The signal transduction inhibitors, 2-methyl imidazolium iodides, are substrates for P-glycoprotein

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Calmodulin-mediated signal transduction is essential for normal physiology and is often abnormal in malignancy. Calmodulin-dependent kinase III, also known as elongation factor 2 kinase (eEF-2K), is overexpressed in many cancers. We recently reported that effective targeting of eEF-2K by a novel class of histidine kinase inhibitors (HKIs) decreased enzyme activity and cell viability at nanomolar concentrations (Arora et. al., Cancer Res. 2003). Multidrug resistance (MDR) mediated by P-glycoprotein (P-gp) is a deterrent to effective cancer treatment. Since the 2-methyl imidazolium derivatives possess structural features consistent with other P-gp interacting agents, we investigated whether or not these HKIs were transported by P-gp. We focussed this work on NH125 as a representative compound because we found it to be most potent against both histidine kinase and eEF-2K among the series. Sensitive and MDR MCF-7, A2780, and P388 cells were exposed to 0-100 μM NH125 for 48 hrs and cell viability assayed by MTT. P-gp (+) cell lines were 2-20 fold resistant to NH125 compared to P-gp (-) counterparts. The IC₅₀ of MCF-7/AdrR, A2780-Dx5 and P388/VMDRC.04 were 5μM, 8.1μM and 0.01μM respectively, whereas the IC₅₀ of the parental lines, MCF-7, A2780 and P388-S, were 2.5μM, 1.6μM and 0.005μM respectively. Incubation of MDR cells with the HKIs increased accumulation of paclitaxel and doxorubicin as a function of hydrophobicity, but these HKIs had no effect on sensitive lines or on the accumulation of a non-P-gp substrate, methotrexate. P-gp modulators, verapamil or trans-flupenthixol, re-sensitized MCF-7/AdrR and A2780-Dx5 cells to NH125. Prolonged exposure of P-gp (-) cells to NH125 induced P-gp expression and the MDR phenotype. Finally, NH125 increased survival of mice bearing P388/S, P-gp (-) leukemia but had no effect on the survival of P-gp (+) P388/VMDRC.04 bearing mice. In conclusion, HKIs represent a novel class of signal transduction inhibitors that are effective versus P-gp (-) cell lines and can enhance the sensitivity of MDR cells to other P-gp substrates.