Background -- In concert with oxidative metabolism, intestinal absorption and renal elimination determine the pharmacokinetics of anti-viral drugs, including both HIV protease inhibitors and nucleoside phosphonates. At both sites, specific transporters, e.g., the classical organic anion and ATP-driven excretory ATPases, govern transepithelial drug flux. These same processes are also found at the blood/brain and blood/cerebrospinal fluid barriers, where they limit penetration of drugs into the central nervous system. In addition, carrier-mediated drug accumulation within excretory organs, liver and kidney, often result in dose-limiting cellular toxicity. These studies are designed to use the cloned transporters in combination with cell and tissue flux studies to define the critical steps in AIDS anti-viral drug pharmacokinetics and to evaluate the effectiveness of transport modulation in enhancing intestinal drug entry, increasing CNS penetration, and reducing nephrotoxicity. Project Summary -- Active transport of anionic and cationic drugs plays a critical role in their intestinal absorption, their penetration across the blood-CSF and blood-brain barriers, and their accumulation within excretory organs. Using the cloned organic anion transporters, rOAT1 and hOAT1, we have demonstrated that the nucleoside phosphonate antiviral drugs are excellent substrates for both transporters, with particularly high affinity for hOAT1, explaining their extensive renal accumulation and toxicity. We have also demonstrated the presence of OAT1 at the apical (CSF-facing) membrane of the choroid plexus (i.e., the blood-CSF barrier), suggesting that CSF to blood transport may contribute to the limited ability to achieve therapeutic levels of these drugs in the brain and thus, to their limited effectiveness in combating CNS viral infection. However, recent data indicates that, despite the presence of OAT1 in choroid plexus, the nucleotide phosphonates are much less effectively transported by this tissue than by the kidney. Since other OAT1 substrates, e.g., the herbicide 2,4-dichlorophenoxyacteic acid, are very well transported by choroid plexus, this is a very surprising finding. Its basis is under investigation. Finally, in collaboration with Dr. David Miller, we have begun to use confocal microscopy to examine the cell to blood step in drug and xenobiotic transport. It appears that the ATP-driven drug pump, MRP2, is found at the apical (facing the vascular lumen) membrane of the blood-brain barrier and the apical (CSF facing) membrane of choroid plexus. The basolateral (blood-side) transporter in choroid plexus appears to be MRP1. Since Miller has also shown that the nucleoside phosphonates are substrates for ATP-driven MRP2-mediated transport at other sites, it is likely that the MRPs play a role in the barrier function of choroid plexus for the nucleoside phosphonates.
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