P-Glycoprotein Mediated Drug Transport in the Bbb

Elmquist, William

Abstract

The long-term objective of this research is to develop better therapeutic strategies to use multidrug resistance (MDR) modulators in combination with antitumor agents (e.g., doxorubicin, vincristine, taxol) without causing undue CNS toxicity. MDR cancers represent an important challenge in cancer chemotherapy. Several modulators of the MDR1 gene product (p-glycoprotein, P-gp) are currently in clinical trials, and several more compounds are under development specifically for MDR modulation. P-gp, a membrane-bound transporter, is found in many normal tissues that have barrier functions and is expressed on the luminal side of the brain capillary endothelial cells, indicating that P-gp may be an important functional component of the blood-brain barrier (BBB). The long-term objective of this research relates to the hypothesis that the transport of P-gp substrates to the brain depends on the expression and functional capacity of P-gp in the BBB, and that this function is critical in avoiding dose-limiting CNS toxicity of some antitumor agents. The goal will be to characterize the functional capacity of the P-gp efflux pump in the BBB in an in vivo model using a pharmacokinetic analysis of the effect of P-gp inhibition on P-gp substrate distribution to the brain.
The specific aims are as follows: 1) to examine the effect of P-gp substrate dose on the transport of a model substrate into the CNS to determine the linear range and the capacity of the transport system, 2) to quantify the effect of a specific P-gp inhibitor on the distribution of a model substrate across the BBB and blood-CSF barrier, and 3) to determine if exposure to a P-gp inhibitor will change the capacity of the transport system due to increased expression of P-gp in the BBB. The capacity of P-gp in the brain will be determined by examining the effect of cyclosporin A, a potent P-gp inhibitor, on the CNS transport kinetics of a fluorescent P-gp substrate, rhodamine-123 (R-123), in the freely moving rat model using in vivo brain microdialysis. Using pharmacokinetic modeling, the fraction of the total transport that is subject to inhibition and the kinetic parameters characterizing the capacity (Tmax), substrate affinity (Kt) and inhibitor affinity (Ki) of the transport system will be determined. The effect of acute and chronic dosing of a P-gp inhibitor on the expression of P-gp in the brain capillary endothelial cells will be determined by immunoblotting using a specific P-gp monoclonal antibody. Characterizing the role P-gp plays in limiting drug delivery to the brain will enable the future design of several therapeutic strategies, including the comparison of different P-gp inhibitor classes in the search for a tumor-selective inhibitor. New strategies may reduce the incidence of untoward effects resulting from the coadministration of a potentially neurotoxic antitumor agent with a P-gp modulator when treating a peripheral MDR cancer, such as leukemia or breast cancer. Moreover, these insights may allow for using P-gp inhibitors to specifically target and enhance the brain delivery of compounds to treat diseases of the CNS, including brain tumors, with relatively non-neurotoxic drugs.