Treatment of Bcr/Abl caused leukemias with FTIs

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DESCRIPTION (provided by applicant): Despite substantial advances in the treatment of acute lymphoblastic leukemia (ALL), the rate of long-term survival has reached a plateau, at approximately 80 percent for children and 55 percent for adults. We developed a transgenic mouse model for a subtype of ALL caused by a fusion between the BCR and ABL genes and can treat the leukemic cells in co-culture with a fibroblast feeder layer, to model the in vivo circumstances of drug treatment and emerging drug resistance. How BCR/ABL leukemic cells precisely differ from their corresponding normal non-malignant counterparts is a matter of continued intense investigation. Protein glycosylation takes place on a massive scale but has been largely neglected because of the difficulty in studying and identifying these often exceedingly complex side chains. However, all cells are covered by a dense network of carbohydrates that are attached to proteins, lipids and other carbohydrates- the so-called glycocalx- and the presence, absence or type of glycosylation can have a tremendous impact on biological function. Our preliminary data show that expression of many "glyco-genes" differs significantly between drug-resistant BCR/ABL lymphoblasts and non-resistant counterparts, and between non-leukemic and leukemic pro-B cells. We hypothesize, that BCR/ABL lymphoblasts and drug-resistant BCR/ABL lymphoblasts differ from their non-leukemic normal and drug-sensitive counterparts, respectively, in the glycocalyx that covers their cell surface, resulting in abnormalities in adhesive interactions. To test this, we propose to analyze our own microarray data for "glyco-genes" and to validate the differential expression of Clec4d, Galectin-3, St3gal1, St8sia4 and HexA and other genes forthcoming from this analysis. We will also investigate the expression of 9-O-acetylation and of sialyl-Lex during the development of leukemia in BCR/ABL transgenic mice and during the evolution of drug resistance in vitro co-culture of cells treated with the tyrosine kinase inhibitor nilotinib. Finally, we will compare the sugars and lectins expressed on the surface of normal pro-B cells with their leukemic counterparts, by using glycome and lectin arrays on flow-sorted AA41+, CD19+ cells from wild type, preleukemic and fully leukemic BCR/ABL transgenic mice and on leukemic cells co-cultured with stroma before, during and after the induction of drug resistance in vitro. Results will be correlated with glycogene expression. These experiments will provide important information on the types of carbohydrates and carbohydrate-binding proteins that are specifically expressed on BCR/ABL ALL cells and if these change during drug treatment. In addition to providing new tools for diagnosis, these experiments may yield very novel targets for drug treatment within a class of biomolecules that have not been studied at all in the field of Ph-positive ALL. Public Health Relevance: Although over 80% of adults who develop acute lymphoblastic leukemia can achieve a complete remission, the majority of such patients relapse. The bone marrow is the place where the development of normal blood cells takes place and also the location where the residual leukemia cells hide, but our understanding of how the surrounding cells promote and control the development of normal blood cell development is incomplete. We here propose to investigate how the normal development of specific blood cells -the B-lineage- occurs in the bone marrow to better understand how leukemia cells are helped by these cells in withstanding drug treatment