RNAi Identifies Survivin as a Potential Target for Therapy In Pediatric Acute Lymphoblastic Leukemia

Pediatric Acute Lymphoblastic Leukemia (ALL) remains the most common pediatric malignancy. Despite advances in treatment and outcomes, there continue to be subsets of patients that are refractory to standard intensive chemotherapy and hematopoietic stem cell transplant. Therefore, novel gene targets for therapy are needed to further advance treatment for this disease. Survivin, a member of the chromosome passenger complex and inhibitor of apoptosis has been shown to be over-expressed in malignant cells and in relapsed disease. Therefore, survivin may be a potential target for therapy in pediatric ALL. Further, RNAi technology can be used as a rapid functional screen to identify target genes crucial for viability in ALL cell lines and in primary patient samples.

Pediatric lymphoblastic cell lines and fresh primary mononuclear cells from newly diagnosed patients with ALL were used for all studies. Survivin was silenced by transfecting cells (via electroporation or transductin (IDT)) with siRNA SMARTpools obtained from Dharmacon. Protein expression was identified 48 hours after treatment with siRNA. Viability was measured using a standard methane-thiosulfonate viability assay. Activation of apoptosis was identified using the Guava nexin Annexin V binding assay.

Survivin is highly expressed in multiple ALL cell lines and in many primary ALL samples. Silencing of survivin expression with siRNA decreases viability and increases apoptosis in ALL cell lines and in primary ALL samples. Decreased protein expression of approximately fifty percent was sufficient to cause an increase in cell death. Finally, inhibition of P53 expression abrogates this phenotype suggesting that the mechanism of cell death induced following survivin silencing involves activation of the P53 pathway.

Gene silencing with siRNA can be used as a rapid functional screen to identify potential targets for therapy. Implementing this technique identified survivin as a potential target for therapy in pediatric ALL. The results of these experiments will be used as a foundation to develop a comprehensive understanding of the mechanisms of survivin dependence in pediatric ALL. Future studies will also be designed to develop a model system for targeting of survivin in a therapeutically-relevant manner such that these findings can be translated into improved treatments for patients with ALL.

No relevant conflicts of interest to declare.