Males have a higher incidence of bladder cancer than females, but the reason remains unknown. Unlike prostate cancer, human bladder cancer is not generally considered to be dependent on hormone activity. We investigated the possible involvement of androgens and the androgen receptor (AR) in bladder cancer. We used N -butyl- N -(4-hydroxybutyl)nitrosamine (BBN) to induce bladder cancer in wild-type male and female mice, with and without castration in males, and in AR knockout (ARKO) male and female mice, with and without dihydrotestosterone (DHT) supplementation in males. We also treated human bladder cancer cell lines, including TCC-SUP and UMUC3, and mouse xenograft models established from these same lines with androgen deprivation therapy (antiandrogen treatment or castration), AR–small-interfering RNA (AR-siRNA), or the anti-AR molecule ASC-J9, which causes selective degradation of the AR. More than 92% of wild-type male and 42% of wild-type female mice treated with BBN eventually developed bladder cancer, whereas none of the male or female ARKO mice did. Treatment with BBN induced bladder cancer in 25% of ARKO mice supplemented with DHT and in 50% of castrated wild-type male mice. Androgen deprivation of AR-positive human bladder cancer cells by androgen depletion in vitro or castration in mice and/or by treatment with the antiandrogen flutamide in vitro or in vivo, as well as AR knockdown by AR-siRNA or by ASC-J9, suppressed cell proliferation in vitro and xenograft tumor growth in vivo. Our findings implicate the involvement of both androgens and the AR in bladder cancer. Targeting AR and androgens may provide novel chemopreventive and therapeutic approaches for bladder cancer.

Background Large epidemiological cohort studies in the United States have indicated that active and passive smoking are associated with increased breast cancer risk. However, there was no direct evidence of an effect of tobacco carcinogens on the cellular molecules involved in breast tumorigenesis. Methods Reverse transcription–polymerase chain reaction was used to determine the expression of all of the nicotinic acetylcholine receptor (nAChR) subunits in 50 human breast cancer samples and to determine the expression of the α9-nAChR subunit in 276 surgical and laser capture microdissected breast tumor vs normal tissue pairs. Stable MDA-MB-231 breast cancer cell lines were established in which expression of the α9-nAChR subunit was inhibited using short interfering RNA. MCF-10A normal human breast epithelial cells were established in which the α9-nAChR subunit could be conditionally overexpressed by removal of doxycycline from the culture fluid. Cell proliferation and soft agar assays and tumor growth in nude mice were used as measures of cell transformation. All statistical tests were two-sided. Results In 186 (67.3%) of the 276 paired samples, α9-nAChR mRNA was expressed at (mean 7.84-fold) higher levels in breast cancers than in surrounding normal tissue. Stable expression of α9-nAChR short interfering RNA in MDA-MB-231 cells attenuated nicotine-stimulated proliferation and growth in soft agar and reduced tumor volume when the cells were introduced as xenografts in SCID mice (n = 5 mice per group; mean tumor volume at 6 weeks treatment in mice injected with Si α9 cells = 995.6 mm³, in mice injected with parental cells = 2993.2 mm³, difference = 1997.6 mm³, 95% confidence interval [CI] = 1705 to 2290.2 mm³, P = .009). Long-term treatment of MCF-10A normal breast epithelial cells with either nicotine or its active metabolite, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, triggered precancerous transformation as defined by soft agar assay. Inducible overexpression of α9-nAChR in MCF-10A cell xenografts in nude mice substantially increased tumor growth (n = 5 mice per group; DOX+, mean tumor volume without nicotine vs with nicotine = 266.2 vs 501.6 mm³, difference = 235.4 mm³, 95% CI = 112.7 to 358 mm³, P = .009; DOX−, mean tumor volume without nicotine vs with nicotine = 621.2 vs 898.6 mm³, difference = 277.4 mm³, 95% CI = 98.1 to 456.7 mm³, P = .016; mean tumor volume in the presence of nicotine, DOX+ vs DOX− = 501.6 vs 898.6 mm³, difference = 397 mm³, 95% CI = 241.3 to 552.6 mm³, P = .009). Conclusion The α9-nAChR is important for nicotine-induced transformation of normal human breast epithelial cells.

Androgens and the androgen receptor (AR) play important roles in the testes. Previously we have shown that male total AR knockout (T-AR^−/y) mice revealed incomplete germ cell development and lowered serum testosterone levels, which resulted in azoospermia and infertility. However, the consequences of AR loss in particular types of testicular cells remain unclear. Using a Cre-loxP conditional knockout strategy, we generated a tissue-selective knockout mouse with the AR gene deleted in testis peritubular myoid cells (PM-AR^−/y). Phenotype analyses showed that PM-AR^−/ymice were indistinguishable from WT AR (AR^+/y) mice with the exception of smaller testes size. PM-AR^−/ymice have serum testosterone concentrations comparable with AR^+/ymice. PM-AR^−/ymice have oligozoospermia in the epididymis; however, fertility was normal. Although normal germ cell distribution ratio was found, total germ cell number decreased in PM-AR^−/ymice. Further mechanistic studies demonstrated that PM-AR^−/ymice have defects in the expression of Sertoli cells' functional marker genes such as tranferrin, epidermal fatty acid-binding protein, androgen-binding protein, and other junction genes including occludin, testin, nectin, zyxin, vinculin, lamininγ3, gelsolin, connection43, and N-cadherin. Furthermore, there were defects in peritubular myoid cell contractility-related genes such as endothelin-1, endothelin receptor A and B, adrenomedullin, adrenomedullin receptor, and vasopressin receptor 1a. Together, our PM-AR^−/ymice providein vivoevidence for the requirement of functional AR in peritubular myoid cells to maintain normal Sertoli cells function and peritubular myoid cell contractility, thus ensuring normal spermatogenesis and sperm output.