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Alectinib (CH5424802) antagonizes ABCB1- and ABCG2-mediated multidrug resistance in vitro , in vivo and ex vivo

View Article: PubMed Central - PubMed

ABSTRACT

Alectinib, an inhibitor of anaplastic lymphoma kinase (ALK), was approved by the Food and Drug Administration (FDA) for the treatment of patients with ALK-positive non-small cell lung cancer (NSCLC). Here we investigated the reversal effect of alectinib on multidrug resistance (MDR) induced by ATP-binding cassette (ABC) transporters, which is the primary cause of chemotherapy failure. We provide the first evidence that alectinib increases the sensitivity of ABCB1- and ABCG2-overexpressing cells to chemotherapeutic agents in vitro and in vivo. Mechanistically, alectinib increased the intracellular accumulation of ABCB1/ABCG2 substrates such as doxorubicin (DOX) and Rhodamine 123 (Rho 123) by inhibiting the efflux function of the transporters in ABCB1- or ABCG2-overexpressing cells but not in their parental sensitive cells. Furthermore, alectinib stimulated ATPase activity and competed with substrates of ABCB1 or ABCG2 and competed with [125I] iodoarylazidoprazosin (IAAP) photolabeling bound to ABCB1 or ABCG2 but neither altered the expression and localization of ABCB1 or ABCG2 nor the phosphorylation levels of AKT and ERK. Alectinib also enhanced the cytotoxicity of DOX and the intracellular accumulation of Rho 123 in ABCB1-overexpressing primary leukemia cells. These findings suggest that alectinib combined with traditional chemotherapy may be beneficial to patients with ABCB1- or ABCG2-mediated MDR.

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Effect of alectinib on the efflux of Rho 123, ATPase activity and the [125I]-IAAP photoaffinity labeling of ABCB1 and ABCG2. The cells were pre-incubated with 5 μM Rho 123 at 37 °C for 30 min and then incubated with alectinib for the indicated time points in Rho 123-free media. Rho 123 retention was measured by flow cytometry in KB, KBv200, S1 and S1-MI-80 cells (a, b). The vanadate-sensitive ABCB1 or ABCG2 ATPase activity was evaluated in the presence of the indicated concentrations of alectinib (c, d). Alectinib competed for photolabeling of ABCB1 or ABCG2 by [125I]-IAAP. A representative autoradiogram is shown for three independent experiments. The relative amount of [125I]-IAAP incorporated was plotted against the concentration of alectinib present. Hundred percent incorporation refers to the absence of alectinib (e, f). The data points represent the mean±s.d. of at least three independent experiments. **P<0.01.
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fig4: Effect of alectinib on the efflux of Rho 123, ATPase activity and the [125I]-IAAP photoaffinity labeling of ABCB1 and ABCG2. The cells were pre-incubated with 5 μM Rho 123 at 37 °C for 30 min and then incubated with alectinib for the indicated time points in Rho 123-free media. Rho 123 retention was measured by flow cytometry in KB, KBv200, S1 and S1-MI-80 cells (a, b). The vanadate-sensitive ABCB1 or ABCG2 ATPase activity was evaluated in the presence of the indicated concentrations of alectinib (c, d). Alectinib competed for photolabeling of ABCB1 or ABCG2 by [125I]-IAAP. A representative autoradiogram is shown for three independent experiments. The relative amount of [125I]-IAAP incorporated was plotted against the concentration of alectinib present. Hundred percent incorporation refers to the absence of alectinib (e, f). The data points represent the mean±s.d. of at least three independent experiments. **P<0.01.

Mentions: To confirm whether the increase in intracellular accumulation of DOX and Rho 123 was due to a blockage of transporter-mediated efflux, the time course of intracellular retention of Rho 123 was examined. As we expected, the intracellular retention of Rho 123 was remarkably less in KBv200 and S1-MI-80 cells than that in their corresponding parental KB and S1 cells in the absence of alectinib (Figure 4a and b). In addition, the intracellular retention of Rho 123 was much more in the presence of alectinib in KBv200 and S1-MI-80 cells than in the absence of alectinib, while there are no significant changes in the presence or absence of alectinib in KB and S1 cells at the same time point (Figure 4a and b). Taken together, the increase in intracellular accumulation of chemotherapeutic agents by alectinib in MDR cells is due to the inhibition of efflux substrate function of ABCB1 and ABCG2.


Alectinib (CH5424802) antagonizes ABCB1- and ABCG2-mediated multidrug resistance in vitro , in vivo and ex vivo
Effect of alectinib on the efflux of Rho 123, ATPase activity and the [125I]-IAAP photoaffinity labeling of ABCB1 and ABCG2. The cells were pre-incubated with 5 μM Rho 123 at 37 °C for 30 min and then incubated with alectinib for the indicated time points in Rho 123-free media. Rho 123 retention was measured by flow cytometry in KB, KBv200, S1 and S1-MI-80 cells (a, b). The vanadate-sensitive ABCB1 or ABCG2 ATPase activity was evaluated in the presence of the indicated concentrations of alectinib (c, d). Alectinib competed for photolabeling of ABCB1 or ABCG2 by [125I]-IAAP. A representative autoradiogram is shown for three independent experiments. The relative amount of [125I]-IAAP incorporated was plotted against the concentration of alectinib present. Hundred percent incorporation refers to the absence of alectinib (e, f). The data points represent the mean±s.d. of at least three independent experiments. **P<0.01.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5382559&req=5

fig4: Effect of alectinib on the efflux of Rho 123, ATPase activity and the [125I]-IAAP photoaffinity labeling of ABCB1 and ABCG2. The cells were pre-incubated with 5 μM Rho 123 at 37 °C for 30 min and then incubated with alectinib for the indicated time points in Rho 123-free media. Rho 123 retention was measured by flow cytometry in KB, KBv200, S1 and S1-MI-80 cells (a, b). The vanadate-sensitive ABCB1 or ABCG2 ATPase activity was evaluated in the presence of the indicated concentrations of alectinib (c, d). Alectinib competed for photolabeling of ABCB1 or ABCG2 by [125I]-IAAP. A representative autoradiogram is shown for three independent experiments. The relative amount of [125I]-IAAP incorporated was plotted against the concentration of alectinib present. Hundred percent incorporation refers to the absence of alectinib (e, f). The data points represent the mean±s.d. of at least three independent experiments. **P<0.01.
Mentions: To confirm whether the increase in intracellular accumulation of DOX and Rho 123 was due to a blockage of transporter-mediated efflux, the time course of intracellular retention of Rho 123 was examined. As we expected, the intracellular retention of Rho 123 was remarkably less in KBv200 and S1-MI-80 cells than that in their corresponding parental KB and S1 cells in the absence of alectinib (Figure 4a and b). In addition, the intracellular retention of Rho 123 was much more in the presence of alectinib in KBv200 and S1-MI-80 cells than in the absence of alectinib, while there are no significant changes in the presence or absence of alectinib in KB and S1 cells at the same time point (Figure 4a and b). Taken together, the increase in intracellular accumulation of chemotherapeutic agents by alectinib in MDR cells is due to the inhibition of efflux substrate function of ABCB1 and ABCG2.

View Article: PubMed Central - PubMed

ABSTRACT

Alectinib, an inhibitor of anaplastic lymphoma kinase (ALK), was approved by the Food and Drug Administration (FDA) for the treatment of patients with ALK-positive non-small cell lung cancer (NSCLC). Here we investigated the reversal effect of alectinib on multidrug resistance (MDR) induced by ATP-binding cassette (ABC) transporters, which is the primary cause of chemotherapy failure. We provide the first evidence that alectinib increases the sensitivity of ABCB1- and ABCG2-overexpressing cells to chemotherapeutic agents in vitro and in vivo. Mechanistically, alectinib increased the intracellular accumulation of ABCB1/ABCG2 substrates such as doxorubicin (DOX) and Rhodamine 123 (Rho 123) by inhibiting the efflux function of the transporters in ABCB1- or ABCG2-overexpressing cells but not in their parental sensitive cells. Furthermore, alectinib stimulated ATPase activity and competed with substrates of ABCB1 or ABCG2 and competed with [125I] iodoarylazidoprazosin (IAAP) photolabeling bound to ABCB1 or ABCG2 but neither altered the expression and localization of ABCB1 or ABCG2 nor the phosphorylation levels of AKT and ERK. Alectinib also enhanced the cytotoxicity of DOX and the intracellular accumulation of Rho 123 in ABCB1-overexpressing primary leukemia cells. These findings suggest that alectinib combined with traditional chemotherapy may be beneficial to patients with ABCB1- or ABCG2-mediated MDR.

No MeSH data available.


Related in: MedlinePlus