Chi-Chi Peng, Utkarsh Doshi, Chandra Prakash and Albert P. Li Pages 3 - 15 ( 13 )
Objective: Development and validation of a novel assay, the Plated Hepatocyte Relay Assay (PHRA), for the determination of the metabolic fates of slowly metabolized compounds. Method: Cryopreserved human hepatocytes were cultured for 4 h followed by incubation with slowly metabolized compounds for 24 h (initial incubation). On the next day, the incubated media were collected and added to hepatocytes was similarly prepared on the day of incubation (48 h incubation; 1st relay). The procedures were repeated on the next days (72 h (2nd relay), 96 h (3rd relay), and 120 h (4th relay) incubations). Results: A proof-of-concept study with two low clearance compounds, diazepam and tolbutamide, and a validation study with 15 ultra-low clearance compounds (CLnon-renal < 1 mL/min/kg) and low clearance compounds (CLnon-renal 1- 5.1 mL/min/kg) were performed. Linear time-dependent disappearance of the parent compounds was observed for all compounds. Application of published free fraction values in combination with a correction factor with in vitro hepatic clearance results obtained with the PHRA accurately predicted in vivo hepatic clearance. Conclusion: PHRA represents a useful experimental system for the evaluation of the metabolic fates of low clearance compounds in drug development..
Cryopreserved hepatocytes, hepatic clearance, human hepatocytes, low clearance compounds, metabolic clearance, relay assay, slowly metabolized compounds.
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