Enhancement of Imatinib-Induced Apoptosis through Catalase Activity Modulation in Imatinib-Resistant Chronic Myelogenous Leukemia Blastic Phase Cells.

Therapeutic selectivity and drug resistance are two major issues in cancer chemotherapy. Strategies to improve therapeutic selectivity largely rely on our understanding of the biological difference between cancer and normal cells, and on the availability of therapeutic agents that target biological events critical for cancer cells but not essential for normal cells. Imatinib is a selective inhibitor of the BCR-ABL tyrosine kinase activity in chronic myelogenous leukemia (CML). Resistance to imatinib is a major problem in CML patients especially in blast phase (BP). Recently, it was also described that reactive oxygen species (ROS) is another mechanism through which imatinib induces apoptosis. To evaluate the possible role of catalase (a well known ROS enzyme scavenger) in imatinib resistant cells, we studied through apoptosis/necrosis analysis, K562 cells in presence or absence of 3-amino-1,2,4-triazole (AT), an inhibitor of catalase activity. Cell apoptosis index was tested in cells labeled with Annexin V-FITC/PI and analysed by flow cytometry. In order to examine the efficacy of imatinib apoptosis induction we evaluated firstly the apoptosis index in K562 cells after 24 hours of incubation with imatinib at the less cytotoxic concentrations of 0.5, 1.0 and 5.0 μM. Induced imatinib early apoptosis, was significantly higher, (25%, p < 0.05) than control in all concentrations tested. After that, we conducted the same assay in K562 cells previously treated with aminotriazole (AT-K562) and, to address the question of whether the resistance to imatinib could also be counteracted by glutathione modulation, another ROS scavenger, K562 cells previously treated with buthionine S,R-sulphoximine (BSO) a specific inhibitor of g-glutamylcysteine synthetase the rate limiting enzyme in glutathione synthesis, were also analysed. Untreated K562 cells and those previously treated with AT or BSO were incubated with imatinib (0.5, 1.0 and 5.0 μM) and hydrogen peroxide (H₂O₂) 1.0 μM, a well known apoptosis inductor inhibited by catalase. Only AT-K562 cells had their apoptosis significantly enhanced when compared with non treated cells (p < 0.05). Indeed, we showed that inhibition of catalase activity in K562 cells overcame imatinib resistance in 35% of these cells in the final concentration of 0.5 μM which is equivalent to the half mean plasma concentration used in clinical practice. The present findings suggest that combination of catalase inhibitors with imatinib in lower dosage represent an emerging therapeutic concept for preventing or overcoming resistance formation in CML. Therefore, further studies with drugs clinically available that inhibit catalase activity should be explored.