Engineered mitochondria-derived vesicles suppress colorectal cancer through metabolic reprogramming and mtDNA-cGAS-STING signaling.

Metabolic reprogramming and innate immune signaling are closely involved in colorectal cancer (CRC) progression, but the effects of extracellular mitochondria-derived vesicles (MDVs) on these processes remain unclear, partly because of their low natural yield. In this study, we overexpressed MDV formation-related genes in mesenchymal stem cells and identified OPA1 and PINK1 as regulators that increased the recovery of extracellular MDVs. The resulting OPA1- and PINK1-MDV preparations were characterized and then incubated with LOVO and SW480 CRC cells to assess their uptake, metabolic effects, inflammatory signaling, and antitumor activities. OPA1-MDVs contained a TOMM20-positive/PDHE1α-positive vesicle population and shifted CRC cell metabolism away from glycolysis toward oxidative phosphorylation, as shown by reduced glycolytic enzyme activity and lactate production, together with increased oxygen consumption, respiratory-chain activity, and mitochondrial membrane potential. These changes were accompanied by reduced reactive oxygen species levels and impaired CRC cell proliferation, migration, and invasion. In contrast, PINK1-MDVs impaired mitochondrial function, increased reactive oxygen species production, and were associated with activation of cGAS-STING-related inflammatory signaling, possibly through the delivery of mitochondrial DNA. Both MDV preparations suppressed tumor growth in an immunocompetent syngeneic MC38 tumor model. These findings suggest that MDVs derived from OPA1- or PINK1-overexpressing cells suppress CRC growth through distinct metabolic and innate immune mechanisms and provide a potential strategy for engineering extracellular mitochondrial vesicles for cancer research.