We have identified the first in vivo and in vitro small molecule inhibitor of Vav3, a signaling hub that is overex- pressed in many cancers and an activator of the small GTPase Rac. Rac is a major mediator of oncogenic and non-oncogenic addiction in cancer stem/progenitor cells. At low dose, our inhibitor eliminates TKI-resistance in vivo, prolongs the survival of a mouse model of pre-B-ALL, and eradicates cancer stem cell propagation in a model of mouse serial transplantation. It induces apoptosis of primary pediatric Philadelphia-positive (Ph+) and Ph-like B-ALL and chemotherapy-resistant RAM immunophenotype primary pediatric AML cells. It specifically targets leukemic cells while sparing normal hematopoiesis in vivo and shows no toxicity. In addition, it is active in oncogenic Ras xenografts mouse models of human solid tumors. Given this broad activity and the wide in- volvement of Vav3 and Rac in human disease, it is likely that our inhibitor will be efficacious in several human cancers resistant to current therapies. Even though we focus on Ph+ B-cell acute lymphoblastic leukemia (Ph+ B-ALL) as a simpler cancer model to validate the mechanism of action of our inhibitor, we will test its efficacy in models of pediatric TKI-resistant B- ALL and AML. Despite the introduction of ABL tyrosine kinase inhibitor (TKI) therapy and more recently highly- toxic immunotherapies, Ph+ B-ALL and AML remain poor prognosis diseases, especially in adults, as a result of frequent relapse and resistance to current therapies. The long-term goal of this grant application is a multidrug approach consisting of a TKI and our drug or an optimized derivative as a new therapy for ALL and AML without the toxicity associated with current salvage therapy approaches. We postulate that multitarget approaches in ALL and AML are necessary to prevent resistance to single-agent TKI therapy. Based on our preliminary data, we hypothesize that our drug increases death of leukemia initiating and propa- gating cells and overcomes TKI-resistance by targeting Vav3. The goal of the proposed research is to (1) validate the Vav3/Rac signaling axis as our drug?s target using biochemical and genetic approaches and determine in vivo implications; (2) to identify our drug?s binding site on Vav3 using biophysical, structural, and genetic ap- proaches, and validate the site using site-directed mutagenesis. Finally, (3) we will take advantage of the CCHMC Oncology Leukemia/Solid Tumor Repository to test our drug?s efficacy in PDX models of chemotherapy- resistant pediatric ALL and AML alone and in combination with existing TKI approaches and validate metallothi- onein as a biomarker. If successful, we would like to see our drug, or a more potent analog, move into pre-clinical safety analysis and potentially into a Phase I clinical trial in ALL and AML resistant to TKI therapies. Allosteric targeting of the Vav3 autoinhibited conformation as proposed here could be generalized to other ?undruggable? protein-protein interfaces.!
Philadelphia-positive (Ph+) acute lymphoblastic leukemia and acute myeloid leukemia remain poor prognosis diseases. One small molecule has been identified with in vivo activity on BCR- ABL B-ALL and on Ras-driven solid tumors. This project intends to validate the mechanism of action of our inhibitor in well-established murine models of primary BCR-ABL B-ALL and in pa- tient-derived xenografts of pediatric ALL and AML leukemias, thus paving the way for novel ALL and AML therapeutics that can also be translated to other cancers.
