Use of the probasin promoter ARR₂PB to express bax in androgen receptor-positive prostate cancer cells

Background: Adenovirus-mediated overexpression of the apoptosis-inducing protein Bax can induce apoptosis in prostate cancer cell lines. Constitutive overexpression of Bax could result in unwanted apoptosis in every site of accidental Bax accumulation in vivo. Therefore, we developed an adenoviral construct (Av-ARR₂PB-Bax) in which the probasin promoter, modified to contain two androgen response elements, drives Bax expression. This promoter would be expected to limit expression of Bax to cells expressing the androgen receptor. Methods: A variety of androgen receptor (AR)-positive and -negative cell lines of prostatic or nonprostatic origin were infected with Av-ARR₂PB-Bax or a control virus, Av-ARR₂PB-CAT, in which the same promoter drives expression of the chloramphenicol acetyl transferase-reporter gene. Bax expression and apoptosis in vitro were assessed by western blot analysis. Tumor size and apoptosis in vivo were assessed after four weekly injections of Av-ARR₂PB-Bax or Av-ARR₂PB-CAT into subcutaneous LNCaP xenografts growing in uncastrated male mice. All statistical tests were two-sided. Results: Bax was overexpressed in an androgen-dependent way in AR-positive cell lines of prostatic origin but not in AR-positive cells of nonprostatic origin or in AR-negative cell lines of either prostatic or nonprostatic origin. The androgen dihydrotestosterone activated apoptosis in LNCaP cells infected with Av-ARR₂PB-Bax but not in those infected with Av-ARR₂PB-CAT. Av-ARR₂PB-Bax-injected LNCaP xenograft tumors decreased in tumor size from 34.1mm³ (95% confidence interval [CI] = 25.1 mm³ to 43.1 mm³) to 24.6 mm³ (95% CI = -2.5 mm³ to 51.7 mm³), but the difference was not statistically significant (P = .5). Tumors injected with Av-ARR₂PB-CAT increased in size, from 28.9 mm³ (95% CI = 12.7 mm³ to 45.1 mm³) to 206 mm³ (95% CI = 122 mm³ to 290 mm³) (P = .002) and contained statistically significant more apoptotic cells (23.3% [95% CI = 21.1% to 25.6%] versus 9.5% [95% CI = 8.0% to 11.1]) (P<.001). Conclusions: Av-ARR₂PB-Bax induces androgen-dependent therapeutic apoptosis in vitro and in vivo by activating apoptosis in AR-positive cells derived specifically from prostatic epithelium and does not affect nonprostatic cells.