Background:

In recent years, numerous efforts have been dedicated to reducing the side effects of doxorubicin (DOX). Exosomes (EXOs), as extracellular vesicles (EVs), can play a role in the safe transport of DOX in breast cancer treatment. The aim of this study was to alleviate the adverse effects associated with DOX while enhancing its targeted delivery to cancer cells through the codelivery of melatonin (MEL) as an antioxidant and DOX into EXOs-derived from human adipose tissue mesenchymal stem cells (A-MSCs). Methods MSCs were isolated from liposuction samples using collagenase II enzyme, and stemness markers were evaluated by flow cytometry. EXOs were extracted from conditioned A-MSCs media through ultracentrifugation, and surface markers were evaluated by western blotting, DLS and TEM. The absorption and release of EXOs in cells were investigated using PKH-26 dye and UV–Vis spectrophotometry, respectively. DOX and MEL were loaded into EXOs using the sonication method, and their cytotoxic effects on normal and cancer cells were evaluated using the MTT test. Additionally, the expression of p53, NANOG, and miR-34a genes was analyzed using qRT-PCR, and apoptosis was assessed using flow cytometry and acridine/orange dye. Results It was observed that they exhibited remarkable stability under pH ~ 7.4 while displaying a high release rate under low pH conditions commonly found within cancerous environments (pH ~ 5.0). Cellular uptake experiments revealed a substantial percentage of internalization. Cytotoxicity evaluation demonstrated that co-delivery of DOX and MEL into EXOs (Exo-DOX-MEL) enhanced their toxicity towards normal MCF-10A and A-MSC cells, while exhibiting greater lethality towards MCF-7 and MDA-MB231 cancer cells. In normal cells, Exo-DOX-MEL augmented the effects of DOX, leading to increased expression of p53 and miR-34a and decreased expression of NANOG, particularly in MCF-7 and MDA-MB231 cells. Apoptotic analysis validated the favorable outcomes associated with Exo-DOX-MEL, which enhanced DOX efficacy in cancer cells while reducing apoptosis in normal cells compared to the administration of free DOX. Conclusions Exo-DOX-MEL appears to enhance the destructive effects of DOX in cancer cells, particularly those resistant to chemotherapy such as MDA-MB231 cells. It also plays a protective role in normal cells, which could be crucial in the treatment of drug resistance and the side effects caused by DOX.