Most patients with acute myeloid leukemia (AML) achieve complete remission (CR) after induction chemotherapy, however, in some patients, the disease subsequently relapses and may lead to death. Leukemia stem cells (LSC) have been identified as the main cause for recurrence. Increased aldehyde dehydrogenase (ALDHhigh) activity in a variety of cancer stem cells prevents effective action of chemotherapeutic drugs. In this study, we found that approximately 50.7% of AML patients had ALDHhigh, and the presence of ALDHhigh stem cells was associated with poor cytogenetic prognosis. Lentiviral vector transduced ALDHhigh leukemia cell lines are insensitive to the conventional chemotherapy drug cytarabine, and inhibition of ALDH activity by disulfiram (DSF) can increase the sensitivity of ALDHhigh leukemia cells to cytarabine. Unlike traditional chemotherapy drugs, DSF is not toxic to healthy umbilical cord blood stem cells. An ALDHhigh leukemia cell xenograft model was established using immunodeficient mice to mimic the disease environment, and DSF and cytarabine were found to eliminate the ALDHhigh leukemia cells in transplanted mice while not affecting the healthy blood cells of mice. These findings suggest that DSF may have therapeutic potential by inhibiting ALDH activity and thereby increasing chemosensitivity.Keywords: AML; Aldehyde dehydrogenase; Cytarabine; Disulfiram; Leukemia stem cells.