Anti-proliferative effect of a novel purine-template based dual Src/Abl tyrosine kinase inhibitor AP23464 on native and Imatinib-resistant Bcr-Abl cell lines

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Bcr-Abl is a dysregulated tyrosine kinase that causes chronic myeloid leukemia and is targeted by the signal transduction inhibitor Imatinib mesylate (IM). Imatinib is an effective treatment choice for Bcr-Abl-positive malignancies. However, resistance toward IM develops frequently in advanced-stage (Blast crisis), and is even observed in chronic-phase of leukemia. Point mutations within the ABL kinase and SH3 domain emerged as a major mechanism of resistance toward imatinib. Frequently, mutations occur at positions that determine specific contacts of imatinib to the ATP-binding site. However, an interesting class of mutants appears to act allosterically by influencing the confirmation of the kinase domain. These mutants alter the “breathing” of the N-lobe of the kinase domain relative to the C-lobe. Emergence of resistance to IM warrants the exploration of novel well-tolerated anticancer agents. We evaluated a novel purine template-based dual Src/Abl kinase inhibitor AP23464 (ARIAD Pharmaceuticals) for their potential to overcome resistance in various imatinib-resistant cell lines. This compound effectively suppressed the activity of 30 different kinase domain mutants resistant to IM. The common P-loop mutants are sensitive to AP23464, and only the L267R (type Ib) remains resistant. Interestingly, the most frequent and robust mutant T334I (type Ib) was effectively suppressed; whereas, F336L (type Ib) is 30 fold resistant over native Bcr-Abl. Surprisingly, eight of the 30 kinase domain mutants (D295V, E300K, V308S, E311Q, Q319H, T334S, G340W and M370T; type Ib) were highly sensitive as compared to the native Bcr-Abl. These observations led us to conclude that AP23464 is an effective compound against most of the frequently observed mutations of Bcr-Abl in connection with IM resistance. Understanding the underlying molecular mechanisms involved in drug sensitivity is important to exploiting this novel Src/Abl kinase inhibitor and its analogs. Moreover, combination studies of AP23464 and other drugs may also be a promising therapeutic strategy to overcome IM resistance. Our data support the development of AP23464 for the treatment of CML and provide impetus to test AP23464 in combination with other drugs relative to further understanding the molecular mechanisms involved in Bcr-Abl-positive malignancies.