3.5 Effects of ningetinib and gefitinib on M1 mediated by efflux transporters.
M1 exhibited a longer plasma elimination half-life than the parent drug in ICR mice and in NSCLC patients, possibly because the efflux pathway of M1 was likely to be blocked. The influence of efflux transporters on the disposition of ningetinib and M1 was further studied.
To determine whether ningetinib or M1 was a substrate of efflux transporters, the MDCKII cells stable-overexpressed with human P-gp (MDCKII-MDR1), BCRP (MDCKII-BCRP) or MRP2 (MDCKII-MRP2) were used for substrate screening (Table 3). The ER values of ningetinib/M1 were all greater than 2 apart from that of ningetinib in MDCKII-MRP2 cells. Then the addition of GF120918 (P-gp inhibitor), KO143 (BCRP inhibitor) and TAK875 (MRP2 inhibitor) markedly reduced the ER values of M1 in MDCKII-MDR1, MDCKII-BCRP and MDCKII-MRP2 cells, respectively, while ningetinib displayed a similar result except for that in MDCKII-MRP2 cells. Thus, both ningetinib and M1 were substrates of P-gp and BCRP. MRP2 mediated the efflux of M1 but not of ningetinib.
The inhibitory effect of ningetinib and gefitinib on P-gp, BCRP and MRP2-mediated M1 efflux was further investigated. The logarithm concentration of ningetinib or gefitinib was plotted on the abscissa, and the remaining activity of P-gp, BCRP or MRP2 (percentage of efflux ratio in the inhibitor group to that in the control group) was plotted on the ordinate, as shown in Fig. 6. Ningetinib displayed a concentration-dependent inhibition on P-gp-, BCRP- and MRP2-mediated M1 efflux with IC50 values of 0.413 (0–100 μM), 18.7 μM (0–100 μM) and 3.06 μM (0–30.0 μM), respectively. Similarly, gefitinib exhibited a weak concentration-dependent inhibition on P-gp- and BCRP-mediated M1 efflux with IC50 of 5.40 (0–100 μM) and 9.09 μM (0–30.0 μM), respectively, but showed no inhibition on MRP2-mediated M1 efflux (0-100 μM). These results, especially the P-gp inhibition phenomenon, indicated that the parent drug ningetinib could potently inhibit the canalicular efflux of its metabolite M1 and thereby elevate its plasma exposure. The inhibitory effect of gefitinib on M1 efflux was significantly weaker than that of ningetinib.