Abstract
Our group and others have previously shown that genistein combined polysaccharide (GCP), an aglycone isoflavone-rich extract with high bioavailability and low toxicity, can inhibit prostate cancer (CaP) cell growth and survival as well as androgen receptor (AR) activity. We now elucidate the mechanism by which this may occur using LNCaP and PC-346C CaP cell lines; GCP can inhibit intracrine androgen synthesis in CaP cells. UPLC-MS/MS and qPCR analyses demonstrated that GCP can mediate a ~3-fold decrease in testosterone levels (p < 0.001) and cause decreased expression of intracrine androgen synthesis pathway enzymes (~2.5-fold decrease of 3βHSD (p < 0.001), 17βHSD (p < 0.001), CYP17A (p < 0.01), SRB1 (p < 0.0001), and StAR (p < 0.01)), respectively. Reverse-phase HPLC fractionation and bioassay identified three active GCP fractions. Subsequent NMR and LC-MS analysis of the fraction with the highest level of activity, fraction 40, identified genistein as the primary active component of GCP responsible for its anti-proliferative, pro-apoptotic, and anti-AR activity. GCP, fraction 40, and genistein all mediated at least a ~2-fold change in these biological activities relative to vehicle control (p < 0.001). Genistein caused similar decreases in the expression of 17βHSD and CYP17A (2.5-fold (p < 0.001) and 1.5-fold decrease (p < 0.01), respectively) compared to GCP, however it did not cause altered expression of the other intracrine androgen synthesis pathway enzymes; 3βHSD, SRB1, and StAR. Our combined data indicate that GCP and/or genistein may have clinical utility and that further pre-clinical studies are warranted.
Highlights
Between 20 and 50% of prostate cancer (CaP) patients will experience disease recurrence within 10 years following first line treatment and be placed on leutenizing hormone releasing hormone (LHRH) agonist-based androgen deprivationBiomedicines 2020, 8, 282; doi:10.3390/biomedicines8080282 www.mdpi.com/journal/biomedicinesBiomedicines 2020, 8, 282 therapy (ADT) [1,2,3]
Subsequent quantitative real-time PCR analysis revealed that genistein combined polysaccharide (GCP) treatment had a statistically significant impact on expression of all of the key molecules which are required for intracrine androgen synthesis in CaP cells (Figure 2); GCP mediated a ~2–3-fold decrease in the transcript levels of SRB-1, StAR, Cyp17, 3β HSD, 17β HSD
The combined data demonstrate that GCP treatment can significantly reduce testosterone levels in CaP cells and indicate that this occurs via its ability to inhibit enzymes required for intracrine androgen synthesis
Summary
Between 20 and 50% of prostate cancer (CaP) patients will experience disease recurrence within 10 years following first line treatment (typically prostatectomy or radiation therapy) and be placed on leutenizing hormone releasing hormone (LHRH) agonist-based androgen deprivation. The majority of patients fail ADT and develop castration-resistant prostate cancer (CRPC) within 2 years [5] At this point, most patients are treated with cytotoxic chemotherapy and/or AR signaling pathway inhibitors and the median progression-free survival is estimated at only ~14 months [6]. Intracrine androgen synthesis by CaP cells has been shown to contribute to failure of ADT by allowing for autocrine activation of AR. Therapies which inhibit intracrine synthesis have the potential to extend the timeframe between disease recurrence following first line treatment and ADT failure, and thereby improve CaP survival rates. A disadvantage is that the implementation of clinical studies using CAMs can be challenging because often they are a complex mix and the active component may be unknown To address this issue, we performed HPLC fractionation and bioassays followed by NMR analysis to allow for identification of the active component of GCP
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