Pharmacological functions of multidrug transporters: studies employing combination transporter knockout mice

Abstract

ATP-binding cassette (ABC) multidrug transporters are drug efflux pumps located in the plasma membrane that utilize the energy of ATP hydrolysis to extrude a wide spectrum of endogenous and exogenous compounds from cells, including numerous (anticancer) drugs and/or their metabolites. The studies described in this thesis focus on the pharmacological functions of the ABC transporters: P-glycoprotein (P-gp/ABCB1), the Multidrug Resistance Proteins 2 and 3 (MRP2/ABCC2 and MRP3/ABCC3) and the Breast Cancer Resistance Protein (BCRP/ABCG2). Most results presented in this thesis were obtained by studying single and combination ABC multidrug transporter knockout mice. As ABC multidrug transporters do not only have very broad, but also substantially overlapping substrate specificities, they can often partially, or sometimes even fully, compensate for the loss of each other. Combination ABC drug transporter knockout mice are therefore invaluable tools to study the separate roles and functional overlap of ABC multidrug transporters. We generated and characterized combination P-gp/Mrp2 knockout mice and used these to assess the distinct roles of P-gp and Mrp2 in the pharmacokinetics of the anticancer drug paclitaxel. Although paclitaxel is an excellent P-gp substrate, Mrp2 was found to almost exclusively mediate the excretion of paclitaxel from the liver into the bile, whereas P-gp had little effect. This finding is especially interesting because Mrp2 was thus far thought to mainly affect organic anionic drugs in vivo. However, we show that Mrp2 can also be a major determinant of the pharmacokinetic behavior of highly lipophilic anti-cancer drugs, even in the presence of other efficient transporters. P-gp and BCRP combination knockout mice enabled us to demonstrate that both multidrug transporters act in concert at the blood-brain barrier in restricting the brain penetration of the novel tyrosine kinase inhibitor anticancer drugs dasatinib and sorafenib. Brain penetration of dasatinib was primarily restricted by P-gp, whereas loss of BCRP had no effect. However, when both transporters were absent a disproportionate increase in brain penetration of dasatinib was observed. In contrast, for sorafenib it was the other way around, i.e. absence of P-gp had no effect while BCRP deficiency resulted in markedly elevated brain levels. Again, simultaneous loss of both transporters resulted in a highly increased brain penetration. When we combined dasatinib with the dual P-gp and BCRP inhibitor elacridar we found that the brain penetration in wild-type mice could be increased to P-gp/BCRP knockout levels. These findings might be clinically relevant for patients with intracranial tumors, as concomitant administration of an inhibitor of P-gp and ABCG2 with dasatinib, sorafenib and possibly other tyrosine kinase inhibitors might result in better therapeutic responses in these patients. In conclusion, the studies described in this thesis demonstrate the power of combination ABC multidrug transporter knockout mouse models to study the pharmacological functions of ABC multidrug transporters. We expect that combination ABC transporter knockout mice will be extensively used as preclinical research tools.