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Pharmacological and genetic studies over the past decade suggest that FSH is an actionable target for diseases affecting millions, notably osteoporosis, obesity and Alzheimer's disease (AD). Blocking FSH action prevents bone loss (1, 2), fat and energy metabolism (3) and AD–like features in mice (4). We recently developed a first–in–class, humanized, epitope–specific FSH blocking antibody that binds to a 13–amino–acid–long sequence of FSHβ—"MS-Hu6"—with a KD of 7.52 nM (5). We showed that MS-Hu6 bound specifically to FSHβ and its different glycosylated forms, namely FSHβ21/18 and FSHβ24, without binding to LH and TSH. Here, using a GLP–compliant platform, we report the efficacy of MS-Hu6 in preventing obesity, osteoporosis and AD in mice. Notably, MS-Hu6-treated mice showed lower body weight and fat mass, increased lean mass (qNMR) and evidence of beiging in ThermoMice (IVIS imaging) compared with IgG–treated mice. Consistent with this, the thermogenic genes Ucp1 and Cidea were upregulated, whereas Pparg expression was attenuated in fat depots. Treatment of ThermoMice for 8 weeks also increased bone mineral density (BMD), improved microstructure (micro-CT), elevated bone formation (dynamic histomorphometry), and upregulated the osteoblastic genes Alp and Col1a1. The increase in bone mass and improved microstructure were replicated in C.J.R's lab using female mice 24 weeks post–ovariectomy. Preliminary testing using AD mice, namely APP/PS1 mice, showed that MS-Hu6 prevented the impairment in recognition and contextual memory. Biodistribution studies using 89Zr–labelled, biotinylated or unconjugated MS-Hu6 in mice and monkeys showed localization to bone, bone marrow and fat depots. MS-Hu6 displayed a β phase t½ of 13 days (316 hours) in humanized Tg32 mice, and bound endogenous FSH. In monkeys, an acute single injection of MS-Hu6 did not affect vitals, and biochemical parameters remained within the normative range. We tested 215 variations of excipients using the protein thermal shift assay to generate a final formulation that rendered MS-Hu6 stable in solution upon freeze–thaw and at different temperatures, with minimal aggregation, and without self–, cross–, or hydrophobic interactions or appreciable binding to relevant human antigens. MS-Hu6 showed the same level of "humanness" as human IgG1 in silico, and was non–immunogenic in ELISPOT assays for IL-2 and IFNγ in human peripheral blood mononuclear cell cultures. In conclusion, MS-Hu6 is efficacious, durable and manufacturable, and is therefore poised for future human testing as a multipurpose therapeutic for obesity, osteoporosis, and perhaps for AD.References: 1Sun et al., Cell, 2006, PMID: 16630814; Ji et al, PNAS, 2018, PMID: 29440419; 3Liu et al., Nature, 2017, PMID: 28538730; 4Xiong et al., Nature (In press); 5Gera et al., PNAS, 2020, PMID: 33127753 Presentation: Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.

Menopause is associated with the loss of LH ovulatory surges and enhanced visceral adiposity. Visceral fat depots increase from 5-8% at premenopause to 15-20% of total body fat at postmenopause. Here, we report that high-dose LH, hCG, or small molecule LH/CGR agonist ORG43553 injected twice-a-week into 14-weeks-old C57BL/6 male mice protects them from diet-induced obesity. Testosterone levels were elevated in mice treated with LH or hCG, but not with ORG43553. Notably, the anti-obesity action of LH/hCG is independent of testosterone, as blocking the androgen receptor using flutamide yielded similar results. Importantly, male Lhcgr knockout mice on a high-fat diet treated with LH failed to display a reduction in adiposity, confirming the in vivo specificity of action. Furthermore, our data phenocopied Lhcgr haploinsufficiency in mice. We confirmed the presence of Lhcgr in mouse genital and inguinal fat pads, adipose-derived stromal vascular cells, as well as in differentiated and undifferentiated 3T3-L1 murine adipocytes by qPCR, RNAscope in situ hybridization, and immunohistochemistry. Sanger sequencing showed that the extracellular domain of Lhcgr in genital fat depot was identical to the ovarian receptor. Similarly, we identified LHCGR in human subcutaneous and visceral fat depots. Binding of intraperitoneally injected AlexaFluor-488-labeled hCG was found not only in mouse ovary, but also in genital and subcutaneous fat pad, further confirming the presence of LHCGR in adipose tissue. This binding could be competitively displaced in 3T3-L1 cells using unlabeled hCG. LH, hCG and ORG43553 activated ERK1/2 in a dose-dependent manner in undifferentiated and differentiated 3T3-L1 cells, suggesting that the adipose LHCGR is fully functional. LH, hCG, and ORG43553 reduced adipogenic differentiation in 3T3-L1 cells, which is further confirmed by RNA sequencing. Moreover, we observed, that LH and hCG also alters several aspects of immune response in adipose tissue, including inflammatory response and adaptive immunity. In conclusion, we demonstrated that LH/CG receptors are present and fully functional in adipose tissue, and that high-dose intermittent activation of LHCGR in mouse fat depots protects mice from diet-induced obesity and modifies adipose tissue immune response. Presentation: Saturday, June 11, 2022 1:42 p.m. - 1:47 p.m., Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.