Human hepatic HepaRG cells maintain high intrinsic CYP450 activity/metabolism and significantly outperform standard HepG2/C3A cells used in drug pharmacology applications

L. J. Nelson, P. Treskes, C. J. Henderson, N. Homer, K. Morgan, C. LeBled, M. H. Grant, J. N. Plevris

Research output: Contribution to journalMeeting abstract

Abstract

Introduction: Conventional in vitro human hepatic models for drug testing are based on the use of cell lines or primary human hepatocytes (PHHs). However, limited availability, inter-donor functional variability and early phenotypic alterations of PHHs in vitro restrict their use; whilst cell lines such as HepG2/C3As lack a substantial and variable set of liver-specific functions, specifically, CYP450 activity. In this study we compared CYP450 activity/ metabolism between HepG2/C3A and human HepaRG cells as hepatic models for pre-clinical drug testing. Methods: Human hepatic cell lines [HepG2/C3A or HepaRG] were grown to >80% confluence on collagen-I-coated plates and treated (in triplicates) 24 h with prototypical inducers rifampicin (CYP3A4) and omeprazole (CYP1A2), [n=3]. 50μM testosterone or phenacetin were added and supernatant and cell samples taken after 2 hours of incubation at 37°C. CYP1A2/3A4 activity [P450-Glo™-Luminometry; Promega] was determined (Figure 1). Relative turnover of testosterone [HPLC] and phenacetin [LC-MS/MS] metabolites was also measured. Cell phenotype was assessed by light-microscopy, histology (PAS-Glycogen), CYP3A4, F-actin/phalloidin, and JC-1 fluorescent-staining. Results: Figure 1 shows HepaRG CYP1A2/3A4 activity was 40-80x fold >> HepG2/C3A cells [P<0.001]; Drug profiling revealed HepaRGs had both enhanced production of major metabolites of phenacetin and testosterone and more intact drug metabolism compared with HepG2/C3A. In contrast with HepG2/C3A, HepaRGs displayed a more intact hepatic phenotype, including: Strong positive glycogen, CYP3A4 staining, high JC-1-positive intrinsic metabolic activity (ΔΨm) and organotypic gross morphology. Discussion / Conclusion: HepaRG cells may represent a more physiologically-relevant pre-clinical platform for CYP450 activation/ inhibition, safety pharmacology, as well as drug-drug interaction studies.

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Hep G2 Cells
Hepatocytes
Phenacetin
Cytochrome P-450 CYP1A2
Pharmacology
Testosterone
Cytochrome P-450 CYP3A
Pharmaceutical Preparations
Glycogen
Cell Line
Liver
Staining and Labeling
Phenotype
Phalloidine
Omeprazole
Rifampin
Drug Interactions
Actins
Microscopy
Histology

Keywords

  • pre-clinical drug testing
  • human HepaRG cells
  • human hepatic models
  • HepG2/C3A
  • CYP450 activation/inhibition

Cite this

Nelson, L. J. ; Treskes, P. ; Henderson, C. J. ; Homer, N. ; Morgan, K. ; LeBled, C. ; Grant, M. H. ; Plevris, J. N. / Human hepatic HepaRG cells maintain high intrinsic CYP450 activity/metabolism and significantly outperform standard HepG2/C3A cells used in drug pharmacology applications. In: Journal of Hepatology. 2014 ; Vol. 60, No. 1 Supplement. pp. S176-S177.
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title = "Human hepatic HepaRG cells maintain high intrinsic CYP450 activity/metabolism and significantly outperform standard HepG2/C3A cells used in drug pharmacology applications",
abstract = "Introduction: Conventional in vitro human hepatic models for drug testing are based on the use of cell lines or primary human hepatocytes (PHHs). However, limited availability, inter-donor functional variability and early phenotypic alterations of PHHs in vitro restrict their use; whilst cell lines such as HepG2/C3As lack a substantial and variable set of liver-specific functions, specifically, CYP450 activity. In this study we compared CYP450 activity/ metabolism between HepG2/C3A and human HepaRG cells as hepatic models for pre-clinical drug testing. Methods: Human hepatic cell lines [HepG2/C3A or HepaRG] were grown to >80{\%} confluence on collagen-I-coated plates and treated (in triplicates) 24 h with prototypical inducers rifampicin (CYP3A4) and omeprazole (CYP1A2), [n=3]. 50μM testosterone or phenacetin were added and supernatant and cell samples taken after 2 hours of incubation at 37°C. CYP1A2/3A4 activity [P450-Glo™-Luminometry; Promega] was determined (Figure 1). Relative turnover of testosterone [HPLC] and phenacetin [LC-MS/MS] metabolites was also measured. Cell phenotype was assessed by light-microscopy, histology (PAS-Glycogen), CYP3A4, F-actin/phalloidin, and JC-1 fluorescent-staining. Results: Figure 1 shows HepaRG CYP1A2/3A4 activity was 40-80x fold >> HepG2/C3A cells [P<0.001]; Drug profiling revealed HepaRGs had both enhanced production of major metabolites of phenacetin and testosterone and more intact drug metabolism compared with HepG2/C3A. In contrast with HepG2/C3A, HepaRGs displayed a more intact hepatic phenotype, including: Strong positive glycogen, CYP3A4 staining, high JC-1-positive intrinsic metabolic activity (ΔΨm) and organotypic gross morphology. Discussion / Conclusion: HepaRG cells may represent a more physiologically-relevant pre-clinical platform for CYP450 activation/ inhibition, safety pharmacology, as well as drug-drug interaction studies.",
keywords = "pre-clinical drug testing, human HepaRG cells, human hepatic models, HepG2/C3A, CYP450 activation/inhibition",
author = "Nelson, {L. J.} and P. Treskes and Henderson, {C. J.} and N. Homer and K. Morgan and C. LeBled and Grant, {M. H.} and Plevris, {J. N.}",
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Human hepatic HepaRG cells maintain high intrinsic CYP450 activity/metabolism and significantly outperform standard HepG2/C3A cells used in drug pharmacology applications. / Nelson, L. J.; Treskes, P.; Henderson, C. J.; Homer, N.; Morgan, K.; LeBled, C.; Grant, M. H.; Plevris, J. N.

In: Journal of Hepatology, Vol. 60, No. 1 Supplement, 30.04.2014, p. S176-S177.

Research output: Contribution to journalMeeting abstract

TY - JOUR

T1 - Human hepatic HepaRG cells maintain high intrinsic CYP450 activity/metabolism and significantly outperform standard HepG2/C3A cells used in drug pharmacology applications

AU - Nelson, L. J.

AU - Treskes, P.

AU - Henderson, C. J.

AU - Homer, N.

AU - Morgan, K.

AU - LeBled, C.

AU - Grant, M. H.

AU - Plevris, J. N.

PY - 2014/4/30

Y1 - 2014/4/30

N2 - Introduction: Conventional in vitro human hepatic models for drug testing are based on the use of cell lines or primary human hepatocytes (PHHs). However, limited availability, inter-donor functional variability and early phenotypic alterations of PHHs in vitro restrict their use; whilst cell lines such as HepG2/C3As lack a substantial and variable set of liver-specific functions, specifically, CYP450 activity. In this study we compared CYP450 activity/ metabolism between HepG2/C3A and human HepaRG cells as hepatic models for pre-clinical drug testing. Methods: Human hepatic cell lines [HepG2/C3A or HepaRG] were grown to >80% confluence on collagen-I-coated plates and treated (in triplicates) 24 h with prototypical inducers rifampicin (CYP3A4) and omeprazole (CYP1A2), [n=3]. 50μM testosterone or phenacetin were added and supernatant and cell samples taken after 2 hours of incubation at 37°C. CYP1A2/3A4 activity [P450-Glo™-Luminometry; Promega] was determined (Figure 1). Relative turnover of testosterone [HPLC] and phenacetin [LC-MS/MS] metabolites was also measured. Cell phenotype was assessed by light-microscopy, histology (PAS-Glycogen), CYP3A4, F-actin/phalloidin, and JC-1 fluorescent-staining. Results: Figure 1 shows HepaRG CYP1A2/3A4 activity was 40-80x fold >> HepG2/C3A cells [P<0.001]; Drug profiling revealed HepaRGs had both enhanced production of major metabolites of phenacetin and testosterone and more intact drug metabolism compared with HepG2/C3A. In contrast with HepG2/C3A, HepaRGs displayed a more intact hepatic phenotype, including: Strong positive glycogen, CYP3A4 staining, high JC-1-positive intrinsic metabolic activity (ΔΨm) and organotypic gross morphology. Discussion / Conclusion: HepaRG cells may represent a more physiologically-relevant pre-clinical platform for CYP450 activation/ inhibition, safety pharmacology, as well as drug-drug interaction studies.

AB - Introduction: Conventional in vitro human hepatic models for drug testing are based on the use of cell lines or primary human hepatocytes (PHHs). However, limited availability, inter-donor functional variability and early phenotypic alterations of PHHs in vitro restrict their use; whilst cell lines such as HepG2/C3As lack a substantial and variable set of liver-specific functions, specifically, CYP450 activity. In this study we compared CYP450 activity/ metabolism between HepG2/C3A and human HepaRG cells as hepatic models for pre-clinical drug testing. Methods: Human hepatic cell lines [HepG2/C3A or HepaRG] were grown to >80% confluence on collagen-I-coated plates and treated (in triplicates) 24 h with prototypical inducers rifampicin (CYP3A4) and omeprazole (CYP1A2), [n=3]. 50μM testosterone or phenacetin were added and supernatant and cell samples taken after 2 hours of incubation at 37°C. CYP1A2/3A4 activity [P450-Glo™-Luminometry; Promega] was determined (Figure 1). Relative turnover of testosterone [HPLC] and phenacetin [LC-MS/MS] metabolites was also measured. Cell phenotype was assessed by light-microscopy, histology (PAS-Glycogen), CYP3A4, F-actin/phalloidin, and JC-1 fluorescent-staining. Results: Figure 1 shows HepaRG CYP1A2/3A4 activity was 40-80x fold >> HepG2/C3A cells [P<0.001]; Drug profiling revealed HepaRGs had both enhanced production of major metabolites of phenacetin and testosterone and more intact drug metabolism compared with HepG2/C3A. In contrast with HepG2/C3A, HepaRGs displayed a more intact hepatic phenotype, including: Strong positive glycogen, CYP3A4 staining, high JC-1-positive intrinsic metabolic activity (ΔΨm) and organotypic gross morphology. Discussion / Conclusion: HepaRG cells may represent a more physiologically-relevant pre-clinical platform for CYP450 activation/ inhibition, safety pharmacology, as well as drug-drug interaction studies.

KW - pre-clinical drug testing

KW - human HepaRG cells

KW - human hepatic models

KW - HepG2/C3A

KW - CYP450 activation/inhibition

UR - http://2014.ilc-congress.eu/#&panel1-1

U2 - 10.1016/S0168-8278(14)60493-1

DO - 10.1016/S0168-8278(14)60493-1

M3 - Meeting abstract

VL - 60

SP - S176-S177

JO - Journal of Hepatology

T2 - Journal of Hepatology

JF - Journal of Hepatology

SN - 0168-8278

IS - 1 Supplement

ER -