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Growth arrest and DNA-damage-inducible 45 beta (GADD45β) is a member of the gene family associated with cell growth control, apoptosis, and DNA damage repair. The aim of present study was to determine the potential effects of GADD45β deletion on prostate hyperplasia progression.LNCaP cells were incubated with testosterone propionate (1 μM) for 48 h and specific siRNA used to suppress GADD45β expression in vitro. For in vivo experiments, testosterone (3 mg/kg, IP) was injected into wild-type (WT) and GADD45β knockout (GADD45β−/−) C57BL/6J mice for 21 consecutive days, and serum and prostate tissues subjected to biological and histochemical analyses.GADD45β-silenced LNCaP cells showed suppressed testosterone-induced 5α-reductase 2 and androgen receptor expression compared to control LNCaP cells. Moreover, after 21 days of testosterone treatment, prostate weight and stromal tissue increment were relatively lower in GADD45β−/− than WT counterpart mice. Inhibition of testosterone-induced 5α-reductase 2 and proliferating cell nuclear antigen expression in the GADD45β−/− group was confirmed via immunohistochemistry analyses.Although the exact correlation between GADD45β and prostate hyperplasia remains to be established, the present GADD45βdeletion suppressed testosterone-induced prostate hyperplasia which was accompanied by inhibition of 5α-reductase 2-related protein expression.