PROJECT SUMMARY Chronic myelomonocytic leukemia (CMML) is an aggressive hematologic malignancy with extremely poor survival and no effective pharmacologic therapies. The CBL gene, which encodes an E3 ubiquitin ligase and signaling adaptor protein, is frequently mutated in CMML patients. We identified hyperactivation of the SRC family kinase LYN as a key oncogenic driver in CBL-mutant cells. Consistent with this finding, we have also demonstrated the in vitro and in vivo anti-proliferative effects of LYN inhibition by dasatinib in CBL-mutant cell lines and patient-derived CMML samples, providing rationale to explore the therapeutic potential of dasatinib in patients with CBL-mutant CMML. The development of drug resistance represents a possible challenge to the efficacy of dasatinib in CMML patients. Thus, an understanding of dasatinib-resistance mechanisms is essential for the design of durable therapeutic approaches in CBL-mutant CMML. Moreover, discovery of other targetable pathways in CBL-mutant disease will guide the selection of combination therapies that prevent the emergence of drug resistance. Indeed, we have observed that an inhibitor of SYK, fostamatinib, also has anti-proliferative activity against CBL-mutant cell lines, presenting an opportunity to test the efficacy of combined LYN/SYK inhibition. In Aim 1 of this proposal, I will assess dasatinib resistance and intracellular signaling in CBL-mutant cells expressing a series of LYN and SYK alleles, including wild-type, kinase-dead, and drug-binding mutants. I will also define genetic mechanisms of dasatinib resistance via a genome-wide CRISPR-Cas9 knockout screen in dasatinib- or vehicle-treated cells. In Aim 2, I will assess the biological effects of combined treatment with dasatinib and a panel of FDA-approved inhibitors of SYK in vitro and test the efficacy of dasatinib plus fostamatinib in patient-derived xenograft murine models of CBL-mutant CMML. Altogether, the experiments proposed here will build significantly on our previous work and lead to further progress in the discovery of effective treatments for CMML patients. The applicant, Dr. Roger Belizaire, is a clinical pathologist and laboratory-based investigator in the Department of Oncologic Pathology at the Dana-Farber Cancer Institute (DFCI). He spends 80% of his time in translational research and 20% in clinical practice as a transfusion medicine physician. He has proposed a five-year career development plan to enable the development of his independent laboratory at the DFCI. Dr. Belizaire has assembled an Advisory Committee of world-renowned experts to provide scientific and career mentorship. He has established collaborations with experts in myeloid neoplasia, oncogenic signaling, and targeted cancer therapeutics to provide experimental guidance and specific training in the field. Dr. Belizaire will conduct this research under the mentorship of Dr. Benjamin Ebert at the DFCI and leverage the exceptional research and teach...