Distinct Interaction of Nilotinib and Imatinib with P-Glycoprotein in Intracellular Accumulation and Cytotoxicity in CML Cell Line K562 Cells

Abstract

Nilotinib, a second-generation tyrosine kinase inhibitor (TKI), has been approved for first-line chronic myeloid leukemia (CML) treatment. The improved clinical response of nilotinib over that of the first generation TKI, imatinib, has been thought to be a result of its high potency of inhibition of BCR-ABL kinase. This study aimed to characterize differences between nilotinib and imatinib in the intracellular accumulation and cytotoxic effect on the CML cell line K562. Accumulation of nilotinib in K562 cells was from 4.7-to 9.0-fold higher than that of imatinib. The cytotoxic effect of nilotinib on K562 cells was 14.2-fold higher than that of imatinib. Inhibition experiments in K562 cells, and examination of the cellular uptake using influx transporter-transfected human embryonic kidney (HEK) 293 cells, suggested that the influx transporters OCT1 and OATP1A2, which have been reported to mediate accumulation of imatinib in CML cells, contributed little to the uptake of nilotinib. Nilotinib was found to accumulate in imatinib-resistant K562 (K562/ IM) cells overexpressing the efflux transporter P-glycoprotein (P-gp), although cytotoxic assays showed that K562/IM cells displayed 20000-fold greater resistance to nilotinib over the parent K562 cells. In conclusion, the present findings suggest that intracellular accumulation of nilotinib in CML cells contributes to its clinical response and efficacy in CML patients. Although nilotinib has been reported to be effective against imatinib-resistant ABL kinase mutants, the drug could not overcome imatinib resistance acquired by P-gpoverexpression. These results imply that classification of mechanisms of drug resistance is important for suitable strategies to treat imatinib-resistant CML patients.

Continuous Cytostatic Effects of BCR-ABL Tyrosine Kinase Inhibitors (TKIs) after Washout in Human Leukemic K562 Cells

“…These results are consistent with results of previous studies showing that nilotinib was weaker than imatinib or dasatinib as a substrate for MDR-1. 13,14,31,32) Our results also showed that nilotinib has a stronger and longer continuous cytostatic effect than that of dasatinib. Nilotinib has a longer half-life than that of dasatinib, and MDR-1 is thought to have a lower affinity than dasatinib as a substrate.…”

“…Effects of ABC transporters (MDR-1 (P-gp), BCRP) on the cytostatic effects of TKIs used clinically for CML therapy were observed in previous studies, but sufficient consideration was not given to clinically confirmed plasma concentrations and half-lives of drugs in most studies. 13,14,33) Our results showed that the experimental conditions of our study closely reflect clinical conditions and that dasatinib and nilotinib had sustained cytostatic effects after drug washout with short-term exposure. We also investigated how MDR-1 expression affects the continuous cytostatic effects of TKIs and we found that dasatinib had an attenuated inhibitory effect on cell proliferation due to decreased intracellular accumulation but that nilotinib did not.…”

“…Since previous studies showed that TKIs including imatinib, nilotinib and dasatinib were substrates of MDR-1, the therapeutic effects of TKIs are greatly affected by MDR-1 expression. [13][14][15][16][17] Indeed, our study showed that dasatinib completely suppressed proliferation of parent K562 cells after short-term exposure at a final concentration of 100 ng/mL, but did not suppress proliferation of hMDR-1-transfected cells (Fig. 4D).…”

“…Previous studies have also shown a relationship between therapeutic effect and MDR-1 overexpressions. [13][14][15][16][17] Therefore, it is also necessary to consider the possibility that MDR-1, which is responsible for tolerance of treatment with TKIs, affects maintenance of the therapeutic effect by administration every other day.…”

Selective Inhibition of Human Equilibrative and Concentrative Nucleoside Transporters by BCR-ABL Kinase Inhibitors

“…Organic cation transporter 1 and organic-anion transporting polypeptide 1A2 were shown to mediate imatinib uptake in chronic myelogenous leukemia cells (13)(14)(15). Nilotinib uptake was shown to be enhanced by organic cation transporter 1 but not by organicanion transporting polypeptide 1A2 (16). The cellular uptake of dasatinib was mainly passive and not dependent on organic cation transporters (9).…”