Abstract 3538: An atomic absorption spectroscopy assay (AASA) as a surrogate method to measure imatinib (IM) uptake via the organic cation transporter 1 (OCT1)
Authors: Ringel Frauke , Schwarz Michaela , Dörken Bernd , le Coutre Philipp 
Published: February 2011

Background. In chronic myeloid leukaemia (CML) 15 to 30% of patients (pts) treated with the ABL specific tyrosine kinase inhibitor IM may develop resistance. In these pts clonal evolution, amplification of the BCR-ABL gene or kinase domain mutations have been identified as underlying mechanisms. Additionally, pharmacokinetic alterations leading to reduced intracellular imatinib levels may also exist. IM is transported into the cell via the OCT1. The activity of this transporter could play a pivotal role for the intracellular IM concentration. Until now, uptake efficacy of IM is primarily measured by using radioactive labelled IM. Due to the limitations of these methods, we tried to establish alternative methods to measure the activity of OCT1 using cellular OCT1 dependent oxaliplatin (OX) uptake as a surrogate.

Materials & Methods. To quantitatively analyse OX uptake in human cell lines an AASA was established. Conversely, radioactive IM uptake was measured by scintillation analysis. We compared the uptake of OX and IM in different human cell lines, including JURKAT, Molt-4, K-562, KU-812, Lama-87, KCL-22 and OCI-AML-2. Ten different transport inhibitors were used to block IM or OX uptake, respectively. Furthermore, we investigated the expression levels of different influx (OCT1, OCT2, OCT3) and efflux transporters (ABCB1, ABCC1, ABCG2) in those cell lines.

Results. IM and OX both were taken up linearly into cells within a range from 0,4 to 2 µM (IM) or 1 to 10 µM (OX). 150 mM verapamil and 100 µM prazosin could effectively inhibit cellular uptake of both drugs (21%±8% and 27%±8%). However, a differential uptake pattern between OX and IM was observed in relation to the different cell lines. In contrast, we could show that 15 µM citalopram inhibited the transport of 5 µM OX in a similar type as the uptake of 1-2 µM IM into all cell lines tested yet (1,14±0,15*). In this regard citalopram may be most useful for further studies on OCT1 activity. Using real-time PCR, an adequate expression of OCT1 mRNA could be detected in all cell lines tested, albeit the levels differed significantly from 150 to 27.000 fold less than in the internal control (ß-actin). However, the mRNA expression level of OCT1 does not seem to be a predictor for IM or OX uptake. The expression pattern of the efflux transporters showed no significant distribution, too.

Conclusions. Here, we demonstrate the feasibility and accuracy of AASA testing of oxaliplatin as a surrogate marker for IM uptake via OCT1. As this methodology is secure, fast and sensitive, this approach may substitute conventional testing of OCT1 activity by radio labelled IM. On a clinical level, this procedure could be helpful as a clinical tool to test IM pharmacokinetics in CML patients. However, patient sample derived data will be needed to further evaluate this approach.

* 1 would stand for 100% equality of uptake of imatinib or oxaliplatin

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3538.