Abstract 136: Evaluation of reversible BTK inhibitors in cell lines with clinically relevant Ibrutinib resistance mutations

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Abstract Background Ibrutinib, a small molecule inhibitor that covalently binds Cys481 of Bruton's Tyrosine Kinase (BTK), has recently been approved in Chronic Lymphocytic Leukemia (CLL) and Mantle Cell Lymphoma (MCL). Although the majority of patients respond to this drug, eventually resistance develops. Known causes for resistance include the outgrowth of cancer cells with mutations in either the drug binding pocket of BTK or activating mutations in the downstream effector, PLCγ2. Objective To explore mechanisms of resistance to ibrutinib and evaluate the potential of next generation BTK inhibitors on ibrutinib resistant cancer, we selected for resistance in cultured lymphoma cells. We chose TMD8, a cell line model of diffuse large B-cell lymphoma (DLBCL) that has constitutive B-cell receptor signaling, and REC-1, an ibrutinib sensitive MCL line, to model resistance. Methods TMD8 and REC-1 were cultured in increasing concentrations of ibrutinib to generate resistant cell lines. Genetic changes that may account for resistance were evaluated using whole exome sequencing. Resistant cell lines were evaluated for sensitivity to a small panel of BTK inhibitors. Results We generated a TMD8 derived cell line that is 1000-fold resistant to ibrutinib. Whole exome sequencing revealed a C481Y mutation in BTK. Reversible inhibitors that are not reliant on this site may function as alternative inhibitors of C481 mutant BTK. Ibrutinib-resistant TMD8 cells were tested for response to novel, reversible BTK inhibitors. The potency was similar to or modestly reduced (3 to 10 fold) on the ibrutinib-resistant cells compared to the parental line. In addition, we generated ibrutinib-resistant cells derived from the REC-1 line. Unlike resistance generated in TMD8, REC-1 resistant cells harbor a L527W mutation in BTK and are resistant to all BTK inhibitors that we evaluated. Conclusions C481Y and L527W BTK mutations have been identified in CLL patients that have relapsed after response to ibrutinib, and C481S mutations have been identified in ibrutinib relapsed CLL and MCL patients. Our data suggest that reversible, next generation BTK inhibitors have the potential to be efficacious on ibrutinib relapsed patients that harbor mutations at C481 of BTK. Citation Format: Regina Wai-Yan Choy, Luciana Burton, Wendy Young, James J. Crawford, Elicia Penuel, Lisa D. Belmont. Evaluation of reversible BTK inhibitors in cell lines with clinically relevant Ibrutinib resistance mutations. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 136. doi:10.1158/1538-7445.AM2015-136