An in vitro tool to assess cytochrome P450 drug biotransformation-dependent cytotoxicity in engineered HepG2 cells generated by using adenoviral vectors.
Many adverse drug reactions leading to hepatotoxicity are caused by the cytochrome P450-dependent activation of non-toxic drugs or chemicals into reactive metabolites. To this end, adenoviruses were used as a tool to efficiently deliver specific CYP genes into cultured cells (i.e., human hepatoma cell line HepG2). Recombinant-defective adenoviral vectors encoding for genes CYP3A4 (Adv-CYP3A4), CYP2E1 (Adv-CYP2E1), CYP2A6 (Adv-CYP2A6) and CYP1A2 (Adv-CYP1A2) were used to confer specific CYP drug metabolic capabilities to HepG2 cells. Upgraded cells transiently expressed single specific cytochrome P450 enzymatic activities in terms of the number of the infecting virus particles used in their transduction. HepG2 cells transduced with adenoviruses and wild HepG2 cells cultured in 96 well-plates were incubated in the presence of model compounds, some of which can be metabolized to reactive metabolites. After compound exposure, cell viability was assessed by the commonly used MTT assay. The results confirm that the cell-based assay is a valuable tool in toxicology assessments and high-throughput screenings to detect cytotoxicity mediated by cytochrome P450 biotransformation in preclinical drug development. The assay also has a potential applicability in other industrial sectors such as the chemical industry.
Unidad de Hepatología Experimental, Centro de Investigación, Hospital La Fe, Avda Campanar 21, 46009 Valencia, Spain. agustin.lahoz@uv.es
Agustín
A
Lahoz
Maya R
MR
Vilà
Myriam
M
Fabre
Josep M
JM
Miquel
Manuel
M
Rivas
Jessica
J
Maines
José V
JV
Castell
M José
MJ
Gómez-Lechón
D000256:Adenoviridae / Q000235:genetics*; D001711:Biotransformation; D002470:Cell Survival; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism*; D005822:Genetic Vectors; D056945:Hep G2 Cells; D006801:Humans; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D023382:Toxicity Tests, Acute / Q000379:methods*; D014161:Transduction, Genetic
D004364:Pharmaceutical Preparations; D003577:Cytochrome P-450 Enzyme System