Protein Abundance of Phase I and II Drug Metabolizing Enzymes in Alcoholic and Hepatitis C Cirrhosis Livers
Authors: Prasad Bhagwat , Johnson Katherine , Xiao Guangqing , Lai Yurong , Lee Caroline , Ray Adrian S. , Liao Mingxiang , Chu Xiaoyan , Salphati Laurent , Mathias Anita , Humphreys William , Hop Cornelis E. C. A. , Kumer Sean , Unadkat Jashvant D 
Published: November 2023

Background and Purpose

Liver cirrhosis is associated with the loss of functional hepatocytes and dysregulation of protein expression which can lead to a significant decrease in hepatic drug clearance. In order to predict the impact of liver cirrhosis on hepatic drug clearance using physiologically based pharmacokinetic (PBPK) models, we compared the protein abundance of various Phase I and Phase II drug metabolizing enzymes (DMEs) in alcoholic or hepatitis C (HCV) cirrhotic livers vs. those in non‐cirrhotic livers (control).

Methods

LC‐MS/MS quantifiable surrogate peptides for the major human hepatic DMEs were selected and MRM proteomic methods were developed and validated for their quantification. S9 fractions were isolated from control (n=25), HCV cirrhosis (n=30), and alcoholic cirrhosis (n=27) livers obtained from sources described previously (1). Total protein concentration was determined and the samples were denatured, reduced, alkylated, desalted, and digested as previously reported (1, 2, 3). Eighteen DMEs were quantified by AB Sciex 6500 LC‐MS/MS using optimum conditions of reproducibility (1,2, 3). Protein abundance of DMEs in cirrhotic vs. control livers was compared using Kruskal‐Wallis test followed by Dunn's multiple comparison test.

Results

The total protein concentration was significantly lower in alcoholic and HCV cirrhosis vs. control livers (i.e., 38.3, 32.3 vs. 51.1 mg/g liver, respectively). In general, alcoholic cirrhosis was associated with a larger decrease in the DME abundance than HCV cirrhosis. When normalized to liver weight, the abundance of seven DMEs (UGT1A6, UGT1A4, CYP3A4, UGT2B7, CYP1A2, AOX1, and CES1) in alcoholic cirrhosis and five DMEs (UGT1A6, UGT2B7, CYP1A2, CYP3A4 and UGT1A4) in HCV cirrhosis was below 25% of that in control livers. The abundance of the majority of the DMEs, i.e., CYP2E1, FMO3, EPHX1, CYP2A6, EPHX2, POR, CYPC8 and CYP2C9 (alcoholic cirrhosis) and AOX1, CYP2A6, CES1, FMO3, CYP2E1, EPHX1, CYP2C9, EPHX2, UGT2B15 and CYPC8 (HCV cirrhosis) was between 25 to 50% of that in control livers. UGT1A4 was ~66% lower in alcoholic cirrhosis vs. HCV cirrhosis livers. CES2 expression was not different across the three groups. Integration of UGT2B7 abundance in cirrhotic livers into the Simcyp PBPK model significantly improved the prediction of zidovudine PK in patients with liver cirrhosis.

Conclusions

Liver cirrhosis caused by alcoholism vs. HCV had a differential impact on the abundance of hepatic DMEs. These data are important for improved predictions of drug disposition in patients with liver cirrhosis. While the mechanisms of dysregulation of DME abundance in hepatic cirrhosis (alcoholic vs. HCV vs. control) is not clearly understood, these data can help generate hypotheses to elucidate these mechanisms.

Support or Funding Information

UWRAPT (JDU) and NICHD R01HD081299 (BP).