Altered drug disposition in vivo as a consequence of murine organic anion transporter 3 (Oat3/Slc22a8) deletion

Transport across barrier epithelia is critical for efficient removal of organic anions from the body. Eight organic anion transporter proteins have been cloned within the past ten years; OATs1-7 and URAT1. Of these, six are expressed in the renal cortex, and Oat1 and Oat3 are highly expressed in the basolateral membrane of renal proximal tubule cells, where they are poised to remove organic anions from the blood. The substrate profile of these two multispecific transporters is extensive, encompassing clinically relevant xenobiotic and endogenous compounds. Furthermore, many substrates interact with both transporters, although usually with different affinities. Prior to the studies described in this dissertation, nothing was known regarding the in vivo role of Oat3 drug disposition. This was an important undertaking, as heterologous expression systems and isolated tissues, while extremely versatile and essential for initial characterization of transporter function, do not precisely recapitulate the dynamic nature of an intact renal epithelium in vivo, where multiple transporters on both aspects of the plasma membrane contribute to the total organic anion flux. Furthermore, these experimental systems exclude vital components of excretion which may compete with Oat3 in vivo. Such competing modes of elimination include glomerular filtration, hepatic transport, and metabolism. Therefore, in this dissertation, Oat3 knockout (Oat3(-/-)) mice were employed to investigate the specific effects of targeted ablation of Oat3 in vivo. The major findings are that Oat3 deletion (1) resulted in markedly impaired penicillin G elimination, to such an extent that plasma levels were increased approximately 5-fold relative to wild-type mice 30 minutes after administration; (2) did not severely impact elimination of the prototypical in vitro OAT substrates, p-aminohippurate or estrone-3-sulfate; (3) significantly reduced methotrexate elimination in female mice; (4) caused a loss of male-specific enhancement of methotrexate elimination mediated by reduced folates; and (5) resulted in impaired ciprofloxacin elimination with heightened accumulation of putative anionic metabolites. Furthermore, in this dissertation it was determined for the first time that Oat3 transports carboxyfluoroquinolones, with a marked affinity for ciprofloxacin. The major impact of Oat3 deletion on penicillin G elimination likely stems from three major factors. That is, ∼80% of penicillin G is eliminated renally, it is ∼65% protein bound (reducing filtration substantially), and exhibits a relatively weak interaction with the other renal basolateral transporter, Oat1. Collectively, the findings from this dissertation imply that known human OAT3 dysfunctional polymorphisms and previously unrecognized drug interactions on OAT3 are likely sources of inter- and intra-patient variability in the efficacy and toxicity of commonly prescribed drugs.