The growth and development of the prostate gland are regulated by androgens. Despite our understanding of molecular actions of 5α-dihydrotestosterone (5α-DHT) in the prostate through the trans-activation of the androgen receptor (AR), comprehensive analysis of androgen responsive genes (ARGs) has just been started. Moreover, expression changes induced by the androgen effects of 5α-androstane-3α,17β-diol (3α-diol), a metabolite of 5α-DHT through the action of 3α-hydroxysteroid dehydrogenases (3α-HSDs), remain undefined. We demonstrated that both 5α-DHT and 3α-diol stimulated similar levels of androgen sensitive human prostate cancer LNCaP cell proliferation. However, consistent with the fact that 3α-diol has low affinity toward the AR, 3α-diol did not elicit the same levels of AR trans-activation activity as that of 5α-DHT. Since LNCaP cells respond to androgen stimulation by transcriptionally activating the AR downstream genes, gene expression patterns between 0 and 48h following 3α-diol and 5α-DHT stimulation were analyzed using cDNA-based membrane arrays to define the temporal regulation of ARGs. Array analysis identified 217 and 219 androgen-modulated genes in at least one time point following 3α-diol and 5α-DHTstimulation, respectively, including key regulators of cell proliferation. Only a subset of these genes (143) was regulated by both androgens. These data suggest that 3α-diol exerts androgenic effects independent of the action of 5α-DHT in steroid target tissues. Accordingly, 3α-diol might activate cell proliferation cascades independent of AR pathway in the prostate.