Acridine Orange is an Effective Anti-Cancer Drug that Affects Mitochondrial Function in Osteosarcoma Cells

Acridine orange (AO) is an antimalarial drug that accumulates into acidic cellular compartments. Lysosomes are quite acidic in cancer cells, and on this basis we have demonstrated that photoactivated AO is selectively toxic in sarcomas. However, photodynamic therapy is only locally effective, and cannot be used to eradicate systemic residual disease. In this study, we have evaluated the activity of non-photoactivated AO on sensitive and chemoresistant osteosarcoma (OS) cells to be considered for the systemic delivery. Since lysosomes are even more acidic in chemoresistant cells (MDR), we found that AO accumulation was significantly higher in the lysosomes of MDR in respect to parental cells, and in both cell types, therapeutic doses of AO significantly inhibited cell growth. However, the level of growth inhibition was inversely related to the level of lysosomal uptake of AO, suggesting that the main target of this agent is indeed extralysosomal. A significant reduction of intracellular ATP content and of the expression of mitochondrial complex III suggests a mitochondrial targeting. Notably, MDR cells showed a lower mitochondrial activity. Finally, the combined treatment of AO with the anticancer agent doxorubicin (DXR) significantly increased chemotoxicity by promoting DXR mitochondrial targeting, as revealed by the further reduction in ATP intracellular content. In conclusion, AO is able to effectively target both sensitive and resistant OS cells through mitotoxicity.