Abstract 5596: Altered efficacy of tyrosine kinase inhibitors in chronic myeloid leukemia cells expressing wild type or polymorphic ABCG2

Abstract

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; therefore it would be advantageous to identify predictive markers for failure on imatinib in order to enable a change of therapeutic strategy in an early stage of CML. Imatinib is metabolized by hepatic CYP3A and we have previously reported that high CYP3A activity is associated with a better therapeutic response, indicating a clinical significance of imatinib metabolites (Gréen et al. 2010). Furthermore, imatinib and the second generation TKI's (dasatinib and nilotinib) are substrates for the efflux transporter ABCG2 which might be of importance for the elimination and resistance to TKIs. Several ABCG2 polymorphisms have been reported and some have been suggested to influence imatinib therapeutic outcome. In this study, we aimed to investigate the effects of expressing wild type (wt) or polymorphic ABCG2 on the in vitro resistance to imatinib, dasatinib, nilotinib, bosutinib (currently in clinical trials) and the most abundant CYP3A imatinib metabolite, CGP74588. The ABCG2 polymorphisms 34G>A, 421C>A, 623T>C, 886G>C, 1574T>G and 1582G>A were constructed from wt human ABCG2 cDNA. ABCG2 wt or variant cDNA was retrovirally infected to the CML cell line K562. Cell-surface ABCG2 expression was evaluated using FACS. In addition, K562/ABCG2/623T>C and K562/ABCG2/1574T>G were permeabilized to detect cytoplasmic ABCG2 protein. The influence of wt and variant ABCG2 on the cytotoxic effect of imatinib, CGP74588, dasatinib, nilotinib and bosutinib was assessed using MTT assays. Over-expression of ABCG2 wt in K562 cells conferred resistance to all TKI's except bosutinib. The most striking effect was seen in cells treated with CGP74588 where ABCG2 wt expression induced a 7.8-fold increase in resistance compared to a moderate 2-fold increase in cells treated with imatinib. ABCG2 polymorphisms affected the cytotoxicity of imatinib, CGP74588, dasatinib and nilotinib in a similar way and did not seem to alter substrate specificity of the transporter. Polymorphisms 623T>C, 886G>C and 1574T>G completely sensitized cells to the level of non-ABCG2 expressing cells. In accordance, FACS analysis revealed that 623T>C and 1574T>G caused a failure of ABCG2 protein to localize to the cell surface but was yet detected in the cytoplasm. We conclude that CGP74588 cytotoxicity is more affected by ABCG2 expression than imatinib itself, indicating that ABCG2 activity might affect CGP74588 to a greater extent than imatinib in terms of plasma and target cell concentrations in vivo. ABCG2 with the polymorphisms 623T>C and 1574T>G does not incorporate into the cell membrane, affecting not only TKI transport but most likely all ABCG2 substrates. 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 5596. doi:1538-7445.AM2012-5596