Increased ABCG2 Expression Could Be Responsible for Resistance to Imatinib Mesylate in Patients with Chronic Myeloid Leukemia Who Do Not Have Mutations in BCR-ABL Kinase Domain

Mutation in the Bcr-Abl kinase domain is one of the most common mechanism of resistance to imatinib mesylate (IM), seen in 30– 90% of patients with chronic myeloid leukemia (CML). A total of 57 patients with CML, 41 in chronic phase, 10 in accelerated phase and 6 in blast crisis, were analyzed for bcr-abl kinase domain (KD) mutations using reverse transcriptase polymerase chain reaction (RT-PCR) amplification of the bcr-abl KD followed by direct sequencing. Twelve different mutations including three novel mutations were identified in the bcr-abl KD in 18 patients. Resistance to IM was considered if the patient had rising white blood cell counts or did not achieve hematological remission at 6 months; or had rising bcr-abl levels (as determined by fluorescence in situ hybridization or real-time quantitative PCR) after the start of IM treatment. In order to evaluate other potential mechanisms of resistance in patients without mutations in the bcr-abl kinase domain, we tested the expression levels of transporter genes known to be involved in IM influx and efflux. mRNA expression levels of efflux transporters, ABCG2 and ABCB1, and influx transporter, hOCT1 were measured by real time quantitative PCR using ABL to normalize the expression levels of the same. Serially diluted cDNA made from HepG2 RNA was used to make the standard curve and amplification efficiencies of the target and the house keeping genes were similar. Each sample was analyzed in duplicate and the experiment was repeated twice. In the patients without mutations in the bcr-abl KD, significantly higher ABCG2 mRNA levels were observed compared to patients with mutations (Table). Transcript levels of ABCB1, hOCT1 or bcr-abl were not significantly different between the two groups. This study suggests that over expression of ABCG2 may be one of the mechanisms of resistance to imatinib in patients without mutations in bcr-abl. Future studies should not only compare the expression of these transporters at diagnosis (before the start of IM treatment) but also at the time of clinical resistance. This will help understand the influence of expression levels of these transporters in achieving haematological or molecular response to increased IM doses. Bcr-abl mutation positive (n=18) Bcr-abl mutation negative (n=39) p value ABCG2: median (range) 0.0126 (0.0004–0.45) 0.051 (0.0041–0.51) 0.034 ABCB1: median (range) 0.1842 (0.00349–0.51) 0.1819 (0.0463–0.68) NS hOCT1: median (range) 488 (3.4–3394) 486 (62–3059) NS BCR-ABL: median (range) 53.96 (5.73–179.1) 51.3 (2.36–129) NS