Directed evolution generates an oncolytic Sindbis virus with enhanced tumor-killing capacity in osteosarcoma

Osteosarcoma, a common cancer in adolescents, is characterized for its strong propensity for metastasis, which complicates treatment and results in a poor survival rate. Despite established therapies, the challenge remains to effectively target metastatic tumors. Oncolytic viruses offer a novel therapeutic approach by selectively infecting and destroying cancer cells while triggering antitumor immune responses. Sindbis virus (SINV), although effective in various tumor types, exhibits limited efficacy in osteosarcoma due to insufficient replication within these cells. This study aimed to improve therapeutic efficacy of SINV through directed evolution. By passaging the virus in osteosarcoma cell lines (HOS and U2OS), we developed an adaptive variant, designated SINV-P438L, with a mutation in nonstructural protein 2 (nsP2) that increased viral replication and transcription efficiency. SINV-P438L exhibited improved infectivity and cytotoxicity in osteosarcoma cells, inducing significantly activated apoptosis-associated signals compared with wild-type SINV. In animal models, SINV-P438L demonstrated superior antitumor effects in osteosarcoma and other tumor models, without causing detectable damage to normal tissues. These findings support SINV-P438L as a promising candidate for osteosarcoma treatment, with potential applications across additional cancer types.