<p>Background: One of the most significant problems in the treatment of leukemia is the expansion of resistance to chemotherapeutic agents. Therefore, assessing the drug resistance and especially the drug resistance genes of leukemic cells is important in any treatment. The impact of Mesenchymal Stem Cells (MSCs) and hypoxic condition have been observed in the biological performance of majority of leukemic cells.<br /> Methods: MOLT-4 cells were co-cultured with MSCs in the hypoxic condition induced by Cobalt Chloride (CoCl₂) for 6 and 24 <em>hr</em>. Then, apoptosis of cells was analyzed using annexin-V/PI staining and expression of the drug resistance genes including MDR1, MRP, and BCRP along with apoptotic and anti-apoptotic genes, including BAX and BCL2, was evaluated by real-time PCR.<br /> Results: The hypoxic condition for MOLT-4 cells co-cultured with MSCs could significantly increase the expression of MDR1 and BCRP genes (p&lt;0.05) which are involved in drug resistance. Also, the results indicated that this condition significantly increases the expression of BCL2 (p&lt;0.05) and reduces the apoptosis in MOLT- 4 cells co-cultured with MSCs in the hypoxic condition.<br /> Conclusions: These effects can demonstrate the important role of hypoxia and MSCs on the biological behavior of Acute Lymphoblastic Leukemia (ALL) cells that may lead to particular treatment outcomes.</p>