Construction and Evaluation of a Novel Organic Anion Transporter 1/3 CRISPR/Cas9 Double-Knockout Rat Model

Background: Organic anion transporter 1 (OAT1) and OAT3 have an overlapping spectrum of substrates such that one can exert a compensatory effect when the other is dysfunctional. As a result, the knockout of either OAT1 or OAT3 is not reflected in a change in the excretion of organic anionic substrates. To date, only the mOAT1 and mOAT3 individual knockout mouse models have been available. Methods: In this study, we successfully generated aSlc22a6/Slc22a8double-knockout (KO) rat model using CRISPR/Cas9 technology and evaluated its biological properties. Results: The double-knockout rat model did not expressionmRNAfor rOAT1 or rOAT3 in the kidneys. Consistently, the renal excretion ofp-aminohippuric acid (PAH), the classical substrate of OAT1/OAT3, was substantially decreased in theSlc22a6/Slc22a8double-knockout rats. The relativemRNAlevel ofSlco4c1was up-regulated in KO rats. No renal pathological phenotype was evident. The renal elimination of the organic anionic drug furosemide was nearly abolished in theSlc22a6/Slc22a8knockout rats, but elimination of the organic cationic drug metformin was hardly affected. Conclusions: These results demonstrate that this rat model is a useful tool for investigating the functions of OAT1/OAT3 in metabolic diseases, drug metabolism and pharmacokinetics, and OATs-mediated drug interactions.Keywords:OAT1/OAT3;CRISPR/Cas9;gene knockout;pharmacokinetic