Toxicity Screening of a Combinatorial Library: Correlation of Cytotoxicity and Gene Induction to Compound Structure

Combinatorial chemistry has increased the number of compounds available for efficacy and safety assessment by several orders of magnitude and has made high throughput assays essential. To test whether higher throughput toxicity assays could be of utility in screening compounds in early development, a selected set of combinatorial chemistry compounds was screened for induction of 70-Kd heat shock protein (HSP70) and 45-Kd growth arrest and DNA damage protein (GADD45) mRNA levels as well as cytotoxicity, in HepG2 cells, using a 96-well microtiter plate format. Both assays, the branched DNA (Quantigene) assay for mRNA levels and MTT for cytotoxicity, were robust enough to be incorporated into a screening format using a single replicate and a single concentration of compound. Significantly, a structure/toxicity correlation was established with this set of compounds with cytotoxicity and gene induction patterns linked to compound structure. Therefore, this type of early screening may be useful in identifying toxic substituents, enabling the design of libraries with less potential for toxicity. While structure/toxicity correlations were observed, no relationship was observed between GADD45 gene induction and mutagenesis as measured by the Ames bacterial reverse mutation assay.