2579-P: Temporal Mechanistic Shifts Underlying Sustained Muscle Growth by HM17321 Revealed by Tissue Proteome Analysis

Introduction and Objective: HM17321, a novel CRFR2-selective UCN2 analog, has demonstrated dual effects of fat reduction and lean gain in preclinical models. This study aims to characterize the temporal mechanistic transition underlying muscle augmentation by comparing skeletal muscle proteomes in early-phase (2-weeks) and sustained-phase (4-weeks) of HM17321 administration. Methods: DIO mice were administered either vehicle or HM17321 (30 or 100 nmol/kg) for 2 or 4 weeks. Global and phospho-proteomic data of tibialis anterior muscle were acquired using DIA-based LC-MS/MS. Protein and pathway-level analyses were performed using Limma and Gene Set Variation Analysis (GSVA), respectively. Effect sizes (Hedges’ g; DIO vs. HM17321) were then calculated from the resulting GSVA scores. To identify temporal shifts, these effect sizes were compared between the two timepoints. Results: HM17321 upregulated muscle growth signals with distinct temporal patterns. Sarcomere organization peaked at 2 weeks, characterized by structural protein enrichment, indicating early activation of assembly programs. In contrast, thick filament production was more prominent at 4 weeks, while thin filament expression remained comparable at both timepoints. Regarding mTOR signaling, most downstream steps were more enriched at 2 weeks than 4 weeks, whereas S6K activity reached its peak at 4 weeks. This 4-week S6K increase coincided with the dose-dependent IRS-1 phosphorylation, suggesting S6K-mediated negative feedback. However, stable mTOR phosphorylation suggests CRFR2-associated signaling may contribute to sustain muscle augmentation, overcoming potential feedback inhibition. Conclusion: HM17321 induces sustained muscle augmentation via early enrichment of sarcomere assembly and later-phase stabilized hypertrophic signaling. By maintaining the mTOR-mediated muscle growth, HM17321 ensures sustained weight loss while improving muscular mass and function, highlighting its potential to protect musculoskeletal health in obesity management. Disclosure B. Kim: Employee; Current; Hanmi Pharm. Co., Ltd. Y. Na: Research Support; Current; Hanmi Pharm. Co., Ltd. H. Kwon: Employee; Current; Hanmi Pharm. Co., Ltd. S. Lee: None. J.H. Kim: None. Y. Kim: None. H. Chon: Employee; Current; Hanmi Pharm. Co., Ltd. H. Kim: Research Support; Current; Hanmi Pharm. Co., Ltd. I. Choi: None.