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Background: PSA mediates growth factor responses that stimulate proliferation of prostatic and other cellular types. Androgen-sensitive TE85 human osteosarcoma cells were used to study PSA as a potential mediator of prostatic cancer growth and osseous metastasis., Materials and Methods: TE85 cells were probed for PSA mRNA and protein levels under testosterone (T)-replete and--depleted conditions. TE85 proliferative responses to PSA were evaluated in the absence and presence of LY312340, a monocyclic beta-lactam inhibitor of PSA enzymatic activity., Results: A 3.1-fold increase in PSA mRNA was observed following T stimulation. Low levels of immunoreactive PSA (iPSA) were detected in media of androgen-stimulated TE85 cells while iPSA was not found in control media. Conversely, iPSA was detected in TE85 cell pellets from control but not in androgen-stimulated cell cultures. Exogenously added enzymatically active PSA stimulated TE85 proliferation in a bi-phasic manner. LY312340 inhibited PSA-induced increases in TE85 cell numbers but had no effect on basal or T- stimulated cellular proliferation., Conclusion: While the PSA levels produced by TE-85 cells in response to androgen stimulation are too low to be biologically active, PSA produced by metastatic PCa cells may mediate paracrine stimulation of osteogenic PCa metastasis. Inhibitors of PSA enzymatic activity could be useful therapeutic agents.

The conversion of testosterone (T) to dihydrotestosterone (DHT) has been demonstrated to be catalysed by at least two isoforms of human steroid 5 alpha-reductase, designated types I and II. Type II 5 alpha-reductase expression predominates in human accessory sex tissues, localized to the fibromuscular stromal compartment. The type I isoform predominates in skin, prostatic epithelia and, to a lesser extent, in prostatic fibromuscular stroma. The significance of the type I isoform to prostatic cellular growth and function remains undefined. In cultured DU145 cells, we evaluated the metabolism of [14C]-T and demonstrated the time-dependent formation of [14C]-DHT. Oxidative metabolism (conversion of [14C]-T to [14C]-androstenedione) and the formation of conjugated androgen metabolites occurred at a relatively low rate in the DU145 cells. Using human type I 5 alpha-reductase cDNA, Northern blot analysis of DU145 cell mRNA revealed high levels of type I isoform expression. Analogous probing of the DU145 cells with a human 5 alpha-reductase II cDNA failed to reveal expression of the type II isoform. The expression of functional type I activity has been confirmed pharmacologically using isoform-selective 5 alpha-reductase inhibitors. Reductive metabolism of [3H]-T in the DU145 cells was inhibited in a concentration-dependent manner by LY306089, a potent non-steroidal type I-selective inhibitor (IC50 = 10.0 nM). SKF105657, a steroidal type II-specific inhibitor was distinctly less active at inhibiting [3H]-DHT formation. LY306089 was a non-competitive inhibitor of type I 5 alpha-reductase in DU145 cellular homogenates with an apparent Ki value of 4.0 nM. These studies have identified and pharmacologically defined type I 5 alpha-reductase activity in an androgen-insensitive prostatic cancer cell line and provide the basis for additional investigations into the significance of type I 5 alpha-reductase to human prostatic pathophysiology.

We evaluate the metabolic inhibitory, antiproliferative, and antisecretory effects of LY300502, a benzoquinolinone human-specific type I-selective steroid 5alpha-reductase inhibitor in LNCaP human prostatic adenocarcinoma cell cultures. Reductive metabolism of [3H-T] in the LNCaP cells was inhibited in a concentration-dependent manner by LY300502 (IC50 approximately 5.77 nM). The proliferative responses of LNCaP cells to LY300502 were examined in the presence of 0.1 NM testosterone (T), a concentration that stimulates maximal LNCaP cell numbers 40% above control levels. LY300502 significantly anatagonized T-induced stimulation of LNCaP cellular proliferation at concentrations greater that 10 nM (P<0.05), and at 1,000 nMcompletely blocked the mitogenic effects of T on LNCaP cells. In the absence of androgen, LY300502 had no effect on LNCaP cellular proliferation. In the presence of 100 nM T, an androgen concentration that maximally stimulates in vitro PSA production, LY300502 significantly antagonized T-induced PSA secretion at a concentration equal to or greater than 30 nM (P<0.05). These studies provide the basis for additional investigations into the pathophysiologic significance of type I 5alpha-reductase to prostatic cancer and the potential utility of selective inhibitors as therapeutic agents.