Simple SummaryPancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers worldwide. Treatment of PDAC remains a major challenge. Recently, exosomes derived from mouse skin fibroblasts were modified to deliver siRNA to specifically target mutant KRAS. This approach resulted in disease suppression and increased overall survival in an animal model. This study aims to evaluate, in vitro, the use of human umbilical cord mesenchymal stromal cell (UC-MSC)-derived extracellular vesicles (EVs) to deliver molecules (siRNAs or drugs) to specifically target pancreatic cancer cells.AbstractPancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers worldwide. Treatment of PDAC remains a major challenge. This study aims to evaluate, in vitro, the use of human umbilical cord mesenchymal stromal cell (UC-MSC)-derived EVs to specifically target pancreatic cancer cells. EVs were isolated from the FBS-free supernatants of the cultured UC-MSCs by ultracentrifugation and characterized by several methods. EVs were loaded with scramble or KRASG12D-targeting siRNA by electroporation. The effects of control and loaded EVs on different cell types were evaluated by assessing cell proliferation, viability, apoptosis and migration. Later, the ability of EVs to function as a drug delivery system for doxorubicin (DOXO), a chemotherapeutic drug, was also evaluated. Loaded EVs exhibited different kinetic rates of uptake by three cell lines, namely, BxPC-3 cells (pancreatic cancer cell line expressing KRASwt), LS180 cells (colorectal cell line expressing KRASG12D) and PANC-1 cells (pancreatic cell line expressing KRASG12D). A significant decrease in the relative expression of the KRASG12Dgene after incubation with KRAS siRNA EVs was observed by real-time PCR. KRASG12DsiRNA EVs significantly reduced the proliferation, viability and migration of the KRASG12Dcell lines compared to scramble siRNA EVs. An endogenous EV production method was applied to obtain DOXO-loaded EVs. Briefly, UC-MSCs were treated with DOXO. After 24 h, UC-MSCs released DOXO-loaded EVs. DOXO-loaded EVs were rapidly taken up by PANC-1 cells and induced apoptotic cell death more efficiently than free DOXO. In conclusion, the use of UC-MSC-derived EVs as a drug delivery system for siRNAs or drugs could be a promising approach for the targeted treatment of PDAC.