Received for publication October 11, 2007.
Revised December 6, 2007.
Accepted for publication December 7, 2007.

A novel high-throughput screening system identifies a small molecule repressive for matrix metalloproteinase-9 expression

Abstract

Aberrant gene expression is one of the driving forces for cancer progression, and is considered an ideal target for chemical intervention. While emerging bioluminescence reporter systems allow high-throughput searches for small molecules regulatory for gene expression, frequent silencing of reporter genes by epigenetic mechanisms hinders wide application of this drug discovery strategy. Here we report a novel system that directs the integration of a promoter-reporter construct to an open chromosomal location by Flp-mediated homologous recombination thereby overcoming reporter-gene silencing. Employing this system, we have screened more than 8000 compounds in the DIVERSet chemical library for repressors of a MMP-9 promoter, and identified 5-methyl-2-(4-methylphenyl)-1H-benzimidazol (MPBD) inhibitory for MMP-9 gene expression. Consistent with this effect, MPBD inhibits MMP-9-dependent invasion of UMSCC-1 oral cancer cells, pre-osteoclast migration and RANKL-induced osteoclast activity over concentration ranges that repressed MMP-9 expression. Mechanistic studies indicated that MPBD antagonizes AP-1 function by inhibiting its trans-activation activity. We conclude that the Flp-mediated homologous recombination system to direct reporter integration into open chromatin regions represents a novel strategy allowing for the development of high-throughput systems screening for lead compounds targeting aberrant gene expression in cancer.

Key words: AP-1, Regulation of gene expression, Transcription targets