HUMAN CELL-LINES, DERIVED FROM AHH-1 TK+/- HUMAN LYMPHOBLASTS, GENETICALLY-ENGINEERED FOR EXPRESSION OF CYTOCHROMES-P450

We are developing a panel of human B lymphoblastoid cells which have been engineered to express specific human cDNAs for cytochrome P450 and other xenobiotic metabolizing enzymes. The recipient cells are of a human B lymphoblastoid cell line, designated AHH-1 TK+/-. These cells are transfected using two extrachromosomal vectors both containing OriP sequences derived from Epstein Barr virus but containing independent means of selection in mammalian cells. Using this system, the level of cDNA expression is nearly always stable and consistent from one transfection to another. Thus, once the level of expression has been characterized, cell lines with potentially interesting combinations of xenobiotic-metabolizing enzymes can be predictably developed. cDNAs encoding the following human enzymes have been expressed in this system: CYP1A1, CYP1A2, CYP2A6, CYP2B8, CYP2C6, CYP2C9, CYP2D6, CYP2E1, CYP3A4 and microsomal epoxide hydrolase. We have expressed all of these enzymes individually and have developed cell lines which express combinations of the xenobiotic metabolizing enzymes. The expression of multiple enzymes is important for generalized use of engineered cells as toxicology screening tools. We have primarily used the cell lines in applications to toxicology focusing on procarcinogen activation as detected in assays for the induction of gene locus mutations. In this chapter we discuss the general properties of the system and applications to toxicology testing.