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Ponatinib is an oral BCR-ABL1 inhibitor for treatment of advanced leukemic diseases that carry the Philadelphia chromosome, specifically containing the T315I mutation yielding resistance to previously approved BCR-ABL1 inhibitors. Using in vitrotransport assays and knockout mouse models, we investigated whether the multidrug efflux transporters ABCB1 and ABCG2 transport ponatinib and whether they, or the drug-metabolizing enzyme CYP3A, affect the oral availability and brain accumulation of ponatinib and its active N-desmethyl metabolite (DMP). In vitro, mouse Abcg2 and human ABCB1 modestly transported ponatinib. In mice, both Abcb1 and Abcg2 markedly restricted brain accumulation of ponatinib and DMP, but not ponatinib oral availability. Abcg2 deficiency increased DMP plasma levels ∼3-fold. Cyp3a deficiency increased the ponatinib plasma AUC 1.4-fold. Our results suggest that pharmacological inhibition of ABCG2 and ABCB1 during ponatinib therapy might benefit patients with brain (micro)metastases positioned behind an intact blood-brain barrier, or with substantial expression of these transporters in the malignant cells. CYP3A inhibitors might increase ponatinib oral availability, enhancing efficacy but possibly also toxicity of this drug.

P-glycoprotein (P-gp/ABCB1) and breast cancer resistance protein (BCRP/ABCG2) combination knockout mice display disproportionately increased brain penetration of shared substrates, including topotecan and several tyrosine kinase inhibitors, compared to mice deficient for only one transporter. To better study the interplay of both transporters also in vitro, we generated a transduced polarized MDCKII cell line stably coexpressing substantial levels of human ABCB1 and ABCG2 (MDCKII-ABCB1/ABCG2). Next, we measured concentration-dependent transepithelial transport of topotecan, sorafenib and sunitinib. By blocking either one or both of the transporters simultaneously, using specific inhibitors, we aimed to mimic the ABCB1-ABCG2 interplay at the blood−brain barrier in wild-type, single or combination knockout mice. ABCB1 and ABCG2 contributed to similar extents to topotecan transport, which was only partly saturable. For sorafenib transport, ABCG2 was the major determinant at low concentrations. However, saturation of ABCG2-mediated transport occurred at higher sorafenib concentrations, where ABCB1 was still fully active. Furthermore, sunitinib was transported equally by ABCB1 and ABCG2 at low concentrations, but ABCG2-mediated transport became saturated at lower concentrations than ABCB1-mediated transport. The relative impact of these transporters can thus be affected by the applied drug concentrations. A comparison of the in vitroobserved (inverse) transport ratios and cellular accumulation of the drugs at low concentrations with in vivobrain penetration data from corresponding Abcb1a/1b−/−, Abcg2−/−and Abcb1a/1b;Abcg2−/−mouse strains revealed very similar qualitative patterns for each of the tested drugs. MDCKII-ABCB1/ABCG2 cells thus present a useful in vitromodel to study the interplay of ABCB1 and ABCG2.

Vemurafenib (PLX4032) is a novel tyrosine kinase inhibitor that has clinical efficacy against metastatic melanoma harboring a BRAFV600Emutation. We aimed to establish whether oral availability and brain penetration of vemurafenib could be restricted by the multidrug efflux transporters P-glycoprotein (P-gp/ABCB1) and breast cancer resistance protein (BCRP/ABCG2), as these might limit therapeutic efficacy, especially against brain metastases. In vitro, vemurafenib was efficiently transported by both human ABCB1 and ABCG2, and very efficiently by mouse Abcg2, but not by mouse Abcc2. Upon oral administration of vemurafenib (5 mg/kg), Abcb1a/1b–/–mice had a 1.6-fold increased, Abcg2–/–mice a 2.3-fold increased, and Abcb1a/1b–/–;Abcg2–/–mice a 6.6-fold increased plasma AUC, respectively, compared to wild-type (WT) mice, indicating a marked and additive role of these transporters in limiting vemurafenib oral availability. Brain-to-plasma ratios of vemurafenib (oral, 25 mg/kg) were not increased in Abcg2–/–mice, only 1.7-fold in Abcb1a/1b–/–mice, but 21.4-fold in Abcb1a/1b–/–;Abcg2–/–mice, indicating pronounced overlapping functions of these transporters in reducing vemurafenib brain accumulation. Oral coadministration of the dual ABCB1 and ABCG2 inhibitor elacridar almost completely eliminated the roles of Abcb1 and Abcg2 in restricting oral availability and brain accumulation of vemurafenib. As predicted by previously described pharmacokinetic modeling, halving the amount of active efflux transport at the WT blood–brain barrier by testing heterozygous Abcb1a/1b+/–;Abcg2+/–mice had little impact on vemurafenib brain accumulation. Our data suggest that elacridar coadministration may be considered to improve the therapeutic efficacy of vemurafenib, especially for brain metastases located behind a functional blood–brain barrier.