Imatinib mesylate (IM) induces remission in chronic myelogenous leukemia (CML) patients but does not eliminate leukemia stem cells (LSCs), which remain a potential source of relapse. Here we investigated the ability of HDAC inhibitors (HDACis) to target CML stem cells. Treatment with HDACis combined with IM effectively induced apoptosis in quiescent CML progenitors resistant to elimination by IM alone, and eliminated CML stem cells capable of engrafting immunodeficient mice. In vivo administration of HDACis with IM markedly diminished LSCs in a transgenic mouse model of CML. The interaction of IM and HDACis inhibited genes regulating hematopoietic stem cell maintenance and survival. HDACi treatment represents an effective strategy to target LSCs in CML patients receiving tyrosine kinase inhibitors.