Identification of the major oxidative 3α-hydroxysteroid dehydrogenase in human prostate

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Prostate cancer (PCA) and benign prostatic hyperplasia (BPH) are the most common androgen-dependent prostate diseases among adult males. Disease progression is dependent upon 5α-dihydrotestosterone (DHT) a potent ligand for the androgen receptor. DHT is formed by the reduction of testosterone by type 2 5α-reductase which is inactivated by the mechanism based inactivator, finasteride. Although finasteride reduces both the volume and size of the prostate by more than 90% the intraprostatic concentration of DHT is only decreased by 50%. Several mammalian studies showed that 5α-androstane-3α, 17β-diol (3α-diol), an inactive metabolite of DHT, but not the epimer 5α-androstane-3β, 17β-diol, can be converted back to DHT by an unidentified oxidative 3α-hydroxysteroid dehydrogenase (3α-HSD). The putative 3α-HSD candidates that can oxidize 3α-diol to DHT include type 3 3α-HSD (AKR1C2), 11-cis retinol dehydrogenase (RODH 5), L-3-hydroxyacyl Coenzyme A dehydrogenase (ERAB), RODH like 3α-HSD, new type of human microsomal 3α-HSD and retinol dehydrogenase 4 (RODH 4). The enzymes were compared side-by-side to determine the ability of each to oxidize 3α-diol to DHT by transient transfection in either COS-1 cells or the androgen independent prostatic PC-3 cell line. Construction of bicistronic constructs, whereby both the 3α-HSD of interest and β-galactosidase were expressed by the same promoter, were created to normalize transfected 3α-HSD activity. Results indicate that all the 3α-HSDs were able to oxidize 3α-diol back to DHT except for AKR1C2. RODH 5, RODH 4 and RODH like 3α-HSD were the most robust at converting 3α-diol to DHT at all substrate concentrations tested. By contrast, AKR1C2 reduced the potent androgen DHT to the inactive metabolite 3α-diol and is important in inactivating excess DHT. Although multiple 3α-HSDs can catalyze the formation of DHT, expression levels will identify the dominant isoform in human prostate. Using real-time RT-PCR the expression of the candidate oxidative 3α-HSD isoforms were compared. In whole normal prostate all the candidate enzymes were expressed; however ERAB and RODH like 3α-HSD were elevated 8-fold when compared to the other candidates. Expression levels are now being quantified in cultures of normal and diseased primary epithelial and stromal cells. Based on the activity and expression of RODH like 3α-HSD this isoform maybe the major oxidase in human prostate and inhibitors that target this enzyme may have therapeutic value. (Supported by R01 CA-90744 awarded to T.M.P., DAMD PC040420 awarded to D.M.P. and Training Grant 1R25-CA 101871-D1 Support for D.R.B)