TNF-related apoptosis inducing ligand (TRAIL) is a novel anticancer agent with selective apoptosis-inducing activity on cancer cells. However, many malignant tumors still remain unresponsive. Although cells can bypass apoptosis by different functions, the defect in the blocking role of second mitochondria-derived activator of caspase (Smac) on X-linked inhibitor of apoptosis protein (XIAP) is known to be an important hub for immortal characteristic of malignant cells. Actually, XIAP is known as an apoptosis inhibitor. To date, the sensitization of cancerous cells to TRAIL was successfully performed with different protocols, mainly through blocking XIAP with Smac administration. However, all these sensitization methodologies need to be performed prior to TRAIL administration on cancerous cells which in turn limit their practical application in clinics. Therefore, we hypothesized that concurrent expression of Smac and TRAIL on human adipose-derived mesenchymal stem cells (hA-MSC-ST) could both sensitize and destroy cancerous cells. To this aim, we generated hA-MSC-ST, secreting a novel cell penetrable form of Smac and a trimeric form of TRAIL. Indeed, the cell penetrable form of Smac obviates the need for any pretreatment of cancerous cells. Our data depicted that individual overexpression of TRAIL or Smac in different breast cancer cell types induced limited or no apoptosis, respectively. Conversely, their concomitant overexpression markedly increased cell death even for a resistant type of breast cancer cells, MCF-7. Notably, we observed no cytotoxicity of our methodology on normal cells. In summary, this is the first demonstration that a dual approach using simultaneous overexpression of a cell penetrable form of Smac and TRAIL sensitize and promote apoptotic process even in resistant breast cancer cells.