Stable expression of human 5 alpha-reductase type II in COS1 cells due to chromosomal gene integration: a novel tool for inhibitor identification

Inhibitors of human 5 alpha -reductase type II are promising drug candidate s for the treatment of benign prostatic hyperplasia which is associated wit h high prostatic DHT levels. In this study we describe the evaluation of po tential inhibitors in a new cell assay. First a plasmid (pRcCMV-5 alpha II) for the expression of human 5 alpha -reductase type II was constructed by the use of the vector pRcCMV and transfected into the African green monkey fibroblast-like cell line COS1. By selection with G418 sulfate, ten COS1 si ngle cell clones were obtained of which three stably exhibited high 5 alpha -reductase activity. One single cell clone (COS1-5 alpha IIST) was selecte d for further investigations. By Southern blot analysis, fluorescence in si tu hybridization (FISH) and comparative PCR experiments it turned out that the expression plasmid pRcCMV-5 alpha II has been integrated into the chrom osome, resulting in a long-term stable expression of the foreign 5 alpha -r eductase gene. The newly established cell line was used for testing novel c ompounds on their inhibitory effect on human 5 alpha -reductase type IL Usi ng this whole cell assay, inhibitors with IC50 values in the nanomolar rang e could be identified. (C) 2001 Elsevier Science Ltd. All rights reserved.