Pomegranate polyphenols down-regulate expression of androgen-synthesizing genes in human prostate cancer cells overexpressing the androgen receptor

Abstract

Prostate cancer is dependent on circulating testosterone in its early stages and is treatable with radiation and surgery. However, recurrent prostate tumors advance to an androgen-independent state in which they progress in the absence of circulating testosterone, leading to metastasis and death. During the development of androgen independence, prostate cancer cells are known to increase intracellular testosterone synthesis, which maintains cancer cell growth in the absence of significant amounts of circulating testosterone. Overexpression of the androgen receptor (AR) occurs in androgen-independent prostate cancer and has been proposed as another mechanism promoting the development of androgen independence. The LNCaP-AR cell line is engineered to overexpress AR but is otherwise similar to the widely studied LNCaP cell line. We have previously shown that pomegranate extracts inhibit both androgen-dependent and androgen-independent prostate cancer cell growth. In this study, we examined the effects of pomegranate polyphenols, ellagitannin-rich extract and whole juice extract on the expression of genes for key androgen-synthesizing enzymes and the AR. We measured expression of the HSD3B2 (3beta-hydroxysteroid dehydrogenase type 2), AKR1C3 (aldo-keto reductase family 1 member C3) and SRD5A1 (steroid 5alpha reductase type 1) genes for the respective androgen-synthesizing enzymes in LNCaP, LNCaP-AR and DU-145 human prostate cancer cells. A twofold suppression of gene expression was considered statistically significant. Pomegranate polyphenols inhibited gene expression and AR most consistently in the LNCaP-AR cell line (P=.05). Therefore, inhibition by pomegranate polyphenols of gene expression involved in androgen-synthesizing enzymes and the AR may be of particular importance in androgen-independent prostate cancer cells and the subset of human prostate cancers where AR is up-regulated.

Fig. 1

Effects of FBS, CS-FBS and SF on cell proliferation in LNCaP (A), LNCaP-AR (B) and DU-145 (C) human prostate cancer cells. CS-FBS decreased cell proliferation in LNCaP but not in LNCaP-AR and DU-145 cells. As a negative control, SF treatment decreased cell proliferation in all cell lines. FBS: fetal bovine serum, CS-FBS: charcoal-stripped fetal bovine serum, SF: serum-free. Mean ± SEM (n=4-6). *** Significantly different from FBS at P <.001.

Fig. 2

Antiproliferative effects of PA, EA, POMx and PJ in LNCaP (A), LNCaP-AR (B) and DU-145 (C) cells. Exposure to PA, EA, POMx or PJ for 72 h inhibited prostate cancer cell growth in a dose-dependent manner (P <.01). PA: punicalagin, EA: ellagic acid, POMx: pomegranate extract, PJ: pomegranate juice. Mean ± SEM (n=4-6).

Fig. 3

Antiproliferative effects of PA, EA, POMx and PJ in hormone (DHT, β-estradiol or IGF-1)-treated LNCaP (A), LNCaP-AR (B) and DU-145 (C) cells. PA, EA, POMx or PJ inhibited hormone exposed prostate cancer cell growth in a dose-dependent manner (P <.01). Mean ± SEM (n=4-6).

Fig. 3

Antiproliferative effects of PA, EA, POMx and PJ in hormone (DHT, β-estradiol or IGF-1)-treated LNCaP (A), LNCaP-AR (B) and DU-145 (C) cells. PA, EA, POMx or PJ inhibited hormone exposed prostate cancer cell growth in a dose-dependent manner (P <.01). Mean ± SEM (n=4-6).

Fig. 4

Proapoptotic effects of PA, EA, POMx and PJ in LNCaP (A), LNCaP-AR (B) and DU-145 (C) cells. Cells exposed to PA, EA, POMx or PJ for 48 h enhanced apoptosis. Cells were treated 50 μg/ml of PA, EA or POMx, or 100 μg/ml of PJ. Mean ± SEM (n=4-6). *, ** Significantly different from control at P <.05 or P <.01 respectively.