Despite the recent advances in chemotherapy for acute lymphoblastic leukemia (ALL), drug resistance resulting in relapse and long-term side effects of current treatments warrant new treatment modalities. Survivin/BIRC5, an inhibitor of apoptosis (IAP) protein, is critical for the survival and proliferation of cancerous cells and has become the target of an increasing number of preclinical novel therapies against primarily solid tumors. Survivin is expressed in AML and ALL cells and has been implicated in leukemia relapse. We test the hypothesis that survivin is critical to the pathway of self-renewal and maintenance of drug resistant ALL cells. To address this hypothesis, we have developed a murine xenograft model of patient-derived ALL cells, which are referred to here as primary ALL cells, allowing us to assess novel therapies targeting survivin using non-invasive monitoring of leukemogenesis by bioluminescent imaging. Our data suggest that overexpression of survivin increases self-renewal and drug-resistance of patient-derived ALL cells in vitro and accelerates leukemogenesis in vivo. In addition, in vitro inhibition of survivin using shRNA strongly synergizes with conventional chemotherapy in patient-derived ALL cells and decreases self-renewal. In vivo inhibition of survivin prolongs survival of mice engrafted with drug resistant leukemia. Taken together, we show that survivin is a key component in drug-resistance and stem cell self-renewal of drug resistant ALL cells.