Abstract 814: P-glycoprotein transport of the active imatinib metabolite, CGP74588, in chronic myeloid leukemia cells
Authors: Karin Skoglund, Samuel Boiso Moreno, Jan-Ingvar Jönsson, Henrik Gréen, Kourosh Lotfi
Journal: Cancer Research (2012)

Abstract The tyrosine kinase inhibitor (TKI) imatinib is, despite the introduction of second generation TKI's, the standard first-line therapy in chronic myeloid leukemia (CML). Early response to therapy is correlated to long-term therapeutic effect in CML. To be able to switch to second generation TKI's in an early stage of CML, it would be advantageous to identify predictive markers for failure on imatinib. Imatinib is metabolized by hepatic CYP3A4 and CYP3A5, forming approximately 30 metabolites. The main metabolite, CGP74588, is pharmacologically active with similar potency to that of imatinib. CGP74588 is present in ∼20% of imatinib plasma concentrations but with a large inter-individual variation. We have previously reported that high CYP3A activity is associated with a better therapeutic outcome of imatinib, indicating a clinical significance of imatinib metabolites (Gréen et al. 2010). Furthermore, imatinib is a substrate for the efflux transporter P-glycoprotein which is the product of the ABCB1 gene. Several ABCB1 polymorphisms have been described and some have been shown to influence the therapeutic response to imatinib in CML patients. The aim of this study was to investigate the effects of ABCB1 over-expression on the in vitro resistance to imatinib and its CYP3A metabolite CGP74588. ABCB1 wild type human cDNA was infected to the CML cell line K562 using a retroviral system. Co-expression of ABCB1 and the reporter gene for enhanced yellow fluorescent protein (EYFP) was used for the assessment of ABCB1 expression by the analysis of EYFP in FACS. The influence of ABCB1 expression on the cytotoxic effects of imatinib and CGP74588 was assessed using MTT assays. FACS analysis of EYFP confirmed an over-expression of ABCB1 in infected K562 cells with a mean fluorescence intensity of 23.5 compared to 0.55 in parental K562 cells. ABCB1-expressing cells (K562/ABCB1) were slightly, but not significantly, more resistance to imatinib than K562 cells (IC50 K562 = 0.41µM, K562/ABCB1 = 0.50µM). However, ABCB1 expression induced a 12-fold increase in resistance when cells were treated with the imatinib metabolite CGP74588 (IC50 K562 = 0.72µM, K562/ABCB1 = 8.53µM, p = &amp;lt;0.000). Recent reports have shown that CGP74588 accumulates in cell lines with acquired multi-drug resistance and high P-gp expression. Our studies on cells with ABCB1 expression as a single resistance mechanism proves that P-gp is indeed confer resistance to CGP74588 in CML cells. Furthermore, CGP74588 cytotoxicity is affected by ABCB1 expression to a greater extent than imatinib itself, indicating that ABCB1 activity could be important for CGP74588 plasma and target cell concentrations in vivo and might contribute to the large intra-individual variation seen in patient plasma concentrations. Future studies will be needed to determine the clinical significance of P-gp activity and CGP74588 pharmacokinetics in relation to TKI therapy of CML. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 814. doi:1538-7445.AM2012-814