Rate-Determining and Rate-Limiting Steps in the Clearance and Excretion of a Potent and Selective p21-Activated Kinase Inhibitor: A Case Study of Rapid Hepatic Uptake and Slow Elimination in Rat.

backgroundSignificant under-prediction of in vivo clearance in rat was observed for a potent p21-activated kinase (PAK1) inhibitor, GNE1.

objectiveRate-determining (rapid uptake) and rate-limiting (slow excretion) steps in systemic clearance and elimination of GNE1, respectively, were evaluated to better understand the cause of the in vitro-in vivo (IVIV) disconnect.

methodsA series of in vivo, ex vivo, and in vitro experiments were carried out: 1) the role of organic cation transporters (Oct or Slc22a) was investigated in transporter knock-out and wild-type animals with or without 1-aminobenzotriazole (ABT) pretreatment; 2) the concentration-dependent hepatic extraction ratio was determined in isolated perfused rat liver; and 3) excreta were collected from both bile duct cannulated and non-cannulated rats after intravenous injection.

resultsAfter intravenous dosing, the rate-determining step in clearance was found to be mediated by the active uptake transporter, Oct1. In cannulated rats, biliary and renal clearance of GNE1 accounted for only approximately 14 and 16% of the total clearance, respectively. N-acetylation, an important metabolic pathway, accounted for only about 10% of the total dose. In non-cannulated rats, the majority of the dose was recovered in feces as unchanged parent (up to 91%) overnight following intravenous administration.

conclusionBecause the clearance of GNE1 is mediated through uptake transporters rather than metabolism, the extrahepatic expression of Oct1 in kidney and intestine in rat likely plays an important role in the IVIV disconnect in hepatic clearance prediction. The slow process of intestinal secretion is the rate-limiting step for in vivo clearance of GNE1.