Species difference in the excretion routes of Ciprofol (HSK3486) in rats and humans: Contribution of hepatic transporters and renal UDP-glucuronosyltransferase enzymes

The primary metabolic pathways of the anesthetic Ciprofol in both humans and rats are glucuronidation (M4) and 4-hydroxylation, followed by glucuronidation (M5-1). However, there are significant interspecies differences in their excretion routes. M4 and M5-1 are primarily excreted into the bile (61.4% of the dose) in rats, while they are mainly cleared through the kidneys (70.9% of the dose) in humans. This study aimed to elucidate the reasons for this discrepancy between humans and rats. The in vitro microsomal incubations revealed that Ciprofol could be metabolized by both human hepatic and renal UDP-glucuronosyltransferases (UGTs) but only by rat hepatic UGTs. The hepatocyte uptake assays showed that the organic anion transporting polypeptides were involved in the uptake of M4 and M5-1 into rat liver, not human liver. Further, only in sandwich-cultured rat hepatocytes did M4 and M5-1 exhibit significant biliary excretion potential. Vesicular transporter assays indicated that M4 and M5-1 are sensitive substrates of human/rat multidrug resistance protein (hMRP/rMrp) 2 and hMRP3. Considering their protein expression levels in the liver and intrinsic transport clearance, hMRP3 contributed more to the efflux of M4 and M5-1 than hMRP2 in humans, leading to a greater tendency to enter the blood circulation. Conversely, in rat liver, rMrp2-mediated efflux of M4 and M5-1 was the major driving force of their biliary excretion. Overall, our study showed that the differences in the protein expression and activity of hepatic hMRP3/rMrp3 and hMRP2/rMrp2, as well as renal UGT activities, led to species differences in the excretion routes of Ciprofol glucuronide metabolites. SIGNIFICANCE STATEMENT: This research characterized the interspecies differences in the excretion routes of the 2 major glucuronide-conjugated metabolites of Ciprofol (M4 and M5-1) in humans and rats. Bile excretion and urine excretion are the main excretion pathways in rats and humans, respectively. Our findings revealed that different transport activities and expression levels of hepatic human/rat multidrug resistance protein 3 and human/rat multidrug resistance protein 2, as well as different renal UDP-glucuronosyltransferase enzyme activities, are underlying reasons responsible for the differences.