Hypoxia Induces Imatinib Resistance in CML Cell Lines.

Background: Imatinib is a potent tyrosine kinase inhibitor that is highly effective in the treatment of chronic myeloid leukemia. However, some patients are resistant to imatinib. Some of the mechanisms leading to imatinib resistance include amplification of the BCR-ABL gene, determining overexpression of the protein, and mutations in the BCR-ABL protein with alteration of imatinib binding sites. Imatinib uptake is an active process mediated by a group of transporters that includes the organic cation transporters (hOCT) and it has been shown that different expression of OCT1 may play a critical role on intracellular drug levels and, hence, resistance to imatinib. Hypoxia is another important factor that may contribute to drug resistance. Hypoxia-Inducible Factor (HIF-1α) and its downstream target, Vascular endothelial Growth Factor (VEGF), have been shown to be overexpressed in leukemic bone marrow specimens compared to normal bone marrow. The purpose of this study is to determine if the hypoxic conditions and OCT1 inhibition affect imatinib sensitivity.

Methods: Chronic myeloid leukemic cell line K562 and LAMA84 were cultured in 10% serum RPMI medium under hypoxic (3% O₂) or normoxic (21% O₂) conditions. All samples were treated with imatinib 1μM ± prazosin 13 μM (OCT1 reversible inhibitor) for 24 hs.

Results: Cells treated with imatinib and cultured under hypoxic conditions demonstrated decreased apoptosis and increased cell viability compared to normoxic conditions (K562 Annexin + cells 62% vs 94%, p= 0.003, LAMA84 Annexin + cells 61% vs 92%, p=0.0028). The addition of prazosin almost abrogated imatinib efficacy in normoxic environment but did not modify the effect of imatinib under hypoxic conditions.

Conclusions: Our data confirm that OCT1 is the most important imatinib carrier. Exposure of CML cell lines to an hypoxic environment results in reduced sensitivity to imatinib and this effect wasn’t affected by OCT1 inhibition. Search for underlying mechanisms of these findings are in progress.