Low Baseline Intrinsic Sensitivity To Imatinib (High Ic50) In Cml Patients Is Due To Reduced Oct-1 Mediated Influx. Intrinsic Sensitivity To Amn107 Does Not Correlate With That Of Imatinib And Uptake Of Amn107 Is Not Oct-1 Mediated.

Abstract We have recently demonstrated that there is variation in the intrinsic sensitivity of de novo CML patients to imatinib. Patients with low intrinsic sensitivity to imatinib, have a lower probability of achieving major molecular response by 12 months. Intrinsic sensitivity was determined pre-therapy using western blot, with the IC50imatinib defined as the dose of drug required in vitro to reduce phosphorylation of the adaptor protein Crkl (p-Crkl) by 50%. In the current study we show that the observed variation in IC50imatinib between patients can be explained by the intracellular concentration of imatinib. In 16 de novo CML patients the IC50imatinib strongly correlated (R2 = −0.6, p = 0.002) with the intracellular concentration of 14C-labelled imatinib after 2 hours exposure at 37°C. In keeping with the notion that the organic cation transporter Oct-1 is responsible for imatinib influx, the addition of Prazosin, an inhibitor of Oct-1, to 14C imatinib experiments greatly reduced the inter-patient variation in intracellular concentration. Intracellular drug concentration. Median IC50imatinib uM [imatinib] in-vitro * [imatinib] in-vitro* + Prazosin Median IC50AMN107 uM [AMN107] in-vitro * [AMN107] in-vitro* + Prazosin * ng of intracellular imatinib or AMN107 per 200000 cells when 2uM drug added in vitro. low IC50 imatinib (n=8) 0.5 28.4 6.9 0.035 28.6 27.4 p<0.001 p>0.05 high IC50 imatinib (n=8) 1.375 14.8 8.4 0.04 28.5 27.6 p value <0.001 0.04 >0.05 >0.05 >0.05 >0.05 p=0.03 p>0.05 We have now demonstrated that IC50 values for the novel, more selective Bcr-Abl inhibitor AMN107 are on average 22-fold lower than those observed for imatinib and there is no correlation between the IC50 values for imatinib and AMN107 (R2 = 0.375, p>0.05). There was also less inter-patient variation in AMN107 IC50 (range 0.07uM) when compared to imatinib (range 3.1uM). In contrast to imatinib, there is no correlation between IC50AMN107 and the intracellular concentration of drug achieved in 14C-uptake studies (R2 = 0.337, p>0.05). Further to this the addition of prazosin had no effect on influx of AMN107 suggesting that Oct-1 may not be a rate limiting determinant of intracellular drug concentration. In conclusion, variable Oct-1 mediated influx of imatinib may explain the heterogeneous IC50imatinib values we have observed in de novo CML patients. Oct-1 mediated influx may therefore be a key determinant of molecular response to imatinib. The causes of inter-patient variability in Oct-1 mediated influx requires further investigation. By contrast, poor Oct-1 mediated uptake is unlikely to have any impact on response to AMN107. This may be of relevance to patients with low intrinsic sensitivity to imatinib.