SCREENING STRATEGIES FOR THE DETECTION OF ANTICARCINOGENIC ENZYME INDUCERS

The induction of electrophile-processing Phase II enzymes (i.e., gluta thione S-transferases, UDP-glucuronosyltransferases, and quinone reduc tase) is a major mechanism whereby a large group of heterogeneous comp ounds prevent the toxic, mutagenic, and neoplastic effects of carcinog ens. This paper reviews the direct assay of quinone reductase in cultu red cells as a method to rapidly detect potentially anticarcinogenic s ubstances. Cells (usually Hepa lclc7 murine hepatoma cells) growing in 96-well microtiter plates are exposed to test compounds for 24 to 72 hrs and are then lysed and assayed for quinone reductase activity by m easuring the formation of formazan dye from the menadione-dependent re duction of MTT -(4,5-dimethylthiazo-2-yl)-2,5-diphenyltetrazolium brom ide). Reaction velocities are linearly dependent on quinone reductase over a wide range of enzyme concentrations, and the specificity of MTT reduction can be ascertained with dicoumarol, a potent inhibitor of q uinone reductase. This in vitro technique reliably detects compounds t hat effectively induce Phase II enzymes in vivo, and the screening ass ay has identified hitherto unrecognized and novel inducers from synthe tic and natural sources. As inducers of Phase II enzymes are receiving serious consideration as potential human anticarcinogens, the assay d escribed offers the opportunity to rapidly identify isolate, and chara cterize inducers of medicinal interest.