Rutaecarpine Suppresses Osteosarcoma Progression and Tumor-Induced Bone Destruction by Targeting the PRAS40-associated Signaling Network in an Ex Vivo Metastasis Model

Despite advances in current therapeutic strategies, osteosarcoma remains a highly aggressive malignancy with limited treatment options. Rutaecarpine (Rut), an indolopyridoquinazolinone alkaloid with selective cyclooxygenase-2 inhibitory activity, has been reported to possess diverse pharmacological properties; however, its anti-osteosarcoma mechanisms remain poorly defined. Here, we examined the anti-tumor activity of Rut and the signaling pathways underlying its biological effects. Rut reduced MG63 cell viability in a concentration-dependent manner and promoted apoptotic cell death, as demonstrated by TUNEL staining and changes in apoptosis-related proteins. Rut also impaired cell-cycle progression and induced reactive oxygen species-mediated mitochondrial dysfunction, accompanied by modulation of the NRF2/HO-1 pathway and autophagic responses. Proteome profiling identified PRAS40 as a major signaling component associated with the cellular response to Rut. Consistent with this finding, Rut suppressed PRAS40 phosphorylation, leading to activation of GSK3β-associated signaling and subsequent inhibition of cell migration, invasion, extracellular matrix degradation, and anchorage-independent colony formation through reduced expression of MMP-2, MMP-9, and MMP-13. The biological relevance of these findings was further supported using a newly established ex vivo calvaria-osteosarcoma metastasis model, in which Rut attenuated tumor-associated bone destruction while preserving newly formed bone, collagen matrix organization, and the viability of osteoblasts and osteocytes along the periosteal surface. Overall, the present findings suggest that Rut suppresses osteosarcoma progression by modulating the PRAS40-associated signaling network and supports its potential as a therapeutic agent for osteosarcoma-associated bone destruction.