A "tissue culture" method for testing the cytotoxic activity of compounds active through the formation of metabolites is described and cyclophosphamide (CPA), which is known to exert its effect only after hepatic biotransformation, has been chosen as "model" substance. A monolayer of cultured KB cells is treated for one hour with CPA at 37°C, in the presence of different amounts of microsomal enzymes (105,000 × g fraction) and their co-factors and at the end of the incubation the medium is removed and changed. A dose-effect relation is observed by measuring the inhibition of growth or the destruction of the KB cells after 24 hr. The significance of this procedure for screening purposes or for studying the possible cytotoxicity of unstable metabolites is outlined.
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