A6 TRANSPORTER EXPRESSION IN NON-CANCEROUS AND CANCEROUS LIVER TISSUE FROM SUBJECTS WITH HEPATOCELLULAR CARCINOMA AND CHRONIC HEPATITIS C INFECTION QUANTIFIED BY LC-MS/MS PROTEOMICS

Sarah F Billington , University of Washington, Seattle, WA
Adrian S. Ray , Departments of Clinical Pharmacology and Drug Metabolism and Pharmacokinetics, Gilead Sciences, Inc., Foster City, CA
Laurent Salphati , Genentech Inc., South San Francisco, CA
Guangqing Xiao , Preclinical PK and In Vitro ADME, Biogen Idec, Cambridge, MA
Xiaoyan Chu , Pharmacokinetics, Pharmacodynamics and Drug Metabolism, Merck & Co., Rahway, NJ
W. Griffith Humphreys , Pharmaceutical Candidate Optimization, Bristol-Myers Squibb Company, Lawrenceville, NJ
Mingxiang Liao , Drug Metabolism and Pharmacokinetics, Takeda Boston, Cambridge, MA
Caroline Lee , Ardea Biosciences, Inc, San Diego, CA
Anita Mathias , Departments of Clinical Pharmacology and Drug Metabolism and Pharmacokinetics, Gilead Sciences, Inc., Foster City, CA
Cornelis E.C.A. Hop , Drug Metabolism and Pharmacokinetics, Genentech Inc., South San Francisco, CA
Christopher Rowbottom , Biogen Idec, Cambridge, MA
Raymond Evers , Drug Metabolism and Pharmacokinetics, Merck & Co., Rahway, NJ
Yurong Lai , Department of Metabolism and Pharmacokinetics, Departments of Clinical Pharmacology and Drug Metabolism and Pharmacokinetics, Gilead Sciences, Inc., Foster City, CA
Edward J. Kelly , University of Washington, Seattle, WA
Bhagwat Prasad , University of Washington, Seattle, WA
Jashvant D. Unadkat , University of Washington, Seattle, WA
Purpose: Hepatocellular carcinoma is clinically diagnosed by areas of reduced intensity by magnetic resonance imaging as a result of decreased hepatic uptake of the contrast agent gadolinium-ethoxybenzyl-diethylenetriamine-pentaacetic-acid (Gd-EOB-DTPA). Gd-EOB-DTPA is a substrate of hepatobiliary transporters (OATP1B1, OATP1B3, NTCP, MRP2 and MRP3). To understand the impact of this disease on hepatobiliary transporter expression, the protein levels of hepatic ABC (P-gp, BCRP, BSEP, MRP2-4) and SLC (OATPs, NTCP, OCT1, MATE1) transporters in cancerous (C) and non-cancerous (NC) liver tissues obtained from hepatitis C patients with hepatocellular carcinoma (HCV-HCC) were quantified by LC-MS/MS proteomics. Methods: NC (n=25) and matched C and NC (n=8) liver resections from subjects with HCV-HCC, obtained at the time of liver transplant, were subjected to membrane protein extraction, trypsin digestion, and LC-MS/MS proteomic analysis. The protein expression of hepatobiliary transporters in NC and C HCV-HCC tissues was compared on a per gram of liver basis with our previous data from non-cirrhotic (n=36) and HCV-cirrhotic (n=30) [1] livers using the Kruskal-Wallis test with Dunn’s multiple comparison correction. Differences in transporter expression in matched NC and C HCV-HCC tissues were assessed by the non-parametric Wilcoxon signed-rank test. Results: Compared with the control livers, the yield of total membrane protein from NC and C HCV-HCC tissues were reduced by 31% and 67%, respectively. With the exception of NTCP, MRP2 and MATE1, transporter protein expressions were reduced in both NC (38-76%) and C (56-96%) HCV-HCC tissues in comparison to control livers. NTCP expression was higher (113%), MATE1 expression was lower (58%) and MRP2 expression was unchanged in NC HCV-HCC tissues relative to control livers. In C HCV-HCC tissues, NTCP and MRP2 expression were lower (63%, 56%) than control livers, and MATE1 expression was unchanged. Similarly, transporter protein expression was decreased in both NC (41-71%) and C (54-89%) HCV-HCC tissues compared to HCV-cirrhotic livers, with the exception of NTCP, OCT1, BSEP and MRP2. In NC HCV-HCC tissues, NTCP (362%) and MRP2 (142%) expression were increased, whilst OCT1 and BSEP expression were unchanged. In C HCV-HCC tissues expression of OCT1 (90%) and BSEP (80%) were lower than HCV-cirrhotic livers, while NTCP and MRP2 expression were no different. Hepatobiliary transporter protein expression of OATP2B1, BSEP, MRP2 and MRP3 were lower (56-72%) in C than NC HCV-HCC tissues (n=8, matched tissues available from the same subject) but the expression of the remaining transporters were not significantly different. Conclusions: The expression of hepatic transporters was differentially, but modestly, affected in C and NC HCV-HCC tissues. Decreased expression of OATPs and NTCP in C vs. NC tissue could explain the deceased uptake of Gd-EOB-DTPA in C tissue. These data will be useful in the future to predict transporter-mediated drug disposition (including hepatocellular concentrations) in patients with HCV-HCC. Acknowledgement: This work was supported by UWRAPT through funding from Ardea Biosciences, Biogen, Bristol-Myers Squibb, Genentech, Gilead Sciences, Merck and Takeda.

1. Wang, L., et al., Transporter Expression in Liver Tissue from Subjects with Alcoholic or Hepatitis C Cirrhosis Quantified by Targeted Quantitative Proteomics. Drug Metab Dispos, 2016. 44(11): p. 1752-1758.