SUN-026 Selective Androgen Receptor Modulator S42 Has Anabolic and Anti-Catabolic Effects on Cultured Myotubes

Introduction: In an effort to develop a selective androgen receptor modulator (SARM) with beneficial activity in energy homeostasis, we previously screened 119 steroid analogs for the ability to induce uncoupling protein-1 (UCP-1) mRNA without increasing prostate-specific antigen promoter activity in human prostate cancer cell line, LNCap. As a result, we identified a novel SARM, S42, which is structural analog of testosterone. S42 dose not stimulate prostate growth but has a beneficial effect on lipid metabolism and also increased muscle weight of lavator ani in orchiectomized Sprague-Dawley rats. These findings prompted us to investigate whether S42 has a direct effect on cultured C2C12 myotubes. Method: Mouse myoblast C2C12 were seeded into 6well-plates at 2×10(5)cells/well and maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS). When the cultured cells became 80-90% confluent, the medium was changed to DMEM supplemented with 2% horse serum (HS) and cells were further cultured for 4 days to induce differentiation into myotubes. Next, the medium was changed to DMEM containing 2% dextran-coated-stripped HS from one day before treatment with various drugs: S42, 5α-dihydrotestosterone (DHT), insulin and rapamycin were tested. Protein and mRNA expression levels were detected by Western blotting and Real time RT-PCR, respectively. Results: Muscle atrophy is mainly controlled by two specific genes encoding muscle ubiquitin ligase, atrogin1 and Muscle RING-Finger Protein1 (MuRF1). S42 significantly lowered expression levels of atrogin1 and MuRF1 in C2C12 myotubes, as determined by Real time RT-PCR. Phosphorylation of p70 S6 kinase (p70S6K), an essential factor for promoting protein synthesis in skeletal muscle, was significantly increased by S42 to almost the same extent as by insulin, but this was prevented by treatment with rapamycin, an inhibitor of mechanistic target of rapamycin complex 1 (mTORC1). However, phosphorylation of Akt upstream regulator mTORC1, was not changed by S42. S42 did not increase insulin-like growth factor 1 (IGF1) mRNA levels in C2C12 myotubes. Conclusion: These results suggest that S42 may have an anabolic effect through activation of mTORC1-p70S6K signaling, independent of IGF1-Akt signaling and may exert an anti-catabolic effect through inhibition of the degradation pathway in cultured C2C12 myotubes.