, a 10% loss in overall weight results in a 30-40% reduction in energy expanded during physical activity above resting energy expenditure.

[0009] [5] In addition to more invasive procedures such as gastric bypass surgery, a class of incretin-based weight loss medications have been approved by regulatory authorities (e.g., FDA) to treat overweight and obesity (see, e.g., Williams. Diabetes Ther (2020) 11 : 1199-1216). These include orlistat (XENICAL®, ALLI®), phentermine and topiramate (QSYMIA®), naltrexone, HCI / bupropion, HCI (CONTRAVE®), liraglutide (SAXENDA), semaglutide (WEGOVY®, OZEMPIC®, RYBELSUS®), dulaglutide (TRULICITY®), and setmelanotide (IMCIVREE®).

[0010] [6] Undoubtedly, the availability of incretin-based therapies has revolutionized obesity treatment in recent years. However, it has become clear that a significant portion of the weight loss achieved by these drugs - in fact as high as 40% - may be due to muscle loss, rather than fat loss. Furthermore, many patients experience side effects from incretin therapies that are severe enough to cause discontinuation of these therapies. Upon discontinuation, the weight that was lost during incretin therapy often rebounds quickly. Moreover, by this time, the patients have lost significant amounts of muscle tissue, leaving them at an unhealthier status than started out with. In addition to muscle loss, bone health may also be negatively affected. [7] Many patients on incretin-based therapies achieve 10-15% weigh loss in a relatively short period (e g., several months), but the effect often plateaus after about a year, at which point weight loss progress begins to wane (Medical News Today https: / / www.medicalnewstoday.com / articles / why-weight-loss-drugs-stop-working-how-to-break-past- ozempic-plateau). This typically means switching to a stronger medication or making dietary or exercise changes, although these measures do not always resolve the plateau. Upon discontinuing the incretin-based therapy, the lost weight is quickly regained.

[0011] [8] Myostatin is a negative regulator of muscle mass and is considered a potential target for enhancing muscle mass or preventing muscle loss. Myostatin, along with other related growth factors such as Activin A, signals through the common ActRII receptors to activate the downstream signal transduction pathway that promotes muscle breakdown. Therefore, inhibition of this pathway leads to increased muscle mass. Consistent with this notion, evidence in the literature supports that there is added muscle-enhancing benefit to inhibiting both Activin A and myostatin (see, for example, Latres, et al. (2017, Nature Communications, 8: 15153) demonstrated in mice and non-human primates that inhibition of both Activin A and myostatin leads to a greater increase in skeletal muscle than inhibition of the latter alone. More recently, additive effects of myostatin and Activin A inhibition were confirmed in obese male cynos, where reduction in fat mass was greater when the animals were treated with both anti-myostatin antibody and anti-Activin A antibody in conjunction with a GLP-1 receptor agonist (GLP-1 RA), as compared to anti-myostatin and GLP-1 RA (without anti-Activin A). See, for example, Mastaitis, et al. 2023, Diabetes, 72:207-OR, see also WO 2024 / 064842. These data support the notion that enhanced efficacy may be obtained by inhibiting both myostatin and Activin A.

[0012] [9] Bimagrumab is a monoclonal antibody that binds the ActRIIA / B receptors thereby preventing ligand-induced receptor activation. The rationale for this approach is that it should inhibit all ligands (e.g. , growth factors) that signal through the same receptors, and therefore, the sweeping blockade of the receptor activation should result in augmented efficacy. For example, both myostatin and Activin A are known to negatively affect muscle growth through this pathway. Inhibiting both of these ligands by targeting the shared receptors has been considered a more effective approach than inhibiting only one of the ligands.

[0013]

[0010] In a 2021 article, Heymsfield et al published clinical trial results showing that intravenous administration of bimagrumab to obese, diabetic adult patients every four weeks for 48 weeks, can achieve a significant reduction in total body fat mass, as compared to placebo (Heymsfield et al. Effect of Bimagrumab vs Placebo on Body Fat Mass Among Adults With Type 2 Diabetes and Obesity: A Phase 2 Randomized Clinical Trial. JAMA Netw Open. 2021 ;4(1 ):e2033457.). Subsequently, in preclinical findings by Versanis, bimagrumab dosed at 20 mg / kg was shown to enhance fat mass loss in combination with semaglutide and prevent lean mass loss induced by semaglutide or tirzepatide over the course of 3 weeks in diet-induced obesity (DIO) mice, further confirming that antagonizing the ActRII receptor pathway can have positive impact on body compositions when used in conjunction with GLP-1 RAs. These results have established bimagrumab as the benchmark for myostatin inhibitors for treating metabolic conditions, such as obesity.

[0014]

[0011] Previously, Applicant sought to address the challenges associated with incretin-based weight loss approaches by selectively targeting myostatin to enhance weight management in combination with agents that activate the GLP-1 signaling pathway, aimed to improve body composition ratherthan focusing solely on overall weight loss. A combination of a myostatin-selective inhibitor and a GLP-1 RA treatment enhanced weight loss, while maintaining or increasing lean mass. The combination treatment was also found to enhance oxygen consumption as compared to GLP-1 RA alone, suggesting improved metabolism (WO 2022 / 271867). The myostatin-selective approach was at least in part aimed to avoid potential adverse effects of inhibiting other ligands, Activin A in particular, making this a safer alternative to the broader-spectrum inhibition achieved by antagonizing the ActRII receptors altogether. However, a question remained whether selective inhibition of myostatin alone would be sufficient to produce meaningful therapeutic effects, relative to bimagrumab or other non-selective inhibitors of myostatin that also target Activin A.

[0015] SUMMARY

[0016]

[0012] The present disclosure aims to take into consideration both the safety and efficacy aspects of weight management by addressing the question regarding relative contributions of myostatin and Activin A inhibition in muscle enhancement and whether the improved safety profile of the myostatin-selective approach might overweigh the added benefit of Activin A inhibition in the context of obesity treatment.

[0017]

[0013] Because Activin A plays a role in diverse biological processes, such as reproductive biology, wound healing, neuroprotective function and immune response, in addition to being a negative regulator of muscle mass, systemic inhibition of this pathway raises safety concerns, with respect to interfering with an array of important biological functions mediated by Activin A. To that end, Applicant sought to evaluate relative effects of a myostatin-selective inhibitor and bimagrumab on fat mass and lean mass in DIO mice treated with a GLP-1 RA.

[0018]

[0014] The present disclosure provides, inter alia, surprising data demonstrating that selective inhibitors of myostatin can achieve equivalent or superior efficacy as compared to bimagrumab, in combination with a GLP-1 RA in preclinical DIO mouse models. Data presented herein show that SRK-439 has overall greater potency than bimagrumab as measured by dose-dependent effects on fat mass loss and preservation of lean mass when used in combination with semaglutide. SRK-439 also shows greater durability of effects over 28 days when dosed at 10 mg / kg.

[0019]

[0015] Indeed, in a 12-week study, the effects of the combination treatment on fat loss persisted well beyond the timepoint of plateau observed with a GLP-1 RA monotherapy, demonstrating that concurrent inhibition of myostatin can prolong (e.g., extend) the effect of GLP-1 RA therapy. Moreover, SRK-439 treatment improved the parameters of bone evaluated by micro-CT scan analysis in DIO mice. A high-fat diet was shown to be sufficient to cause a reduction in bone density, and this was largely prevented by SRK-439 treatment, indicating that myostatin-selective inhibition can protect against diet-induced bone loss.

[0020]

[0016] Furthermore, pharmacokinetics and related modeling studies in non-human primates indicate that SRK-439 is predicted to have favorable bioavailability, rendering it amenable to subcutaneous administration in human patients.

[0021]

[0017] Taken together, myostatin-selective inhibitors such as SRK-439 can not only provide a safer weight management approach than ActRII antagonists such as bimagrumab, but also achieve competitive efficacy, despite targeting myostatin alone.

[0022]

[0018] Also provided herein is the identification of a particularly effective epitope on proMyostatin. Targeting the epitope with antibodies can effectuate potent inhibition of myostatin activation.

[0023] BRIEF DESCRIPTION OF THE FIGURES

[0024]

[0019] The patent or application file contains at least one drawing executed in color. Copies of this patent application with color drawing(s) will be provided by the Office upon request and payment of the necessary fee.

[0025]

[0020] FIG. 1 provides two graphs showing the effect of SRK-439 on fat mass and lean mass, respectively, in combination with high and low doses of semaglutide. FIG. 1A shows percent change in lean mass from baseline when DIO mice were treated with muSRK-439 at 0.1 mg / kg (mpk), 1.0 mg / kg, 3.0 mg / kg, and 10 mg / kg in conjunction with either high dose (0.04 mg / kg) or low dose (0.01 mg / kg) semaglutide. FIG. 1 B shows percent change in fat mass from baseline when DIO mice were treated with muSRK-439 at 0.1 mg / kg, 1.0 mg / kg, 3.0 mg / kg, and 10 mg / kg in conjunction with either high dose (0.04 mg / kg) or low dose (0.01 mg / kg) semaglutide.

[0021] FIG. 2 provides data showing fasted glucose levels in mice treated with varying doses of muSRK-439 in combination with semaglutide.

[0026]

[0022] FIG. 3 provides two graphs showing effects of SRK-439 treatment in a paradigm where semaglutide treatment is discontinued (e.g., “regain” study). FIG. 3A shows changes in lean mass as measured by qNMR over the study duration. FIG. 3B shows changes in fat mass as measured by qNMR over the study duration.

[0027]

[0023] FIG. 4 provides three graphs showing the effect of SRK-439 treatment at study end on relative fat mass (FIG. 4A), weight in inguinal fat pad (FIG. 4B), and serum leptin levels (FIG. 4C), respectively, in a study where semaglutide treatment was discontinued.

[0028]

[0024] FIG. 5 provides pharmacokinetic data in cynomolgus monkeys showing serum concentrations of SRK-439 over time in groups that received single-dose administrations of SRK-439 subcutaneously at 0.3 mg / kg (Group 1 ), 3 mg / kg (Group 2), 10 mg / kg (Group 3), and 30 mg / kg (Group 4), and another group that received a single-dose administration of SRK-439 intravenously at 10 mg / kg (Group 5).

[0029]

[0025] FIG. 6 provides two graphs showing effects of a murine version of SRK-439 or a murine version of bimagrumab in DIO mice treated with semaglutide. All antibodies were of murine lgG1 subtype. FIG. 6A shows changes in body weight over the duration of 28 days. FIG. 6B shows equivalent effects achieved by high-dose murine SRK-439 (at 10 mg / kg) and high-dose murine bimagrumab (at 20 mg / kg). “Bima” refers to an anti-ActRII antibody comprising the VHA / L regions of bimagrumab fused to the murine lgG1 + lambda constant domains; “SRK-439” refers to an anti-pro / latent myostatin comprising the VHA / L regions of SRK-439 fused to murine lgG1 constant domains.

[0030]

[0026] FIG. 7 provides three graphs showing the effect of murine SRK-439 or murine bimagrumab at low doses (0.3 mg / kg, 1 mg / kg, or 3 mg / kg) in combination with semaglutide. FIG. 7A shows percent changes in lean mass as measured by qNMR over 28 days. FIG. 7B demonstrates efficacy of murine SRK-439 at 0.3 mg / kg in preventing further lean mass loss and lack of efficacy of murine bimagrumab at the same dose. FIG. 7C shows changes in lean mass over time as measured by qNMR. “Bima” refers to an anti-ActRII antibody comprising the VHA / L regions of bimagrumab fused to the murine lgG1 + lambda constant domains; “SRK-439” refers to an anti-pro / latent myostatin comprising the VH / VL regions of SRK-439 fused to murine lgG1 constant domains.

[0031]

[0027] FIG. 8 provides two graphs showing the effect of murine SRK-439 at 10 mg / kg or murine bimagrumab at 20 mg / kg on their ability to preserve lean mass during semaglutide treatment at study end. FIG. 8A shows percent changes in lean mass at the end of the 28-day study over baseline. FIG. 8B shows percent changes in lean mass as measured by qNMR in mice treated with 10 mg / kg of murine SRK-439 and mice treated with 20 mg / kg murine bimagrumab over 28 days. “Bima” or “bimagrumab” refers to an anti-ActRII antibody comprising the VH / VL regions of bimagrumab fused to the murine lgG1 + lambda constant domains; “SRK-439” refers to an anti-pro / latent myostatin comprising the VH / VL regions of SRK-439 fused to murine I gG1 constant domains.

[0032]

[0028] FIG. 9 provides two graphs showing the effect of murine SRK-439 or murine bimagrumab treatment on individual muscle weights in mice treated with semaglutide. FIG. 9A shows absolute weight of gastroc at the end of the 28-day study, and FIG. 9B shows absolute weight of quad at the end of the 28-day study. “Bima” or “Bimagrumab” refers to an anti-ActRII antibody comprising the VHA / L regions of bimagrumab fused to the murine lgG1 + lambda constant domains; “SRK-439” refers to an anti-pro / latent myostatin comprising the VHA / L regions of SRK-439 fused to murine lgG1 constant domains. ****P<0.0001 (One-Way Anova, Tukey’s multiple comparison analysis).

[0033]

[0029] FIG. 10 provides four graphs showing the effect of murine SRK-439 or murine bimagrumab on fat mass in mice treated with semaglutide. FIG. 10A shows fat mass as measured by qNMR over the study duration in mice receiving varying concentrations of murine SRK-439, murine bimagrumab, or control mouse IgG, in combination with 0.04 mg / kg if semaglutide. FIG. 10B shows changes in fat mass as measured by qNMR over study duration in mice receiving either 10 mg / kg murine SRK-439 or 20 mg / kg murine bimagrumab, in combination with semaglutide. FIG. 10C shows percent changes in fat mass over 28 days in mice receiving either 10 mg / kg murine SRK-439 or 20 mg / kg murine bimagrumab, in combination with semaglutide. FIG. 10D shows percent change in fat mass measured at the end of the 28-day study in mice treated with varying doses of murine SRK-439 or murine bimagrumab in combination with semaglutide. “Bima” or “Bimagrumab” refers to an anti-ActRII antibody comprising the VH / VL regions of bimagrumab fused to the murine lgG1 + lambda constant domains; “SRK-439” refers to an anti-pro / latent myostatin comprising the VH / VL regions of SRK-439 fused to murine I gG1 constant domains.

[0034]

[0030] FIG. 11 provides data summarizing body composition, expressed as percent of body weight, as determined at the end of the 28-day study in mice treated with varying doses of murine SRK-439 or murine bimagrumab in combination with 0.04 mg / kg semaglutide. “Bimagrumab” refers to an anti-ActRII antibody comprising the VH / VL regions of bimagrumab fused to the murine lgG1 + lambda constant domains; “SRK-439” as used in this figure refers to an anti-pro / latent myostatin comprising the VH / VL regions of SRK-439 fused to murine lgG1 constant domains.

[0035]

[0031] FIG. 12 provides a graph showing relative serum concentrations (micrograms per milliliter) of murine SRK-439 in mice treated with 0.3 mg / kg, 1.0 mg / kg, 3.0 mg / kg, or 10 mg / kg of murine SRK-439, measured at the outset and the end of the 28-day study.

[0036]

[0032] FIG. 13 provides data showing effects of a combination of a myostatin-selective inhibitor and metformin on fat mass (FIG. 13A) and lean mass (FIG. 13B), as measured by qNMR, in mice on high-fat diet.

[0037]

[0033] FIG. 14 provides data showing effects of a combination of myostatin-selective inhibitor and metformin on body weight (FIG. 14A), lean body mass (FIG. 14B), and fat mass (FIG. 14C) at day 63 of treatment as compared to the pretreatment baseline, as measured by qNMR in mice on high-fat diet, ns, not significant; *P<0.05, ****P<0.0001 .

[0038]

[0034] FIG. 15 provides a graph showing changes in body weight (grams) over time. DIO mice treated with tirzepatide (0.045 mg / kg) lost body weight, with or without SRK-439-mlgG1 , and mice receiving IgG control or SRK-439-mlgG1 alone did not lose body weight over the course of the study period.

[0039]

[0035] FIG. 16 provides data showing effects of SRK-439-mlgG1 and tirzepatide combination on fat mass and lean mass in DIO mice. FIG. 16A shows that SRK-439-mlgG1 (3 mg / kg or 10 mg / kg) in combination with tirzepatide enhanced fat mass loss, as compared to tirzepatide alone. FIG. 16B shows that SRK-439-mlgG1 was able to rescue lean mass lost during tirzepatide treatment. FIG. 16C shows that SRK-439-mlgG1 increased lean mass and enhanced fat mass loss when used in combination with tirzepatide, as determined by qNMR.

[0040]

[0036] FIG. 17 provides three graphs showing tissue weights by qNMR in mice treated with SRK-439-mlgG1 with or without tirzepatide: left side fat pads (infuinal / perigonadal) (FIG. 17A); right side fat pad (inguinal / perigonadal) (FIG. 17B); and left and right gastroc (average) (FIG. 17C).

[0041]

[0037] FIG. 18 provides data showing effects of SRK-439-mlgG1 on body composition in mice treated with tirzepatide.

[0042]

[0038] FIG. 19 changes in fat mass relative to total body weight in DIO mice treated with tirzepatide, SRK-439-mlgG1 , or in combination.

[0043]

[0039] FIG. 20 provides data showing that tirzepatide treatment causes a reduction in liver weight in DIO mice, which is not affected by SRK-439-mlgG1.

[0044]

[0040] FIG. 21 provides data showing free latent myostatin (latent myostatin not bound by antibody) levels following treatment for 22 days with Ab109 or Ab141 when dosed at 0.1 mg / kg to 3 mg / kg alone or in combination with semaglutide.

[0045]

[0041] FIG. 22 provides data showing the level of free latent myostatin in mice treated with Ab109, Ab133, and Ab141 compared to control IgG antibody. Data for individual animals are shown in each plot.

[0042] FIG. 23 provides data showing levels of free latent myostatin in mice following treatment with Ab109 at 2 mg / kg or 20 mg / kg.

[0046]

[0043] FIG. 24 provides two graphs showing the effect of muSRK-439 on lean mass (FIG. 24A) and on fat mass (FIG. 24B) in mice on a high-fat diet, either alone or in combination with semaglutide over the course of 35 days.

[0047]

[0044] FIG. 25 provides two graphs showing the effect of muSRK-439 on forelimb grip strength measured on days 0 and 21 , in mice on a high-fat diet, either alone or in combination with semaglutide. Changes in forelimb grip strength from Day 1 to Day 21 are shown in FIG. 25A. The same data are expressed as % change from the baseline in FIG. 25B.

[0048]

[0045] FIG. 26 provides two graphs showing the effect of muSRK-439 on forelimb-hindlimb grip strength measured on days 0 and 21 , in mice on a high-fat diet, either alone or in combination with semaglutide. Changes in forelimb + hindlimb grip strength from Day 1 to Day 21 are shown in FIG. 26A. The same data are expressed as % change from the baseline in FIG. 26B.

[0049]

[0046] FIG. 27 provides a graph showing the rate of glucose infusion (mg / kg / minute) as a function of time in study groups treated with muSRK-439 or IgG control, either alone or in combination with semaglutide or vehicle.

[0050]

[0047] FIG. 28 provides two graphs showing sex-dependent effects of SRK-439 and / or tirzepatide on body composition in DIO mice. Relative amounts of fat mass and lean mass at study end are expressed as percent of body weight (FIG. 28A) and absolute mass in grams (FIG. 28B) in male and female cohorts.

[0051]

[0048] FIG. 29 provides two graphs showing durable effects of a myostatin-selective inhibitor on lean mass as measured by qNMR in DIO mice in a 12-week study. FIG. 29A shows changes in lean mass over 12 weeks, and FIG. 29B shows percent change from baseline in lean mass over 12 weeks. Semaglutide monotherapy caused a 6.8% loss in lean mass at study end, which was rescued by SRK-439 administration.

[0052]

[0049] FIG. 30 provides two graphs showing prolonged and enhanced fat mass loss in DIO mice treated with a combination of SRK-439 and semaglutide, as measured by qNMR. FIG. 30A shows changes in fat mass over 12 weeks, and FIG. 30B shows percent change from baseline in fat mass over 12 weeks. Semaglutide monotherapy caused rapid and significant loss in fat mass, which appeared to plateau at around 3 weeks. Combination treatment with SRK-439 prolonged this effect through about 9 weeks. SRK-439 / semaglutide combination treatment further enhanced fat mass loss.

[0053]

[0050] FIG. 31 provides two graphs summarizing percent change from baseline in lean mass (FIG. 31A) and fat mass (FIG. 31B) at study end (12 weeks or day 84). SRK-439 treatment increased lean mass and decreased fat mass as monotherapy. SRK-439 in combination with semaglutide prevented lean mass loss and enhanced fat mass loss.

[0054]

[0051] FIG. 32 provides two graphs showing sustained effects of SRK-439 on body composition. Relative body composition was evaluated at midpoint (day 42) (FIG. 32A) and at endpoint (day 84) (FIG. 32B) of the study, demonstrating the effects of SRK-439 used in combination with a GLP-1 RA were maintained over the duration of the study.

[0055]

[0052] FIG. 33 provides four graphs showing results from tissue weight analysis at study end. FIG. 33A: gastrocnemius (gastroc) (left and right average); FIG. 33B: right femur; FIG. 33C: inguinal fat pad average; and FIG. 33D: epididymal fat pad average, in grams. Tissue weights corroborate the qNMR findings. SRK-439 treatment increased lean mass and enhanced fat pad loss (in combination with semaglutide).

[0056]

[0053] FIG. 34 provides images and structural components of bone. FIG. 34A depicts the architecture of a long bone.

[0057] Representative cross-sectional images are shown on the right. FIG. 34B shows the sections / segments of femur analyzed in this study. FIG. 34C provides cross-sectional images of representative sections from the metaphyseal segment and the diaphyseal segment evaluated in the study.

[0058]

[0054] FIG. 35 provides data showing the effects of SRK-439 treatment, with or without semaglutide, on bone volume in DIO mice from a 12-week study. FIG. 35A provides a graph showing bone volume as measured by micro-CT at metaphysis in study groups treated with or without SRK-439 and / or semaglutide. SRK-439 monotherapy caused an increase in bone volume as compared to control I gG treatment, and the effect was augmented by combination treatment of SRK-439 and semaglutide. FIGs. 35B and 35C show bone volume fraction measured at metaphysis and diaphysis, respectively. SRK-439 treatment, with or without semaglutide, increased bone volume fractions.

[0059]

[0055] FIG. 36 provides analyses of trabecular bone from DIO mice. FIG. 36A provides images adapted from Gautam et al. (2014, British Journal of Nutrition, 111 (10): 1811 -1812) showing that in both males and females, high-fat diet (HFD) induces greater porosity (less density) in trabecular bone, as compared to normal chow. FIG. 36B provides a graph showing trabecular thickness in study groups treated with or without SRK-439 and / or semaglutide. SRK-439 treatment, with or without semaglutide, caused an increase in trabecular thickness. FIG. 36C provides a graph showing trabecular separation in study groups. SRK-439 treatment caused decreased separation, indicative of higher bone density, and this effect was further enhanced in mice treated with a combination of SRK-439 and semaglutide. FIG. 36D provides a graph showing trabecular number in study groups. HFD alone caused a significant reduction in trabecular number indicating that the quality of bone is compromised by the diet high in fat. SRK-439 treatment partially restored trabecular number, and the combination therapy further improved this effect.

[0060]

[0056] FIG. 37 provides micro-CT data showing the effects of SRK-439 on bone mineral density (BMD) with or without semaglutide in DIO mice, as measured by micro-CT. FIG. 37A provides a graph showing percent porosity measured at the metaphysis, a region of bone growth. FIG. 37B provides a graph showing bone mineral density in the surrounding cortical bone. SRK-439 treatment led to less porous metaphysis and corresponding increases in BMD.

[0061]

[0057] FIG. 38 provides micro-CT data showing the effects of SRK-439 on diaphysis with or without semaglutide. FIG. 38A provides a graph showing cortical bone area at the diaphysis in study groups. FIG. 38B provides a graph showing bone volume fraction at diaphysis. SRK-439 treatment increased cortical bone area and bone volume fraction with or without semaglutide.

[0062]

[0058] FIG, 39 provides micro-CT data showing the effects of SRK-439 on cortical bone at diaphysis, with or without semaglutide. FIG. 39A provides a graph showing percent cortical bone porosity in study groups, and FIG. 39B provides a graph showing cortical thickness at diaphysis. SRK-439 treatment increased bone density and thickness, indicative of enhanced bone strength.

[0063]

[0059] FIG. 40 provides two graphs showing the effect of SRK-439 on whole body lean mass in non-human primates, as measured by dual-energy X-ray absorptiometry (DEXA) scan. FIG. 40A shows whole body lean muscle over the duration of the study. FIG. 40B shows percent change in whole body lean muscle from baseline.

[0064]

[0060] FIG. 41 provides two graphs showing the effects of SRK-439 on trunk lean mass (trunk lean mass + BMC) in non-human primates, as measured by DEXA scan. FIG. 41A shows trunk lean mass over the duration of the study. FIG. 41 B shows percent change in trunk lean mass from baseline.

[0065]

[0061] FIG. 42 provides two graphs showing the effect of SRK-439 on hindlimb lean mass (FIG. 42A) and forelimb lean mass (FIG. 42B) in non-human primates as measured by DEXA scan.

[0066]

[0062] FIG. 43 provides four graphs showing the effect of SRK-439 on bicep muscle weight (FIG. 43A), gastrocnemius (gastroc) muscle weight (FIG. 43B), tibialis muscle weight (FIG. 43C), and soleus muscle weight (FIG. 43D) in non- human primates at the study end compared to a vehicle control.

[0063] FIG. 44 provides a graph showing the pharmacokinetics of SRK-439 in non-human primates dosed with 0.3 mg / kg, 3 mg / kg, or 10 mg / kg of the antibody over the study duration.

[0067]

[0064] FIG. 45 provides two graphs showing the pharmacodynamics of SRK-439 in non-human primates dosed with 0.3 mg / kg, 3 mg / kg, or 10 mg / kg of the antibody over the study duration. Serum concentrations of total latent myostatin (FIG. 45A) and free latent myostatin (FIG. 45B) are shown.

[0068] DETAILED DESCRIPTION OF CERTAIN EMBODIMENTS

[0069] Definitions

[0070]

[0065] Adjunct therapy. The terms “adjunct therapy” and “add-on” therapy are used interchangeably herein and are intended to refer to a therapeutic regimen in which a second agent (used as an adjunct therapy) is administered to a subject who is on, has received, or to be treated with, a first agent (e.g., background therapy). The terms “in conjunction with” and “complementary to” are used interchangeably herein and intended to refer to therapies used together, whether concurrent or partially overlapping in time. In various embodiments of the present disclosure, an incretin mimetic can be used as an adjunct or add-on therapy to myostatin inhibitor therapy, and conversely, a myostatin inhibitor can be used as an adjunct or add-on therapy to an incretin mimetic for treating metabolic disorders, such as obesity.

[0071]

[0066] Administer / administration: The terms “administer,” “administering,” or “administration” include any method or act of delivery of a pharmacological agent (e.g., a medicament) to an intended subject (e.g., a patient). The pharmacological agent may be any suitable therapeutic agent, such as a biologic agent, such as an antibody or an antigen-binding fragment thereof (e.g., a pharmaceutical composition comprising such an antibody or antigen-binding fragment), a peptide agent (e.g., a hormone or a modified analog thereof), or a low molecular weight agent (e.g., a structurally defined small molecule or chemical entity). The administration can be systemic or local administration. In some embodiments, administration involves one or more agents that can be administered concurrently, simultaneously, or sequentially.

[0072]

[0067] Antibody. As used herein, the term “antibody” refers to full-length immunoglobulin molecules comprising four polypeptide chains, two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds. Each heavy chain is comprised of a heavy chain variable region (abbreviated herein as HCVR or VH) and a heavy chain constant region. The heavy chain constant region is comprised of three domains, CH1 , CH2 and CH3. Each light chain is comprised of a light chain variable region (abbreviated herein as LCVR or VL) and a light chain constant region. The light chain constant region is comprised of one domain, CL. The VH and VL regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR). Each VH and VL is composed of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1 , CDR1 , FR2, CDR2, FR3, CDR3, and FR4. Antibodies provided in the present disclosure include human antibodies and humanized antibodies.

[0073]

[0068] Antigen-binding fragment'. The terms “antigen-binding fragment,” “antigen binding fragment,” “antigen-binding portion,” “antibody fragment,” or “antibody portion” are used interchangeably herein and refer to one or more fragments of an antibody that retain the ability to specifically bind to an antigen (e.g., pro / latent myostatin). It has been shown that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody. Examples of binding fragments encompassed within the term “antigen-binding fragment” of an antibody include (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CL and CH1 domains; (ii) a F(ab')2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; (iii) a Fd fragment consisting of the VH and CH1 domains; (iv) a Fv fragment consisting of the VL and VH domains of a single arm of an antibody, (v) a dAb fragment (Ward et al., (1989) Nature 341 :544-546), which consists of a VH domain; (vi) an isolated complementarity determining region (CDR); and (vii) an adnectin. Furthermore, although the two domains of the Fv fragment, VL and VH, are coded for by separate genes, they can be joined, using recombinant methods, by a synthetic linker that enables them to be made as a single protein chain in which the VL and VH regions pair to form monovalent molecules (known as single chain Fv (scFv); see e.g., Bird et al. (1988) Science 242:423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883). Such single chain antibodies are also intended to be encompassed within the term “antigenbinding portion” of an antibody. Other forms of single chain antibodies, such as diabodies are also encompassed. Diabodies are bivalent, bispecific antibodies in which VH and VL domains are expressed on a single polypeptide chain, but using a linker that is too short to allow for pairing between the two domains on the same chain, thereby forcing the domains to pair with complementary domains of another chain and creating two antigen binding sites (see e.g., Holliger, P. et al. (1993) Proc. Natl. Acad. Sci. USA 90:6444-6448; Poljak, R. J. et al. (1994) Structure 2:1121-1123). Antigenbinding fragments may be incorporated into engineered constructs, such as multi-functional constructs comprising the antigen-binding fragment. Non-limiting examples of such engineered constructs include multi-specific antibodies, such as bispecific antibodies. In some embodiments, the bispecific antibody comprises a Fab fragment of any one of the novel antibodies disclosed herein, which allows single-arm binding to pro / latent myostatin.

[0074]

[0069] Augment, augmentation (of effect)'. As used herein in the context of combination therapy, the term “augment,” “augmentation” or the like refers to improved effects achieved by using two or more therapies in conjunction with each other to produce an increase in the magnitude of effects (efficacy measures) and / or an increase in the duration of effects (durability), as compared to a monotherapy. Preferably, a myostatin-selective inhibitor is said to augment the effect of an incretin-based therapy, when used in combination, by enhancing the effect of the incretin-based therapy in driving fat mass loss and also extending or prolonging the effect of the incretin-based therapy as compared to when incretin-based therapy is used alone.

[0075]

[0070] Bioavailability: As used herein, the term bioavailability (e.g., of a test article, such as antibodies) refers to serum exposure achieved by a particular route of administration. Bioavailability is typically considered relative to serum exposure achieved by direct infusion of the test article, e.g., intravenous infusion, as 100%. Therefore, percentage of serum exposure achieved by subcutaneous administration of a test article (e.g., monoclonal antibody) relative to serum exposure achieved by intravenous administration of the same test article (e.g., monoclonal antibody) when dosed equal amounts to a subject, can be calculated. Commonly, serum exposure can be expressed as area under the curve (AUCt) (i.e., serum concentration of the test article for a predetermined duration of time, t). As an example, if 50% of subcutaneously administered antibody reaches circulation, the antibody is said to have 50% bioavailability. The bioavailability of subcutaneously administered monoclonal antibodies in humans is typically 50-85% (Reviewed in Davis et al, 2024, Clin Pharm & Therapeutics, 115(3): 422-439).

[0076]

[0071] Body composition: The term “body composition” refers to relative components that make up a body, including fat mass, muscle (lean) mass, bone, and water etc. In particular, in the context of weight management, body composition refers to the ratio of muscle mass to fat mass in the body. Unless explicitly stated otherwise, body composition refers to the body composition of the whole body. Body composition can be measured by various suitable methods known in the art, including but not limited to body density, dual energy X-ray absorptiometry (DEXA), air displacement plethysmography (ADP), bioelectrical impedance analysis (BIA), body volume indicator (BVI), skin folds (with measuring caliper), ultrasound, quantitative magnetic resonance (QMR), and measurement of circumferences (e.g., as measured at waistline).

[0077]

[0072] Body mass index (BMI): The term “body mass index” or “BMI” is a numerical value derived from the mass and height of a person and is defined as weight in kilograms divided by height in meters squared (expressed in units of kg / m2). BMI provides general body weight-height relationships which can be used to categorize a person as “underweight” (BMI of < 18.5), “normal / healthy” weight (BMI of 18.5 to 24.9), “overweight” (BMI of 25 to 29.9), “obese” (BMI of 30 and above), based on tissue mass (muscle, fat and bone) relative to height. In some embodiments, BMI of 40 and above is separately classified as “extremely obese” (BMI of 40+), in which case “obese” is classified as BMI of

[0078] 30 and 39.9.

[0079]

[0073] Bone density: Bone density provides information as to how porous bones are. High bone density typically correlates with stronger bone, and low bone density generally correlates with weakened bone with greater fracture risk. Bone density is typically measured as bone mineral density. Bone mineral density, or BMD, is the amount (e.g., mass) of mineral per volume of bone. A BMD test measures calcium and other minerals such as phosphorus in bone. Bones containing more minerals are denser, so they tend to be stronger and less likely to break. Clinically bone density is usually measured by proxy; for example, according to optical density per area of bone surface upon imaging. Bone densitometry is a test of bone density and may be used to measure bone mineral content and density. Known techniques for measuring bone density include DEXA scan, solid-state MRI, and certain CT scan (e.g., cone beam CT (CBCT)). See Jones et al. Radiology. 2023 Jan 24;307(2):e221810. DEXA scan is often considered the gold standard to provide accurate information on bone density. Combination therapy: As used herein, “combination therapy” refers to a therapeutic regimen involving administration of two or more active agents (e.g., two or more pharmacological agents) intended to treat a predetermined indication and / or conditions associated therewith. The two or more agents may be formulated as separate compositions (e.g., formulations) or may be formulated as a single composition (formulation). “Combination therapy” encompasses therapies used in conjunction with each other and complementary to each other.

[0080]

[0074] Decrease / reduce: The term “decrease” or “reduce,” as used herein, in the context of a disease symptom refers to a statistically significant decrease in such level. The decrease can be, for example, at least 1 %, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95%. The decrease can also be, for example, about 1-10%, 10-20%, 1-30%, 20-50%, 30-60%, 40-70%, 50-80%, or 60-90%. In certain embodiments, an individual with a disorder achieves a level of reduced effect that is comparable or within the normal range for that effect in an individual without such disorder.

[0081]

[0075] Dieting / diet regimen: In the context of the present disclosure, certain dieting may be incorporated as part of weight management, e.g., obesity treatment which includes a pharmacological intervention. Dieting or diet regimen may include caloric / calorie restriction (i.e., reduced calorie intake or reduced absorption of calories) as well as alterations in choices about the type of food consumed (e.g., high protein, lower fat, and / or lower carbohydrate regimens), and / or regimented timing / schedules of food intake (e.g., intermittent fasting). Thus, a patient is on a “diet or reduced calorie regimen” when the patient incorporates or is instructed by or in consultation with a physician or equivalent to incorporate dieting into overall therapeutic regimen, e.g., as part of weight management.

[0082]

[0076] Dose: Dose of a therapeutic to be administered to subjects can be flat dose or weight-based dose.

[0083]

[0077] Effective amount: As used herein, the terms “effective amount,” “effective dose,” and “therapeutically effective amount” are used interchangeably and refer to any amount or dose of a compound or composition that is sufficient to result in a desired biological or medicinal effect in a tissue or subject. For example, in certain embodiments of the present disclosure, the intended purpose is to inhibit activation of myostatin in vivo or to achieve a clinically meaningful outcome associated with the myostatin inhibition. For any particular pharmaceutical agent, a therapeutically effective amount (and / or an appropriate unit dose within an effective dosing regimen) may vary, for example, depending on route of administration, on combination with other pharmaceutical agents. In some embodiments, the specific therapeutically effective amount (and / or unit dose) for any particular patient may depend upon a variety of factors including the disorder being treated and the severity of the disorder; the activity of the specific pharmaceutical agent employed; the specific composition employed; the age, body weight, general health, sex, and diet of the patient; the time of administration, route of administration, and / or rate of excretion or metabolism of the specific pharmaceutical agent employed; the duration of the treatment; and like factors as is well known in the medical arts. In some embodiments, an effective amount refers to an amount that, when administered according to a particular regimen, produces a positive physiological or clinical outcome with a reasonably acceptable level of adverse effects (e.g., toxicity), such that the adverse effects, if present, are tolerable enough to continue the experiment or tolerable enough for a patient to continue the therapeutic regimen, and the benefit of the therapy outweighs the toxicity. Those of ordinary skill in the art will appreciate that in some embodiments of the disclosure, the administered amount is considered an effective amount if it contains an amount appropriate for administration that is correlated with a positive outcome.

[0084]

[0078] Exercise / exercise regimen'. As used herein, the term “exercise” includes any physical activities. The term “exercise regimen” refers to a treatment regimen that incorporates physical activity as a component.

[0085]

[0079] GLP-1 analog'. As used herein, the term “GLP-1 analog” refers to a peptide or modified peptide bearing structural similarities to the naturally occurring GLP-1 and is capable of binding to and activating the GLP-1 receptor. A GLP-1 analog may be an extendin-based therapy, a DPP-IV-resistant analog. Non-limiting examples of GLP-1 analogs include, albigl utide, beinaglutide, cotadutide, danuglipron, dulaglutide, exenatide, exenatide ER, liraglutide, lixisenatide, PEG-loxenatide, mazdutide, MEDI0382, noiiglutide, orforglipron, pemvidutide, PF-07081532, retatrutide, semaglutide, taspoglutide, tirzepatide, and XW003. GLP-1 analogs shall include analogues of human GLP-1 conjugated to substances that slow renal excretion, e.g., fatty acids, albumin, alpha-aminoisobutyric acid, etc.; they may be acylated. GLP-1 analogs shall include peptides or modified peptides comprising an amino acid sequence GTFTSD (SEQ ID NO: 1000). In some embodiments, the GLP-1 analog comprises the sequence EGTFTSD (SEQ ID NO: 116). In some embodiments, the GLP-1 analog comprises the sequence QGTFTSD (SEQ ID NO: 1001 ). GLP-1 analogs may also include peptides or modified peptides comprising an amino acid sequence HXXGXFTXD (SEQ ID NO: 117), wherein X is any amino acid residue.

[0086]

[0080] GLP-1 receptor agonist'. The term “GLP-1 receptor agonist” or “GLP-1 R agonist” or “GLP-1 RA” as used herein refers to an agent capable of binding to and activating the GLP-1 receptor. GLP-1 is a naturally occurring agonist of the GLP-1 receptor. GLP-1 receptor agonists encompass GLP-1 analogs. A GLP-1 receptor agonist may be a small molecule GLP-1 receptor agonist. In some embodiments, the GLP-1 receptor agonist is a long-acting small molecule GLP-1 receptor agonist. In some embodiments, the GLP-1 receptor agonist is a GLP-1 analog.

[0087]

[0081] GLP-1 pathway activator. The terms “GLP-1 pathway activator” and “activator of the GLP-1 signaling pathway” are used interchangeably herein and encompass any agent that increases or enhances the activity of the GLP-1 signaling pathway, irrespective of the mechanism of action. Increased or enhanced activity of the GLP-1 signaling pathway may be a result of, for example, a greater degree of activity, longer duration of activity, increased availability of one or more components of the signaling pathway, etc. In some embodiments, GLP-1 pathway activators include agents that modulate upstream regulators of GLP-1 (e.g., dipeptidyl peptidase (DPP-IV) inhibitors). The term GLP-1 pathway activator as used herein encompasses GLP-1 receptor agonists and GLP-1 analogs. In some embodiments, GLP-1 pathway activators include agents that regulate the amount or activity of GLP-1 (e.g., agents that increase production of or secretion of GLP-1 ; GLP-1 stabilizers). In some embodiments, GLP-1 pathway activators include agents that increase activation of the GLP-1 receptor (GLP-1 R). In some embodiments, agents that increase activation of the GLP-1 R include GLP-1 agonists, which include GLP-1 analogs. In some embodiments, GLP-1 pathway activators include agents that activate signaling downstream of the GLP-1 R (e.g., activators of PI3K, PKC, cAMP, etc.). In some embodiments, GLP-1 pathway activators may include agents that modulate receptor GLP-1 R expression and / or trafficking (e.g., inhibitors of GLP-1 R internalization; see, e.g., Jones et al., Nat. Comm. (2018)9: 1602). GLP-1 pathway activators encompass activators of the GLP-1 R. In preferred embodiments, the GLP-1 pathway activator is a GLP-1 receptor agonist. GLP-1 pathway activators include, but are not limited to, antibodies and antigen-binding fragments thereof, engineered protein constructs (such as Fc conjugates and multi-functional molecules comprising a GLP-1 analog), peptides, GLP-1 gene therapy, and small molecules.

[0088]

[0082] Incretin'. Incretins are gut hormones that aid in digestion and blood glucose control. In human, incretins include glucagon-like peptide-1 (GLP-1 ) and glucose-dependent insulinotropic polypeptide (GIP). The term “incretin mimetics” refer to medications that mimic incretin hormones and are functional equivalent of naturally occurring incretins. Incretin mimetics include treatments currently approved for Type 2 diabetes or chronic weight management in certain people. Non-limiting examples of incretin mimetics include: GLP-1 receptor agonists (GLP-1 RAs), GLP-1 / GIP dual agonists, GLP-1 / glucagon agonists, GLP-1 / glucagon dual receptor agonists, GLP-1 / GIP dual agonists, and GLP-1 / GIP / glucagon triple receptor agonists. In some embodiments, incretin therapy is selected from a GLP-1 RA (Dulaglutide (Trulicity®), exenatide (Byetta®), exenatide extended release (Bydureon®), liraglutide (Saxenda® I Victoza®), lixisenatide (Adlyxin®), semaglutide (Ozempic® / Rybelsus®), and tirzepatide (Mounjaro® / Zepbound®); a GLP-1 / GIP dual agonist (e.g., tirzepatide), a GLP-1 / glucagon agonist (DD01 , cotadutide, mazdutide, noiiglutide, oxyntomodulin, pemvidutide, setmelanotide, survodutide), GLP-1 / glucagon dual receptor agonist (e.g., ALT-801 ), GLP-1 / GIP dual agonist (e.g., CT- 388 or CT-868), GLP-1 agonist / GIP antagonist (e.g., AMG-133) or a GLP-1 / GIP / glucagon triple receptor agonist (e.g., retatrutide).

[0089]

[0083] Incretin-based therapy. The term “incretin-based therapy” refers to a therapy that comprises at least one incretin or incretin mimetic.

[0090]

[0084] Inhibit or inhibition of. The term “inhibit” or "inhibition of," as used herein, means to reduce by a measurable amount, and can include, but does not require, complete prevention or inhibition.

[0091]

[0085] Insulin sensitivity; insulin resistance'. The term “insulin sensitivity” refers to the metabolic actions of insulin to promote glucose disposal in a subject’s body. A subject is said to have increased insulin sensitivity if the subject requires smaller amounts of insulin to lower blood glucose levels as compared to the average in a human population. In contrast, a subject is said to have decreased insulin sensitivity if the subject requires higher amounts of insulin to lower blood glucose levels. A subject is said to have “insulin resistance” if the quantity of exogenous or endogenous insulin required to increase glucose uptake and utilization in a subject is significantly higher than that in a healthy subject. For instance, a subject is said to have “insulin resistance” if the quantity of exogenous or endogenous insulin required to increase glucose uptake and utilization in a subject is 10%, 20%, 30%, 40% 50%, 60%, 70%, 80%, 90%, 100%, or higher as compared to that in a healthy subject.

[0092]

[0086] Lean / lean mass'. As used herein, “lean” mass or tissue refers to muscle mass or muscle tissue, as opposed to fat mass or fat tissue (e.g., adipose).

[0093]

[0087] Metabolic disorder. The term “metabolic disorder” is used interchangeably with the terms “metabolic disease” or “metabolic condition” and encompasses any conditions involving dysregulation of the body’s metabolic function, resulting in perturbation of the normal physiological state of homeostasis due to an alteration in metabolism (anabolism and / or catabolism). Metabolic disorders may be inherited or acquired. Non-limiting examples of metabolic disorders include obesity or overweight, type 2 diabetes mellitus, type 2 diabetes mellitus associated with obesity, and metabolic syndrome.

[0094]

[0088] Metabolic rate'. The term “metabolic rate” refers to the amount of energy expended over a specific period of time. It is typically measured in calories, kilocalories, or joules. Metabolic rate may be expressed as oxygen consumed or carbon dioxide produced per unit time.

[0095]

[0089] Metabolism'. The term “metabolism” refers to the processes involved in the biosynthesis and breakdown of components that make up a body, such as fats (e.g., adipose tissue), muscle and bones. “Fat metabolism” therefore means the process of biosynthesis and breakdown of fats.

[0096]

[0090] Microcomputed topography (micro-CT): Microcomputed tomography (sometimes referred to as micro-

[0097] CT, microCT or pCT) is a non-destructive imaging tool for the production of high-resolution three-dimensional (3D) images composed of two-dimensional (2D) trans-axial projections, or 'slices', of a target specimen. Micro-CT is a 3D imaging technique that uses X-rays to create detailed images of internal structures, similar to a regular CT scan but with significantly higher resolution, allowing visualization down to the microstructural level (1-100 pm). For example, micro-CT can be employed to analyze various parameters of bone (e.g., femur) such as bone mass and density; microarchitecture (e.g. , trabecular bone structure, such as trabecular thickness, trabecular separation, and trabecular connectivity); cortical bone properties (e.g., cortical bone thickness, porosity, etc.); bone remodeling (e.g., bone formation, resorption); and fracture healing.

[0098]

[0091] Myostatin: In the context of the present disclosure, unless explicitly defined otherwise, the term “myostatin” can refer to any forms of the myostatin protein, such as pro-myostatin, latent myostatin and mature myostatin, each of which exists in dimers in vivo.

[0099]

[0092] Myostatin inhibitor. As used herein, the term “myostatin inhibitor” refers to any agent that inhibits one or more forms of myostatin (e.g., pro-myostatin, latent myostatin, and / or mature myostatin). The term myostatin inhibitor encompasses any molecular modalities such as large molecules (biologies, such as antibodies and engineered protein constructs) and small molecules (such as structurally-defined low molecular weight chemical entities). The term myostatin inhibitor encompasses both selective inhibitors of myostatin and non-selective inhibitors of myostatin. A myostatin inhibitor may be an anti-myostatin antibody, or antigen-binding fragment thereof, that binds pro- and / or latent myostatin and / or mature myostatin. In some embodiments, the myostatin inhibitor is an anti-pro / latent myostatin antibody, or antigen-binding fragment thereof, that preferentially (e.g., selectively) binds pro- and / or latent myostatin over mature myostatin. In various embodiments, the myostatin inhibitor is an antibody (such as a neutralizing antibody), an activation inhibitor (e.g., an antibody that inhibits activation of pro- and / or latent-myostatin), an adnectin, a peptibody, a receptor trap, or a ligand trap. In some embodiments, the myostatin inhibitor is a small molecule inhibitor. In other embodiments, the myostatin inhibitor refers to a gene therapy.

[0100]

[0093] Myostatin-selective inhibitor. The term “myostatin-selective inhibitor” is used interchangeably with a “selective myostatin inhibitor” and refers to myostatin inhibitor that inhibits myostatin but does not inhibit other members of TGFp superfamily (e.g., GDF11 or Activin A). In some embodiments, a myostatin-selective inhibitor inhibits at least one activity of myostatin signaling (e.g., inhibits myostatin activation and / or inhibits or prevents subsequent downstream signaling by myostatin) with at least 100-fold, 200-fold, 500-fold, 1 ,000-fold, or greater potency (e.g., affinity) toward myostatin as compared to another member of the TGFp superfamily (e.g., GDF11 or Activin A) at a biologically or clinically relevant concentration, as measured by any suitable in vitro assays, such as functional enzyme-linked immunosorbent assay (ELISA). In preferred embodiments, a myostatin-selective inhibitor exhibits no detectable binding towards other TGFp family members. In some embodiments, myostatin-selective inhibitors are neutralizing antibodies that bind mature myostatin and inhibit its activity. In some embodiments, myostatin-selective inhibitors are antibodies that bind to pro / latent myostatin, and inhibit the activation step of myostatin. In some embodiments, the myostatin-selective inhibitor is an antibody or antigen-binding fragment provided herein (e.g. any one of Ab101-Ab141 ). In some embodiments, the myostatin-selective inhibitor is an antibody or antigen-binding fragment thereof comprising the heavy and light chain variable domains of any one of Ab101-Ab141 , e.g., the pair of SEQ ID NOs identified for a specific antibody in Table provided herein. In some embodiments, the aforementioned antibody sequences are those of Ab109, Ab133 or Ab141 or an antigen-binding fragment thereof. In most preferred embodiments, the antibody is SRK-439.

[0101]

[0094] Overweight / obesity: A person whose weight is higher than what is considered as a normal weight adjusted for height is described as being overweight or having obesity. Thus, obesity is a chronic condition defined by an excess amount of body fat. Typically, the normal amount of body fat (expressed as percentage of body weight) is between 25- 30% in women and 18-23% in men. Women with over 30% body fat and men with over 25% body fat are considered obese. Obesity can be defined using any clinically relevant definitions. For example, in adults, body mass index (BMI, kg / m2) is frequently used as a measure of overweight and obesity, with overweight being defined as a BMI 25-29.9 kg / m2, obesity as a BMI equal to or greater than 30 kg / m2, and morbid obesity being defined as BMIs over 40 kg / m2. Obesity can also be defined in adults by central adiposity as measured by waist circumference, with raised waist circumference defined as equal to or greater than 102 cm in men and equal to or greater than 88 cm in women. Based on the CDC guidelines, using the BMI-based classification, for human adults (ages 20 and older), BMI of 18.5 to 24.9 is considered normal weight' BMI of 25 to 29.9 is considered overweight' BMI of 30+ is considered obese (including extreme obesity); and BMI of 40+ is considered extremely obese. For children and adolescents (ages 2-19), BMI at or above the 85thpercentile on the CDC growth chart is considered overweight or obese; BMI at or above the 95thpercentile on the CDC growth charts is considered obese (including extreme obesity); and, BMI at or above 120 percent of the 95thpercentile on the CDC growth chart is considered extremely obese. Notwithstanding the foregoing, as used herein, the term “obesity” or “obese” may be collectively refer to both obese and overweight populations by medical definitions, unless otherwise specified. Thus, the expression “treatment of obesity” and the like is intended to encompass therapeutic interventions intended to provide weight management of a subject who benefits therefrom, where the subject may be medically classified as overweight, obese, or extremely obese.

[0102]

[0095] Prevent / preventing: The terms “preventing” and “prevent” as used herein refer to preventing or delaying the onset of a condition or disease in a subject or preventing or delaying the onset of at least one symptom of the condition or disease in the subject. In the context of the present disclosure, prevention of a manifestation (such as muscle loss) is considered a form of treatment.

[0103]

[0096] Serum clearance: As used herein, the term “serum clearance” or “clearance” refers to a relative pharmacokinetic / pharmacodynamic behavior pertaining to changes in serum concentrations (e g., circulating levels) of an analyte (e.g., target protein or protein complex) over time. When the analyte being measured accumulates in the serum, it is said to have slow clearance. By contrast, when the analyte being measured is removed (“cleared”) rapidly from the serum, it is said to have fast clearance. Serum clearance in vivo may occur via multiple mechanisms, including, for example, targeted degradation, Fc-mediated internalization, etc. For example, serum clearance of circulating latent myostatin may be measured with an assay for determining binding of an antibody to free latent myostatin in serum (circulating latent myostatin that is not bound by the antibody). Such an assay may involve immobilizing a biotinylated capture antibody known to bind pro and latent myostatin, before adding a sample comprising myostatin to test the ability of the antibody to bind to the free latent myostatin and adding a detection antibody with a detectable marker. The capture antibody known to bind latent myostatin may be a biotinylated antibody capable of binding latent myostatin, and the detection antibody may be a ruthenium labeled (“ruthenylated”) antibody known to bind both latent and mature myostatin.

[0104]

[0097] Subject: As used herein, the term “subject” is a target to whom the therapy or therapies described herein may be administered. In a clinical context, the terms “subject” and “patient” may be used interchangeably. In some embodiments, a subject is a mammalian subject, e.g., companion animals (e.g., dogs, cats and the like), farm animals (e.g., cows, pigs, horses, sheep, goats, poultry and the like), and laboratory animals (e.g., rats, mice, guinea pigs and the like). In preferred embodiments, the subject is a human subject.

[0105]

[0098] Total fat mass: The term “total fat mass” refers to the cumulative fat content in a subject’s body. Total fat mass includes fat made up of various types of fat cells, such as white fat, brown fat, and beige fat, and includes fat storage in different body compartments, such as essential fat, subcutaneous fat, and visceral fat. Total fat mass may be measured or estimated by any method known in the art, including by skinfold measurements using calipers, measuring the circumference of certain body parts, dual-energy X-ray absorptiometry (DEXA), hydrostatic weighing, air displacement plethysmography, bioelectric impedance analysis, bioimpedance spectroscopy, electrical impedance myography, 3-dimensional body scanners, multi-compartment models, or magnetic resonance imaging. The term “fat mass gain” or “fat mass loss” refers to a change in the amount of fat mass measured as compared to a baseline measurement. For example, a subject having a metabolic disorder may exhibit fat mass loss or visceral fat mass loss after a treatment for the metabolic disorder (e.g., treatment with a myostatin inhibitor). The term “subcutaneous fat” refers to fat found just beneath the skin. The term “visceral fat” refers to fat content that is predominantly made of fat found deep within the abdominal organs, e.g., in the abdominal area of a subject’s body and around the subject’s major organs, such as the liver, kidneys, pancreas, intestines, and heart.

[0099] Treating or preventing'. The term “treating” a condition or disease in a subject refers to the act of providing a therapeutic regimen aimed to cure, heal, alleviate, relieve, alter, remedy, delay progression, ameliorate, improve, or affect a medical condition or at least one symptom of the condition, including slowing or delaying its onset or progression. Thus, the term treating does not necessarily require complete treatment of the disease or disorder. In terms of prevention of muscle loss, maintenance of muscle, or lesser degree of muscle loss over time, enabled by a therapeutic represents a form of treatment, relative to control arm / cohort that show net loss. Thus, treatment does not necessarily require an increase in muscle mass. In one embodiment, treating a subject alleviates symptoms of a disease or disorder by at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, or at least 50%.

[0106]

[0100] Weight loss'. Weight loss refers to a reduction of body weight, irrespective of particular tissue(s) being lost. For example, weight loss per se does not distinguish between loss in fat mass vs muscle mass. Overall loss of total body weight does not necessarily reflect improved body composition,

[0107]

[0101] Weight management'. As used herein, the term “weight management” encompasses measures taken to lose weight, sustain weight, as well as to reduce adipose tissue, increase lean mass, or otherwise to improve or sustain body composition. Successful weight management that is clinically meaningful may or may not accompany overall weight loss. Thus, weight management can include diet (e.g., a calorie restriction diet, e.g., reduced calorie intake or reduced absorption of calories), an exercise regimen, and / or medication (e.g., a treatment comprising a myostatin inhibitor) in order to reduce the amount of total body weight, reduce the amount of total fat mass, reduce the amount of visceral fat mass, increase the metabolic rate, increase the amount of lean mass, and / or increase the ratio of muscle to fat, or otherwise to improve body composition, in a subject.

[0108]

[0102] Weight-related condition / comorbidity. The term “weight-related condition” or “weight-related problem” as used herein, refers to one or more medical condition(s) (e.g., comorbidities or co-morbidities) associated with excess fat mass (i.e., in addition to being overweight or obese), where the excess fat mass of the subject is a contributing factor. Non-limiting examples of weight-related conditions include type 2 diabetes mellitus, high blood pressure, high triglyceride or cholesterol level (e.g., dyslipidemia), metabolic syndrome, vitamin D deficiency, cardiovascular / heart disease (e.g., hypertension, heart failure, coronary artery disease, atrial fibrillation, stroke, idiopathic intracranial hypertension (IIH), venous thromboembolism), osteoarthritis, kidney disease, fatty liver (e.g., nonalcoholic fatty liver disease (NAFLD), nonalcoholic steatohepatitis (NASH) also known as metabolic dysfunction associated steatohepatitis (MASH)), obesity hypoventilation syndrome (OHS), weakened bone, and sleep apnea (e.g., obstructive sleep apnea or OSA), etc. Reviewed in, for example, Lim Y, Boster J. Obesity and Comorbid Conditions. [Updated 2023 Aug 28], In: StatPearls [Internet], Treasure Island (FL): StatPearls Publishing; 2024 Jan-. The present disclosure is based, at least in part, on the surprising finding that selectively targeting myostatin alone with a potent anti-myostatin antibody can achieve equivalent or superior efficacy as a broad-spectrum inhibitor such as anti-ActRI I antibody, for the treatment of obesity, in combination with an incretin therapy. Whilst superior safety profile has been well recognized, the myostatin-selective approach has been considered less than optimal as to efficacy, as compared to non-selective inhibitors of myostatin which can also inhibit Activin A. The general assumption has been thatfor the benefit of improved safety associated with myostatin-selective inhibitors over non-selective inhibitors, overall efficacy would have to be compromised. For example, bimagrumab is an anti-ActRII antibody that inhibits ligand-induced activation of the receptor, which inhibits, not only myostatin but also other ligands for the receptor, including Activin A. Both myostatin and Activin A are recognized as negative regulators of muscle mass. Lines of evidence suggest that inhibition of both myostatin and Activin A can produce additive effects in enhancing muscle mass. Bimagrumab has been shown to achieve clinically beneficial effects in improving body weight, reducing adiposity and preserving lean mass (Heymsfield et al. (JAMA Network Open. 2021 ; 4(1 ):e2033457). Because multiple ligands (including Activins, myostatin, GDF11 and others) share this receptor, bimagrumab presumably blocks both myostatin and Activin A, hence is capable of producing the observed anti-obesity effects. For the same reason, however, such broad inhibitory activities of bimagrumab come with liabilities. In particular, known risk of Activin A inhibition includes perturbation of reproductive hormone regulation. Moreover, because Activin A serves diverse biological functions in normal tissues, systemic inhibition of this pathway raises safety concerns beyond reproductive biology. These risks should also apply to other non-selective inhibitors of myostatin, such as neutralizing antibodies, ligand traps and anti-myostatin Adnectins™ because they too can inhibit multiple ligands not limited to myostatin.

[0109]

[0103] To address these limitations, the inventors of the present disclosure sought to investigate whether highly potent, highly selective inhibitors of myostatin could achieve equivalent efficacy as the benchmark anti-ActRII antibody, bimagrumab, in diet-induced obesity model, in combination with an incretin mimetic. This notion goes against the widely accepted view that inhibition of myostatin alone is insufficient to produce optimal muscle-enhancing effects because myostatin contributes to only a portion of the ActRII-mediated signaling. Indeed, Latres et al. previously suggested that Activin A, not myostatin, is the more prominent regulator of muscle mass in primates (Latres et al., 2017, Nature Communications 8, 15153, “Activin A more prominently regulates muscle mass in primates than does GDF8”). Notwithstanding, the Inventors reasoned that by carefully selecting a highly potent myostatin-selective inhibitor, competitive efficacy may be achieved without a significant compromise, while retaining desirable safety profile at the same time.

[0110]

[0104] To that end, SRK-439, an antibody that selectively inhibits myostatin, was selected for further evaluation. SRK- 439 is a fully human monoclonal antibody that specifically binds pro / latent myostatin with a sub-nanomolar affinity and has an inhibitory potency of below 1 nM (ICso < 1 nM) as determined for its ability to block protease-dependent activation of myostatin in vitro, in conjunction with an incretin mimetic to treat metabolic disorders, such as obesity. One selection criterion is that such myostatin-selective inhibitor shows in vivo efficacy that is at least equivalent to bimagrumab or its variant, when tested in a preclinical DIO mouse model.

[0111]

[0105] Incretin mimetics such as GLP-1 RAs are effective weight loss drugs. While these therapies have been shown to significantly reduce body weight in both preclinical and clinical settings, substantial portions of the weight loss induced by these drugs are in fact due to muscle loss. Applicant has previously recognized that the focus should be shifted from weight loss per se to the quality of weight loss and that part of the healthier weight management should incorporate preserving and enhancing muscle. When the focus is on total body weight, which reflects most of the traditional weight loss approaches, the benefit of preserving muscle for overall health can be lost. Applicant reasoned that muscle enhancement by myostatin inhibition may provide more durable weight management with overall improvement in metabolic health, particularly during rapid weight loss triggered by interventions such as incretin-based therapies. Rapid weight loss typically refers to at least a 10% reduction of body weight from the baseline (e.g. , at least 10%, 15%, 20%, 25%, etc.), over a 6-month period or less (e.g., over 6 months, 5 months, 4 months, 3 months, etc.).

[0112]

[0106] SRK-439 and its variants have been shown to potently and selectively inhibit myostatin and are capable of preventing muscle loss during incretin-induced weight loss. As low as 0.3 mg / kg of SRK-439 is found to be effective in preventing the loss of lean mass in DIO mice treated with GLP-1 receptor agonist. Importantly, such myostatin-selective approach should retain the normal functions of the other growth factors that share the same receptors, such as GDF11 and Activin A, so that the diverse biological roles they play (discussed further below) are preserved.

[0113] Metabolic disorders and diseases

[0114]

[0107] In various embodiments, disclosed herein are methods for treating or preventing a metabolic disease in a subject. A metabolic disease (also referred to as ametabolic disorder or metabolic condition) is generally associated with aberrant glucose, lipid / fat and / or protein / nitrogen metabolism, or osmotic dysregulation, and has pathological consequences arising from such a condition. A number of metabolic disorders of the disclosure share certain characteristics, e.g., they are associated with a loss of fat-free or lean muscle mass, an excess of fat mass, a lower metabolic rate, insulin resistance, lack of ability to regulate blood sugar, weight gain, and / or increase in body mass index. In some embodiments, such metabolic diseases or disorders are triggered or exacerbated by medication that the patients receive.

[0115]

[0108] The present disclosure is based, at least in part, on the discovery that administration of a myostatin inhibitor described herein (e.g., an anti-myostatin antibody or antigen-binding fragment thereof disclosed herein) to a subject having a metabolic disease, e.g., via a subcutaneous route, may improve both the physiological and the functional characteristics of the subject. In particular, selective inhibitors of myostatin (e.g., those that do not inhibit GDF11 or Activin A) may be advantageously used to treat subjects with one or more conditions detailed below, who benefit from GDF11 and / or Activin A functions. Such subjects include, for example, those who suffer from or are at risk of developing anemia, neuropathy, chronic wounds, bone disorders, frequent bone fractures, etc. Such subjects may also include those who are under the age of 18, who are not on contraceptives / birth control, who are pregnant, and / or who are nursing.

[0116]

[0109] Further examples of metabolic diseases that may be treated or prevented by the methods of the present disclosure include but are not limited to, pre-diabetes and diabetes (e.g., type 1 or type 2 diabetes, or diabetes associated with obesity), obesity (e.g., adult-onset obesity, diet-induced obesity, pediatric obesity, etc.), obesity syndrome (e.g., diet-associated or diet-induced obesity), insulin resistance, insulin insufficiency, hyperinsulinemia, impaired glucose tolerance (IGT), abnormal glycogen metabolism, hyperlipidemia, hypoalbuminemia, hypertriglyceridemia, kidney disease, e.g., chronic kidney disease, syndrome X, fatty liver disease, metabolic bone diseases, spinal cord injury (SCI) (e.g., complete or incomplete / partial SCI), hypo-metabolic states, double diabetes, and Cushing’s disease (also referred to as Cushing’s syndrome). In some embodiments, metabolic diseases include diseases associated with impaired neurological signaling or partial denervation. In some embodiments, metabolic diseases include conditions triggered by or associated with certain medication (e.g., side effects).

[0117]

[0110] Additional diseases or conditions related to metabolic disorders and / or body composition that would be apparent to the skilled artisan and are within the scope of this disclosure.

[0118]

[0111] As discussed in more detail herein, metabolic disorders can occur secondarily to, or occur as a result of, a muscle condition or disorder. Because muscle homeostasis is correlated with amino acid / protein metabolism, it is further contemplated that myostatin inhibition may in turn regulate nitrogen metabolism and nitrogen mobilization in the body. In muscle catabolism, a muscle tissue breaks down into its building blocks, amino acids, which may be considered as a major reservoir (and thus a source) of nitrogen. Nitrogen is an element of ammonia, which is highly toxic to the body and is excreted in a form of urea in humans. When nitrogen metabolism is dysregulated, a possible outcome includes an imbalance of fluid retention, which may manifest as systemic or local edema (e.g., congestion or fluid overload).

[0119]

[0112] Diabetes is a prevalent metabolic disease that refers to a group of metabolic diseases characterized by high blood sugar (glucose) levels which result from defects in insulin secretion or action, or both. Insulin is a hormone released by the pancreas in response to increased levels of blood sugar (glucose) in the blood. There are two most common types of diabetes, namely type 1 diabetes and type 2 diabetes, which both result from the body’s inability to regulate insulin.

[0120]

[0113] In type 2 diabetes mellitus (T2DM), also referred to as noninsulin-dependent diabetes mellitus, NDDM), the pancreas continues to manufacture insulin, sometimes even at higher-than-normal levels. However, the body develops resistance to its effects, resulting in a relative insulin deficiency. As the pancreas continues to manufacture insulin and the body becomes insulin-resistant, the insulin-producing cells in the Islets of Langerhans in the pancreas may wear out, limiting or losing their ability to continue to produce insulin. Type 2 diabetes may occur in children and adolescents but usually begins after age 30 and becomes progressively more common with age: about 15 percent of people over age 70 have type II diabetes. Obesity is a risk factor for type 2 diabetes, and 80 to 90 percent of the people with this disorder are obese.

[0121]

[0114] In some embodiments, diabetes includes pre-diabetes. “Pre-diabetes” refers to one or more early diabetic conditions including impaired glucose utilization, abnormal or impaired fasting glucose levels, impaired glucose tolerance, impaired insulin sensitivity and insulin resistance. Prediabetes is a major risk factor for the development of type 2 diabetes mellitus, cardiovascular disease, and mortality. Much focus has been given to developing therapeutic interventions that prevent the development of or progression to type 2 diabetes by effectively treating prediabetic patients.

[0122]

[0115] In some embodiments, diabetes includes double diabetes, which is a combination of type 1 diabetes with features of insulin resistance and type 2 diabetes.

[0123]

[0116] Diabetes and prediabetes can be diagnosed by the administration of a glucose tolerance test, which can be determined by venous blood draws from a fasting or non-fasting subject. They can also be diagnosed by measuring blood levels of hemoglobin A1C (A1 C), a glycosylated form of hemoglobin that reflects the average amount of glucose in the blood over the past two to three months. Normal, prediabetic, and diabetic A1 C levels are known in the art and can vary with age. In some embodiments, the expected values for normal fasting blood glucose concentration can be between about 70 mg / dL (3.9 mmol / L) and about 100 mg / dL (5.6 mmol / L). In some embodiments, pre-diabetes is associated with hemoglobin A1 C levels of about 100 mg / dL to about 125 mg / dL or about 5.6 mmol / L to about 6.9 mmol / L. In some embodiments, diabetes is associated with fasted glucose levels above about 125 mg / dL or hemoglobin A1 C levels above about 6.9 mmol / L.

[0124]

[0117] Clinically, diabetes is often divided into several basic categories. Primary examples of these categories include, autoimmune diabetes mellitus, non-insulin-dependent diabetes mellitus (type 1 NDDM), insulin-dependent diabetes mellitus (type 2 IDDM), non-autoimmune diabetes mellitus, non-insulin-dependent diabetes mellitus (type 2 NIDDM), and maturity-onset diabetes of the young (MODY). A further category, often referred to as secondary, refers to diabetes brought about by some identifiable condition which causes or allows a diabetic syndrome to develop. Examples of secondary categories include, diabetes caused by pancreatic disease, hormonal abnormalities, drug- or chemical- induced diabetes, diabetes caused by insulin receptor abnormalities, diabetes associated with genetic syndromes, and diabetes of other causes, (see e.g., Harrison’s (1996) 14th ed., New York, McGraw-Hill).

[0125]

[0118] Obesity is another prevalent metabolic disease that can be treated or prevented by the methods of the present disclosure.

[0126]

[0119] Subjects with obesity may exhibit other symptoms such as increased fasting plasma glucose, glucose intolerance, heart failure, hypertension, insulin resistance, increased fasting plasma triglycerides, decreased fasting high-density lipoprotein (HDL) level, prediabetes, increased blood pressure, stroke, obstructive sleep apnea, reproductive hormone impairment, obstructive sleep apnea, osteoarthritis, gallstones, gastroesophageal reflux, or renal disease. Obesity may also cause various orthopedic problems, skin disorders and swelling of the feet and ankles. Severe complications of obesity include a much higher risk of coronary artery disorder and of its major risk factors type II diabetes, hyperlipidemia and hypertension. Much of the morbidity associated with obesity is associated with type II diabetes, as poorly controlled diabetes and obesity lead to a constellation of symptoms that are together known as syndrome X, or metabolic syndrome. In some embodiments, the obesity is sarcopenic obesity. In some embodiments, the subject having obesity is on a caloric restriction regimen.

[0127]

[0120] In an aspect, the methods of the present disclosure are suitable for treating all forms of obesity. Obesity includes obesity associated with diabetes, obesity associated with metabolic syndrome, obesity associated with a monogenetic disorder, obesity associated with anti-psychotic drug use, glucocorticoid-associated obesity, and hypothalamic obesity.

[0121] In a further aspect, the methods of the present disclosure are suitable for treating (e.g., preventing) inflammatory bone loss. Indeed, the link between the effect of inflammation and bone loss is well documented (Hardy and Cooper, 2009, Journal of Endocrinology 201 (3):309-20; Epsley et al., 2021 ; Front Physiol., 11 :511799; Redlich and Smolen, 2012, Nat. Rev. Drug Discovery 11 :234-250). A net loss of bone can result at least in part from an imbalance of bone resorption and bone formation. The cause of systemic inflammation can vary and may include inflammatory bowel disease, chronic obstructive lung disease, cystic fibrosis, periodontitis, rheumatoid arthritis and other inflammatory conditions such as obesity.

[0128]

[0122] As already alluded to herein, obesity can be regarded as an inflammatory condition. Obesity is often associated with the consumption of a diet high in fat, i.e. , a high-fat diet (HFD), which can cause not only unhealthy weight gain (e.g., overweight and obesity) but also an increased risk of systemic inflammation and weakened bone (e.g., inflammatory bone loss). Indeed, this is consistent with the observations presented herein. A diet high in fat led to a reduction in bone density in DIO mice, as compared to mice on a normal diet. High-fat diet-induced weakening of bone can be at least partially prevented by selectively inhibiting the myostatin pathway, demonstrating a protective effect of myostatin-selective inhibitors against bone loss associated with high-fat diet. In some embodiments, protective effects of myostatin inhibitors on bone comprise increased bone volume, increased bone density, reduced porosity, increased cortical thickness, increased trabecular thickness, increased bone mineral density, reduced trabecular separation, increased trabecular number, increased bone volume fractions, increased cortical bone area, increased cortical bone porosity, increased bone regeneration or formation, and / or reduced bone resorption (breakdown of bone tissue).

[0129]

[0123] According to the present disclosure, myostatin inhibitors (e.g., myostatin-selective inhibitors) can therefore be used to prevent bone loss (i.e., bone mass loss) in a subject. In some embodiments, the subject is at risk of bone loss from an inflammatory condition. In some embodiments, the subject is at risk of bone loss from an atrophy condition. In some embodiments, the subject is on a high-fat diet. In some embodiments, the subject has diabetes. In some embodiments, the subject is overweight or obese. In some embodiments, the subject has a BMI greater than 25 but less than 30. In some embodiments, the subject has a BMI of 30 or greater. In some embodiments, the subject is on an incretin-based therapy. Effects on bone may be monitored by known techniques, such as DEXA in patients. In preferred embodiments, a myostatin-selective inhibitor is selected for preventing bone loss in these patient populations, such as SRK-439, GYM329, and trevogrumab. In most preferred embodiments, the myostatin-selective inhibitor is SRK-439 or antigen-binding fragments thereof.

[0130]

[0124] In another aspect, the methods of the present disclosure are suitable for treating or preventing metabolic disease such as obesity syndromes. The term “obesity syndrome” refers to any disorder or condition causing a subject to be grossly fat or overweight. Like other metabolic diseases, people with obesity syndrome are usually associated with a loss of fat-free or lean muscle mass, an excess of fat mass, a lower metabolic rate, insulin resistance, lack of ability to regulate blood sugar, weight gain, and increase in body mass index. In some embodiments, the obesity syndrome is selected from the group consisting of Prader Willi syndrome, an obesity syndrome associated with a genetic disorder, and an obesity syndrome associated with a hypothalamic disorder.

[0131]

[0125] The methods of the present disclosure are also suitable for treating or preventing metabolic disease such as metabolic syndromes. As used herein, “metabolic syndrome” refers to the concept of a clustering of metabolic risk factors that come together in a single individual and lead to a high risk of developing diabetes and / or cardiovascular diseases. The main features of metabolic syndrome include insulin resistance, hypertension (high blood pressure), cholesterol abnormalities, dyslipidemia, triglyceride abnormalities, an increased risk for clotting and excess body weight, especially in the abdomen, or obesity. In some embodiments, metabolic syndrome can be diagnosed by the presence of three or more of the following components: (1 ) an elevated waist circumference (men, equal to or greater than 40 inches (102 cm); women, equal to or greater than 35 inches (88 cm); (2) elevated triglycerides (equal to or greater than 150 mg / dL); (3) reduced high density lipoprotein cholesterol (HDL) (men, less than 40 mg / dL; women, less than 50 mg / dL); (4) elevated blood pressure (equal to or greater than 130 / 85 mm Hg); and (5) elevated fasting glucose (equal to or greater than 100 mg / dL).

[0132]

[0126] Body composition may be measured by a variety of methods, including dual energy X-ray absorptiometry (DEXA). Total body scans using DEXA provide generally accurate and precise measurements of body composition, including bone mineral content, bone mineral density, lean tissue mass, fat tissue mass, and fractional contribution of fat.

[0133]

[0127] Obesity is a risk factor for the development of cardiovascular disease. Obese individuals experience cardiovascular disease events at an earlier age, live with cardiovascular disease for a greater proportion of their lifetime and have a shorter average lifespan than individuals with normal weight. Obesity contributes directly to cardiovascular risk factors, including dyslipidemia, type 2 diabetes, hypertension, and sleep disorders (e.g., sleep apnea). Obesity accelerates atherosclerotic changes through multiple mechanisms, including insulin resistance and inflammation. Obesity also leads to the development of cardiovascular disease and cardiovascular disease mortality independently of other cardiovascular risk factors. Visceral adiposity promotes systemic and vascular inflammation, which is fundamental to the atherosclerotic process. Inflammation induced by obesity increases the likelihood of low-density lipoprotein (LDL) oxidation, which in turn promotes atherogenesis. Insulin resistance is associated with dyslipidemia and metabolic syndrome, which are linked to atherosclerosis. Endothelial function in obesity, e.g., due to decreased bioavailability of nitric oxide in the setting of inflammation and oxidative stress, also contributes to the progression of atherosclerosis. Obesity also has been linked to abnormalities in the coronary microvasculature and on epicardial coronary vessels. Another aspect of the disclosure includes a method of treating a subject having a metabolic disease or condition related to aging. Exemplary diseases and conditions related to ageing include, without limitation, sarcopenia (age-related muscle loss), frailty, and androgen deficiency. In addition, obesity (e.g., being overweight) is linked to higher risk of certain types of cancer. According to the CDC, 13 types of cancers, which make up 40% of all cancers that are diagnosed in the United States each year, are associated with overweight and obesity. CDC Obesity and Cancer (2025). These include meningioma, thyroid cancer, adenocarcinoma of the esophagus, breast cancer, multiple myeloma, liver cancer, kidney cancer, gallbladder cancer, upper stomach cancer, pancreatic cancer, uterus cancer, colorectal cancer, and ovarian cancer. Therefore, improving health outcomes, or otherwise treating obesity, by the use of myostatin inhibitors as described herein (e.g., myostatin inhibitors that spare Activin A) may in turn reduce the incidence and / or improve the prognosis of such cancers.

[0134]

[0128] Accordingly, the methods of the present disclosure are suitable for treating or preventing metabolic diseases such as cardiovascular disease, e.g., cardiovascular disease associated with metabolic syndrome. The term “cardiovascular disease” refers to any disease of the heart or blood vessels. Cardiovascular or heart disease includes but is not limited to, for example, angina, arrhythmia, coronary artery disease (CAD), coronary heart disease, cardiomyopathy (including dilated cardiomyopathy, restrictive cardiomyopathy, arrhythmogenic right ventricular cardiomyopathy, and diabetic cardiomyopathy), heart attack (myocardial infarction), heart failure (e.g., acute heart failure (AHF), chronic heart failure (CHF) or heart failure with preserved ejection fraction (HfpEF)), hypertrophic cardiomyopathy, mitral regurgitation, mitral valve prolapse, and pulmonary stenosis. Blood vessel disease includes but is not limited to, for example, peripheral vascular disease, artery disease, carotid artery disease, deep vein thrombosis, venous diseases, and atherosclerosis. In some embodiments, a subject having heart failure is resistant to diuretic therapy. In another embodiment, a subject having heart failure responds poorly to diuretic therapy. In addition to various metabolic impairments, patients with cardiovascular diseases, such as heart failure, can develop muscle wasting and weakness. It is contemplated herein that myostatin inhibitors may provide beneficial effects in reducing risk of major cardiovascular events particularly in patients with metabolic disorders, such as obesity and diabetes. Such effects may be synergistic when used in conjunction with a GLP-1 pathway activator, such as GLP-1 receptor agonists. Importantly, myostatin inhibitors used in conjunction with incretin therapies such as GLP-1 receptor agonists may also protect against muscle loss / atrophy associated with cardiovascular disorders. In preferred embodiments, the myostatin inhibitor is a myostatin-selective inhibitor. In particularly preferred embodiments, the myostatin inhibitor is selected from the novel antibodies disclosed herein, e.g., any one of Ab101-Ab141. In most preferred embodiments, the myostatin inhibitor is selected from Ab109, Ab133 and Ab141.

[0135]

[0129] Pulmonary edema, as well as renal congestion, for example, is frequently observed in patients with heart failure, associated with decreased cardiac output. Pulmonary congestion is in fact the most frequent cause of hospitalization in this clinical setting and correlates with poor prognosis. Similarly, in pathologic conditions that involve impaired osmoregulation, the affected individual may be particularly sensitive to salt intake, which may cause or exacerbate fluid overload. Therefore, for subjects with fluid retention or volume-overload, such as subjects having impaired osmoregulation and subjects with heart failure, e.g., chronic heart failure, current guidelines suggest that decongestion should be attempted using diuretic therapy (see, e.g., Regolisti et al., Nephrology @ Point of Care 2016; 2(1 ):e73-e87). However, in many cases, diuretic treatment is ineffective, or the subject is refractory to diuretic therapy. Myostatin inhibition according to the present disclosure may provide such patients with clinical benefits. Specifically, the methods of the present disclosure are suitable for increasing responsiveness of subjects who are refractory to diuretic treatment, or poorly responsive to diuretic treatment. For example, administration of a myostatin inhibitor reduces the diuretic dose needed and / or offers improved control of symptoms, such as CHF symptoms; improves cardiac function; and / or prevents pathologic cardiac remodeling or other worsening of cardiac function chronically. Myostatin inhibition using an inhibitor described herein also reduces the risk of CHF exacerbations, such as episodes of acute pulmonary edema.

[0136]

[0130] For subjects at higher risk for developing acute pulmonary edema, such as subjects receiving IV fluids, blood transfusions, orfluid shifts, the myostatin inhibitors disclosed herein may be administered prophylactically. For example, a subject with congestive heart failure who needs to receive a blood transfusion can be prophylactically administered a myostatin inhibitor during the blood transfusion to prevent the onset of acute pulmonary edema during the transfusion. For subjects having CHF and / or other volume-overload states who develop hyponatremia, either due to the volumeoverload, itself, or from diuretics used to treat the volume-overload, myostatin inhibitors disclosed herein can be administered to treat the hyponatremia and / or enable higher doses of diuretics to be used, when diuretic dosing is limited by hyponatremia as a side effect. Generally speaking, however, the myostatin inhibitors disclosed herein may be used to treat hyponatremia, irrespective of the underlying etiology.

[0137]

[0131] For other volume-overload states, e.g., renal failure or liver disease, which require high-dose diuretics, a myostatin inhibitor disclosed herein reduces the diuretic dose needed; offer improved control of symptoms, such as peripheral edema or congestion within the body internally (including pleural effusions, ascites, hepatic congestion, or volume overload within the eyes, which can lead to retinal detachment, and / or reduce the risk of pulmonary edema.

[0138]

[0132] Metabolic disorders and diseases for treatment according to the methods provided herein also include metabolic conditions that affect the liver. Nonalcoholic fatty liver disease (NAFLD) is a spectrum of hepatic diseases associated with metabolic and cardiovascular disorders and is strongly associated with metabolic syndrome (Godoy-Matos (2020) Diabetes Metab Synd 12:50). Metabolic conditions that affect the liver include non-alcoholic steatohepatitis (NASH), NAFLD, hereditary hemochromatosis, alpha-1 antitrypsin deficiency, and Wilson disease. Fibrosis, a thickening of connective tissues, occurs in both NASH and the later stages of NAFLD, as excess tissue deposits lead to fibrotic scarring. Fibroblast growth factor 21 (FGF21 ) and FGF19 have been observed to decrease hepatic steatosis. Accordingly, in some embodiments, the subject receives an FGF21 or FGF19 receptor agonist as well as a myostatin inhibitor of the invention. Zhao et al., Signal Transduction and Targeted Therapy (2022) 7:206. In some embodiments, the subject also receives, or has received, one or more of the following therapeutics to treat the liver disease: Hydronidone, BI089-100, Efruxifermin, Pegbelfermin, Aldafermin, MK-3655, PRI-724, Selonsertib, CC-90001 , Epeleuton, Elafibranor, Saroglitazar, Lanifibranor, Pemafibrate, ZSP0678, Obeticholic Acid, Cilofexor, Nidufexor, TERN-101 , Vonafexor, EDP-305, Tropifexor, JKB-121 , JKB-122, Semaglutide, Tirzepatide, Cotadutide, HM-15211 , Resmetirom, VK2809, Cenicriviroc, Belapectin, GB1211 , Azemiglitazone potassium, Deuterium-Stabilized I- Pioglitazone, Aramchol, PF-05221304, Firsocostat, ZSP1601 , Epeleuton, PXL-770, ALS-L1023, Namodenoson, TVB- 2640, LPCN 1144, HepaStem, BMS-986263, Foralumab, Elobixibat, Apararenone, PF-06835919, ARO-HSD, and CB4211.

[0139]

[0133] Furthermore, patients suffering from liver diseases such as NAFLD and NASH / MASH, are at risk of experiencing muscle weakness or atrophy. Therefore, beyond its metabolic benefits, myostatin inhibitors may be particularly beneficial to counter muscle weakness / atrophy in these patient populations. More recently, incretin therapies such as GLP-1 receptor agonists have been reported to exert clinical benefits in liver diseases. Because these therapies can also cause muscle loss (e.g., incretin-induced muscle loss), patients with liver disease who are on an incretin therapy can particularly benefit from concurrent myostatin inhibitor therapy. Preferred myostatin inhibitors are myostatin- selective inhibitors, such as SRK-439 and Ab101-Ab141.

[0140]

[0134] The compositions and methods of the present disclosure are also suitable for treating or preventing metabolic diseases associated with a hypo-metabolic state. The term “a hypo-metabolic state” refers to a state of reduced metabolism or metabolic activity, where the body is not producing enough energy. Patients with a hypo-metabolic state generally have a lower metabolic rate, a loss of fat-free or lean muscle mass, an excessive gain of fat mass, insulin resistance, lack of ability to regulate blood sugar, weight gain, and an increase in body mass index. In some embodiments, the hypo-metabolic state is selected from the group consisting of a state associated with prolonged immobilization, a state associated with bed rest, a state associated with casting, a state associated with a stroke, a state associated with amputation, and a post-surgery state. In some embodiments, the hypo-metabolic state is a postsurgery state, e.g., paraspinal muscle atrophy after lumbar spine surgery. In one embodiment, the paraspinal muscle atrophy is a nerve injury-dependent muscle atrophy. In one embodiment, the surgery is a spinal surgery. In one embodiment, the spinal surgery is a lumbar spine surgery or a lumbar spine procedure, e.g., a lumbar fusion procedure, a lumbar nonfusion procedure, a posterior lumbar fusion procedure, an anterior lumbar fusion procedure, a minimally invasive (MIS) posterior lumbar decompression procedure, a minimally invasive (MIS) posterior lumbar fusion procedure, a non-MIS equivalent procedure, etc.

[0141]

[0135] In some embodiments, the myostatin inhibitors of the invention are used to attenuate spinal cord injury (SCI)- induced reduction in sub-lesional muscle mass and overall body mass and, while at the same time reducing the mass of undesirable adipose tissue such as white and visceral adipose tissue. Subjects treated with the myostatin inhibitor therapy may also exhibit a significant improvement in their locomotor function, muscle strength, as well as motor coordination and balance skills.

[0142]

[0136] In another aspect, the methods of the present disclosure are suitable for treating or preventing metabolic diseases such as Cushing’s disease, which is also referred to as Cushing’s syndrome or hypercortisolism. The term “Cushing’s disease” refers to a collection of signs and symptoms due to prolonged exposure to cortisol. In some embodiments, the Cushing’s disease is selected from the group consisting of corticosteroid-induced Cushing’s disease and tumor-induced Cushing’s disease. In some embodiments, the antibodies or antigen-binding fragments disclosed herein are useful as an alternative or additive treatment option to the current standard-of-care treatments for patients suffering from Cushing’s disease.

[0143]

[0137] Accordingly, the present disclosure provides methods for treating or preventing metabolic diseases in a human subject. The methods include selecting a human subject suffering from a metabolic disease and administering to the human subject an effective amount of a myostatin inhibitor, thereby treating or preventing the metabolic disease in the human subject. Preferably, the myostatin inhibitor is a myostatin-selective inhibitor. More preferably, the myostatin- selective inhibitor is an antibody, or antigen-binding fragment thereof, that specifically binds to pro / latent myostatin, but does not bind to GDF11 , e.g., Ab101-Ab141 , GYM329, etc. In the most preferred embodiments, the antibody is SRK- 439. Antibodies that specifically recognize pro / latent myostatin, but not GDF11 or Activin A, are beneficial in avoiding undesirable toxicity caused by off-target binding without compromising the normal function of these growth factors in the subject. In one embodiment, the subject is a pediatric subject. In one embodiment, the subject is a subject aged 2- 19 years, inclusive of endpoints. In one embodiment, the subject is a subject aged 12 years or older (e g., 12-17 years), inclusive of endpoints.

[0144] Outcome measures

[0145]

[0138] To assess effects on healthier weight management, one or more of the following parameters may be measured at predetermined time point(s), such as at the outset (Baseline), at 24 weeks, at 30 weeks, at 40 weeks, etc.:

[0146] 1. Change from Baseline in total Lean Body Mass (kg). In some embodiments, dual-energy X-ray absorptiometry (DEXA) may be used to evaluate body composition.

[0147] 2. Change from Baseline in body weight. In some embodiments, total body weight may be assessed via a calibrated scale.

[0148] 3. Change from Baseline in percent lean body mass (%). In some embodiments, dual-energy X-ray absorptiometry (DEXA) may be used to evaluate body composition.

[0149] 4. Change from Baseline in fat body mass (kg and %). In some embodiments, dual-energy X-ray absorptiometry (DEXA) may be used to evaluate body composition.

[0150] 5. Change from Baseline in visceral adipose tissue (VAT), subcutaneous adipose tissue (SAT), and trunkfat body mass (kg and %). In some embodiments, dual-energy X-ray absorptiometry (DEXA) may be used to evaluate body composition.

[0151] 6. Percent (%) of weight loss from baseline due to fat body mass loss. In some embodiments, dual-energy X-ray absorptiometry (DEXA) may be used to evaluate body composition.

[0152] 7. Percent (%) of weight loss from baseline due to lean body mass loss. In some embodiments, dual-energy X-ray absorptiometry (DEXA) may be used to evaluate body composition.

[0153] 8. Concentration of myostatin inhibitor (such as an antibody that inhibits myostatin) in circulation over time. In some embodiments, blood samples may be collected and assessed for circulating / serum concentration of the inhibitor for pharmacokinetic analyses.

[0154] 9. Concentration of latent myostatin in circulation over time. In some embodiments, blood samples may be collected and assessed for circulating / serum concentration of latent myostatin. In some embodiments, changes in serum concentrations of latent myostatin may serve as a pharmacodynamic marker.

[0155] 10. Treatment emergent adverse events (TEAEs) and serious adverse events (SAEs), including incidence and severity of TEAEs and SAEs, are determined to assess safety profile of the myostatin inhibitor.

[0156] 11 . Presence of anti-drug antibodies (ADA) against the myostatin inhibitor (e g., antibody that inhibits myostatin) over time is measured in serum blood samples.

[0157] 12. Change in HbA1c (glycated hemoglobin) from baseline is assessed in blood samples.

[0158] 13. Change from baseline in fasting serum triglycerides in blood samples.

[0159] 14. Change from baseline in total cholesterol is measured from blood samples.

[0160] 15. Change from baseline in low-density lipoprotein cholesterol (LDL-C) is measured in blood samples. GDF11 and patient selection considerations

[0161]

[0139] GDF11 is a member of the TGFp superfamily of growth factors and is most closely related to myostatin (i.e., GDF8) by amino acid sequence (e.g., approximately 90% sequence identity). GDF11 also signals through the same ActRII receptors as myostatin. Non-selective myostatin inhibitors, such as ligand traps and anti-myostatin Adnectins™ , typically also target GDF11 due to structural similarities shared by these ligands. Similarly, most of the known neutralizing antibodies that bind mature myostatin (e.g., mature growth factor / ligand) also bind GDF11 due to their structural similarities, although many such antibodies are often referred to as “anti-myostatin antibodies.” In addition, ActRII antagonists, such as anti-ActRI I antibodies (e.g., bimagrumab), inhibit all such ligands that share the receptor, including myostatin, Activin A and GDF11. Despite structural similarity, GDF11 and myostatin play distinct biological functions (reviewed in: Suh and Lee, 2020, Experimental & Molecular Medicine, 52:1673-1693), and careful selection of inhibitors for a particular patient or a patient population is warranted.

[0162]

[0140] GDF11 is involved in early development of various tissues, and homozygous deletion of Gdf11 generates defects in axial skeletal patterning and organ development in mice. Gdf11-null mice, unlike Mstn-null mice, die shortly after birth.

[0163]

[0141] The postnatal role of GDF11 remains unclear to date. Rogers et al. (Endocrinology 156, 3885-3888 (2015)) reported that the molar concentration of circulating GDF11 was approximately 500 times lower than that of myostatin. This raises the possibility that broad-spectrum (non-selective) inhibitors of myostatin that also inhibit GDF11 may have differential impact on these targets at a given dose. Assuming the relative affinities of such inhibitors to GDF11 and myostatin are equivalent, pharmacokinetic / pharmacodynamic (PK / PD) profiles may vastly differ. Tissue distributions of these growth factors are also notable; while myostatin expression is largely limited to skeletal muscle, GDF11 is expressed more broadly.

[0164]

[0142] In a 2021 article, Muramatsu et al. examined the relative contribution of myostatin and GDF11 to muscle strength in mice, using a myostatin-selective inhibitor (GYM329) and a GDF11-selective inhibitor. The authors found that the muscle strength enhancement induced by GYM329 was significantly attenuated by concurrent treatment with the anti- GDF11 antibody, whereas anti-GDF11 antibody treatment alone did not enhance muscle strength. See Muramatsu et al. Sci Rep. 2021 Jan 25; 11 (1 ):2160. doi: 10.1038 / s41598-021-81669-8. Conversely, treating mice with exogenous recombinant GDF11 significantly alleviated hindlimb suspension-induced muscle weakness. It was concluded that GDF11 and myostatin act in opposite directions in terms of muscle strength enhancement, where inhibition of GDF11 signaling has negative impacts. Based on these findings, concurrent inhibition of myostatin and GDF11 may reduce muscle-enhancing effects of the former, supporting the advantage of the myostatin-selective approach, particularly with respect to GDF11 . Accordingly, the present disclosure provides a myostatin inhibitor for use in the treatment of obesity and other metabolic disorders, where a myostatin inhibitor that does not inhibit GDF11 is selected to carry out various embodiments described herein.

[0165]

[0143] In addition, Suh et al. recently demonstrated that transgenic overexpression of follistatin, an endogenous inhibitor of myostatin, GDF11 , and activins, substantially enhanced muscle mass but induced spontaneous tibial fractures due to a reduction in bone mineral density, implying that inhibition of GDF11 may have adverse effects on bone (Proc. Natl Acad. Sci. USA 117, 4910-4920 (2020)). These findings suggest that those who are at risk of bone fractures or bone disorders in particular should avoid non-selective myostatin inhibitors that also inhibit GDF11. For example, those who have bone disorders or are at risk of developing a bone disorder include those suffering from an underlying condition known to elevate the risk of reduced bone mineral density. Non-limiting examples of conditions associated with reduced bone mineral density include diabetes (e.g., type 1 diabetes and type 2 diabetes) and overweight / obesity. Those undergo rapid weight loss, for instance via medication (such as incretin-based therapies, e.g., GLP-1 RAs, GIP RAs and glucagon RAs), extreme dieting and / or exercise regimen, gastric surgery, etc., are also at risk of weakened bone. In addition, those who are at risk of bone fractures or bone disorders include those with a history of falls and / or bone fractures. Advantageously, in carrying out various embodiments of the present disclosure, myostatin inhibitors that do not inhibit GDF11 may be selected to treat these patient populations. Rapid weight loss typically refers to at least a 10% reduction of body weight from the baseline (e.g., at least 10%, 15%, 20%, 25%, etc.), over a 6-month period or less (e.g., over 6 months, 5 months, 4 months, 3 months, etc.).

[0166]

[0144] In addition to detrimental effects of GDF11 inhibition on enhancing muscle mass, active GDF11 may also provide metabolic benefits (outlined below). This suggests that it is advantageous to select myostatin inhibitors which are capable of retaining GDF11 activities in the context of metabolic health.

[0167] Metabolic benefits of GDF11:

[0168]

[0145] Despite structural similarities, lines of evidence suggest that GDF11 biology is discrete from that of myostatin and that there are in fact certain benefits to maintaining GDF11 (e.g., GDF11 activities) in metabolic contexts, raising the possibility that it can be detrimental to antagonize GDF11 activities in patients who suffer from metabolic dysregulations.

[0169]

[0146] Frohlich et al. (Cell Prolif. 2022; 55(10): e13310) reported that GDF11 inhibits adipogenesis and improves the function of mature adipocytes in part via the SMAD2 / 3 pathway. This suggests that non-selective inhibitors that also target GDF11 can interfere with the ability of GDF11 to improve metabolic function.

[0170]

[0147] In a 2019 publication, Lu et al. (J Transl Med. 2019; 17:422) showed that Gdf11 gene transfer was able to prevent high fat diet-induced obesity and improve metabolic homeostasis in a diabetic mouse model. These findings suggest that active GDF11 plays a positive role in regulating metabolic health. Therefore, the use of myostatin inhibitors that do not target GDF11 would be advantageous in patients who suffer from or are at risk of developing metabolic disorders.

[0171]

[0148] Similarly, exogenous GDF11 , but not GDF8 (i.e., myostatin), was shown to reduce body weight and improve glucose homeostasis / tolerance in mice on a high-fat diet (Walker et al., 2020; Scientific Reports 10: 4561 ). Again, these findings support the notion that GDF11 inhibition can be detrimental to metabolic health and caution against the use of non-selective inhibitors that interfere with GDF11 function.

[0172]

[0149] Taken together, preferred myostatin inhibitors to be used to treat metabolic disorders are those that do not inhibit GDF11 , so that the beneficial effects of GDF11 on metabolism can be retained.

[0173]

[0150] Accordingly, in various embodiments of the present disclosure, a myostatin inhibitor is used to treat metabolic disorders (e.g., obesity / overweight, diabetes, and fatty liver, etc.) in a subject, wherein preferably, the myostatin inhibitor does not inhibit GDF11 . In some embodiments, such myostatin inhibitors are antibodies that bind mature myostatin but not mature GDF11 , wherein optionally the antibody is trevogrumab (also referred to as REGN1033), a variant thereof, or a derivative thereof. In preferred embodiments, the myostatin inhibitor is a selective inhibitor of myostatin (i.e., myostatin-selective inhibitor), which inhibits myostatin but not GDF11 or Activin A. In some embodiments, the myostatin-selective inhibitor is an antibody or antigen-binding fragments thereof that binds a precursor of myostatin (e.g., pro / latent myostatin) thereby inhibiting activation of myostatin. Examples of antibodies that bind pro / latent myostatin thereby inhibiting myostatin activation include: apitegromab, GYM329, SRK-439, as well as Ab101-Ab141 disclosed herein.

[0174] Activin A and patient selection considerations

[0175]

[0151] Activins are widely expressed in many tissues and play diverse biological roles both during development and in adults (reviewed in, for example, Namwanje and Brown; Cold Spring Harb Perspect Biol. 2016 Jul; 8(7): a021881 ). It is generally accepted in the art that, like myostatin, Activin A inhibition can lead to muscle mass enhancement. For example, in mice, inhibition of myostatin alone typically leads to about 15-30% increase in muscle mass, depending on how it is measured. Further inhibition of Activin A in mice leads to greater increase in muscle, around 30-45%. In general, the overall increase in muscle mass seen with inhibition is lower in non-human primates and even lower in humans. In humans, inhibition of only myostatin typically leads to 3-5% increase in muscle mass depending on the particular inhibitor and the patient population being studied. In one clinical study with healthy volunteers, combination of anti-myostatin and anti-Activin A led to greater muscle mass gains (~7% at highest dose tested), indicating additive effects of myostatin inhibition and Activin A inhibition. Consistent with these observations, ActRllb inhibition using the anti-ActRI I antibody bimagrumab leads to muscle mass increase within this range. Thus, it is generally well established that inhibition of both myostatin and Activin A is the preferred approach for achieving optimal effects on muscle enhancement.

[0176]

[0152] However, given the diverse biological roles that Activin A plays in a number of tissues, Activin A inhibition may also lead to undesirable consequences, e.g., interference to or perturbation of normal bodily function of Activin A, notwithstanding its potential benefit in terms of muscle enhancement. Importantly, many of the biological roles of Activin A may be especially sensitive to various cardiometabolic conditions, and therefore, striking a balance between the benefit of Activin A inhibition and risk associated therewith is a crucial consideration in selecting the right kind of myostatin inhibitors as therapeutics for a particular patient or patient population / subpopulation. Data disclosed herein suggest that it is possible to achieve equivalent or in some cases superior efficacy in a well-established DIO model by using highly potent myostatin-selective inhibitors, as compared to broadly antagonizing the ActRII signaling altogether, thus avoiding risk associated with Activin A inhibition without compromising efficacy.

[0177]

[0153] Some of the notable biological functions of Activin A and possible implications of inhibiting Activin A in a cardiometabolic context are further discussed below.

[0178] Effects on reproductive biology:

[0179]

[0154] Activin A serves important roles in the regulation of reproductive hormones and is a major regulator of testicular and ovarian development. In the ovary, Activin A promotes oocyte maturation and regulates granulosa cell steroidogenesis. It is also essential in endometrial repair following menstruation, decidualization and maintaining pregnancy. Activin is produced in the gonads, pituitary gland, placenta, and other organs. In the ovarian follicle, Activin increases follicle-stimulating hormone (FSH) binding and FSH-induced aromatization. It is known that Activin stimulates secretion of FSH from pituitary cells desensitized to gonadotropin-releasing hormone. FSH is a key reproductive hormone which controls menstrual cycle in women and is critical for puberty and reproductive development. In addition, it participates in androgen synthesis enhancing luteinizing hormone (LH) action in the ovary and testis. In the male, activin enhances spermatogenesis. As such, systemic inhibition of Activin A can perturb the normal hormonal regulation and reproductive development mediated by Activin A. Indeed, clinical trials with an ActRII inhibitor (bimagrumab) observed a reduction in FSH in both men and women. (Garito et al. Diabetes Obes Metab. 2018;20(1 ):94-102).

[0180]

[0155] These risks may be particularly undesirable for patients who are either still in development / growth phase, or those who are of reproductive potential. In previous clinical trials of non-selective myostatin inhibitors that also inhibit Activin A, it appears that patient enrollment criteria were carefully considered to circumvent these risks. For example, for anti-myostatin Adnectin™ which was in the clinic for Duchenne muscular dystrophy (DMD) (BMS), the target patient population was limited to boys due to the X chromosome-linked nature of the disorder that affect boys, and most of the healthy volunteers for the phase 1 trial were limited to male subjects. As such, possible risk of Activin A inhibition to girls and women was not addressed by the study design. Eventually, the trial was discontinued. In another example, Novartis conducted a clinical trial of the anti-ActRII antibody bimagrumab, which inhibits multiple ligands that signal through the common receptor, including myostatin, GDF11 and Activin A. They studied the effects of the antibody in adult obese and diabetic patients. In this study, the enrollment criteria were limited to men or postmenopausal women, presumably due to the potential risk of Activin A inhibition for the regulation of reproductive hormones particularly in girls and women of reproductive potential.

[0181]

[0156] To address these limitations associated with Activin A inhibitors, myostatin inhibitors that do not inhibit Activin A can be employed, which are capable of targeting myostatin while retaining normal Activin A activities intact. This approach should circumvent these risks associated with Activin A inhibition and potentially allow the expansion of the eligible population to include those of reproductive potential.

[0182]

[0157] Accordingly, the present disclosure includes therapeutic use of a myostatin inhibitor that does not inhibit Activin A for treating a patient of reproductive potential who suffers from a metabolic disorder (e.g., obesity / overweight, diabetes, etc.), or who otherwise benefit from a myostatin inhibitor therapy. In some embodiments, the patient does not qualify or is not eligible for a non-selective myostatin inhibitor therapy due to one or more of the following factors: i) age; ii) pre- vs. post-menopause; iii) sex; iv) pregnancy / nursing status; and v) use of contraceptives. In some embodiments, a myostatin-selective inhibitor is used in the treatment of a metabolic disorder (e.g., obesity / overweight, diabetes, etc.) in a patient, wherein the treatment comprises administration of a myostatin-selective inhibitor that does not inhibit Activin A, to the patient in an amount effective to treat the disorder, wherein the patient is a male or female of reproductive potential. In some embodiments, the patient is under the age of 55. In some embodiments, the patient is a premenopausal female patient. In some embodiments, the patient is a female below the age of 55. In some embodiments, the patient is trying to become pregnant or is pregnant. In some embodiments, the patient is nursing. In some embodiments, the patient is a female patient not on a contraceptive / birth control.

[0183]

[0158] In some embodiments, suitable myostatin inhibitors for carrying out various embodiments of the disclosure are inhibitors of myostatin and GDF11 (e.g., dual inhibitors of myostatin and GDF11 ), which do not inhibit Activin A. These include, for example, certain ligand traps that bind mature myostatin and GDF11 , but not Activin A; certain neutralizing antibodies that bind both mature myostatin and GDF11 but not Activin A; and certain anti-myostatin Adnectins™ that bind mature myostatin and GDF11 but not Activin A. These myostatin inhibitors would not circumvent the potential detrimental effects of GDF11 inhibition but should avoid those associated with Activin A inhibition.

[0184]

[0159] In preferred embodiments, a myostatin-selective inhibitor is used for treating such patients. This approach has the advantage of both sparing GDF11 for its metabolic benefit, while avoiding Activin A inhibition-associated risks. In some embodiments, the myostatin-selective inhibitor is a neutralizing antibody that binds and inhibits mature myostatin but does not bind or inhibit Activin A, wherein optionally the neutralizing antibody is trevogrumab, a variant thereof or a derivative thereof. In other embodiments, the myostatin-selective inhibitor for treating such patient is an antibody that specifically targets pro / latent myostatin thereby inhibiting its activation, wherein optionally the antibody is apitegromab, GYM329, SRK-439, Ab101-Ab141 disclosed herein, a variant thereof, or derivative thereof. In most preferred embodiments, the myostatin-selective inhibitor is SRK-439.

[0185]

[0160] In carrying out any of these embodiments, the therapeutic regimen comprising a myostatin inhibitor may further include another therapy to address the metabolic condition, such as incretin-based therapy (e.g., incretin mimetics).

[0186] Risk of anemia:

[0187]

[0161] A causal relationship between obesity and iron deficiency has been suggested; see, for example, Wang et al. Front Public Health. 2023; 11 : 1188246. Using a genetic approach, Wang et al provides evidence that obesity can lead to iron deficiency anemia, suggesting that obesity is a risk factor for comorbidities and complications including iron deficiency anemia. (See, also, Alshwaiyat et al. Exp Ther Med. 2021 Nov; 22(5): 1268). This is consistent with the notion that obesity may constitute a form of chronic inflammation. Correlation between chronic inflammation and anemia is well-established in conditions such as chronic kidney disease (CKD). Furthermore, for diabetic individuals, inflammation can occur in the joints, muscles, or excess adipose tissue. Indeed, anemia is also one of the most common and prevalent hematologic disorders among patients with diabetes. Taken together, it is postulated herein that there appears a plausible link between obesity, diabetes, chronic inflammation and anemia.

[0188]

[0162] Lines of evidence suggest that Activin A expression is induced during hematopoietic differentiation and that Activin A enhances erythropoiesis. Given this, it is contemplated herein that it is potentially detrimental to systemically inhibit Activin A in patients with an elevated risk of anemia. Whilst Activin A inhibition may have beneficial effects on weight loss and muscle enhancement per se, careful risk-benefit consideration is warranted. As alluded to above, obese and / or diabetic patients, as well as those with chronic inflammation, are at higher risk of suffering from anemia. Therefore, it is advantageous to use a myostatin-selective inhibitor, over non-selective inhibitors which also inhibit Activin A, in the treatment of these metabolic conditions in order to avoid elevating the risk of anemia.

[0189]

[0163] Anemia is also a frequent comorbidity of heart failure and is associated with poor outcomes. Anemia in heart failure is considered to develop due to a complex interaction of iron deficiency, kidney disease, and cytokine production, although micronutrient insufficiency and blood loss may contribute. In heart failure, serum levels of Activin A are elevated and positively correlate both with severity of disease and age-dependent cardiac dysfunction (MacDonnell et al. Front Cardiovasc Med. 2022 Nov 10:9: 1038114. doi: 10.3389 / fcvm.2022.1038114. eCollection 2022), suggesting that Activin A inhibition may potentially be beneficial; however, such benefit must be carefully weighed against its potential risk.

[0190]

[0164] Accordingly, the present disclosure provides therapeutic use of myostatin inhibitors that do not inhibit Activin A. Such inhibitors encompass inhibitors of myostatin and GDF11 (e.g., dual inhibitors of myostatin and GDF11), which do not inhibit Activin A. These include, for example, certain ligand traps that bind mature myostatin and GDF11 , but not Activin A; certain neutralizing antibodies that bind both mature myostatin and GDF11 but not Activin A; and certain anti- myostatin Adnectins™ that bind mature myostatin and GDF11 but not Activin A. These myostatin inhibitors would not circumvent the potential detrimental effects of GDF11 inhibition but should avoid those associated with Activin A inhibition.

[0191]

[0165] In preferred embodiments, a myostatin-selective inhibitor is used for treating such patients. This approach has the advantage of both sparing GDF11 for its metabolic benefit, while avoiding Activin A inhibition-associated risk of anemia. In some embodiments, the myostatin-selective inhibitor is a neutralizing antibody that binds and inhibits mature myostatin but does not bind or inhibit Activin A, wherein optionally the neutralizing antibody is trevogrumab, a variant thereof or a derivative thereof. In other embodiments, the myostatin-selective inhibitor for treating such patient is an antibody that specifically targets pro / latent myostatin thereby inhibiting its activation, wherein optionally the antibody is apitegromab, GYM329, SRK-439, Ab101-Ab141 disclosed herein, a variant thereof, or derivative thereof. In most preferred embodiments, the myostatin-selective inhibitor is SRK-439.

[0192]

[0166] Accordingly, the present disclosure provides a myostatin-selective inhibitor for use in the treatment of a metabolic disorder in a subject, wherein the treatment comprises administration of a selective inhibitor of myostatin (i.e. , selectively inhibits myostatin and does not inhibit Activin A or GDF11 ), wherein the subject is suffering from or at risk of developing anemia. In some embodiments, the metabolic disorder is obesity, diabetes, fatty liver disease and / or chronic inflammation. In some embodiments, the chronic inflammation is CKD. In some embodiments, the fatty liver disease is NAFLD or NASH / MASH. In some embodiments, the subject suffers from anemia (such as spur cell anemia) associated with severe liver dysfunction, such as NASH / MASH.

[0193]

[0167] In carrying out any of these embodiments, the therapeutic regimen comprising a myostatin inhibitor may further include another therapy to address the metabolic condition, such as incretin-based therapy (e.g., incretin mimetics). Effects on wound healing:

[0194]

[0168] Evidence in literature suggests that Activin A is involved in the process of wound healing (see, for example, Munz et al. 1999; The EMBO J. 18(19): 5205-5215). More specifically, data point to the ability of Activin A to accelerate wound healing (reviewed in, for example: Cangkrama et al., 2020, Trends in Mol. Med., 26(12):1107-1117). Wound healing is a complex biological process that is essential for normal function, and as such, it is detrimental to interfere with the body’s ability to promote wound healing by the use of non-selective myostatin inhibitors that block Activin A.

[0195]

[0169] Patients suffering from cardiovascular disorders, including diabetic patients, are at a higher risk of experiencing impaired wound healing. Indeed, there is significant evidence showing that diabetic patients are at a higher risk for increased wound infections, wound dehiscence, and pathological scarring. Factors such as nutritional status and glycemic control also significantly influence diabetic wound outcomes (Dasari et al., 2021 , Semin Plast Surg., 35(3): 153- 158). The myostatin-selective approach disclosed herein should avoid the risk associated with impaired would healing which may be exacerbated by Activin A inhibition.

[0196]

[0170] Accordingly, the present disclosure includes a myostatin inhibitor for use in the treatment of a cardiovascular disorder in a subject, or a metabolic disorder with a risk of cardiovascular condition, wherein the myostatin inhibitor does not inhibit Activin A, wherein the cardiovascular disorder is associated with higher risk of impaired wound healing. Therapeutic use of myostatin inhibitors that do not interfere with Activin A activities are contemplated herein to treat patients who suffer from a metabolic condition that may elevate the risk of impaired would healing, such as diabetic patients. Those with open skin lesions and / or chronic wounds (e.g. , those with a history of difficulty healing) should benefit from this approach.

[0197]

[0171] Such inhibitors encompass inhibitors of myostatin and GDF11 (e.g., dual inhibitors of myostatin and GDF11 ), which do not inhibit Activin A. These include, for example, certain ligand traps that bind mature myostatin and GDF11 , but not Activin A; certain neutralizing antibodies that bind both mature myostatin and GDF11 but not Activin A; and certain anti-myostatin Adnectins™ that bind mature myostatin and GDF11 but not Activin A. By taking this approach, these myostatin inhibitors would not circumvent the potential detrimental effects of GDF11 inhibition but should spare Activin A for its benefit in promoting wound healing.

[0198]

[0172] In preferred embodiments, a myostatin-selective inhibitor is used for treating such patients (e.g., patients who suffer from a metabolic condition that may elevate the risk of impaired wound healing). This approach has the advantage of sparing both GDF11 for its metabolic benefit and Activin A for its benefit in promoting would healing. In some embodiments, the myostatin-selective inhibitor is a neutralizing antibody that binds and inhibits mature myostatin but does not bind or inhibit Activin A, wherein optionally the neutralizing antibody is trevogrumab, a variant thereof or a derivative thereof. In other embodiments, the myostatin-selective inhibitor for treating such patient is an antibody that specifically targets pro / latent myostatin thereby inhibiting its activation, wherein optionally the antibody is apitegromab, GYM329, SRK-439, Ab101-Ab141 disclosed herein, a variant thereof, or derivative thereof. In most preferred embodiments, the myostatin-selective inhibitor is SRK-439.

[0199]

[0173] Accordingly, the present disclosure provides a myostatin-selective inhibitor for use in the treatment of a metabolic disorder with cardiovascular risk in a subject, wherein the treatment comprises administration of a selective inhibitor of myostatin (i.e. , selectively inhibits myostatin and does not inhibit Activin A or GDF11 ), wherein the subject is suffering from or at risk of cardiovascular disorder / complications. In some embodiments, the metabolic disorder is obesity, diabetes, fatty liver disease and / or chronic inflammation. In some embodiments, the chronic inflammation is CKD. In some embodiments, the fatty liver disease is NAFLD or NASH / MASH.

[0200]

[0174] In carrying out any of these embodiments, the therapeutic regimen comprising a myostatin inhibitor may further include another therapy to address the metabolic condition, such as incretin-based therapy (e.g., incretin mimetics). Effects on bone and bone repair:

[0201]

[0175] Activin A has been shown to promote bone fracture repair (Yao et al., 2022, BioRxiv, 2022.10.05.510962), indicating that Activin A activation may serve a positive role in the process. Indeed, Activin A expression becomes elevated during bone repair, the process of which likely involves Activin A-dependent stimulation of stem cell proliferation and differentiation. In support of the finding, systemic inhibition of Activin A with the use of a neutralizing anti-Activin A antibody was found to delay bone fracture repair. Conversely, local administration of exogenous Activin A accelerated fracture healing. These findings point to an important role of Activin A in bone repair, and it raises concerns regarding the risk of interfering with this process by using non-selective myostatin inhibitors that also target Activin A. This is particularly relevant for patients who suffer from or are at risk of developing metabolic disorders, because weakened bone is a risk factor in many of these patients. Additionally, people with a history of frequent falls and / or bone fractures should also be considered a high-risk category. Those at risk of developing or suffering from bone loss, such as osteoporosis, are also at elevated risk of bone fracture, including, for example, women over 55.

[0202]

[0176] Data from DIO mice presented herein reveal that a diet high in fat can cause detrimental effects on bone. This finding points to the possible risk of Activin A inhibition in patients suffering from diet-induced obesity, again, reinforcing the notion that the use of myostatin-selective inhibitors is advantageous over non-selective inhibitors in this context.

[0203]

[0177] Accordingly, contemplated herein are methods involving therapeutic use of myostatin inhibitors that spare (i. e. , do not inhibit) Activin A, for treating patients suffering from a metabolic disorder, who are at risk of bone fracture or who can benefit from accelerated or enhanced fracture repair. Patients who may benefit from such treatment include those suffering from or at risk of developing a bone disorder, such as more frequent or more severe fractures. In some embodiments, the metabolic disorder is diabetes, e.g., type 2 diabetes. In some embodiments, the bone disorder is osteoporosis. In some embodiments, such myostatin inhibitor is selected to treat a female patient, such as those over the age of 55. In some embodiments, the subject suffers from or at risk of developing a cardiovascular disease. For example, the subject is at risk of developing a cardiovascular disease when the subject has had a prior cardiovascular incident and / or has a family history of cardiovascular disease(s).

[0204]

[0178] Myostatin inhibitors that do not inhibit Activin A encompass inhibitors of myostatin and GDF11 (e.g., dual inhibitors of myostatin and GDF11 ), which do not inhibit Activin A. These include, for example, certain ligand traps that bind mature myostatin and GDF11 , but not Activin A; certain neutralizing antibodies that bind both mature myostatin and GDF11 but not Activin A; and certain anti-myostatin Adnectins™ that bind mature myostatin and GDF11 but not Activin A. By taking this approach, these myostatin inhibitors would not circumvent the potential detrimental effects of GDF11 inhibition but should spare Activin A for its benefit in promoting bone repair.

[0205]

[0179] In preferred embodiments, a myostatin-selective inhibitor is used for treating such patients. This approach has the advantage of sparing both GDF11 for its metabolic benefit and Activin A for its benefit in promoting bone repair. In some embodiments, the myostatin-selective inhibitor is a neutralizing antibody that binds and inhibits mature myostatin but does not bind or inhibit Activin A, wherein optionally the neutralizing antibody is trevogrumab, a variant thereof or a derivative thereof. In other embodiments, the myostatin-selective inhibitor for treating such patient is an antibody that specifically targets pro / latent myostatin thereby inhibiting its activation, wherein optionally the antibody is apitegromab, GYM329, SRK-439, Ab101-Ab141 disclosed herein, a variant thereof, or derivative thereof. In most preferred embodiments, the myostatin-selective inhibitor is SRK-439.

[0206]

[0180] Accordingly, the present disclosure provides a myostatin-selective inhibitor for use in the treatment of a metabolic disorder with fracture risk in a subject, wherein the treatment comprises administration of a selective inhibitor of myostatin (i.e., selectively inhibits myostatin and does not inhibit Activin A or GDF11 ), wherein the subject has a bone fracture or is at risk of frequent or severe bone fracture. In some embodiments, the metabolic disorder is obesity, diabetes, fatty liver disease and / or chronic inflammation. In some embodiments, the chronic inflammation is CKD. In some embodiments, the fatty liver disease is NAFLD or NASH / MASH.

[0207]

[0181] In carrying out any of these embodiments, the therapeutic regimen comprising a myostatin inhibitor may further include another therapy to address the metabolic condition, such as incretin-based therapy (e.g., incretin mimetics).

[0208] Effects on hair follicle regulation; hair loss:

[0209]

[0182] Among the many biological functions of Activin A concerns regulating hair follicles and regrowth of the hair shaft. In a 2011 research article, Qiu et al. reported that conditional knockout of Activin Receptor type 1 B in mice revealed abnormal hair loss (J. Invest. Dermatol. 131 (5): 1067-1076). Using histological analyses, the authors observed that those hair follicles that developed during morphogenesis were later disrupted by delays in hair cycle reentry. Failure in cycling of the hair follicles and regrowth of the hair shaft and the inner root sheath resulted in subsequent severe hair loss.

[0210]

[0183] In the context of combination therapies that include an Activin A inhibitor (such as selective inhibitors of Activin A and non-selective inhibitors of myostatin) and an incretin-based therapy, this is particularly noteworthy because commercially available GLP-1 RAs such as Ozempic® have recently been implicated in possible hair loss (see, for example: Desai et al, Int J Dermatol. 2024, 63(9): 1128-1130. Review, “GLP-1 agonists and hair loss: a call for further investigation”; Nakhla et al. “Risk of Suicide, Hair Loss, and Aspiration with GLP1 -Receptor Agonists and Other Diabetic Agents: A Real-World Pharmacovigilance Study.” Cardiovasc Drugs Ther. 2024 Sep 12.). Whilst the possibility of a direct link is still under investigation, it is nevertheless prudent to avoid combining two active agents, each of which is associated with hair loss, in orderto reduce the risk. Therefore, according to the present disclosure, a myostatin inhibitor that does not inhibit Activin A (preferably myostatin-selective inhibitor) is selected for use in combination with an incretin therapy (e.g., a GLP-1 RA) for treating a patient having a metabolic disorder, such as overweight / obesity. In some embodiments, the patient is at risk of hair loss.

[0211] Effects on neuroprotection / neuro-survival:

[0212]

[0184] Activin has previously been shown to act as a nerve cell survival factor and to have neurotrophic effects on neurons. See, e.g., Bao et al. J Endocrinol. 2005 Mar;184(3):493-504. doi: 10.1677 / joe.1.05978. Activin receptors are highly expressed in neuronal cells, and activin mRNA expression is upregulated by neuronal activity. In Drosophila, Activin signaling has been reported to promote synaptic growth at the neuromuscular junction (Ellis et al. Dev Biol. 2010 Jun 15;342(2): 121 -33. doi: 10.1016 / j.ydbio.2010.03.012. Epub 2010 Mar 24).

[0213]

[0185] Similarly, in mammals, Activin A has been reported to exert powerful neurotrophic effects on striatum GABAergic neurons in rat model of Huntington’s disease (Hughes et al. Neuroscience. 1999;92(1 ):197-209. doi: 10.1016 / s0306- 4522(98)00724-6). While it is unclear whether Activin A has equivalent effects on peripheral neurons, GABA is widely present in peripheral tissues, and moreover, the GABA receptors are expressed in overlapping tissues as Activin A, raising the possibility that Activin A may have similar neurotrophic effects on peripheral neurons as well. See, e.g., Tanaka and Taniyama, (1992). The Role of GABA in the Peripheral Nervous System. In: Erdo, S.L. (eds) GABA Outside the CNS. Springer, Berlin, Heidelberg, https: / / doi.org / 10.1007 / 978-3-642-76915-3_1.

[0214]

[0186] It is well documented that a significant fraction of diabetic patient population suffers from diabetic neuropathy, which is often triggered by high blood sugar that can injure nerves throughout the body, although nerves in the legs and feet are most often affected. Given the possible role of Activin A in promoting survival and growth of neurons, the use of inhibitors that inhibit Activin A raises safety concerns particularly for those at high risk of developing or suffering from nerve injuries, e.g., neuropathies.

[0187] Peripheral neuropathy is nerve damage caused by several different conditions. Health conditions that can cause peripheral neuropathy include but are not limited to the following:

[0215] • Autoimmune diseases, including, without limitation: Sjogren's syndrome, lupus, rheumatoid arthritis, Guillain- Barre syndrome, chronic inflammatory demyelinating polyneuropathy and vasculitis, as well as certain cancers related to the body's immune system can cause polyneuropathy, which are a form of autoimmune disorder called paraneoplastic syndrome);

[0216] • Diabetes and metabolic syndrome: This is the most common cause associated with peripheral neuropathy. Among people with diabetes, more than half will develop some type of neuropathy.

[0217] • Infections: These include certain viral or bacterial infections, including Lyme disease, shingles, hepatitis B and C, leprosy, diphtheria, and HIV.

[0218] • Inherited disorders: Disorders such as Charcot-Marie-Tooth disease are hereditary types of neuropathy that run in families.

[0219] • Tumors: Cancerous growths, also called malignant, and noncancerous growths, also called benign, can grow on or press on nerves.

[0220] • Bone marrow disorders: These include a protein in the blood that isn't usually there, called monoclonal gammopathies, a rare form of myeloma that affects the bones, lymphoma and the rare disease amyloidosis.

[0221] • Other diseases associated with peripheral neuropathy include metabolic conditions such as kidney disease or liver disease, and an underactive thyroid, also known as hypothyroidism.

[0222] • Other causes of neuropathies include: Alcohol use disorder: Unhealthy dietary choices made by people with alcohol use disorder, also known as alcoholism, and poor absorption of vitamins can lead to low amounts of essential vitamins in the body; Exposure to poisons: Toxic substances include industrial chemicals and heavy metals such as lead and mercury; Medicines / drug-induced neuropathy: Certain medicines, especially chemotherapy used to treat cancer, can cause peripheral neuropathy; Injury or pressure on the nerve; and, Low vitamin levels: B vitamins, including B-1 , B-6 and B-12, as well as copper and vitamin E are crucial to nerve health.

[0223]

[0188] Advantageously, myostatin-selective inhibitors such as those disclosed herein can avoid the risk of exacerbating neuropathy triggered by Activin A inhibition. Therefore, it is contemplated that patients who are in need of myostatin inhibitor therapy but who suffer from or are at risk of developing peripheral neuropathy (e.g., those with one or more of the aforementioned health factors) benefit from myostatin inhibitors that can spare Activin A (such as myostatin- selective inhibitors and dual myostatin / GDF11 inhibitors), over non-selective inhibitors that also target Activin A and Activin A-selective inhibitors.

[0224]

[0189] Accordingly, the present disclosure provides a myostatin inhibitor that does not inhibit Activin A for use in the treatment of a cardiovascular or metabolic disorder in a subject wherein the cardiovascular or metabolic disorder is associated with elevated risk of neuropathy (peripheral neuropathy). In some embodiments, the subject has diabetes, e.g., type 2 diabetes.

[0225]

[0190] Myostatin inhibitors that do not inhibit Activin A encompass inhibitors of myostatin and GDF11 (e.g., dual inhibitors of myostatin and GDF11 ), which do not inhibit Activin A. These include, for example, certain ligand traps that bind mature myostatin and GDF11 , but not Activin A; certain neutralizing antibodies that bind both mature myostatin and GDF11 but not Activin A; and certain anti-myostatin Adnectins™ that bind mature myostatin and GDF11 but not Activin A. By taking this approach, these myostatin inhibitors would not circumvent the potential detrimental effects of GDF11 inhibition but should avoid the risk of exacerbating neuropathy triggered by Activin A inhibition in high-risk patient populations.

[0226]

[0191] In preferred embodiments, a myostatin-selective inhibitor is used for treating such patients. This approach has the advantage of both sparing GDF11 for its metabolic benefit and avoiding increased risk of neuropathy associated with Activin A inhibition. In some embodiments, the myostatin-selective inhibitor is a neutralizing antibody that binds and inhibits mature myostatin but does not bind or inhibit Activin A, wherein optionally the neutralizing antibody is trevogrumab, a variant thereof or a derivative thereof. In other embodiments, the myostatin-selective inhibitor for treating such patient is an antibody that specifically targets pro / latent myostatin thereby inhibiting its activation, wherein optionally the antibody is apitegromab, GYM329, SRK-439, Ab101-Ab141 disclosed herein, a variant thereof, or derivative thereof. In most preferred embodiments, the myostatin-selective inhibitor is SRK-439.

[0227]

[0192] Accordingly, the present disclosure provides a myostatin-selective inhibitor for use in the treatment of a metabolic disorder in a subject, wherein the treatment comprises administration of a selective inhibitor of myostatin (i.e. , selectively inhibits myostatin and does not inhibit Activin A or GDF11 ), wherein the subject is suffering from or at risk of neuropathy. In some embodiments, the metabolic disorder is obesity, diabetes, fatty liver disease and / or chronic inflammation. In some embodiments, the chronic inflammation is CKD. In some embodiments, the fatty liver disease is NAFLD or NASH / MASH. In preferred embodiments, the subject is an obese / overweight diabetic patient.

[0228]

[0193] In carrying out any of these embodiments, the therapeutic regimen comprising a myostatin inhibitor may further include another therapy to address the metabolic condition, such as incretin-based therapy (e.g., incretin mimetics). In some embodiments, the subject is on a metformin therapy.

[0229] The subject

[0230]

[0194] In carrying out various embodiments disclosed herein, preferred subjects, who may benefit from myostatin inhibitor therapy (preferably myostatin-selective inhibitor therapy), are human subjects suffering from a metabolic disorder and display one or more of the features / criteria outlined below.

[0231]

[0195] Table 1A and Table 1 B below show BMI ranges that may be used for overweight and obesity. In some embodiments, the subject is male or female over the age of 10 and is overweight or obese as determined by BMI.

[0232] Table 1 A. BMI of adults ages 20 and older

[0233]

[0196] Younger individuals (e.g., children and adolescents) grow at different rates at different times, so it is not always easy to tell if a child is overweight. The CDC BMI growth charts may be used to compare a child’s or adolescent’s BMI with other children or adolescents of the same sex and age. An online tool for gauging the BMIs of children and teens can be found at: https: / / www.cdc.gov / healthyweight / bmi / calculator.html.

[0234] Table 1 B. BMI of children and adolescents ages 2 to 19

[0197] In some embodiments, a subject having obesity is a subject who carries excess weight but does not have any comorbidities or risk factors for comorbid conditions and who does not experience any impairments in their daily feeling or functioning. In other embodiments, a subject having obesity is obese with risk factors, wherein the subject carries excess weight and does not yet have any comorbidities, but wherein the subject has measurable risk factors for comorbid conditions and / or impairments to their daily feeling or functioning. For instance, the subject may be at risk for insulin resistance, glucose intolerance, hypertension, cardiovascular disease, dyslipidemia, hyperuricemia, type 2 diabetes, stroke, fatty liver disease, kidney disease and other health issues. In other embodiments, a subject having obesity is obese and sick, wherein the subject carries excess weight and has one or more obesity-attributable comorbidities and impairments to their daily feeling or functioning. In some embodiments, the subject is overweight or obese but is not diabetic.

[0235]

[0198] In some embodiments, a subject having obesity is evaluated using the Edmonton Obesity Staging System (EOSS), a 5-point ordinal classification system that considers comorbidity and functional status, as shown in Table 2.

[0236] Table 2. The Edmonton obesity staging system.

[0237] (See, e.g., Padwal et al, CMAJ. 2011 Oct 4; 183(14): e1059-e1066.)

[0238]

[0199] In some embodiments, a medicament disclosed herein is suitable for administration in a pediatric population, adult population, and / or an elderly population.

[0239]

[0200] In some embodiments, such medicament is suitable for administration in a subject aged 2-19 years, inclusive of endpoints.

[0240]

[0201] In some embodiments, such medicament is suitable for administration in a subject aged 12 years or older (e.g., 12-17 years), inclusive of endpoints.

[0241]

[0202] The population in need for a myostatin inhibitor (e.g., an anti-pro / latent myostatin antibody or antigen-binding fragment) described herein, may range between 0 and 6 months of age, between 0 and 12 months of age, between 0 and 18 months of age, between 0 and 24 months of age, between 0 and 36 months of age, between 0 and 72 months of age, between 6 and 36 months of age, between 6 and 36 months of age, between 6 and 72 months of age, between 12 and 36 months of age, between 12 and 72 months of age. In some embodiments, the pediatric population suitable for receiving the myostatin inhibitor, e.g., antibody or antigen-binding fragment, described herein who is likely to benefit from such treatment ranges between 0 and 6 years of age, between 0 and 12 years of age, between 3 and 12 years of age, between 0 and 17 years of age. In some embodiments, the population has an age of at least 5 years, e.g., 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, or 17 years. In some embodiments, the pediatric population is aged below 18 years old. In some embodiments, the pediatric population may be (a) at least 5 years of age and (b) below 18 years of age.

[0203] The adult population in need of therapy comprising a myostatin inhibitor (e.g., an anti-pro / latent myostatin antibody or antigen-binding fragment) described herein may have an age of at least 20 years, e.g., at least 20, 25, 30, 35, 40, 45, 50, 55, 60 or 65 years. In some embodiments, the adult population is below 65 years of age. In some embodiments, the adult population is below 75 years of age.

[0242]

[0204] The elderly population in need of therapy comprising a myostatin inhibitor (e.g., an anti-pro / latent myostatin antibody or antigen-binding fragment) described herein have an age of 65 years or older (i.e. , 2 65 years old), e.g., at least 70, 75 or 80 years.

[0243]

[0205] In some embodiments, a human subject who is likely to benefit from the treatment comprising a myostatin inhibitor (e.g., an anti-pro / latent myostatin antibody or antigen-binding fragment) described herein is a human patient having, at risk of developing, or suspected of having a metabolic disease. In some embodiments, a human subject who is likely to benefit from the treatment comprising a myostatin inhibitor (e.g., an anti-pro / latent myostatin antibody or antigen-binding fragment) described herein is a human patient having a metabolic condition associated with a muscle disorder (e.g., spinal muscular atrophy, spinal cord injury, a muscle dystrophy). A subject having a metabolic disease or condition (e.g., obesity, type 2 diabetes mellitus (T2DM), etc.) can be identified by routine medical examination, e.g., laboratory tests, organ functional tests, CT scans, or ultrasounds. A subject suspected of having a metabolic disease or condition might show one or more symptoms of the metabolic disease or condition. A subject at risk for a metabolic disease or condition can be a subject having one or more of the risk factors for the metabolic disease or condition.

[0244]

[0206] In some embodiments, a human subject who is likely to benefit from the treatment comprising a myostatin inhibitor is a human subject having T2DM with a hemoglobin A1 C (HbA1 C) between 5% and 15% (e.g., between 6.5% and 10%, inclusive of endpoints).

[0245]

[0207] In some embodiments, a human subject who is likely to benefit from the treatment comprising a myostatin inhibitor (e.g., an anti-pro / latent myostatin antibody or antigen-binding fragment) described herein is a human subject having T2DM who has been on an anti-diabetic treatment for at least 3 months. In some embodiments, the subject has been on an anti-diabetic treatment for approximately 3 months. In some embodiments, the subject has been on an antidiabetic treatment for at least 6 months.

[0246]

[0208] In some embodiments, a human subject who is likely to benefit from the treatment comprising a myostatin inhibitor (e.g., an anti-pro / latent myostatin antibody or antigen-binding fragment) described herein is a human subject having a body weight of at least 70 kg. In some embodiments, the subject has a body weight of at least 80 kg. In some embodiments, the subject has a body weight of 80 kg to 140 kg, inclusive of endpoints. In some embodiments, the subject has a body weight of more than 140 kg.

[0247]

[0209] In some embodiments, the subject has maintained stable body weight (e.g., ±5 kg) with a predetermined time period (e.g., 90 days) of screening (e.g., being considered as candidate for receiving the therapy).

[0248]

[0210] In some embodiments, the subject is an adult male or female of 218 and 265 years of age and has a BMI of 230.0 kg / m2to 245 kg / m2, or 227.0 kg / m2to 30 kg / m2, with the presence of one or more weight-related comorbidity.

[0249]

[0211] In some embodiments, assessment of weight and health risk involves using three key measures: i) Body mass index (BMI); ii) Waist circumference; and, iii) Risk factors for diseases and conditions associated with obesity.

[0250]

[0212] Body weight is a factor to be considered. For example, difference from ideal body weight is >35% in males and >45% in females may be indicative of the subject being overweight / obese.

[0251]

[0213] Body mass index (BMI), which is the ratio of body weight in kg and surface area in m2, of >30 kg / m2(25 kg / m2in Asians) may indicate overweight / obesity.

[0214] Waist circumference, which is measured at the end of expiration in a standing position at the midpoint of the lowest rib case and the iliac crest, which exceeds 102 cm in men and 88 cm in women (90 cm in males and 80 cm in Asian females) may indicate high risk.

[0252]

[0215] Waist-hip ratio, which is the ratio of waist circumference and the hip circumference, which exceeds 1.0 for men and 0.85 for women may indicate high risk.

[0253]

[0216] In some embodiments, the subject is diabetic. In some embodiments, the subject has type 2 diabetes (T2D).

[0254]

[0217] In some embodiments, the subject is nondiabetic.

[0255]

[0218] In some embodiments, the subject is pre-diabetic. In some embodiments, prediabetes is managed with non- pharmacologic approaches, such as exercise and / or diet.

[0256]

[0219] In some embodiments, the subject has type 1 diabetes (T1 D). Evidence in literature suggests that bone mineral density (BMD) of total body and lumbar spine can be significantly lower in patients with T1 DM as compared to controls. Similarly, patients with T1 DM are reported to show about 10% less bone mineral content (BMC) in comparison with controls. These observations support the notion that patients with T 1 DM are at higher risk for fractures.

[0257]

[0220] In some embodiments, the subject has a history of cardiovascular disease (e g., heart failure) either in the patient or in family history (increased propensity).

[0258]

[0221] In some embodiments, the subject suffers from a fatty liver disease (e.g., NAFLD, NASH / MASH). Individuals with hepatic steatosis often display several metabolic abnormalities including insulin resistance and muscle atrophy. Several recent studies have shown frequent loss of muscle mass associated with poor prognosis in various chronic liver diseases. Overall, studies demonstrate muscle loss in nearly 60% of patients with end-stage liver diseases and this is associated with a worse prognosis. However, muscle loss is already present in the early stages of liver disease and worsens with its severity. This is particularly the case of nonalcoholic fatty liver disease (NAFLD), which encompasses a broad spectrum of disorders ranging from simple steatosis to nonalcoholic steatohepatitis (NASH), cirrhosis, and hepatocellular carcinoma, and where loss of muscle mass can occur very early during the disease. Indeed, in addition to the disturbances in muscle homeostasis related to metabolic disorders, several pieces of evidence indicate a specific role of the alterations in liver function in muscle loss (reviewed in: De Bandt et al (2018) Nutrients. 10(9): 1195). Accordingly, it is contemplated herein that patients suffering from liver diseases such as NAFLD and NASH / MASH may benefit from myostatin inhibitor therapy, preferably myostatin-selective therapy, such as SRK-439. Recently, incretins such as GLP-1 receptor agonists have been shown to be effective in promoting significant weight loss and decreasing liver fat contents in individuals with NAFLD or NASH. Despite these hepatoprotective benefits, incretin therapies can also trigger significant muscle loss (as discussed herein), and given the existing vulnerability of these patients to muscle atrophy / weakness, it is desirable to treat these patients with myostatin inhibitors (preferably myostatin-selective inhibitors such as SRK-439) in conjunction with the incretins. It is contemplated that the myostatin inhibitors may prevent or reduce the severity of muscle loss in patients with liver diseases, particularly when the patients are treated with incretins such as GLP-1 receptor agonists, which can further induce muscle loss.

[0259]

[0222] In some embodiments, the subject has a chronic disease, wherein optionally the chronic disease is associated with anemia (e.g., impaired erythropoiesis). In some embodiments, the chronic disease is chronic kidney disease (CKD). Notably, CKD patients often exhibit a low muscle mass and strength, leading to physical impairment and an increased mortality, and the myostatin pathway has been implicated in the process (reviewed in, for example, Bataille et al, (2021 ) Nephrol Dial Transplant. 36(11 ): 1986-1993). Thus, it is contemplated herein that patients suffering from muscle atrophy associated with CKD can benefit from a myostatin inhibitor therapy. It is particularly advantageous to employ a myostatin-selective inhibitor that does not inhibit GDF11 or Activin A in such patient population because of possible detrimental effects of GDF11 as discussed above, as well as the potential negative impact of Activin A inhibition on erythropoiesis in this population that is prone to developing anemia. Furthermore, more recently, GLP-1 analog therapy (e.g., semaglutide) has been reported to cause a dramatic reduction in the risk of kidney complications, heart issues and death in individuals with type 2 diabetes and CKD. Thus, the data support the use of incretins such as GLP-1 RAs as a treatment option for those suffering from CKD. However, as presented herein, GLP-1 receptor agonist therapy such as semaglutide can also induce significant muscle loss, and this effect may be especially undesirable and detrimental to CKD patients who are already at risk of muscle atrophy / weakness. It is contemplated herein that myostatin inhibitors (preferably myostatin-selective inhibitors such as SRK-439) can be used concurrently (e.g., in conjunction) with incretin therapies to counter (e.g., prevent or reduce severity of) muscle atrophy or weakness in CKD patients who receive incretin therapy.

[0260]

[0223] In some embodiments, the subject has physical limitations (e.g., disability such as spinal cord injury and muscular disorders) that lead to inability to exercise.

[0261]

[0224] In some embodiments, the subject has peripheral neuropathy, wherein optionally the peripheral neuropathy is associated with diabetes.

[0262]

[0225] In some embodiments, the subject has a chronic wound that is difficult to heal, wherein optionally the chronic wound is associated with cardiovascular disease such as diabetes.

[0263]

[0226] In some embodiments, the subject has a bone fracture, has a history of frequent falls, and / or, is prone to frequent and / or severe bone fractures.

[0264]

[0227] In some embodiments, the subject is on an incretin-based therapy, wherein optionally the incretin-based therapy comprises a GLP-1 receptor agonist, wherein further optionally the subject responds poorly to the incretin-based therapy, e.g., has adverse events (AEs).

[0265]

[0228] In some embodiments, the subject has been on an incretin-based therapy and is discontinuing the incretinbased therapy; or, the subject was on an incretin-based therapy and has discontinued the incretin therapy.

[0266]

[0229] In some embodiments, the subject is on an incretin-based therapy (such as incretins, e.g., GLP-1 analogs), wherein the subject is at risk of developing or has experienced hair loss.

[0267]

[0230] In some embodiments, the subject is on a metformin therapy. In some embodiments, the subject is a diabetic subject who is on a metformin therapy. In some embodiments, the subject is a prediabetic subject who is on a metformin therapy. In some embodiments, the subject is a diabetic or prediabetic subject who is overweight or obese, who is on a metformin therapy. In some embodiments, the subject is on a metformin therapy, wherein optionally the subject has experienced side effects with an incretin therapy, wherein optionally the incretin therapy comprises a GLP-1 receptor agonist, such as GLP-1 analogs,

[0268]

[0231] In some embodiments, the subject has received an incretin-based therapy (such as GLP-1 RAs, e.g., GLP-1 analogs) for weight management, but the effect of the incretin-based therapy has diminished over time. For example, the rate of fat mass loss or weight loss has decreased over time (e.g., following an initial phase of rapid weight loss) despite continuing the same dosing regimen, at which point, the risk of continued incretin-based therapy (e.g., side effects) may outweigh the benefits, leading to discontinuation of the therapy. Data from a longer-term preclinical study (see Example 14 below) show that the addition of a myostatin-selective inhibitor (e.g., SRK-439) to a semaglutide treatment paradigm can augment (e.g., prolong) the effect on fat mass loss, as compared to semaglutide monotherapy. More specifically, data in mice provided herein show that the effect of semaglutide on fat mass loss peaked at around 3 weeks after the start of the study, plateaued, then gradually declined over time, despite continued treatment. In the presence of SRK-439 (as a combination therapy with semaglutide), however, the peak effect occurred at around 9 weeks, indicating that myostatin inhibition can markedly prolong the fat loss effect in combination with GLP-1 receptor activator. In addition to this enhanced durability, the combination therapy further enhanced fat mass loss, as compared to semaglutide monotherapy, demonstrating an increased magnitude of efficacy. Accordingly, in some embodiments, a patient is administered a myostatin inhibitor, e.g., a myostatin-selective inhibitor, after the effects of an incretin-based therapy alone would have been expected to diminish. In some embodiments, the patient receiving the myostatin inhibitor also received prior doses of the myostatin inhibitor earlier, when the effects of the incretin-based began to diminish. In other embodiments, the patient receiving the myostatin inhibitor did not receive doses of the myostatin inhibitor prior to when the effects of the incretin-based therapy began to diminish. In some embodiments, the subject is a female subject of child-bearing potential.

[0269]

[0232] In some embodiments, the subject is not on a contraceptive.

[0270]

[0233] In some embodiments, the subject is pregnant or is trying to become pregnant.

[0271]

[0234] In some embodiments, the subject is nursing.

[0272]

[0235] In some embodiments, the subject has or is at risk of developing a bone disorder.

[0273]

[0236] In some embodiments, the subject has (e.g., is recovering from) a bone fracture.

[0274]

[0237] In some embodiments, the subject has a history of frequent falls or multiple bone fractures.

[0275]

[0238] In some embodiments, the subject is recovering from an injury.

[0276]

[0239] In some embodiments, the subject has or is at risk of developing anemia.

[0277]

[0240] In some embodiments, the subject lacks peripheral venous access hence benefits from subcutaneous administration of the myostatin inhibitor.

[0278]

[0241] In some embodiments, the subject has limited access to a medical facility that provides infusion (e.g., IV administration), hence benefits from subcutaneous administration of the myostatin inhibitor.

[0279]

[0242] In some embodiments, myostatin inhibitor is used in conjunction with another therapy to treat a metabolic disease. In some embodiments, said “another therapy” is or comprises a GLP-1 receptor agonist. In some embodiments, the “another therapy” includes, without limitation, albiglutide, taspoglutide, semaglutide, exenatide, BPI- 3016, GW002, glutazumab, exendin-4, exenatide, GLP-1 (7-36)NH2, everestmab, liraglutide, lixisenatide, tirzepatide, dulaglutide, danuglipron (Pfizer), PF-07081532, or orforglipron. In some embodiments, other therapies comprising a GLP-1 receptor agonist include, but are not limited to, GLP-1 receptor agonist / GIP receptor antagonist combination such as AMG 133 (Amgen); GLP-1 / GIP dual agonists such as tirzepatide (LY3298176; Eli Lilly), and CT-388; amylin / GLP-1 combination such as cagrilintide / semaglutide combination (Novo Nordisk); GLP-1 / glucagon combination such as DD01 (Neuraly); GLP-1 / glucagon receptor (GCG) agonist combinations such as ALT-801 (Altimmune), GLP- 1 / GIP such as CT-388 (Carmot); GLP-1 / glucagon dual agonist such as IBI362 (LY-330567 (Innovent / Eli Lilly), cotadutide, DD01 , danuglipron (PF-06882961 ) (Pfizer), mazdutide (IBI362; LY-330567), MEDI0382; noiiglutide, oxyntomodulin, pemvidutide, setmelanotide (Rhythm), survodutide, and GLP-1 / GIP / Glucagon triple receptor agonist such as retatrutide and LY343794. In some embodiments, the intended therapeutic goal or expected therapeutic outcome is a reduction of body weight (percent change from baseline) of at least 5% or at least 10%, reduced BMI, reduced waist circumference, reduced fat mass, increased lean-to-fat mass ratio, and / or improved insulin sensitivity.

[0280]

[0243] In some embodiments, said “additional therapy” or “another therapy” is or comprises a diet and / or exercise regimen. In some embodiments, the “additional therapy” or “another therapy” is or comprises a biguanide (e.g., metformin). In some embodiments, the “additional therapy” or “another therapy” is or comprises a sulfonylurea (e.g., glipizide, glimepiride, or glyburide), a bile acid sequestrant, a dopamine-2 agonist (e.g., bromocriptine), a dipeptidyl peptidase 4 inhibitor (e.g., alogliptin, linagliptin, saxagliptin, or sitagliptin), a meglitinide (e.g., nateglinide or repaglinide), a sodium-glucose cotransporter-2 (SGLT2) inhibitor (canagliflozin, dapagliflozin, or empagliflozin), a thiazolidinediones (e.g., troglitazone, rosiglitazone or pioglitazone), a weight-loss drug such as phentermine, benzphetamine, diethylpropion, or phendimetrazine.

[0244] In some embodiments, the “additional therapy” or “another therapy" comprises a healthy diet, exercise, or a thyroid hormone receptor-beta (THR-beta) selective agonist e.g., an aryloxyphenyl-based thyromimetic such as resmetirom or eprotirome, or a diphenylmethane based thyromimetic such as sobetirome, Sob-AM2, VK2809 (MB08711 ), MB07344, IS25, and TG68, e.g., wherein the metabolic disease is a liver disease, e.g., nonalcoholic steatohepatitis (NASH) or nonalcoholic fatty liver disease (NAFLD).

[0281]

[0245] In some embodiments, the “additional therapy” or “another therapy” is not a TGFp inhibitor (e.g., TGFpl inhibitor) if the subject has or is at risk of having NASH or NAFLD. In some embodiments, a subject having NAFLD or NASH or is at risk of developing NAFLD or NASH is not receiving and / or has not been treated with a TGFp inhibitor (e.g., TGFpl inhibitor).

[0282] Subjects suffering from muscle disorders or spinal cord injury:

[0283]

[0246] Patients with motor function impairment are at higher risk of developing certain metabolic dysregulation. These include muscle disorders associated with one or more genetic mutations that affect neuromuscular or motor functions, disuse, as well as injury-derived conditions such as spinal cord injury. Such metabolic dysregulation associated with muscle or motor impairment may include obesity (e.g., overweight or obese), diabetes, and fatty liver disorders.

[0284]

[0247] Incretin-based therapies, such as GLP-1 receptor agonists, have been shown to be effective in reducing body weight, controlling glucose metabolism and in some cases improve liver function. Patients with muscle / motor function impairment could benefit from such therapeutic effects of incretin-based therapies to address metabolic dysregulation. However, any benefit should be balanced against potential risk of further muscle breakdown induced by incretin-based therapies because any loss of lean mass could be especially detrimental to these patient populations who already suffer from compromised muscle / motor function. Moreover, due to disabilities, these patients may have limited ability to carry out physical exercise, rendering them even more susceptible to developing conditions such as obesity. To address such risk, concurrent treatment of myostatin inhibitors may provide protective effects against muscle loss when the patient is treated with an incretin-based therapy. Furthermore, in the event the incretin-based therapy is to be discontinued, the patient may further benefit from the myostatin inhibitor therapy in order to counter the effects (e.g., fat rebound / regain) induced by the incretin discontinuation.

[0285]

[0248] Accordingly, in some embodiments, a myostatin inhibitor is used in the treatment of a metabolic disorder in a subject, wherein the subject suffers from a muscle disorder and is on an incretin-based therapy. In some embodiments, the subject suffers from a myopathy, such as a muscular dystrophy. In some embodiments, the subject suffers from a neuromuscular disorder, such as spinal muscular atrophy (SMA) and amyotrophic lateral sclerosis (ALS). In preferred embodiments, the myostatin inhibitor is a myostatin-selective inhibitor, such as neutralizing antibodies that bind and mature myostatin and activation inhibitor antibodies that bind pro / latent myostatin thereby preventing its activation. In particularly preferred embodiments, the myostatin-selective inhibitor is selected from Ab101-Ab141. In most preferred embodiments, the myostatin-selective inhibitor is SRK-439.

[0286] Subjects with low bone mass or at risk of bone fracture:

[0287]

[0249] Data from DIO mice presented herein reveal that a diet high in fat (HFD) can cause detrimental effects on bone health. This finding points to the possible risk of Activin A inhibition in patients suffering from diet-induced obesity, again, reinforcing the notion that the use of myostatin-selective inhibitors is advantageous over non-selective inhibitors in this context. Furthermore, the findings discussed in the Example section herein suggest that certain parameters of bone health are negatively affected by incretin-based therapies, such as GLP-1 RAs. For example, data disclosed herein showed that trabecular thickness of the femur was slightly decreased by semaglutide treatment (as monotherapy) as compared to vehicle control in DIO mice. Similarly, cortical bone area measured at the diaphysis of the femur was also found to decrease in response to semaglutide monotherapy in DIO mice. Both of these parameters (i.e. , trabecular thickness and cortical bone area) are important indicators of bone health, which can provide information as to bone strength and changes in bone structure, such as those that occur in osteoporosis. Therefore, the potentially detrimental effects on bone observed with semaglutide treatment on these bone parameters raise the possibility that incretin-based therapies may negatively impact bone health. Selective inhibition of the myostatin pathway with the use of a myostatin-selective inhibitor antibody (e g., SRK-439) used in conjunction has shown to counter the detrimental effects of semaglutide. This, together with the findings that the combination therapy can enhance fat mass loss and prevent muscle loss, points to an improved therapeutic approach of incorporating a myostatin-selective inhibitor as part of the therapeutic regimen for patients who receive incretin-based therapies, such as GLP-1 RAs. This may be particularly advantageous for those who undergo rapid weight loss, which can induce muscle and bone atrophies. Rapid weight loss typically refers to at least a 10% reduction of body weight from the baseline (e.g., at least 10%, 15%, 20%, 25%, etc.), over a 6-month period or less (e.g., over 6 months, 5 months, 4 months, 3 months, etc.).

[0288]

[0250] Accordingly, the invention provides a myostatin-selective inhibitor for use in the treatment of a metabolic disorder (e.g., prevention of bone atrophy) in a subject who is treated with an incretin-based therapy or who undergoes a rapid weight loss, wherein the treatment comprises administration of a myostatin-selective inhibitor in an amount effective to maintain or enhance bone, wherein optionally, the maintenance or enhancement of bone is assessed by measuring bone density and / or cortical bone area. In some embodiments, the incretin-based therapy is or comprises a GLP-1 RA, such as GLP-1 analog. As used herein, “bone atrophy” refers to a reduction in bone density due to excessive breakdown of the bone substance and structure.

[0289]

[0251] In some embodiments, the subject who undergoes rapid weight loss receives or has received surgical interventions, such as bariatric surgery. In some embodiments, the bariatric surgery is sleeve gastrectomy, Roux-en-Y gastric bypass surgery, biliopancreatic diversion with duodenal switch, vertical banded gastroplasty, and / or gastric plication. In some embodiments, the surgery comprises the use of intragastric balloon, adjustable gastric band, and / or implantable gastric stimulation. With or without incretin-based therapy, atrophies of muscle and / or bone can accompany rapid weight loss associated with such surgical intervention. Incorporation of the myostatin-selective inhibitor therapy is aimed to prevent the muscle / bone atrophy as part of healthier weight management.

[0290]

[0252] Bone density can indicate bone fracture risk. Bone density tests can determine the relative bone health of an individual (e.g., patient) as compared to known standard values (e.g., what is considered healthy / normal within a particular cohort). This process involves comparing the test subject’s bone mineral density (BMD) to at least one of the following two norms: i) healthy 25- to 35-year-old adults of the same sex and ethnicity (“T-score”); and, ii) age-matched adults (“Z-score”). (Johns Hopkins Medicine at https: / / www.hopkinsmedicine.org / health / treatment-tests-and- therapies / bone-densitometry#:~:text=What%20is%20a%20bone%20density,of%20the%20hip%20or%20spine). The T-scores are calculated based on a standard deviation-based scale (positive T-scores indicate the bone is stronger than normal; negative T-scores indicate the bone is weaker than normal).

[0291]

[0253] The World Health Organization (WHO) sets the criteria for osteoporosis based on the following bone density levels:

[0292] • A T-score within 1 standard deviation (SD) (+1 or -1 ) of the young adult mean indicates normal bone density.

[0293] • A T-score of 1 to 2.5 SD below the young adult mean (-1 to -2.5 SD) indicates low bone mass.

[0294] • A T-score of 2.5 SD or more below the young adult mean (more than -2.5 SD) indicates the presence of osteoporosis.

[0295]

[0254] In general, the risk for bone fracture doubles with every SD below normal. Thus, a person with a BMD of 1 SD below normal (e.g., T-score of -1 ) has twice the risk for bone fracture as a person with a normal BMD. When this information is known, people with a high risk for bone fracture can be treated with the goal of preventing future fractures. Severe (established) osteoporosis is defined as having a bone density that is more than 2.5 SD below the young adult mean with one or more past fractures due to osteoporosis.

[0296]

[0255] On the other hand, to calculate a Z-score, the test subject’s BMD is compared to an age-matched norm. Z- scores are calculated in the same way, but the comparisons are made to someone of the test subject’s age, sex, race, height, and weight.

[0297]

[0256] Accordingly, the present invention encompasses the use of myostatin-selective inhibitors for the maintenance of bone mass or prevention of bone mass loss in a subject. In some embodiments, the subject has normal bone density, e.g., a T-score within 1 SD (between +1 and -1 ) at baseline (prior to commencing myostatin-selective inhibitor therapy), wherein the subject undergoes rapid weight loss. Optionally, the rapid weight loss is driven by an incretin-based therapy, such as GLP-1 RA therapy; and / or, the rapid weight loss is associated with a surgical intervention such as gastric bypass surgery. Rapid weight loss typically refers to at least a 10% reduction of body weight from the baseline (e.g., at least 10%, 15%, 20%, 25%, etc.), over a 6-month period or less (e.g., over 6 months, 5 months, 4 months, 3 months, etc.).

[0298]

[0257] In other embodiments, the subject has low bone mass (e.g., a T-score of between -1 and -2.5 SD) at baseline (i.e., prior to commencing myostatin-selective inhibitor therapy) and therefore is at risk of developing osteoporosis.

[0299]

[0258] In some embodiments, the present disclosure encompasses the use of myostatin-selective inhibitors for preventing bone fracture in a subject. In some embodiments, the subject has normal bone density, e.g., a T-score within 1 SD (between +1 and -1 ) at baseline (i.e., prior to commencing myostatin-selective inhibitor therapy), wherein the subject has another risk factor for bone fracture, e.g., an inflammatory condition such as obesity or diabetes, and / or undergoes rapid weight loss. In some embodiments, the subject has below-normal bone density, e.g., a T-score of -1 SD or below at baseline (prior to commencing myostatin-selective inhibitor therapy), wherein, optionally, the subject has undergone rapid weight loss. Optionally, the rapid weight loss is driven by an incretin-based therapy, such as GLP- 1 RA therapy; and / or, the rapid weight loss is associated with a surgical intervention such as gastric bypass surgery. Rapid weight loss typically refers to at least a 10% reduction of body weight from the baseline (e.g., at least 10%, 15%, 20%, 25%, etc.), over a 6-month period or less (e.g., over 6 months, 5 months, 4 months, 3 months, etc.).

[0300] Subjects suffering from injury or undergoing recovery therefrom

[0301]

[0259] In skeletal muscle, damage to muscle fibers leads to satellite cell activation - muscle stem cells located between myofibers that, upon stimulation, can convert to myoblasts that subsequently form myotubes to facilitate muscle repair. Applicant of the present disclosure recently found in two preclinical models of muscle diseases that selective inhibition of the myostatin signaling pathway using an antibody that selectively inhibits myostatin activation resulted in enhanced muscle strength (as evidenced by increased specific force indicative of improved muscle quality), as well as increased expression of muscle-specific protein (data not shown). These observations point to dual benefits of myostatin-selective inhibition: i) enhancing muscle function; and, ii) promoting or upregulating gene expression of muscle proteins which are building blocks for new muscle tissue.

[0302]

[0260] Therefore, it is contemplated herein that myostatin-selective inhibitors may be used to enhance (e.g., accelerate) the process of recovery from an injury. The selection of a myostatin-selective inhibitor, as opposed to non-selective inhibitor, is advantageous in this context for at least two reasons, Firstly, Activin A is said to play a positive role in would healing and bone repair, and therefore, it is undesirable to employ a non-selective inhibitor that inhibits not only myostatin but also Activin A. And secondly, concurrent inhibition of GDF11 has been shown to cause detrimental effects, such as causing weakened bone (Proc. Natl Acad. Sci. USA 117, 4910-4920 (2020)), indicating that there is a benefit in avoiding GDF11 inhibition. In particular, injury to a limb may involve at least partial immobilization during recovery, which can result in substantial muscle and bone atrophy, which may be at least in part due to disuse. The injury may or may not involve muscle damage. For example, the injury may be to a tendon, such as Achilles; a ligament, such as anterior cruciate ligament (ACL); a joint; a bone (e g., fractures); a muscle; and / or to a surrounding connective tissue. Myostatin inhibitor therapy is aimed to facilitate recovery from the injury by preventing atrophy and promoting muscle and / or bone regeneration. Myostatin inhibitor therapy may further improve recovery from an injury to achieve (e g., maintain or restore) a degree of flexibility and / or a range of motion that is as close to pre-injury as possible.

[0303]

[0261] Accordingly, the invention includes a myostatin-selective inhibitorfor use in the treatment of an injury in a patient, wherein the treatment comprises administration of a myostatin-selective inhibitor. In some embodiments, the patient does not have an underlying muscle disorder but suffers from an injury that can interfere with mobility or otherwise a normal level of physical activity due to the injury. In some embodiments, the injury is to a limb, such as upper limbs and lower limbs. In some embodiments the injury is to a shoulder, arm, elbow, forearm, wrist, hand, hip, thigh, knee, leg, ankle, and / or foot. In some embodiments, the injury involves a muscle, a tendon, a ligament, a bone, a joint, or surrounding connective tissue thereof. In some embodiments, the injury is a sports injury, such as a torn Achilles and torn ACL.

[0304]

[0262] In any of the embodiments, administration of a myostatin-selective inhibitor (e g., SRK-439) achieves one or more of the following effects: recovery time is shortened (e.g. , accelerated), muscle is retained (i.e. , muscle atrophy is prevented or the degree of muscle atrophy is lessened), and / or, bone atrophy is prevented or the degree of bone atrophy is lessened,

[0305]

[0263] In some embodiments, a myostatin-selective inhibitor (such as SRK-439) is administered to a patient who has suffered an injury immediately following the injury, e.g., within 24-48 hours of the injury. Such administration can be carried out at the first post-injury office visit with a physician. Depending on the injury (such as certain join injuries), patients undergo pre-surgery physical therapy in order to build up and strengthen the muscle before surgery can be operated on. Myostatin-selective inhibitor therapy during this period (pre-surgery) may facilitate the preparation for surgery and in turn shorten the overall recovery time.

[0306]

[0264] In some embodiments, myostatin-selective inhibitor is administered to a patient with an injury post-surgery in order to facilitate recovery. Such administration may be carried out within 24-48 hours after the surgery.

[0307]

[0265] In some embodiments, myostatin-selective inhibitor therapy is administered to a patient who has suffered an injury as part of the recovery period comprising physical therapy. Myostatin inhibition, coupled with suitable physical therapy, may synergistically facilitate the recovery process.

[0308]

[0266] Beyond the effects on facilitating injury recovery, it is contemplated that myostatin-selective inhibitor can help prevent metabolic dysregulation induced by disuse (e.g., immobilization) associated with the injury.

[0309] Drug-induced muscle-conditions; side effects:

[0310]

[0267] Certain medications are known to cause undesirable side effects, which may include muscle weakness, muscle cramps, and overall fatigue. It is therefore contemplated herein that myostatin inhibitors (myostatin-selective inhibitors in particular), used in conjunction with such medication, may be beneficial in preventing or lessening the severity and / or frequency of such side effects associated with the medication. In some embodiments, such medications are intended to treat a cardiometabolic / cardiovascular disorder. To give one example, commonly prescribed cholesterol-lowering medicine, such as statins, are associated with risk of muscle weakness and fatigue, among others. These patients may benefit from a myostatin inhibition therapy, which may not only improve metabolic health, but also may prevent drug- induced muscle weakness. Non-limiting examples of medications which may cause muscle weakness are further discussed below.

[0311]

[0268] Statins are a class of drugs prescribed to lower cholesterol levels. Common side effects include muscle pain, muscle weakness, muscle cramps, fatigue and tendon pain. In rare circumstances, statins may also cause rhabdomyolysis, which is a serious condition involving rapid muscle breakdown. Up to about one in three statin users experience one or more muscle-related side effects. Examples of statins include but are not limited to: atorvastatin (Lipitor), simvastatin (Zocor), rosuvastatin (Crestor), fluvastatin (Lescol) and pravastatin (Pravachol). Accordingly, the present disclosure provides a myostatin inhibitor for use in the treatment of hypercholesterolemia in a subject, wherein the treatment comprises administration of a myostatin inhibitor to the subject suffering from hypercholesterolemia, wherein the subject is on a statin therapy. In preferred embodiments, the myostatin inhibitor is a myostatin-selective inhibitor, such as SRK-439, Ab101-Ab141 (disclosed herein), apitegromab, GYM329 and trevogrumab. In particularly preferred embodiments, the myostatin inhibitor is SRK-439.

[0312]

[0269] Corticosteroids are a class of steroid hormones used to reduce inflammation and pain by suppressing the immune system. Use of corticosteroids, particularly high-dose and / or prolonged use, is reported to cause muscle weakness in some patients. Examples of corticosteroids include prednisone, triamcinolone, fluocinonide and dexamethasone.

[0313]

[0270] Fluoroquinolone antibiotics, which include, for example, ciprofloxacin, ofloxacin and levofloxacin, can in some cases cause muscle-related side effects.

[0314] Myostatin inhibitors

[0315]

[0271] Muscle provides more than motor function - in fact, muscle is critical for overall health. For example, muscle can increase basal metabolic rate (BMR); enhance glucose homeostasis and better insulin sensitivity and lower risk of prediabetes; signal to other systems as an endocrine organ impacting overall health; reduce visceral fat; increase caloric expenditure post-exercise; and increase bone density, strength, function and longevity and decrease risk of injury and disability. Historically, a wide variety of myostatin inhibitors (predominantly non-selective inhibitors of myostatin) have been evaluated in the clinic for a number of muscle disorders. Applicant previously further demonstrated in vivo effects of myostatin inhibition on metabolic conditions (see, for example, WO 2018 / 129395 and WO 2022 / 271867). In carrying out various embodiments of the present disclosure, myostatin inhibitors are used to treat metabolic disorders such as obesity (e g., being overweight, obese, extremely obese, as determined by BMI or other well-accepted methods / criteria), diabetes, fatty liver diseases (e g., NAFLD and NASH / MASH), kidney disease, and cardiovascular diseases.

[0316]

[0272] In some embodiments, non-selective inhibitors of myostatin are used. In some embodiments, myostatin inhibitors that do not inhibit GDF11 are selected, in order to circumvent detrimental effects of GDF11 inhibition. In some embodiments, myostatin inhibitors that do not inhibit Activin A are selected in order to circumvent detrimental effects of Activin A inhibition. In preferred embodiments, myostatin inhibitors that are selective to myostatin, such that they do not inhibit GDF11 or Activin A, are selected.

[0317]

[0273] Examples of non-selective myostatin inhibitors include a ligand trap (e.g., ACE-031 , ACE-083, and BIIB- 110 / ALG-801 ); an anti-ActRllb (e.g., bimagrumab); neutralizing antibody that binds mature myostatin (e.g., stamulumab (MYO-029), domagrozumab (PF-06252616), Landogrozumab (LY2495655), AMG-745 / PINTA-745 (Myostatin peptibody), RG6206 (an anti-myostatin adnectin, which is a single-strand fusion protein containing domains of fibronectin), and BMS-986089 (anti-myostatin adnectin also known as taldefgrobep alfa).

[0318]

[0274] To achieve favorable product profiles, preferably, the myostatin inhibitor is a myostatin-selective inhibitor, which is highly potent (e.g., IC50 of below 0.5 nM), highly stable, with favorable pharmacokinetic (PK) profile that allows excellent bioavailability. These include, for example, antibodies that bind mature myostatin (e.g., neutralizing antibodies) that do not bind GDF11 or Activin A.

[0319]

[0275] Non-limiting examples of antibodies and antigen-binding fragments that selectively bind and inhibit myostatin activation are provided below. These antibodies or antigen-binding fragments bind the antigen with high affinities (KD of less than 1 nM) and potently inhibit myostatin with an IC50 of below 1 nM as measured by in vitro assay in which protease-induced activation of myostatin is measured by the amount of mature myostatin released from latent complex.

[0320]

[0276] In preferred embodiments, such antibodies and antigen-binding fragments bind an epitope that includes a region within the prodomain of the pro / latent myostatin complex FVQILRLIKPMKDGTRYTGIRSLKLD (SEQ ID NO: ) (amino acid positions 147-172 of human pro-myostatin, as numbered according to SEQ ID NO: 52; provided below) and / or KALDENG (SEQ ID NO: ) (amino acid positions 205-211 of human pro-myostatin, as numbered according to SEQ ID NO: 52; provided below). In some embodiments, the antibody or anti gen-binding fragment thereof is selected from: SRK-439, Ab101 , Ab102, Ab103, Ab104, Ab105, Ab106, Ab107, Ab108, Ab109, Ab111 , Ab114, Ab115, Ab116, Ab117, Ab118, Ab121 , Ab122, Ab123, Ab124, Ab125, Ab127, Ab128, Ab129, Ab133, and Ab141. Thus, the invention encompasses the therapeutic use of an antibody that binds an epitope on pro-myostatin, wherein the epitope comprises one or more amino acid residues of the amino acid sequence FVQILRLIKPMKDGTRYTGIRSLKLD and / or the amino acid sequence KALDENG, where putative contact residues are shown in bold. In various embodiments, such antibodies are used in the treatment of a metabolic disorder disclosed herein. Patients who may particularly benefit from such therapy include those who are at risk of muscle atrophy or weakness, whether it is disease-associated, or drug-induced. Those who suffer from or at risk of developing anemia, chronic wounds (e.g. , difficult-to-heal wounds), and those with reproductive potential, also benefit from such therapy over non-selective myostatin inhibitor therapies.

[0321]

[0277] proGDF8 (human):

[0322] NENSEQKENVEKEGLCNACTWRQNTKSSRIEAIKIQILSKLRLETAPNISKDVIRQLLPKAPPLRELIDQYDVQRDDSSD GSLEDDDYHATTETIITMPTESDFLMQVDGKPKCCFFKFSSKIQYNKVVKAQLWIYLRPVETPTTVFVQILRLIKPMKDG TRYTGIRSLKLDMNPGTGIWQSIDVKTVLQNWLKQPESNLGIEIKALDENGHDLAVTFPGPGEDGLNPFLEVKVTDTP KRSRRDFGLDCDEHSTESRCCRYPLTVDFEAFGWDWIIAPKRYKANYCSGECEFVFLQKYPHTHLVHQANPRGSAG PCCTPTKMSPINMLYFNGKEQIIYGKIPAMVVDRCGCS (SEQ ID NO: 52).

[0323] Table 3. Exemplary antigen-binding fragments of myostatin-selective inhibitory antibodies

[0324]

[0278] In some embodiments, myostatin-selective antibody or antigen-binding fragment thereof is a MST1032 variant. A “MST1032 variant”, as used herein, refers to an antibody or antigen-binding fragment comprising any of the sequences disclosed in PCT / JP2015 / 006323, including but not limited to sequences of MS1032L001-SG1 , MS1032L006-SG1 , MS1032LO11-SG1 , MS1032LO18-SG1 , MS1032LO19-SG1 , MS1032LO21-SG1 , MS1032LO25- SG1 , and sequences provided in Table 2a, Table 11 a, Table 11 b, or Table 13 of PCT / JP2015 / 006323. In certain embodiments, a MST1032 variant comprises an antibody or antigen-binding fragment comprising a heavy chain variable domain comprising three CDR sequences of HCDR1 , HCDR2, and HCDR3, and a light chain variable domain comprising three CDR sequences of LCDR1 , LCDR2, and LCDR3, wherein the heavy chain CDRs comprise the amino acid sequences of: X1X2DIS (HCDR1 ; SEQ ID NO: 17); IISYAGSTYYASWAKG (HCDR2; SEQ ID NO: 18); GVPAYSX3GGDL (HCDR3; SEQ ID NO: 19), respectively; and the light chain CDRs comprise amino acid sequences of: X4X5SQSVYX6X7NWLS (LCDR1 ; SEQ ID NO: 20); WASTLAX8 (LCDR2; SEQ ID NO: 21 ); and AGGYGGGX9YA (LCDR3; SEQ ID NO: 22), respectively, wherein each of X1-X9 is any amino acid residue. In certain embodiments, X1 is S or H; X2 is Y, T, or D; X3 is T or H; X4 is Q or T; X5 is S or T; X6 is D or H; X7 is N or E; X8 is S or Y; X9 is L or R. In certain embodiments, a MST1032 variant comprises the six CDR sequences of SEQ ID NOs: 18, 66, 67, 68-70; SEQ ID NOs: 119, 18, 120, 68, 121 , 122; or SEQ ID Nos: 123, 18, 120, 124, 121 , 122. In certain embodiments, a MST1032 variant comprises a heavy chain variable domain comprising the amino acid sequence of any one of SEQ ID NOs: 125- 128. In certain embodiments, a MST1032 variant comprises a light chain variable domain comprising the amino acid sequence of any one of SEQ ID NOs: 129, 126, 127 and 130. In certain embodiments, a MST1032 variant comprises a set of six CDRs (e.g., from the same MST1032 variant antibody) or a paired VHA / L (e.g., from the same MST1032 variant antibody) from those listed in the Tables 4A-4F below. Table 4A.

[0325] Table 4B.

[0326] Table 4C.

[0327] Table 4D.

[0328] Table 4E.

[0329] Table 4F.

[0330] Light Chain Variable Domain (VL)

[0331]

[0279] In some embodiments, a myostatin antibody or antigen binding fragment thereof includes or is engineered to include a mutation or modification that causes an extended half-life of the antibody. In some embodiments, such mutations or modifications are within the Fc domain of the antibodies (e.g., Fc-modified antibodies), e.g., to promote circulating half-life or other PK properties. In some embodiments, the mutation is a YTE mutation (e.g., M252Y / S254T / T256E) (see, e.g., Saunders, KO (2019) “Conceptual Approaches to Modulating Antibody Effector Functions and Circulation Half-Life,” Frontiers in Immunology 10:1296 and U.S. Patent No. 7,083,784, each of which is herein incorporated by reference). In some embodiments, the mutation is an LS mutation (e.g., M428L / N434S) (Zalevsky et al., Nat Biotechnol. 2010, 28:157-159). In some embodiments, the mutation is an AAA mutation (e.g., T307A / E380A / N434A) (Petkova et al., Int Immunol. 2006, 18:1759-1769). In some embodiments, the mutation is a QL mutation (e.g., T250Q / M428L) (Hinton et al., J Biol Chem. 2004, 279:6213-6216). In some embodiments, the mutation comprises V308P (Datta-Mannan et al., Drug Metab Dispos. 2012, 40:1545-1555). Thus, in some embodiments, a variant of SRK-439, any one of Ab101-Ab141 , Trevogrumab, or GYM329, comprises one or more mutations in the Fc domain. In some embodiments, such mutation(s) include a YTE mutation, LS mutation, AAA mutation, QL mutation, and / or V308P.

[0332]

[0280] In some embodiments, the myostatin-selective inhibitor which may be selected to carry out one or more of the embodiments of the present disclosure is a monoclonal antibody referred to as GYM329 (also known as RO7204239 and RG6237), which is an anti-latent myostatin antibody with Fc modifications, currently being developed by Chugai / Roche / Genentech. In some embodiments, the myostatin-selective inhibitor is a variant of GYM329. In some embodiments, the myostatin-selective inhibitor is a derivative of GYM329, wherein optionally the derivative is a recombinant construct that comprises an antigen-binding fragments of GYM329.

[0333]

[0281] In some embodiments, the myostatin-selective inhibitor which may be selected to carry out one or more of the embodiments of the present disclosure is trevogrumab, which is also referred to as REGN1033. In some embodiments, the myostatin-selective inhibitor is a variant of trevogrumab. In some embodiments, the myostatin-selective inhibitor is a derivative of trevogrumab, wherein optionally the derivative is a recombinant construct that comprises an antigenbinding fragments of trevogrumab.

[0334]

[0282] In most preferred embodiments, the myostatin inhibitor is SRK-439. The full amino acid sequences of SRK-439 are as follows (the CDR sequences are shown underlined below; putative paratope residues are shown in bold below):

[0335]

[0283] Heavy chain amino acid sequence:

[0336] EVQLVESGGGLVQPGGSLRLSCAASGFTFSSYGMSWVRQAPGKGLELVASFTGSGGTYYPDSVKGRFTISRDNAKN SLYLQMNSLRAEDTAVYYCA RDLLIRFLEWSHYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCL VKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPP CPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRV VSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVE WESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG (SEQ ID NO: 503)

[0284] Light chain amino acid sequence:

[0337] DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSSGHNFLHWYLQKPGQSPQLLIYEVSNRVSGVPDRFSGSGSGTDFTL KISRVEAEDVGVYYCMQQTQYPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKV DNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0338] (SEQ ID NO: 504)

[0339] Variants and derivatives of SRK-439:

[0340]

[0285] Various embodiments of the present disclosure are carried out by the use of a variant of SRK-439. In some embodiments, a variant of SRK-439 comprises the VH and VL sequences of SRK-439 and one or more constant domains (e.g., CH1 , CH2, CH3, and / or CL) that differ from those of SRK-439. In some embodiments, a variant of SRK- 439 comprises the Fab fragment of SRK-439 and a Fc fragment that differs from that of SRK-439. In some embodiments, a variant of SRK-439 comprises the same six CDRs of SRK-439 in a different framework. In some embodiments, a variant of SRK-439 comprises one or more amino acid changes within one or more of the six CDRs but retains substantially similar antigen binding properties as SRK-439. In some embodiments, a variant of SRK-439 comprises one or more Fc modification(s).

[0341]

[0286] In engineering or selecting variants of SRK-439, residues involved in epitope-paratope interactions may be taken into consideration. In some embodiments, one or more mutations or substitutions are introduced on residue(s) that do not form direct contact with the epitope on proMyostatin. The potential paratope residues identified by cryo-EM are shown in bold above.

[0342]

[0287] In some embodiments, an antigen-binding fragment of SRK-439 is used to construct a derivative of SRK-439. Non-limiting examples of derivatives include multi-specific antibodies, such as bispecific antibodies, and any other engineered constructs incorporating the antigen-binding fragment of SRK-439 or a variant thereof. In some embodiments, such engineered constructs include fusion proteins and protein conjugates that comprise an antigenbinding fragment of SRK-439 or a variant of SRK-439.

[0343] Incretin-based therapies

[0344]

[0288] Incretins are a group of gut-derived metabolic hormones. The two most well-known examples of incretins are GLP-1 and GIP. The term “incretin mimetics” refers to synthetic or recombinant molecules that are a functional equivalent of naturally occurring incretins. Incretin mimetics are capable of activating the receptors for incretins. Nonlimiting examples of incretin mimetics include GLP-1 RAs (such as GLP-1 analogs) and GIP RAs (such as GIP analogs).

[0345]

[0289] GLP-1 , an incretin, is a potent stimulator of insulin secretion, an inhibitor of glucagon secretion, and a regulator of energy homeostasis. By increasing the synthesis and release of insulin, it decreases blood glucose. It also inhibits gastric mobility and plays a role in decreasing food intake and modulating the brain-based motivation / reward systems, e.g., to promote satiety (Smith et al. (2019) Neurochem Int 128:94.). GLP-1 is produced by the cleavage of proglucagon in pancreatic alpha cells and endocrine cells in the gastrointestinal tract. It has a short half-life, ranging from two minutes to eleven minutes, thus, longer lasting analogues have been developed which delay its metabolism and increase its circulating half-life. These GLP-1 analogues can act as receptor agonists and improve the clinical efficacy of GLP-1 activity.

[0346]

[0290] GLP-1 receptor agonists are widely in use for the treatment of weight loss, obesity and / or diabetes. Dulaglutide (Trulicity®), exenatide (Byetta®), exenatide extended release (Bydureon®), liraglutide (Saxenda® / Victoza®), lixisenatide (Adlyxin®), semaglutide (Ozempic® / Rybelsus®), and tirzepatide (Mounjaro® / Zepbound®)) have been approved by the United States Food and Drug Administration. However, both tolerability and durability are a concern. They require a prolonged, e.g., six-month, dose escalation in an attempt to improve tolerability and minimize side effects. Even so, gastrointestinal side effects such as nausea, vomiting, and diarrhea are common. In the SURPASS-2 trial (NCT03987919), 40-45% of the subjects on semaglutide or tirzepatide reported mild to moderate gastrointestinal adverse events. Weiss et al (2022) BMJ Open Diabetes Res Care 10:e002517 reported a 45% discontinuation rate at 12 months and a 65% discontinuation rate at 24 months. Discontinuation leads to body weight regain and loss of cardiometabolic benefits, as evidenced by an increase in systolic blood pressure and an increase in HbA1 c levels (STEP1 Trial, NCT03548935).

[0347]

[0291] Furthermore, the FDA currently requires that GLP-1 receptor agonists carry a black box warning on the label regarding the risk of C-cell tumors. It recommends against using GLP-1 receptor agonists in patients with a personal or family history of medullary thyroid cell cancer or multiple endocrine neoplasia type 2a or 2b.

[0348]

[0292] The well-known challenges that occur with weight loss treatment may also occur with the use of GLP-1 receptor agonist treatment. A substantial portion of weight loss is muscle rather than fat; as much as 20-30% of weight loss is estimated to be muscle and this percentage may be higher in individuals with rapid and significant weight loss (Cava et al. (2017) Adv Nutr 8:511 ). Also, weight loss is difficult to sustain because the basal metabolic rate declines to compensate for the body’s calorie deficit. With this lower basal metabolic rate, the subject may need to maintain a reduced calorie intake and increased physical activity level to stay calorie budget neutral, but these lifestyle changes are not always sustainable. Moreover, significant rebound in weight has been reported in patients who discontinue GLP-1 receptor agonist treatment (see, e.g., Wlding et al. Diabetes Obes Metab. 2022 Aug;24(8):1553-1564, reporting that participants regained two-thirds of their prior weight loss one year after discontinuing semaglutide treatment). Similarly, patients who discontinued tirzepatide treatment regained 70% of their lost weight one year after stopping tirzepatide treatment (see, e.g., Aronne et al, JAMA. Published online December 11 , 2023. doi: 10.1001 / jama.2023.24945). These observations highlight unresolved challenges associated with incretin therapies.

[0349]

[0293] The well-known challenges that occur with treatment of type 2 diabetes also may still occur with the use of GLP- 1 receptor agonists. As a progressive disease, type 2 diabetes frequently requires multiple pharmacologic therapies.

[0350]

[0294] GLP-1 receptor agonist therapies have been shown to be associated with unintended loss of lean mass and certain adverse events. For example, a sizable fraction of patients choose to discontinue the GLP-1 receptor agonist therapy due to side effects such as nausea. In addition, upon discontinuation, patients quickly regain most of the weight they lost during GLP-1 receptor agonists therapy. Moreover, prolonged use of GLP-1 receptor agonists therapy may be associated with increased instances of certain cancers, such as thyroid cancer and pancreatic cancer.

[0351]

[0295] To address these shortcomings, the present disclosure encompasses improved therapies comprising a GLP-1 receptor agonist and a myostatin inhibitor (e.g., myostatin-selective inhibitors) or alternative combinations with myostatin inhibitors such as biguanides (e.g., metformin) in lieu of a GLP-1 receptor agonist (as discussed in further detail in the next section below) as novel approaches to achieve synergistic effects for improving metabolic health. First, rather than focusing primarily on body weight (weight loss), the inventors sought to improve overall body composition through fat loss and maintained lean mass at the same time. Second, incorporation of myostatin inhibition into a therapeutic regimen may reduce or slow the observed rebound effects associated with GLP-1 receptor agonists discontinuation. And third, this approach may allow reduced doses for the GLP-1 receptor agonists to achieve efficacy, which as monotherapy would be suboptimal or sub-efficacious, but may still provide efficacy in conjunction with a myostatin inhibitor. Thus, treatments comprising a GLP-1 receptor agonist and a myostatin inhibitor (e.g., a myostatin- selective inhibitor, e.g., an antibody disclosed herein) may reduce the risk, frequency, and / or degree of severity of certain side effects associated with the GLP-1 receptor agonists. In some embodiments, a combination of an antibody or antigen-binding fragment thereof, as disclosed herein, lessens the need for multiple anti-diabetic therapies and / or augment the effect of GLP-1 in weight loss treatment by maintaining lean mass.

[0352]

[0296] Accordingly, the present disclosure provides a myostatin inhibitor for use in conjunction with a GLP-1 receptor agonist in the treatment of obesity or for use in improving body composition in a subject. In some embodiments, the present disclosure provides a myostatin inhibitor and a GLP-1 receptor agonist for use in the treatment of obesity or for use in improving body composition in a subject, wherein the treatment comprises administration of a myostatin inhibitor in conjunction with a GLP-1 receptor agonist to the subject, optionally wherein the myostatin inhibitor is a selective myostatin inhibitor, e.g., an antibody disclosed herein or an antigen-binding fragment thereof (e.g. , Ab109, Ab133, or Ab141 ). The disclosure also includes a myostatin inhibitor for use in the treatment of obesity or for use in improving body composition in a subject, wherein the treatment comprises administration of a myostatin inhibitor to the subject, wherein the subject is treated has been treated with a GLP-1 receptor agonist, optionally wherein the myostatin inhibitor is a selective myostatin inhibitor, e.g., an antibody disclosed herein or an antigen-binding fragment thereof. The disclosure further provides a GLP-1 receptor agonist for use in the treatment of obesity or for use in improving body composition in a subject, wherein the treatment comprises administration of a GLP-1 receptor agonist to the subject, wherein the subject is treated or has been treated with a myostatin inhibitor, optionally wherein the myostatin inhibitor is a selective myostatin inhibitor, e.g., an antibody disclosed herein or an antigen-binding fragment thereof. In any of these embodiments, the myostatin inhibitor is a non-selective inhibitor of myostatin. In any of these embodiments, the GLP-1 receptor agonist is a semaglutide, liraglutide, tirzepatide, or retatrutide. In any of these embodiments, the myostatin inhibitor may be any one of Ab101-Ab141 , such as Ab109, Ab133, or Ab141 or a variant thereof, wherein, preferably, the myostatin inhibitor is Ab109 or a variant thereof. In particularly preferred embodiments, the antibody is SRK-439.

[0353]

[0297] In some embodiments, administration of the myostatin-selective inhibitor (e.g., Ab109) after discontinuing the GLP-1 receptor agonist maintains fat loss and / or prevents loss of lean mass in the subject. In some embodiments, administration of the myostatin-selective inhibitor (e.g., Ab109) reduces the rate or degree of fat regain after discontinuing the GLP-1 receptor agonist.

[0354]

[0298] In various embodiments, incretins or analogs are employed in conjunction with myostatin inhibitors (preferably myostatin-selective inhibitors) in accordance with the present disclosure. In some embodiments, an incretin-based therapy, such as a GLP-1 and / or GIP agonist, is used to treat metabolic disorders in combination with a myostatin inhibitor. In some embodiments, the incretin-based therapy is GLP-1 or a GLP-1 analog, a variant, or a mimetic thereof. In some embodiments, the incretin mimetic comprises an amino acid sequence represented by the formula wherein X is any amino acid, wherein optionally X is E or Q. In some embodiments, the incretin mimetic comprises an amino acid sequence represented by the formula EGTFXSD, wherein X is any amino acid, wherein optionally X is T or I (SEQ ID NO: ). In some embodiments, the incretin mimetic comprises an amino acid sequence represented by the formula FX1X2WL (SEQ ID NO: ), wherein each of Xi and X2 is any amino acid, wherein optionally, Xi is I or V.

[0355]

[0299] In some embodiments, the incretin mimetic is a GLP-1 analog that comprises an amino acid sequence represented by the formula HX1X2GX3FTX4D (SEQ ID NO: ), wherein each of Xi, X2, X3 and X4 is any amino acid. In some embodiments, Xi is G. In some embodiments, X2 is E or Q. In some embodiments, X3 is T. In some embodiments, X4 is S. In some embodiments, the GLP-1 analog comprises HGEGTFTSD (SEQ ID NO: ).

[0356]

[0300] In various embodiments, one or more of the following incretins or analogs are used in conjunction with the myostatin inhibitors to carry out various embodiments of the invention as described herein:

[0357] HAEGTFTSDVSSYLEGQAAKEFIAWLVKG (SEQ ID NO: )

[0358] HAEGTFTSDVSSYLEGQAAKEFIAWLVKGRG (SEQ ID NO: )

[0359] HGEGTFTSDVSSYLEGQAAKEFIAWLVKG (SEQ ID NO: )

[0360] HGEGTFTSDVSSYLEGQAAKEFIAWLVKGRG (SEQ ID NO: )

[0361] HGEGTFTSDVSSYLEEQAAKEFIAWLVKGG (SEQ ID NO: ) HCEGTFTSDVSSYLEGQAAKEFIAWLVKGRG (SEQ ID NO: )

[0362] HGEGTFTSDLSKQMEEECVRLFIEWLKNGGPSSGAPPPGCG (SEQ ID NO: )

[0363] HSQGTFTSDYSKYLDEQAAKEFIAWLMNT (SEQ ID NO: )

[0364] HGQGTFTSDKSKYLDERAAQDFVQWLLDGGPSSGAPPPS (SEQ ID NO: )

[0365] HGQGTFTSDLSKQKDEQRAKLFIEWLGAGGPPSGKPPPK (SEQ ID NO: )

[0366] HGQGTFTSDYSKYLDKRRAQDFVQWLLNTGGPSSGAPPPS (SEQ ID NO: )

[0367] HSQGTFTSDVSEYLDSERARDFVAWLEAGG (SEQ ID NO: )

[0368] YAEGTFISDYSIAMDKIHQQDFVNWLLAQKGKKNDWKHNITQ (SEQ ID NO: )

[0369] YGEGTFISDYSIAMDKIHQQDFVNWLLAQKGKKNDWKHNITQ (SEQ ID NO: )

[0370] HGEGTFTSDVSSYLEGQAAKEFIAWLVKGRG (SEQ ID NO: )

[0371] HGEGTFTSDVSSYLEEQAAKEFIAWLVKGG (SEQ ID NO: )

[0372] HGEGTFTSDVSSYLEEQAAKEFIAWLVKGGG (SEQ ID NO: )

[0373] HGEGTFTSDLSKQMEEECVRLFIEWLKNGGPSSGAPPPGCG (SEQ ID NO: )

[0374] HGEGTFTSDVSSYLEEQAAKEFIANLSKGG (SEQ ID NO: )

[0375] HGEGTFTSDVSSYLEEQNASEFIAWLVKGG (SEQ ID NO: )

[0376] HGQGTFTSDKSKYLDERAAQDFVQWLLDGGPSSGAPPPS (SEQ ID NO: )

[0377] HGEGTFTSDVSSYLEEECVRLFIEWLKNGGPSSGAPPPGCG (SEQ ID NO: )

[0378] HGEGTFTSDVSSYLEEQAAKEFIAWLVKGGHGEGTFTSDVSSYLEEQAAKEFIAWLVKGG (SEQ ID NO: )

[0379] HSQGTFTSDYSKYLDEQAAKEFIAWLMNT (SEQ ID NO: )

[0380] HGQGTFTSDLSKQKDEQRAKLFIEWLGAGGPPSGKPPPK (SEQ ID NO: )

[0381] HGQGTFTSDYSKYLDKRRAQDFVQWLLNTGGPSSGAPPPS (SEQ ID NO: )

[0382] HSQGTFTSDVSEYLDSERARDFVAWLEAGG (SEQ ID NO: )

[0383] HGEGTFTSDLSKQMEEEAVRCFIEWLKNGGPSSGAPPCGG (SEQ ID NO: )

[0384] HGEGTFTSDLSKQMEEEAVRCFIEWLKNGGPSSGAPPC (SEQ ID NO: )

[0385] YGEGTFISDYSIAMDKIHQQDFVNWLLAQKGKKNDWKHNITQ (SEQ ID NO: )

[0386] HGEGTFTSDVSSYLEEQAAKEFIAWLVKGGGGGSGGGGSGGGGHGEGTFTSDVSSYLEEQAAKEFIAWLVKGG (SEQ ID NO: )

[0387] HGEGTFTSDVSSYLEEQAAKEFIAWLVKGGGGAAKEFIAWLVKGG (SEQ ID NO: )

[0388] HGEGTFTSDVSSYLEEQAAKEFIAWLVKGGGGKEFIAWLVKGG (SEQ ID NO: )

[0389] HGEGTFTSDVSSYLEEQAAKEFIAWLVKGGGGAAKEFIAWLGG (SEQ ID NO: )

[0390] HGEGTFTSDYSIYLDKIAQKAFVQWLAIGGPSSGAPPPS (SEQ ID NO: )

[0391] HGEGTFTSDVSSYLEEQAAKECIAWLVKGG (SEQ ID NO: )

[0392] HGEGTFTSDVSSYLEEQAAKCFIAWLVKGG (SEQ ID NO: ) HGEGTFTSDVSSYLEEQCAKEFIAWLVKGG (SEQ ID NO: )

[0393] HGEGTFTSDVSSYLEECAAKEFIAWLVKGG (SEQ ID NO: )

[0394] HGEGTFTSDVSSYLEELAAKRFILWLVLRREGFSPERLAALESRLQALERRL (SEQ ID NO: )

[0395] HGEGTFTSDVSSYLEELAACRFILWLVLRREGFSPERLAALESRCQALERRL (SEQ ID NO: )

[0396] HGEGTFTSDVSSYLEELAACRFILWLVLRSGGFSPERLAALESRCQALERRL (SEQ ID NO: )

[0397] HGEGTFTSDVSSYLEGQAAKEFIAWLVKGRGG (SEQ ID NO: )

[0398] YGEGTFISDYSIAMDKIHQQDFVNWLLAQKGKKNDWKHNITQG (SEQ ID NO: )

[0399] EGTFISDYSIAMDKIHQQDFVNWLLAQK (SEQ ID NO: )

[0400] HGEGTFTSDLSKQMEEEAVRCFIEWLKNGGPSSGAPPGCGG (SEQ ID NO: )

[0401] HGEGTFTSDLSKQMEEEAVRCFIEWLKNGGPSSGAPPCGGG (SEQ ID NO: )

[0402] YGEGTFTSDYSIYLDKIAQKAFVQWLAIGGPSSGAPPPS (SEQ ID NO: )

[0403] YGEGTFTSDYSIQMDKIAQRLFVEWLKAGGPSSGAPPPGG (SEQ ID NO: )

[0404] HGEGTFTSDYSIYLDKICQKAFVQWLAIGGPSSGAPPPGCG (SEQ ID NO: )

[0405] HGEGTFTSDYSIYLDKIAQKCFVQWLAIGGPSSGAPPCS (SEQ ID NO: )

[0406] YGEGTFTSDYSIQMDKIAQRCFVEWLKAGGPSSGAPPCGG (SEQ ID NO: )

[0407] YGEGTFTSDYSIQMDKIAQRCFVEWLKNGGPSSGAPPCGG (SEQ ID NO: )

[0408] YGEGTFTSDYSIQMDKIAVRCFVEWLKNGGPSSGAPPCGG (SEQ ID NO: )

[0409] HGEGTFTSDYSIQMDKIAQRCFVEWLKNGGPSSGAPPCGG (SEQ ID NO: )

[0410] YGEGTFTSDYSIQMDEEAQRCFVEWLKNGGPSSGAPPCGG (SEQ ID NO: )

[0411] YGEGTFTSDYSIQMEEEAQRCFVEWLKNGGPSSGAPPCGG (SEQ ID NO: )

[0412] YGEGTFTSDYSIQMEEEAVRCFIEWLKAGGPSSGAPPCGG (SEQ ID NO: )

[0413] HGEGTFTSCLSKQMEEEAVRLFIEWLKNGGPSSGAPPPSGGGGSGGGGSGC (SEQ ID NO: )

[0414] HGEGTFTSCLSKQMEEEAVRLFIEWLKNGGPSSGAPPPC (SEQ ID NO: )

[0415] HGEGTFTSDVSSYLEELAAKRFILWLVLRSGGFSPERLAALESRLQALERRL (SEQ ID NO: )

[0416] HGEGTFTSDVSSYLEELAAKCFILWLVLRREGFSPERLAALESRCQALERRL (SEQ ID NO: )

[0417] HGEGTFTSDVSSYLEECAAKRFILWLVLRREGFSPERLAALESRLQALCRRL (SEQ ID NO: )

[0418]

[0301] In some embodiments, the present disclosure provides a method of improving liver health (e.g., reducing liver fat) in a subject, e.g., in an obese subject and / or a subject with fatty liver disease, comprising administering to the subject a myostatin-selective inhibitor in an amount effective to improve liver health. In some embodiments, administering the myostatin-selective inhibitor reduces liver weight by at least 10% (e.g., 10%, 20%, 25%, or more). In some embodiments, the subject is receiving or has received metformin. In some embodiments, the myostatin-selective inhibitor is any one of the antibodies or antigen-binding fragments disclosed herein (e.g., Ab109). In some embodiments, a myostatin-selective inhibitor is used in combination with a GLP-1 receptor agonist in the treatment of fatty liver in a subject, wherein the treatment comprises administration of a myostatin-selective inhibitor and a GLP-1 receptor agonist for at least 8 weeks to treat the fatty liver, wherein optionally the myostatin-selective inhibitor is Ab109, Ab133 or Ab141 , and wherein further optionally the GLP-1 receptor agonist is selected from: albiglutide, taspoglutide, semaglutide, exenatide, BPI-3016, GW002, glutazumab, exendin-4, exenatide, GLP-1 (7-36)NH2, everestmab, liraglutide, lixisenatide, tirzepatide, dulaglutide, danuglipron (Pfizer), PF-07081532, or orforglipron. In some embodiments, other therapies comprising a GLP-1 receptor agonist include, but are not limited to, GLP-1 receptor agonist / GIP receptor antagonist combination such as AMG 133 (Amgen); GLP-1 / GIP dual agonists such as tirzepatide (LY3298176; Eli Lilly), and CT-388; amylin / GLP-1 combination such as cagrilintide / semaglutide combination (Novo Nordisk); GLP-1 / glucagon combination such as DD01 (Neuraly); GLP-1 / glucagon receptor (GCG) agonist combinations such as ALT-801 (Altimmune), GLP-1 / GIP such as CT-388 (Carmot); GLP-1 / glucagon dual agonist such as I BI362 (LY-330567 (Innovent / Eli Lilly), cotadutide, DD01 , danuglipron (PF-06882961 ) (Pfizer), mazdutide (IBI362; LY- 330567), MEDI0382; noiiglutide, oxyntomodulin, pemvidutide, setmelanotide (Rhythm), survodutide, and GLP- 1 / GIP / Glucagon triple receptor agonist such as retatrutide and LY343794.

[0419] Assessment of body composition

[0420]

[0302] Clinical effects of the therapies described herein may be measured by any suitable methods or criteria in order to assess benefits on pharmacologic management of excess adiposity and accompanying metabolic disturbances (Heymsfield et al. J Biol Chem. 2020 Apr 17;295(16):5404-5418.).

[0421]

[0303] In some embodiments, various parameters of body composition can be measured, such as changes in total body fat mass, lean mass, waist circumference, HbA1 c levels, and body weight. Any suitable techniques can be employed to assess body composition, including, without limitation, qNMR, dual energy x-ray absorptiometry (DEXA), magnetic resonance imaging (MRI)-derived hepatic fat fraction, hydrodensitometry, air displacement plethysmography (ADP), bioelectrical impedance analysis (BIA), bioimpedance spectroscopy (BIS), electrical impedance myography (EIM), 3D body scanners, and multi-compartment models (e g., 3-compartment and 4-compartment models).

[0422]

[0304] In some embodiments, the combination and adjunct therapies described herein results in a change in total body fat mass in a subject as compared to baseline. In some embodiments, total body fat mass is measured by dual energy x-ray absorptiometry (DEXA) (Garito et al. Diabetes Obes Metab. 2018;20(1 ):94-102). In some embodiments, the combination and adjunct therapies described herein results in a change in diabetes status in a subject as compared to baseline. In some embodiments, diabetes status can be determined by measuring the subject’s HbA1 C level, homeostatic model assessment, quantitative insulin sensitivity check, Matsuda Index (Yokoyama et al. J Clin Endocrinol Metab. 2004;89(3):1481-1484; Hrebicek et al. J Clin Endocrinol Metab. 2002;87(1 ): 144-147; Matsuda et al. Diabetes Care. 1999;22(9): 1462-1470). In some embodiments, the combination and adjunct therapies described herein results in a change in a subject’s body weight, BMI, waist circumference, and / or waist-to-hip ratio. In some embodiments, the combination and adjunct therapies described herein results in a change in a subject’s body composition. In some embodiments, the subject’s body composition is assessed by DEXA-measurements of bone mineral-free lean mass, magnetic resonance imaging (MRI)-derived hepatic fat fraction (Mashood et al. J Magn Reson Imaging. 2013;37(6):1359-1370), and / or subcutaneous and abdominal visceral adipose tissue (Fallah et al. MAGMA. 2017;30 (2): 139-151 ). In some embodiments, the combination and adjunct therapies described herein results in a change in a subject’s metabolic status (Garito et al. Diabetes Obes Metab. 2018;20(1 ):94-102; Goyal et al. N Am J Med Sci. 2012;4(4): 180-184; Rossi et al. Obesity (Silver Spring). 2011 ; 19(9): 1747-1754). In some embodiments, the subject’s metabolic status is assessed using metabolic biomarkers, non-limiting examples of such markers are discussed in, e.g., Robberecht et al., Metab Syndr Relat Disord. 2016 Mar; 14(2):47-93, and Belhayara et al., Nutrients. 2020 Mar; 12(3): 727, each of which is herein incorporated by reference in its entirety. In some embodiments, the subject’s metabolic status is assessed based on the subject’s cardiovascular risk factors, including serum lipid levels, high-sensitivity C-reactive protein level, interleukin 6 level, leptin level, adiponectin level, and blood pressure. In some embodiments, the combination and adjunct therapies described herein results in a change in a subject’s physical performance. In some embodiments, the subject’s physical performance is assessed by measuring hand grip strength by dynamometry (Rooks et al. J Am Geriatr Soc. 2017;65(9):1988-1995). Biological effects of myostatin inhibitors

[0423]

[0305] Methods or treatments comprising administering a myostatin inhibitor are encompassed by the present disclosure. In some embodiments, the myostatin inhibitor is a myostatin-selective inhibitor, e.g., an anti-myostatin antibody or antigen-binding fragment thereof disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141. In some embodiments, the myostatin inhibitor is administered in an effective amount to effectuate one or more beneficial effects (e.g., therapeutic effects) in a subject. Exemplary biologically beneficial effects are provided herein. In some embodiments, beneficial biological effects in a subject can be achieved by administration of a myostatin inhibitor, e.g., an antibody or antigen-binding fragments thereof that specifically binds to pro / latent myostatin as described herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141. In some embodiments, beneficial biological effects in a subject can be achieved by administration of myostatin inhibitor alone. In other embodiments, the myostatin inhibitor is used in conjunction with at least one additional therapeutic agent or regimen to achieve one or more of the beneficial biological effects discussed herein, wherein, optionally, the at least one additional therapeutic agent or regimen includes one or more approved agents for treating a metabolic disorder such as diabetes, obesity, or a liver disease, wherein, optionally, the one or more approved agents include, e.g., a diet and / or exercise regimen, a GLP-1 pathway activator (e.g., dulaglutide, exenatide, semaglutide, liraglutide, lixisenatide, PF-07081532, or orforglipron), a biguanide (e.g., metformin), a sulfonylurea (e.g., glipizide, glimepiride, or glyburide), a bile acid-metformin, sequestrant, a dopamine-2 agonist (e.g., bromocriptine), a dipeptidyl peptidase 4 inhibitor (e.g., alogliptin, linagliptin, linagliptin-empagliflozin, linagliptin saxagliptin, or sitagliptin), a meglitinide (e.g., nateglinide, repaglinide, or repaglinide-metformin), a sodiumglucose cotransporter-2 (SGLT2) inhibitor (canagliflozin, canagliflozin-metformin, dapagliflozin, dapagliflozin-metformin or empagliflozin), a thiazolidinediones (e.g., tosiglitazone or pioglutazone), a weight-loss drug such as phentermine, benzphetamine, diethylpropion, or phendimetrazine, or a thyroid hormone receptor-beta (THR-beta) selective agonist, e.g., an aryloxyphenyl based thyromimetic such as resmetirom or eprotirome, or a diphenylmethane based thyromimetic such as sobetirome, Sob-AM2, VK2809 (MB08711 ), MB07344, IS25, and TG68. In some embodiments, the myostatin inhibitor (e.g., an antibody or antigen-binding fragments thereof that specifically binds to pro / latent myostatin as described herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) is used in conjunction with a GLP-1 pathway activator to achieve one or more of the biological effects discussed herein. In some embodiments, the myostatin inhibitor (e.g., an antibody or antigen-binding fragments thereof that specifically binds to pro / latent myostatin as described herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) is used in conjunction with a GLP-1 pathway activator and a diet and / or exercise regimen to achieve one or more of the biological effects discussed herein.

[0424]

[0306] In some embodiments, the myostatin inhibitor (e.g., a selective myostatin inhibitor, e.g., an anti-myostatin antibody or antigen-binding fragment thereof disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) is administered in an amount effective to cause two or more of the biological effects described below. In some embodiments, the myostatin inhibitor is administered in an amount effective to cause three or more of the biological effects described below. In some embodiments, the myostatin inhibitor is administered in an amount effective to cause four or more of the biological effects described below. In some embodiments, the myostatin inhibitor is administered in an amount effective to cause five or more of the biological effects described below. In some embodiments, the myostatin inhibitor is administered in an amount effective to cause six or more of the biological effects described below. In some embodiments, the myostatin inhibitor is administered in an amount effective to cause seven or more of the biological effects described below. In some embodiments, the myostatin inhibitor is administered in an amount effective to cause eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, or sixteen of the biological effects described below. In some embodiments, the myostatin-selective inhibitor is an antibody or antigen-binding fragment thereof comprising all six CDRs of any one of Ab101-Ab141. In some embodiments, the myostatin-selective inhibitor is an antibody or antigenbinding fragment thereof comprising the heavy and light chain variable domains of any one of Ab101-Ab141 , e.g., the pair of SEQ ID NOs identified for a specific antibody in Table 3. In some embodiments, the myostatin-selective inhibitor is an antibody comprising the heavy and light chains of any one of Ab101-Ab141. In some embodiments, the myostatin- selective inhibitor antibody or antigen-binding fragment is selected from Ab101 , Ab102, Ab103, Ab104, Ab105, Ab106, Ab107, Ab108, Ab109, Ab110, Ab111 , Ab112, Ab113, Ab114, Ab115, Ab116, Ab117, Ab118, Ab119, Ab120, Ab121 , Ab122, Ab123, Ab124, Ab125, Ab126, Ab127, Ab128, Ab129, Ab130, Ab131 , Ab132, Ab133, Ab134, Ab135, Ab136, Ab137, Ab138, Ab139, Ab140 and Ab141. In preferred embodiments, the antibody or antigen-binding fragment thereof is Ab109, Ab133 or Ab141. In most preferred embodiments, the antibody or antigen-binding fragment thereof is SRK- 439.

[0425]

[0307] In some embodiments, the myostatin inhibitor is administered in conjunction with at least one additional therapeutic agent (e.g., an approved therapeutic agent for treating a metabolic disorder, e.g., diabetes and / or obesity), wherein the administration is sufficient to cause two or more of the biological effects described herein.

[0426] Effect on mass and / or function of muscle tissue in the human subject:

[0427]

[0308] In some embodiments, administration of a myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) increases mass and / or function of a muscle tissue in the human subject. In some embodiments, the muscle tissue is selected from the group consisting of a smooth muscle tissue, a skeletal muscle tissue and a cardiac muscle tissue. Smooth muscle tissue is made up from long tapering cells, generally involuntary and differs from striated muscle in the much higher actin / myosin ratio, the absence of conspicuous sarcomeres and the ability to contract to a much smaller fraction of its resting length. Smooth muscle cells are found particularly in blood vessel walls, surrounding the intestine and in the uterus. Cardiac muscle tissue is a striated but involuntary tissue responsible for the pumping activity of the vertebrate heart. The individual cardiac muscle cells are not fused together into multinucleate structures as they are in striated muscle tissue. Skeletal muscle tissue is under voluntary control. The muscle fibers are syncytial and contain myofibrils, tandem arrays of sarcomeres. There are two general types of skeletal muscle fibers: slow-twitch (type I) and fast-twitch (type II) according to the expression of their particular myosin heavy chain (MHC) isoform. Slow- twitch muscles are better equipped to work aerobically and help enable long-endurance feats such as distance running, while fast-twitch muscles fatigue faster but are better equipped to work anaerobically and are used in powerful bursts of movements like sprinting. The differentiation between slow and fast twitch muscle fibers is based on histochemical staining for myosin adenosine-triphosphatase (ATPase) and the type of myosin heavy chain. The slow twitch muscle fiber (type I fiber) is MHC isoform I and the three fast twitch isoforms (type II fibers) are MHC isoform Ila, MHC isoform lid, and MHC isoform lib (S. Schiaffino, J. Muscle Res. Cell. Motil., 10 (1989), pp. 197-205).

[0428]

[0309] In some embodiments, the mass and / or function of a fast twitch muscle tissue in the human subject is increased. In other embodiments, the mass and / or function of a slow twitch muscle tissue in the human subject is increased. In some embodiments, maintaining muscle mass in the context of treating obesity provides beneficial effects for weight loss, e.g., as observed by resistance training. In some embodiments, administration of a myostatin inhibitor according to the present disclosure (e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) preferentially preserves fast-twitch fibers.

[0429]

[0310] In some embodiments, the biological effects of administering an effective amount of the pharmaceutical compositions and therapeutic methods provided herein is associated with a phenotypic change of muscle fiber types, which is a process referred to as fiber type switch. In some embodiments, fiber type switch is triggered by an event, such as an injury and starvation.

[0430]

[0311] In one embodiment, the disclosure provides a method for promoting fiber type switch in a subject. The method comprises administering to the subject a composition comprising an antibody or antigen-binding fragment thereof (e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) that specifically binds pro / latent-myostatin and blocks release of mature myostatin in an amount effective to promote fiber type switch, thereby promoting fiber type switch in the subject.

[0312] In another embodiment, the disclosure provides a method for preferentially increasing type II or fast twitch fibers over type I or slow twitch fibers in a subject. The method comprises administering to the subject a composition comprising a myostatin inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein (e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ), in an amount effective to preferentially increase type II or fast twitch fibers over type I or slow twitch fibers fiber type switch, thereby preferentially increasing type II or fast twitch fibers over type I or slow twitch fibers in the subject.

[0431]

[0313] In some embodiments, administration of an effective amount of a myostatin inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, or Ab133, can cause an increase in muscle mass and / or muscle function. Preferably, such an increase in muscle mass is clinically meaningful to benefit or otherwise improve the health status of the subject. For example, clinically meaningful changes in muscle mass may improve the patient’s mobility, self-care, metabolism, etc. In some embodiments, the increase in muscle mass is an increase in lean muscle or lean muscles. In some embodiments, such increase in muscle mass is a systemic effect such that muscles in the whole body or substantially whole body show the measurable effect. In some embodiments, lean muscle is muscle that is densely packed with contractile tissue and has low fat and connective tissue content. In other embodiments, effects are localized to certain group / type of muscles. In some embodiments, the mass of the muscle tissue, e.g., lean muscle tissue, is increased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the mass of the muscle tissue, e.g., lean muscle tissue, is increased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%. Such increase in muscle mass may be deduced or measured by any suitable known methods, including measurement of cross-sectional area via MRI (e.g., forearm cross section), circumference, diaphragm width (e.g., via ultrasound), qNMR, DEXA, etc.

[0432]

[0314] In some embodiments, administration of an effective amount of a myostatin inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., an antibody selected from Ab101 , Ab102, Ab103, Ab104, Ab105, Ab106, Ab107, Ab108, Ab109, Ab110, Ab111 , Ab112, Ab113, Ab114, Ab115, Ab116, Ab117, Ab118, Ab119, Ab120, Ab121 , Ab122, Ab123, Ab124, Ab125, Ab126, Ab127, Ab128, Ab129, Ab130, Ab131 , Ab132, Ab133, Ab134, Ab135, Ab136, Ab137, Ab138, Ab139, Ab140 and Ab141 or an antigen binding fragment thereof, preferably Ab102, Ab109, Ab130, Ab132, or Ab133 or an antigen binding fragment thereof, can cause an enhancement in muscle function. Muscle function may be assessed by a variety of measures, including, without limitation: force generation, grip strength (e.g., maximum grip strength), endurance, muscle oxidative capacity, dynamic grip endurance, etc. In some embodiments, biomarkers indicative of myostatin inhibition includes a gene or protein expression marker described in PCT / US2018 / 012686, the contents of which are hereby incorporated by reference. In some embodiments, the biomarkers comprise one or more downregulated marker of adipogenesis, adipocytes, or adipokines. The biomarker change observed after administration may indicate an improvement in cardiovascular health, e.g., an increase in Actcl. In some embodiments, the biomarker change indicates a change in fatty acid synthesis, e.g., a reduction in Acc1, Pdk4, and / or an increase in PDP1. In some embodiments, serum creatinine levels are used as a biomarker indicative of muscle mass, e.g., an increase in serum creatinine following administration.

[0433]

[0315] In some embodiments, the biomarker change observed after administration of a myostatin inhibitor indicates a reduction in inflammation. In some embodiments, administration results in a reduction in one or more catabolic markers and / or an increase in one or more anabolic markers, e.g., when measuring protein synthesis, and the opposite when measuring fat synthesis. In some embodiments, administration results in reduced intramuscular fat.

[0434]

[0316] In various embodiments, one or more biomarker change affected by administering an antibody disclosed herein comprises one or more of: (1) a decrease in the expression of pyruvate dehydrogenase kinase 4 (PDK4) and an increase in the expression of pyruvate dehydrogenase phosphatase 1 (PDP1 ); (2) a decrease in the expression of one or more of acetyl-CoA carboxylase (ACC1 ), adipogenin (ADIG), CCAAT / enhancer-binding protein delta (CEBPD), fatty acid binding protein 5 (FABP5), fatty acid synthase (FASN), lipase E (LIPE), perilipin 1 (PLIN1 ), perilipin 4 (PLIN4), angiotensinogen (AGT), angiopoietin-like 14 (ANGPTL14), apolipoprotein C1 (APOC1 ), adiponectin (ADIPOQ), Leptin (LEP), Resistin (RES), and Haptoglobin (HP); (3) an increase in the expression of Sharpl ; (4) a decrease in the expression of MYOG, MYL2, MYL4, and / or TNNC1 ; (5) an increase in expression in actin alpha cardiac muscle 1 (ACTC1 );and / or, (6) a decrease in the expression of PGC1A, NOR1 , UCP1 , and / or NUR77. In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in the expression level of one or more regulators of adipogenesis, adipocyte markers, and adipokines (e.g., AGT, ANGPTL14, APOC1 , ADIPOQ, ACC1 , ADIG, LEP, RES, and HP). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in low-density lipoprotein (LDL) levels and / or decrease in the expression levels of apolipoprotein B (APOB). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises an increase in high-density lipoprotein (DHL) levels and / or increase in the expression levels of apolipoprotein A-1 (APOA-1 ). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in the expression level of one or more inflammatory markers (e.g., IL-6, IL1 Ra, IL1 p, fibrinogen, and TNFa). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in the expression level of GlycA, high-sensitivity C-reactive protein (hs-CRP), and / or N-terminal prohormone of brain natriuretic peptide (NT-proBNP). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises reduced intramuscular fat. In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises an increase in expression levels of insulin-like growth factor 1 (IGF-1 ), procollagen type III N-terminal peptide (P3NP), and / or fibroblast growth factor 21 (FGF-21 ). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises an increase in the level of Ghrelin, total testosterone, growth hormone, and / or dehydroepiandrosterone sulfate (DHEA-S). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in the expression level of creatine kinase (CK) and / or an increase in serum creatinine levels. In some embodiments, biomarker changes affected by administering an antibody disclosed herein comprises a decrease in markers previously identified as targets of myostatin signaling (e.g. ubiquitin ligases Fbxo32 (Atrogin-1 ) and / or Trim63 (MuRF) and / or ubiquitin protein Ubc (polyubiquitin C). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in the expression level of plasminogen activator inhibitor 1 (PAI-1 ). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in the level of 1 ,5-anhydroglucitol, fructosamine, and / or hemoglobin A1 c (HBA1 C). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in serum insulin. In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in cortisol levels. In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in expression level of pancreatic polypeptide family. In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises an increase in expression level of chromatin assembly factor 1 (CAF-1 ). In some embodiments, biomarker change affected by administering an antibody disclosed herein comprises a decrease in leptin and fasting glucose. In some embodiments, treatment is continued if a listed change is detected. In some embodiments, dosages or treatment frequencies are increased if a listed change is not detected. In some embodiments, treatment is discontinued if a listed change is not detected. In some embodiments, the biomarker change is detected in samples collected 1-10 days, e.g., 2-7 days, e.g., 3 days after administration of an antibody disclosed herein. In some embodiments, the biomarker change is detected in a sample collected 7 days after administration of an antibody disclosed herein. In some embodiments, the biomarker change is detected in a sample collected 28 days after administration of an antibody disclosed herein. In some embodiments, the biomarker change serves as a pharmacodynamic biomarker for myostatin inhibition.

[0317] In some embodiments, the function of the muscle tissue is increased by at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the function of the muscle tissue is increased by at least about 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%. In some embodiments, increased muscle function comprises improved rating, for example, from 1 to 2, 2 to 3, 3 to 4, 4 to 5, 5 to 6, 6 to 7, 7 to 8, 8 to 9, or 9 to 10.

[0435]

[0318] In some embodiments, use of a myostatin inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 , in the methods of the present disclosure increases the mass and / or function of the muscle tissue in the subject suffering from a lesion, e.g., due to a spinal cord injury. In some embodiments, the subject is in an acute spinal cord injury phase immediately after injury, where diagnosis between complete and incomplete injury is generally difficult. In other embodiments, the subject is in a sub-acute spinal cord injury phase, where there is a distinction between complete and incomplete spinal cord injury, and recovery is possible through ongoing rehab. In yet another embodiment, the subject is in a chronic spinal cord injury phase. The chronic spinal cord injury phase occurs around 4 months or 6 months from the date of injury, where patients have demonstrated substantial decrease in rate of recovery or when rehab efforts have reached a plateau despite the ongoing standard of care efforts.

[0436]

[0319] In some embodiments, the mass and / or function of the muscle tissue below a lesion is increased in a subject suffering from a lesion, e.g., a spinal cord injury. In other embodiments, the mass and / or function of the muscle tissue above a lesion is increased in a subject suffering from a lesion, e.g., a spinal cord injury. In some embodiments, the muscle is selected from the group consisting of a soleus muscle, a gastrocnemius muscle, a bicep muscle and a tricep muscle. In some embodiments, the mass of the muscle tissue is increased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the mass of the muscle tissue is increased by at least about 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%. In some embodiments, the function of the muscle tissue is increased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the function of the muscle tissue is increased by at least 1-5%, 5-10%, 10- 20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%.

[0437]

[0320] In some embodiments, administration of a myostatin inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 , increases locomotor function in the human subject, e.g., in a subject suffering from a lesion. In some embodiments, the locomotor function of the human subject is increased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the locomotor function of the human subject is increased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10- 50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%.

[0438]

[0321] In some embodiments, administration of the myostatin inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 , increases the motor coordination and balance in the human subject, e.g., in a subject suffering from a lesion. In some embodiments, the motor coordination and balance of the human subject is increased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the motor coordination and balance of the human subject is increased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%.

[0439]

[0322] In another embodiment, the myostatin inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 , is administered to a human subject suffering from a lesion. In some embodiments, the muscle strength of the human subject is increased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the muscle strength of the human subject is increased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%.

[0440]

[0323] In some embodiments, administration of a myostatin inhibitor e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 , can cause clinically meaningful changes in muscle function which corresponds to enhanced functionality of the patient. In some embodiments, enhanced functionality includes improvement in the patient’s mobility, self-care, metabolism, etc. In some embodiments, administration of an effective amount of the myostatin inhibitor facilitates or accelerates recovery from a condition, such as injuries, surgeries, and other medical procedures. Suitable such conditions may involve a condition that is associated with a nerve damage (whether resulting from an injury or a surgical or other clinical procedure).

[0441]

[0324] For example, suitable subjects include generally healthy individuals, such as a patient who: i) has sustained an acute injury involving a nerve damage that affects muscle function; ii) is scheduled to undergo a surgical procedure (therapeutic or corrective) that may cause an unintended nerve injury (e.g., motor neuron injury); iii) has undergone a surgical procedure that has caused an unintended muscle dysfunction; iv) receives a treatment that involves immobilization of a particular muscle or muscle groups (e.g., cast, etc.); v) is on ventilator (e.g., as a result of acute injury). The administration of a myostatin inhibitor described herein (e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) may accelerate recovery in such patients. In some embodiments, such administration is prophylactic. For example, prior to undergoing or immediately following a surgical procedure that may cause a nerve damage and associated muscle dysfunction, the antibody may be administered to prevent muscle dysfunction. Prevention includes alleviating or lessening the severity of such dysfunction. In these embodiments, administration is a local administration at or near the site of the affected area, e.g., injury, surgery, etc.

[0442] Effect on muscle catabolism of protein and / or muscle release of amino acids in the human subject:

[0443]

[0325] In some embodiments, administration of a myostatin inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 , to a human subject decreases muscle catabolism of protein and / or muscle release of amino acids in the human subject. In some embodiments, muscle catabolism of protein and / or muscle release of amino acids is decreased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, muscle catabolism of protein and / or muscle release of amino acids is decreased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%.

[0444] Effect on preventing muscle loss or atrophy in the human subject:

[0445]

[0326] In some embodiments, administration of an effective amount of a myostatin inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 , to a human subject prevents muscle loss or atrophy wherein the subject is at risk of developing muscle loss and / or atrophy. In some embodiments, muscle loss or atrophy is decreased or prevented by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In some embodiments, muscle loss or atrophy is decreased or prevented by at least 1-5%, 5-10%, 10- 20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%. In some embodiments, muscle mass is retained (i.e., no muscle mass gain or loss) as a result of the administration. In some embodiments, muscle mass is retained while fat mass is reduced as a result of the administration. In some embodiments, a subject receiving the myostatin inhibitor exhibits an increase in muscle mass. In some embodiments, administration of the myostatin inhibitor maintains the gain in muscle mass relative to muscle mass prior to treatment.

[0327] In some embodiments, a suitable subject is a human subject who has not yet developed atrophy but is considered at risk of developing atrophy. In some embodiments, the subject is on a weight loss program, such as a program comprising diet restriction and / or an exercise regimen. In some embodiments, the subject is on a weight loss program comprising a diet restriction regimen of at least 20%, e.g., at least 30%, 35%, 40%, 45%, or 50%, calorie reduction. In some embodiments, the subject has type 2 diabetes. In some embodiments, the subject is obese. In some embodiments, the subject is obese and has type 2 diabetes. In some embodiments, the subject has cardiovascular disease.

[0446]

[0328] In some embodiments, a suitable subject is a human subject who has a disease or condition associated with a neurological defect that impairs motor neuron function. In some embodiments, such disease or condition is caused by muscular dystrophy or atrophy. In some embodiments, the neurological defect is caused by a nerve injury. In some embodiments, the nerve injury involves partial denervation of motor neurons, which causes partial impairment of function in the affected muscle. In some embodiments, such disease or condition is caused by SCI. In some embodiments, the subject with SCI is in an acute or sub-acute phase of SCI (e.g., not yet reached a chronic phase).

[0447]

[0329] In some embodiments, when a composition comprising a myostatin inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, or Ab133, is administered to a population of patients who are at risk of developing muscle atrophy associated with partial denervation of motor neurons, the administration is capable of i) preventing onset or progression of the muscle atrophy in a statistically significant fraction of the patient population; or, ii) reducing the severity of the muscle atrophy in the statistically significant fraction of the patient population. In some embodiments, the myostatin inhibitor is administrated at a dose and frequency sufficient to prevent onset or progression of muscle atrophy or reduce the severity of muscle atrophy.

[0448]

[0330] Prevention of muscle loss or atrophy by the use of a myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) can be readily monitored or assessed by any suitable methods to evaluate muscle mass or motor function involving affected muscles.

[0449]

[0331] In some embodiments, administration of an effective amount of a myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) prevents or lessens neuropathy, e.g., diabetic neuropathy, or an early-onset axonal polyneuropathy in affected limbs.

[0450] Effect on intramuscular fat infiltration in the human subject:

[0451]

[0332] In some embodiments, administration of a myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) decreases intramuscular fat infiltration in the human subject. In some embodiments, intramuscular fat infiltration is decreased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, intramuscular fat infiltration is decreased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%.

[0452] Effect on the level of adipose tissue in the human subject:

[0453]

[0333] Brown adipose tissue (BAT) is known to function in the dissipation of chemical energy in response to cold or excess feeding, and also has the capacity to modulate energy balance. Activation of brown adipose tissue has been shown to improve glucose homeostasis and insulin sensitivity in humans suggesting that anyone with impaired insulin function might benefit from BAT activation (Stanford et al., J Clin Invest. 2013, 123(1 ): 215-223).

[0334] Beige adipose tissue is generated as a result of browning of white adipose tissue (WAT), also known as “beiging.” This occurs when adipocytes within WAT depots develop features of BAT. Beige adipocytes take on a multilocular appearance (containing several lipid droplets) and increase expression of uncoupling protein 1 (UCP1 ). In doing so, these normally energy -storing white adipocytes become energy-releasing adipocytes (Harms et al. Nature Medicine. 2013, 19 (10): 1252-63).

[0454]

[0335] Visceral fat or abdominal fat (also known as organ fat or intra-abdominal fat) is located inside the abdominal cavity, packed between the organs (stomach, liver, intestines, kidneys, etc.). Visceral fat is different from subcutaneous fat underneath the skin, and intramuscular fat interspersed in skeletal muscles. Fat in the lower body, as in thighs and buttocks, is subcutaneous and is not consistently spaced tissue, whereas fat in the abdomen is mostly visceral and semi-fluid. An excess of visceral fat is known as central obesity, “central adiposity,” or "belly fat," in which the abdomen protrudes excessively. Measurements such as the Body Volume Index (BVI) are specifically designed to measure abdominal volume and abdominal fat. Excess visceral fat is linked to type 2 diabetes, insulin resistance, inflammatory diseases and other obesity-related diseases (Mokdad et al., JAMA: The Journal of the American Medical Association. 2001 , 289 (1 ): 76-9).

[0455]

[0336] Mass of adipose tissue can be determined by any method known to a person of ordinary skill in the art. For example, adipose tissue may be measured by qNMR, dual-energy X-Ray absorptiometry (DEXA).and other methods known in the art.

[0456]

[0337] In some embodiments, administration of a myostatin inhibitor (e g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) affects the level of adipose tissue in the human subject. As used herein, the term “adipose tissue” refers to fat including the connective tissue that stores fat. Adipose tissue is derived from preadipocytes. Its main role is to store energy in the form of lipids, although it also cushions and insulates the body. The two types of adipose tissue are white adipose tissue (WAT), which stores energy, and brown adipose tissue (BAT), which generates body heat.

[0457]

[0338] In some embodiments, administration of the myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) increases the level of brown adipose tissue and / or the level of beige adipose tissue in the human subject. In some embodiments, the administration decreases the level of white adipose tissue and visceral adipose tissue in the human subject.

[0458]

[0339] In some embodiments, administration of a myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, or Ab133) increases the level of brown or beige adipose tissue by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In some embodiments, the administration increases the level of brown or beige adipose tissue by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%.

[0459]

[0340] In some embodiments, administration of a myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) decreases the level of white or visceral adipose tissue by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the administration decreases the level of white or visceral adipose tissue by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%.

[0460]

[0341] In some embodiments, administration of the myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) has different effects during the weight loss phase as compared to the weight maintenance phase. In some embodiments, administration of the myostatin inhibitor promotes more fat mass loss in the weight maintenance phase as compared to fat mass loss in the active weight loss phase. In some embodiments, administration of the myostatin inhibitor promotes more fat mass loss under moderate calorie restriction (e g., 30% or less calorie restriction, e.g., 20% calorie restriction) as compared to extreme calorie restriction (e.g., 30% or more calorie restriction).

[0461]

[0342] In some embodiments, administration of the myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) slows the rate of fat increase relative to the rate prior to starting the administration. In some embodiments, the administration maintains the reduced rate of fat increase. In some embodiments, the administration stops fat increase during the duration of the administration.

[0462]

[0343] In some embodiments, the above effect(s) are achieved with administration of the myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) alone or in conjunction with at least one additional agent (e.g., a standard of care therapeutic, e.g., for a metabolic disease). In some embodiments, the at least one additional agent is a GLP-1 pathway activator.

[0463]

[0344] In some embodiments, administration of the myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 , e.g., Ab109) prevents the degree or rate of fat regain after discontinuation of a GLP-1 pathway activator.

[0464]

[0345] In some embodiments, administration of the myostatin inhibitor, such as an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133 or Ab141 , e.g., Ab109) reduces fat mass around the subject’s liver.

[0465] Effect on the ratio of adipose-to-muscle tissue in the human subject:

[0466]

[0346] In some embodiments, administration of a myostatin inhibitor (e.g., a myostatin-selective inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, or Ab133) decreases the ratio of adipose-to-muscle tissue in the human subject. In some embodiments, the ratio of adipose-to-muscle tissue is decreased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the ratio of adipose-to-muscle tissue is decreased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20- 30%, 20-60%, 30-80%, 40-90%, or 50-100%. In some embodiments, administration of the myostatin inhibitor maintains the reduction in the ratio between adipose-to-muscle tissue in a human subject who has previously achieved a reduced ratio of adipose-to-muscle tissue.

[0467]

[0347] In some embodiments, administration of a myostatin inhibitor (e.g., a myostatin-selective inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) increases the ratio of muscle tissue to adipose in the human subject. In some embodiments, the ratio of muscle tissue-to-adipose is increased by at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the ratio of muscle tissue-to-adipose is increased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%. In some embodiments, administration of the myostatin inhibitor maintains the increased ratio of muscle tissue to adipose tissue in a human subject who has previously achieved such an increased ratio.

[0468]

[0348] In some embodiments, the above effect(s) are achieved with administration of the myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) alone or in conjunction with at least one additional agent (e.g., a standard of care therapeutic, e.g., for a metabolic disease). In some embodiments, the at least one additional agent is a GLP-1 pathway activator.

[0469] Hormonal control:

[0470]

[0349] Leptin is a hormone produced and secreted by adipose tissue that plays a role in regulating food intake and stimulating energy expenditure. Defects in leptin production have been reported to cause severe hereditary obesity in rodents and humans. In addition to its effects on body weight, leptin has a variety of other functions, including the regulation of hematopoiesis, angiogenesis, wound healing, and the immune and inflammatory response. Leptin acts through the leptin receptor, a single-transmembrane-domain receptor of the cytokine receptor family, which is found in many tissues in several alternatively spliced forms. The LEP gene is the human homolog of the gene (ob) mutant in the mouse 'obese' phenotype. The leptin level in a subject refers to the amount of circulating leptin in the body of the subject, e.g., a mammalian subject, and a reduction in leptin levels refers to a reduction in the amount of circulating leptin as compared to a baseline measurement, e.g., as a result of treatment.

[0471]

[0350] In some embodiments, the therapeutic efficacy of treating a subject (e.g., a mammalian subject) with a myostatin inhibitor (e.g., a myostatin-selective inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) is determined by measuring a reduction in leptin levels in the blood of the subject after administering the myostatin inhibitor as compared to before the administration. In some embodiments, the therapeutic efficacy is determined by (i) determining a blood level of leptin in the subject prior to administering the myostatin inhibitor; (ii) administering the myostatin inhibitor; and (iii) determining a blood level of leptin in the subject after administering the myostatin inhibitor; wherein a reduction in blood leptin level indicates therapeutic efficacy.

[0472]

[0351] Adiponectin is an adipokine involved in the control of fat metabolism and insulin sensitivity with direct antidiabetic, anti-atherogenic and anti-inflammatory activities. Adiponectin stimulates AMPK phosphorylation and activation in the liver and the skeletal muscle, enhancing glucose utilization and fatty-acid combustion and inhibits endothelial NF-kappa-B signaling through a cAMP-dependent pathway. Adiponectin may be involved in brown fat cell differentiation. As used herein, the term “adiponectin level” refers to the amount of circulating adiponectin, e.g., total adiponectin or glycosylated adiponectin. The plasma adiponectin level of a subject, e.g., a mammalian subject, may be measured by, e.g., the total adiponectin assay (ELISA kit EZHADP-61 K, Millipore, St. Charles, Missouri, USA) which captures all forms of circulating adiponectin, with a sensitivity of 0.78 ng / mL and within-batch and between-batch coefficients of variation of 1.8% and 6.2%, respectively. An increase or decrease in adiponectin level refers to a higher or lower amount of circulating adiponectin detected in a subject (e.g., following a treatment for a metabolic disease or disorder) compared to a baseline measurement.

[0473]

[0352] Ghrelin is an appetite-regulating hormone that has an appetite-stimulating effect, induces adiposity, and stimulates gastric secretion. Plasma ghrelin concentration is increased in fasting conditions and reduced after habitual feeding. The normal ghrelin concentration of plasma samples in humans is 10-20 fmol / ml for n-octanoyl ghrelin and 100-150 fmol / ml for total ghrelin, including both acyl-modified and des-acyl ghrelins. Ghrelin levels may be measured using, e.g., the Ghrelin Human ELISA kit, CAT# BMS2192 (Invitrogen).

[0474]

[0353] In some embodiments, the above effect(s) are achieved with administration of the myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) alone or in conjunction with at least one additional agent (e.g., a standard of care therapeutic, e.g., for a metabolic disease). In some embodiments, the at least one additional agent is a GLP-1 pathway activator. Effect on the metabolic rate of the human subject:

[0475]

[0354] In some embodiments, administration of a myostatin inhibitor (e.g., a myostatin-selective inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, or Ab133) increases the metabolic rate of the human subject. In some embodiments, the administration can increase the basal metabolic rate in the subject. Metabolic rates can be calculated by any methods known in the art, for example, by examining the oxygen input and carbon dioxide output, or by indirect calorimetry. In some embodiments, the metabolic rate is increased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, the metabolic rate is increased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30- 80%, 40-90%, or 50-100%.

[0476]

[0355] In some embodiments, the above effect(s) are achieved with administration of the myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) alone or in conjunction with at least one additional agent (e.g., a standard of care therapeutic, e.g., for a metabolic disease). In some embodiments, the at least one additional agent is a GLP-1 pathway activator.

[0477] Effect on glucose uptake in the human subject:

[0478]

[0356] In some embodiments, administration of a myostatin inhibitor (e.g., a myostatin-selective inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, or Ab133) affects glucose uptake by tissues in the human subject. In some embodiments, glucose uptake by muscle tissue is increased. In some embodiments, glucose uptake by the muscle tissue is increased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In some embodiments, glucose uptake by the muscle tissue is increased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%.

[0479]

[0357] In other embodiments, glucose uptake by white adipose tissue, liver tissue and / or blood vessel tissue are reduced. In some embodiments, glucose uptake by white adipose tissue, liver tissue and / or blood vessel tissue are reduced by at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, glucose uptake by white adipose tissue, liver tissue and / or blood vessel tissue are reduced by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30- 80%, 40-90%, or 50-100%.

[0480]

[0358] In some embodiments, treatment of a subject with a myostatin inhibitor (e.g., a myostatin-selective inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, or Ab133) reduces glucose levels (e.g., non-fasted glucose levels orfasted glucose levels) in the serum of the subject relative to baseline prior to treatment. In some embodiments, treatment of a subject comprising a GLP-1 receptor agonist and a myostatin inhibitor (e.g., a myostatin-selective inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) reduces glucose levels (e.g., non-fasted glucose levels or fasted glucose levels) in the serum of the subject relative to a subject treatment with a GLP-1 receptor agonist alone. In some embodiments, the antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin is Ab109 or comprises the CDRs and / or variable domains from Ab109.

[0481]

[0359] The term “non-fasted glucose levels” refers to glucose content in a subject’s blood as measured without having to fast or refraining from eating or drinking for a prolonged period of time, e.g., 8 hours. Blood glucose can be measured using a hemoglobin A1 C test, also known as an A1 C, HbA1 C, glycated hemoglobin, or glycosylated hemoglobin test. Conventional home glucose monitoring can also be used to measure blood sugar. The term “postprandial insulin level” refers to the insulin concentration in a subject’s blood shortly after eating (e.g., two hours after eating). The term “postprandial glucose level” refers to glucose concentrations in a subject’s blood shortly after eating (e.g., two hours after eating). The postprandial insulin and glucose levels may be affected by carbohydrate absorption, insulin and glucagon secretion, and their coordinated effects on glucose metabolism in the liver and peripheral tissues. The magnitude and time of the peak plasma insulin or glucose concentration depend on a variety of factors, including the timing, quantity, and composition of the meal. In non-diabetic human subjects, plasma glucose concentrations peak about 60 min after the start of a meal and return to preprandial levels within 2-3 hour. Postprandial insulin levels may be measured using a postprandial insulin test, such as a two-hour postprandial insulin test. Postprandial glucose levels may be measured using a postprandial blood glucose test, such as a two-hour postprandial blood glucose test.

[0482]

[0360] In some embodiments, the above effect(s) are achieved with administration of the myostatin inhibitor (e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) alone or in conjunction with at least one additional agent (e.g., a standard of care therapeutic, e.g., for a metabolic disease). In some embodiments, the at least one additional agent is a GLP-1 pathway activator.

[0483] Effect on preventing development of a metabolic disorder in the subject:

[0484]

[0361] In some embodiments, administration of an effective amount of a myostatin inhibitor (e.g., a myostatin-selective inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, including SRK-439 and any one of Ab101-Ab141 , e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) prevents development of a metabolic disorder in the subject, e.g., a human subject. In some embodiments, development of a metabolic disorder is decreased by at least 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90% or 100%. In other embodiments, development of a metabolic disorder is decreased by at least 1-5%, 5-10%, 10-20%, 1-30%, 1-40%, 1-50%, 10-50%, 20-30%, 20-60%, 30-80%, 40-90%, or 50-100%.

[0485]

[0362] In some embodiments, administration of an effective amount of a myostatin inhibitor (e.g., a myostatin-selective inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., SRK-439, Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) reduces a risk of a cardiovascular disease, e.g., by reducing levels of cholesterol and or the LDL / HDL ratio. Cholesterol encompasses both low-density lipoprotein (LDL) cholesterol and high-density lipoprotein (HDL) cholesterol. Total cholesterol level refers to a measure of the total amount of cholesterol (e.g., LDL and HDL) in a blood sample collected from a subject. LDL cholesterol contains low- density lipoprotein that makes up most of the cholesterol found in the body. A high level of LDL may cause lipid deposits (e.g., plaques) in arteries and may be associated with or linked to increased risk for cardiovascular conditions, such as heart disease and stroke. By contrast, HDL cholesterol contains high-density lipoprotein, which absorbs cholesterol and carries it back to the liver. Generally, a high level of HDL cholesterol is associated with lower risk of heart disease and stroke. Cholesterol levels can be measured using a blood test, including blood tests commonly known as complete cholesterol tests, lipid panels, or lipid profiles.

[0486]

[0363] Notably, medications such as statins, which are commonly prescribed for managing unhealthy cholesterol levels, may lead to side effects, including muscle weakness and fatigue. Advantageously, myostatin inhibitors may be used in conjunction with cholesterol-lowing medications to prevent or lessen muscle weakness or fatigue associated with such medications.

[0487]

[0364] In some embodiments, a suitable subject is a subject who has not fully developed a metabolic disease but is considered at risk of developing such a condition. In some embodiments, a subject has a disease or condition associated with muscle dysfunction. In some embodiments, the muscle dysfunction is associated with partial denervation of motor neurons, which causes partial impairment of function in the affected muscle. In some embodiments, such conditions are caused by muscular dystrophy or atrophy. In some embodiments, such condition is caused by SCI. In some embodiments, the subject with SCI is in an acute or sub-acute phase of SCI (e.g., not yet reached a chronic phase).

[0488]

[0365] In some embodiments, when a composition comprising an effective amount of a myostatin inhibitor (e.g., a myostatin-selective inhibitor, e.g., an antibody or antigen-binding fragment thereof that binds specifically to pro / latent myostatin as disclosed herein, e.g., Ab102, Ab109, Ab130, Ab132, Ab133, or Ab141 ) is administered to a population of patients who are at risk of developing a metabolic disorder associated with muscle dysfunction, the composition i) prevents manifestation or aggravation of the metabolic disorder in a statistically significant fraction of the patient population; or, ii) lessens the severity of the metabolic disorder in the statistically significant fraction of the patient population. In some embodiments, myostatin inhibitor is administered alone or in conjunction with at least one additional agent (e.g., a standard of care therapeutic, e.g., for a metabolic disease). In some embodiments, the at least one additional agent is a GLP-1 pathway activator.

[0489]

[0366] In some embodiments, effects on metabolism is monitored or measured by insulin resistance, lipid panel / markers (e.g., leptin, adiponectin, ghrelin), inflammatory markers and oxidative stress markers, including, but are not limited to: IL-6, TNF, CRP, plasma total antioxidant status, lipid oxidation and erythrocyte glutathione peroxidase activity.

[0490] Routes of administration

[0491]

[0367] In carrying out various embodiments of the present disclosure, the preferred route of administration is subcutaneous administration.

[0492]

[0368] Products intended for use in the treatment of metabolic disorders such as obesity are preferably offered for selfadministration (home administration). As such, flat dosing is the preferred dosage form, which may reduce the risk of dosing errors. In some embodiments, the product is in a single-use format. In preferred embodiments, the product is available as a prefilled syringe or an injector, such as an autoinjector.

[0493]

[0369] In some embodiments, the formulation to enable subcutaneous administration of the myostatin inhibitor comprises at least about 150 mg / mL of the antibody, e.g., about 160 mg / mL, about 170 mg / mL, about 175 mg / mL, about 180 mg / mL, about 190 mg / mL, about 200 mg / mL. In some embodiments, administration comprises a single subcutaneous injection of up to 2.0 mL, which corresponds to up to about 400 mg of the therapeutic antibody per injection.

[0494]

[0370] Preferably, SRK-439 is subcutaneously administered at flat dose of between about 8 mg and 800 mg, e.g., 8 mg, 24 mg, 80 mg, 240 mg, or 800 mg.

[0495]

[0371] In some embodiments, a loading dose of SRK-439 is administered at the outset of a treatment regimen, followed by maintenance doses which are lower than the loading dose.

[0496]

[0372] In some embodiments, dosing regimen for SRK-439 is Q1W, Q2W, Q3W, Q4W, or once monthly.

[0497] Combination therapy

[0498]

[0373] In addition to combination or add-on therapy with an incretin-based treatment, myostatin inhibitors can be used in conjunction with other interventions intended to aid weight loss or otherwise help improve metabolic conditions. It is documented that almost any drastic weight loss - whether it is achieved by incretins or other nutrition-triggered hormones or their analogs, strict diet, combination of diet and exercise, or surgical means (e.g., bariatric surgery) - is accompanied by significant muscle loss which is typically around 30% of the weight loss, and / or patient-reported muscle weakness. It is contemplated herein that inhibition of myostatin can have a beneficial impact in any of these situations. As such, incorporating myostatin inhibitor therapy into various interventions for metabolic conditions, such as weight management, is encompassed by the disclosure.

[0499]

[0374] Accordingly, the present disclosure provides myostatin inhibitors for use in the treatment of a metabolic disorder in a subject, who benefits from retaining Activin A function, or for whom Activin A inhibition may be particularly detrimental, wherein the myostatin inhibitor does not inhibit Activin A. In various embodiments, the myostatin inhibitor is selected from non-selective inhibitors that spare Activin A, such as ligand traps that inhibit both myostatin and GDF11 but not Activin A, neutralizing antibodies that bind mature myostatin and mature GDF11 but not mature Activin A, and the like. In preferred embodiments, the myostatin inhibitor that does not inhibit Activin A is a myostatin-selective inhibitor, such as anti-mature myostatin antibodies (i. e. , neutralizing antibodies) that selectively bind and inhibit myostatin (such as trevogrumab), and anti-pro / latent myostatin antibodies that selectively inhibit activation of myostatin (such as SRK-439, apitegromab, GYM329, Ab101 , Ab102, Ab103, Ab104, Ab105, Ab106, Ab107, Ab108, Ab109, Ab110, Ab111 , Ab112, Ab113, Ab114, Ab115, Ab116, Ab117, Ab118, Ab119, Ab120, Ab121 , Ab122, Ab123, Ab124, Ab125, Ab126, Ab127, Ab128, Ab129, Ab130, Ab131 , Ab132, Ab133, Ab134, Ab135, Ab136, Ab137, Ab138, Ab139, Ab140, and Ab141 ).

[0500]

[0375] Such myostatin inhibitors that retain Activin A function may be used in conjunction with any one or more of the following agents, which are aimed to improve metabolic health or weight management: GLP-1 , GLP-1 analogs, glucagon, glucagon analogs, GIP, GIP analogs, cholecystokinin (CCK), IGF-1 , BDNF, IL-15, irisin, myonectin, decorin, FGF21 , a GPR119 agonist, a GPR19 agonist, a CB1R agonist, amylin, amylin analog (e g., cagrilintide), apelin, adiponectin, leptin, peptide YY (PYY), parathyroid hormone (PTH), teriparatide, and exendin-4.

[0501]

[0376] Such myostatin inhibitors that retain Activin A function may be used in conjunction with any one or more of the following agents, which are aimed to improve metabolic health or weight management: biologically active peptide that modulates glucose or lipid metabolism (e.g. , comprising a hormone or a cytokine that modulates glucose or lipid metabolism). In some embodiments, the biologically active peptide is an endocrine hormone or a neuroendocrine hormone. In some embodiments, the biologically active peptide is a peptide that regulates the activity of an endocrine or neuroendocrine hormone. In some embodiments, the biologically active peptide is a hormone of the hypothalamus, pituitary gland, thyroid, adrenal gland, parathyroid, pancreas, pineal gland, ovary, or testis. In some embodiments, the biologically active peptide is a hepatokine, wherein, optionally, the hepatokine is a GPR19 agonist, wherein, further optionally, the GPR19 agonist is adropin. In some embodiments, the biologically active peptide is a pituitary hormone (e.g., IGF-1), a GLP-1 pathway activator (e.g., an incretin, e.g., GLP-1 , GIP, exendin-4), a neuropeptide, a neuroendocrine hormone (e.g., amylin), a myokine (e.g., irisin, BDNF, IL-15, myonectin, decorin, or FGF21), an adipokine (e.g., adiponectin, apelin, leptin), a parathyroid hormone (e.g., a parathyroid hormone-related peptide, e.g., teriparatide), a GPR19 agonist, a GPR119 agonist, a CB1 R agonist, or a fragment of any one of the foregoing.

[0502]

[0377] In some embodiments, myostatin inhibitors that retain Activin A function can be used in conjunction with non- pharmacological interventions for aiding weight management, such as diet and / or exercise. In some embodiments, the diet comprises high-protein diet. In some embodiments, the diet comprises caloric restriction, such as reduction of calory intake by certain amount, e.g., by 500 kcal per day. In some embodiments, the exercise comprises resistance exercises.

[0503]

[0378] In some embodiments, myostatin inhibitors that retain Activin A function can be used in conjunction with surgical interventions, such as bariatric surgery. In some embodiments, the bariatric surgery is sleeve gastrectomy, Roux-en-Y gastric bypass surgery, biliopancreatic diversion with duodenal switch, vertical banded gastroplasty, and / or gastric plication. In some embodiments, the surgery comprises the use of intragastric balloon, adjustable gastric band, and / or implantable gastric stimulation. Myostatin assays

[0504]

[0379] Myostatin exists in multiple forms in vivo. The fully processed form that is secreted from cells is a dimer complex, in which the growth factor is “trapped” or in other words protected by the prodomain via non-covalent interactions. This is an inactive form often referred to as latent myostatin. Activation of myostatin involves the dissociation (e.g., release) of the growth factor from the latent complex. Once released, the free, mature growth factor can now bind its cell-surface receptors (e.g., ActRIIA / B) thereby triggering ligand-induced downstream receptor signaling. This activation step is believed to be induced by certain proteases (such as BMP-1 / mTLL proteases), which cleave the prodomain, triggering the growth factor dissociation. Antibodies that specifically target the latent complex (e.g., proMyostatin) and prevent its activation are referred to as “activation inhibitors.” These antibodies therefore do not directly inhibit mature growth factor; rather, they bind the latent (e.g., inactive) complex prior to the activation step. The antibody-antigen complex for this type of inhibitors therefore is comprised of the antibody bound to the dimer complex that includes both the growth factor and the prodomain (latent complex). This immune complex is then “pulled” from the tissue (such as muscle) and enters circulation, where in some cases it can persist for a prolonged period of time. Thus, levels of the immune complex (activation inhibitor antibody bound to the latent myostatin dimer) in serum can be measured and serve as a PD surrogate for subjects who receive myostatin inhibitor therapy. Alternatively, where free latent myostatin is affected by a myostatin inhibitor treatment, levels of free latent myostatin in serum can be measured and serve as a PD surrogate for subjects receiving myostatin inhibitor therapy.

[0505]

[0380] As a potent negative regulator of muscle mass, myostatin availability is tightly controlled in vivo. In healthy individuals, levels of free myostatin growth factor in circulation are low. The majority of circulating myostatin forms are latent myostatin.

[0506]

[0381] When a subject is administered with an activation inhibitor antibody (such as SRK-439, Ab101-Ab141 , apitegromab and GYM329), the total latent myostatin in circulation corresponds to a sum of the pool of latent myostatin that exists as the immune complex (bound by the antibody) and the second pool of latent myostatin that is not bound by antibody (e.g., “free” latent myostatin).

[0507] Free latent myostatin assays:

[0508]

[0382] An assay of the target engagement of the antibodies, e.g., a pharmacodynamic assay, can be performed by measuring the levels of free latent myostatin. In some embodiments, a pharmacodynamic assay can be performed by measuring the level of serum free latent myostatin (i.e. , circulating latent myostatin that is not bound by antibody). Such an assay may immobilize streptavidin, bind a biotinylated antibody known to bind latent myostatin, add a sample comprising latent myostatin, and test the ability of an antibody to bind to the free latent myostatin by labeling the test antibody with a detectable marker. In some embodiments, the capture antibody known to bind latent myostatin is biotinylated, and the detection or test antibody is a ruthenium labeled antibody that binds free latent myostatin. In some embodiments, saturation in target engagement indicates a dosage sufficient to achieve certain therapeutic effects (e.g., efficacy), although it may be possible to obtain efficacy without saturation. Upon administration of a single dose of the novel myostatin inhibitor disclosed herein (e.g., at 2-20 mg / kg) to mice, a rapid reduction in serum free latent myostatin is observed within one day, which remains at undetectable or nearly undetectable levels for at least 42 days, indicating durable target engagement and inhibitory activity of the antibodies.

[0509]

[0383] In some embodiments, the present disclosure encompasses a method of detecting circulating free latent myostatin (i.e., circulating latent myostatin that is not bound by an antibody). Such an assay may immobilize streptavidin, bind a biotinylated anti-pro / latent myostatin mAb (i.e., capture antibody reagent), incubate with a serum or other sample, and detect binding of a ruthenylated anti-myostatin mAb (i.e., detection antibody reagent) to detect the free latent myostatin in the sample that is bound to the capture antibody (and not bound by an antibody in the sample, e.g. by a myostatin inhibitor therapy antibody in the sample). Electrochemiluminescence (ECL) signal detected is proportional to the amount of free latent myostatin in the sample. In some embodiments, the biotinylated anti-pro / latent myostatin mAb (i.e., capture antibody reagent) comprises a biotinylated antibody of the disclosure.

[0510]

[0384] In some embodiments, the biotinylated anti-pro / latent myostatin mAb (i.e., capture antibody reagent) is a biotinylated antibody comprising any one of Ab101-Ab141. In some embodiments, the biotinylated anti-pro / latent myostatin mAb (i.e., capture antibody reagent) is biotinylated Ab101 , Ab102, Ab103, Ab104, Ab105, Ab106, Ab108, Ab109, Ab110, Ab111 , Ab112, Ab113, Ab114, Ab115, Ab116, Ab117, Ab118, Ab119, Ab120, Ab121 , Ab122, Ab123, Ab124, Ab125, Ab126, Ab127, Ab128, Ab129, Ab130, Ab131 , Ab132, Ab133, Ab134, Ab135, Ab136, Ab137, Ab138, Ab139, Ab140, or Ab141. In some embodiments, the biotinylated anti-pro / latent myostatin mAb (i.e., capture antibody reagent) comprises Ab109, Ab133 or Ab141. In some embodiments, the biotinylated anti-pro / latent myostatin mAb (i.e., capture antibody reagent) comprises an antibody disclosed in WO2016 / 073853 or a mouse chimera thereof, e.g., Ab1 from WO2016 / 073853 or a mouse chimera thereof with antigen-binding fragments of Ab1 with a murine lgG1 constant domain. In some embodiments, the biotinylated anti-pro / latent myostatin mAb (i.e., capture antibody reagent) comprises GYM329 or antigen-binding fragments thereof. In some embodiments, the biotinylated anti-pro / latent myostatin mAb (i.e., capture antibody reagent) comprises SRK-439 or antigen-binding fragments thereof.

[0511]

[0385] In some embodiments, the ruthenylated anti-myostatin mAb (i.e., detection antibody reagent) comprises a ruthenium-labeled antibody of the disclosure. In some embodiments, the ruthenylated anti-myostatin mAb (i.e., detection antibody reagent) is a ruthenylated antibody comprising any one of Ab101-Ab141. In some embodiments, the ruthenylated (detection) antibody known to bind latent myostatin is a ruthenylated antibody comprising Ab101 , Ab102, Ab103, Ab104, Ab105, Ab106, Ab108, Ab109, Ab110, Ab111 , Ab112, Ab113, Ab114, Ab115, Ab116, Ab117, Ab118, Ab119, Ab120, Ab121 , Ab122, Ab123, Ab124, Ab125, Ab126, Ab127, Ab128, Ab129, Ab130, Ab131 , Ab132, Ab133, Ab134, Ab135, Ab136, Ab137, Ab138, Ab139, Ab140, or Ab141. In some embodiments, the ruthenylated anti- myostatin mAb (i.e., detection antibody reagent) comprises ruthenylated anti-myostatin mAb, RK22. In some embodiments, the ruthenylated anti-myostatin mAb (i.e., detection antibody reagent) is a ruthenylated antibody comprising an antibody disclosed in WO2016 / 073853 or a mouse chimera thereof, e.g., Ab1 from WO2016 / 073853 or a mouse chimera thereof with antigen-binding fragments of Ab1 with a murine lgG1 constant domain, GYM329, Ab109, Ab133, or Ab141. In some embodiments, the ruthenylated anti-myostatin mAb (i.e., detection antibody reagent) is a ruthenylated antibody comprising an antibody disclosed in WO2016 / 073853 or a mouse chimera thereof, e.g., Ab1 from WO2016 / 073853 or a mouse chimera thereof with antigen-binding fragments of Ab1 with a murine lgG1 constant domain. In some embodiments, the ruthenylated anti-myostatin mAb (i.e., detection antibody reagent) is a ruthenylated antibody comprising GYM329 or a mouse chimera thereof. In some embodiments, the ruthenylated anti-myostatin mAb (i.e., detection antibody reagent) is a ruthenylated antibody comprising SRK-439 or a mouse chimera thereof.

[0512]

[0386] In some embodiments, the capture antibody reagent comprises an antibody disclosed in WO2016 / 073853 or a mouse chimera thereof, e.g., Ab1 from WO2016 / 073853 or a mouse chimera thereof with antigen-binding fragments of Ab1 with a murine lgG1 constant domain, or GYM329, SRK-439, Ab109, Ab133, or Ab141 , and the detection antibody reagent comprises an antibody disclosed in WO2016 / 073853 or a mouse chimera thereof, e.g., Ab1 from WO2016 / 073853 or a mouse chimera thereof with antigen-binding fragments of Ab1 with a murine lgG1 constant domain, or GYM329, SRK-439, Ab109, Ab133, or Ab141.

[0513]

[0387] In some embodiments, the sample is from a subject treated with Ab109 and the capture antibody reagent comprises Ab109, Ab133 or Ab141. In some embodiments, the sample is from a subject treated with SRK-439 and the capture antibody reagent comprises an antibody disclosed in WO2016 / 073853 or a mouse chimera thereof, e.g., Ab1 from WO2016 / 073853 or a mouse chimera thereof with antigen-binding fragments of Ab1 with a murine lgG1 constant domain, SRK-439, Ab109, Ab133 or Ab141. In some embodiments, the sample is from a subject treated with GYM-329 and the capture antibody reagent comprises GYM-329, an antibody disclosed in WO2016 / 073853 or a mouse chimera thereof, e.g., Ab1 from WO2016 / 073853 or a mouse chimera thereof with antigen-binding fragments of Ab1 with a murine lgG1 constant domain, SRK-439, Ab109, Ab133 or Ab141.

[0514]

[0388] In some embodiments, the detection range of the assay may be in the nanogram range, e.g., 0.1 ng / mL-750 ng / mL, 1.0 ng / mL-500 ng / mL, or 3.0 ng / mL-500 ng / mL, e.g., 3.9 ng / mL-500 ng / mL, e.g., 3.9 ng / mL-250 ng / mL.

[0515] LISTING OF CERTAIN EMBODIMENTS

[0516] 1 . A composition comprising a selective inhibitor of myostatin, which is an antibody or antigen-binding fragment thereof, for use in the treatment of a metabolic disorder in a subject, wherein the treatment comprises subcutaneous administration of the composition to the subject, wherein the composition is formulated with the antibody at a concentration of between about 100-200 mg / mL, and wherein the subject is treated with an incretin therapy, wherein optionally the incretin therapy comprises a GLP-1 receptor agonist.

[0517] 2. The composition for use according to embodiment 1 , wherein the subcutaneous administration comprises a flat dose of about 8-800 mg of the antibody, wherein optionally the administration is repeated every 1-8 weeks.

[0518] 3. The composition for use according to embodiment 2, wherein the administration is repeated every 4 weeks.

[0519] 4. The composition for use according to embodiment 1 or 2, wherein the subcutaneous administration comprises a flat dose of about 0.04-4.0 mL of the composition, wherein optionally the administration is repeated every 1-8 weeks.

[0520] 5. The composition for use according to embodiment 4, wherein the administration is repeated every 4 weeks.

[0521] 6. The composition for use according to any one of the preceding embodiments, wherein the composition is formulated with the antibody at a concentration of about 150 mg / mL, about 160 mg / mL, about 170 mg / mL, about 175 mg / mL, about 180 mg / mL, about 190 mg / mL or about 200 mg / mL.

[0522] 7. The composition for use according to any one of the preceding embodiments, wherein the composition is provided as a pre-filled syringe or injector, wherein optionally the injector is an auto-injector.

[0523] 8. The composition for use according to any one of the preceding embodiments, wherein the incretin therapy is selected from a GLP-1 receptor agonist, a dual GLP-1 / GIP receptor agonist, a dual GLP-1 / glucagon receptor agonist, and a triple GLP-1 / GIP / Glucagon receptor agonist.

[0524] 9. The composition for use according to embodiment 8, wherein the GLP-1 receptor agonist is semaglutide.

[0525] 10. The composition for use according to embodiment 8, wherein the dual GLP-1 / GIP receptor agonist is tirzepatide.

[0526] 11 . The composition for use according to embodiment 8, wherein the dual GLP-1 / glucagon receptor agonist is selected from AZD9550, CT-388, CT-868, survodutide (BI456906), pemvidutide (ALT-801), NN1177, and efinopegdutide (MK-6024).

[0527] 12. The composition for use according to embodiment 8, wherein the triple GLP-1 / GIP / Glucagon receptor agonist is selected from retatrutide and LY343794.

[0528] 13. The composition for use according to any one of the preceding embodiments, wherein the selective inhibitor of myostatin is selected from SRK-439, GYM329, trevogrumab, Ab101 , Ab102, Ab103, Ab104, Ab105, Ab106, Ab107, Ab108, Ab109, Ab110, Ab111 , Ab112, Ab113, Ab114, Ab115, Ab116, Ab117, Ab118, Ab119, Ab120, Ab121,

[0529] Ab122, Ab123, Ab124, Ab125, Ab126, Ab127, Ab128, Ab129, Ab130, Ab131, Ab132, Ab133, Ab134, Ab135, Ab136, Ab137, Ab138, Ab139, Ab140, and Ab141.

[0530] 14. A composition comprising SRK-439, a variant thereof or derivative thereof, for use in the treatment of a metabolic disorder in a subject, wherein the treatment comprises subcutaneous administration of the composition to the subject, wherein the composition is formulated with SRK-439 at a concentration of at least 150 mg / mL.

[0531] 15. The composition for use according to embodiment 14, wherein the concentration is about 180 mg / mL

[0532] 16. The composition for use according to embodiment 15, wherein the concentration is about 200 mg / mL.

[0533] 17. A GLP-1 receptor agonist for use in the treatment of a metabolic disorder in a subject, wherein the subject is treated with SRK-439, a variant thereof or derivative thereof, which is formulated as a composition having an antibody concentration of at least 150 mg / mL.

[0534] 18. A method of preventing muscle loss during weight loss in a subject, the method comprising a step of administering to the subject a composition comprising SRK-439, wherein up to 4.0 mL of the composition is administered subcutaneously once every 1-4 weeks, and wherein the composition comprises SRK-439 at a concentration equal to or greater than 150 mg / mL, wherein the subject is on an incretin therapy.

[0535] 19. A method of preventing muscle loss during weight loss in a subject, the method comprising a step of administering to the subject a composition comprising SRK-439, wherein up to 4.0 mL of the composition is administered subcutaneously once every 1-4 weeks, and wherein the composition comprises SRK-439 at a concentration equal to or greater than 150 mg / mL, wherein the subject has been on a incretin therapy and is discontinuing the incretin therapy.

[0536] 20. The composition for use according to any one of the preceding embodiments, wherein the subject is a female subject of child-bearing capacity.

[0537] 21 . The composition for use according to embodiment 20, wherein the subject is not on a contraceptive.

[0538] 22. The composition for use according to embodiment 20, wherein the subject has or is at risk of developing a bone disorder.

[0539] 23. The composition for use according to embodiment 20, wherein the subject has neuropathy, wherein optionally the neuropathy is associated with diabetes.

[0540] 24. The composition for use according to embodiment 20, wherein the subject has or is at risk of developing anemia.

[0541] 25. The composition for use according to embodiment 20, wherein the subject discontinues the incretin therapy.

[0542] 26. An incretin mimetic and a myostatin inhibitor for use in the treatment of a metabolic disease in a subject, wherein the treatment comprises administration of the incretin mimetic and the myostatin inhibitor to the subject, wherein the subject is an adult male or female of 218 and 265 years of age and has a BMI of ...

Claims

1. CLAIMS1 . A myostatin-selective inhibitor for use in the treatment of overweight or obesity in a subject, wherein the subject is on an incretin-based therapy and has a diminished response to the incretin-based therapy, wherein the treatment comprises administration of a myostatin-selective inhibitor to the subject in an amount effective to augment the effect of the incretin-based therapy.

2. The myostatin-selective inhibitor for use of claim 1 , wherein the augmentation comprises prolonging the effect of the incretin-based therapy.

3. The myostatin selective inhibitor for use of claim 1 or claim 2, wherein the augmentation comprises improving the subject’s body composition, wherein the improved body composition comprises increased lean mass, recued fat mass, or both.

4. A myostatin-selective inhibitor for use in the prevention of bone mass loss in a subject who is at risk of bone fracture or bone mass loss, wherein the treatment comprises administration of a myostatin-selective inhibitor to the subject, in an amount effective to protect against bone mass loss, wherein optionally the bone mass loss is assessed by bone mineral density or cortical thickness, wherein further optionally the risk of bone fracture or bone mass loss is associated with a high fat diet, inflammation, medication or injury.

5. The myostatin-selective inhibitor for use of claim 4, wherein the subject is on an incretin-based therapy, optionally wherein the subject is experiencing rapid weight loss.

6. A myostatin-selective inhibitor for use in the prevention of bone mass loss in a subject who is at risk of bone fracture or bone mass loss, wherein the treatment comprises administration of a myostatin-selective inhibitor to the subject, in an amount effective to protect against bone mass loss, wherein optionally the bone mass loss is assessed by bone mineral density or cortical thickness, wherein further optionally the risk of bone fracture or bone mass loss is associated with rapid weight loss comprising at least 10% of initial body weight over 6 months or less.

7. The myostatin-selective inhibitor for use according to claim 6, wherein the rapid weight loss is associated with an incretin-based therapy or a surgical intervention.

8. The myostatin-selective inhibitor for use of any one of claims 4-7, wherein the subject at risk of bone fracture has a T-score of at least -1 SD below the young adult mean prior to receiving the myostatin-selective inhibitor.

9. A method for preventing muscle loss and / or weakened bone during rapid weight loss in a subject, the method comprising a step of administering a myostatin-selective inhibitor to a subject undergoing or expected to undergo a rapid weight loss comprising at least 10% of initial body weight over 6 months or less, wherein optionally the subject is already on an incretin-based therapy or commencing an incretin-based therapy in conjunction with the administration of the myostatin-selective inhibitor.

10. A method for reducing the risk of bone fracture in a subject, the method comprising a step of administering to a subject a myostatin-selective inhibitor in an amount effective to maintain or enhance bone density, wherein optionally the bone density is assessed by bone mineral density or cortical bone porosity as measured by DEXA scan, solid-state MRI, and / or cone beam CT (CBCT).

11. The method of claim 10, wherein the subject is on an incretin-based therapy, wherein optionally the subject is experiencing rapid weight loss.

12. The method of claim 10 or claim 11 , wherein the subject has a T-score of at least -1 SD below the young adult mean prior to receiving the myostatin-selective inhibitor.

13. A method for prolonging an effect of an incretin-based therapy in a subject, the method comprising a step of administering to the subject with a diminished response to an incretin-based therapy, a myostatin-selective inhibitor in an amount effective to restore or increase responsiveness to the incretin-based therapy.

14. The method of claim 13, wherein the response to the incretin-based therapy comprises reduced fat mass.

15. The myostatin-selective inhibitor for use according to any one of claims 1-8, or the method according to any one of claims 9-14, wherein the myostatin-selective inhibitor is selected from: a neutralizing antibody that specifically binds mature myostatin thereby blocking ligand-receptor interactions; an antibody that specifically binds pro / latent myostatin thereby inhibiting activation of myostatin; and a nucleic acid-based agent that inhibits expression of myostatin, wherein optionally the myostatin-selective inhibitor is apitegromab, SRK-439, trevogrumab, GYM329, a variant thereof, or an antigen-binding fragment thereof.

16. The myostatin-selective inhibitor for use or the method according to any one of the preceding claims, wherein the incretin-based therapy is selected from a GLP-1 receptor agonist, a dual GLP-1 / GIP receptor agonist, a dual GLP-1 / glucagon receptor agonist, and a triple GLP-1 / GIP / Glucagon receptor agonist.

17. The myostatin-selective inhibitor for use or the method according to any one of the preceding claims, wherein the GLP-1 receptor agonist is semaglutide.

18. The myostatin-selective inhibitor for use or the method according to any one of the preceding claims, wherein the dual GLP-1 / GIP receptor agonist is tirzepatide.

19. The myostatin-selective inhibitor for use or the method according to any one of the preceding claims, wherein the dual GLP-1 / glucagon receptor agonist is selected from AZD9550, CT-388, CT-868, survodutide (BI456906), pemvidutide (ALT-801), NN1177, and efinopegdutide (MK-6024).

20. The myostatin-selective inhibitor for use or the method according to any one of the preceding claims, wherein the triple GLP-1 / GIP / Glucagon receptor agonist is selected from retatrutide and LY343794.21 . An incretin mimetic and a myostatin inhibitor for use in the treatment of obesity in a subject, wherein the treatment comprises administration of the incretin mimetic and the myostatin inhibitor to the subject in amounts effective to enhance body composition, wherein the subject is an adult male or female of 218 and 265 years of age and, a) has a BMI of 230.0 kg / m2to 245 kg / m2; or, b) has a BMI of 227.0 kg / m2to 230 kg / m2with the presence of one or more weight-related comorbidity; wherein the incretin mimetic is semaglutide or tirzepatide, and wherein the myostatin inhibitor is an antibody that competes or cross-competes with apitegromab or antibody comprising: an HCDR1 sequence of GFTFSSYGMH, an HCDR2 sequence of ISYDGSN, an HCDR3 sequence of DLLVRFLEWSHYYGMDV, an LCDR1 sequence of SSNIGSNT, an LCDR2 sequence of SDN, and an LCDR3 sequence of AAWDDSLNGV.

22. The incretin mimetic and a myostatin inhibitor for use according to claim 21 , wherein the antibody binds human pro / latent myostatin with a KD of less than 1 nM and inhibits mTLL2-induced activation of myostatin with an IC50 of less than 1 nM as determined by in vitro proteolytic assay followed by an ELISA-based immunoassay to detect free myostatin.

23. The incretin mimetic and a myostatin inhibitor for use according to claim 21 or claim 22, wherein the antibody is SRK-439, a variant thereof, or a derivative thereof.

24. The incretin mimetic and a myostatin inhibitor for use according to claim 23, wherein the antibody is SRK- 439.

25. The incretin mimetic and a myostatin inhibitor for use according to any one of claims 21-24, wherein the subject is a diabetic subject.

26. The incretin mimetic and a myostatin inhibitor for use according to any one of claims 21-25, wherein the subject is a pre-diabetic subject.

27. The incretin mimetic and a myostatin inhibitor for use according to any one of claims 21-26, wherein the subject is a non-diabetic subject.

28. The incretin mimetic and a myostatin inhibitor for use according to any one of claims 21-27, wherein the subject has low bone mineral density or is at risk of losing bone mass.

29. The incretin mimetic and a myostatin inhibitor for use according to claim 28, wherein the subject has a T- score of at least -1 SD below the young adult mean prior to receiving the myostatin-selective inhibitor.

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