GSTP1 DNA Methylation and Expression Status Is Indicative of 5-aza-2′-Deoxycytidine Efficacy in Human Prostate Cancer Cells

DNA methylation plays an important role in carcinogenesis and the reversibility of this epigenetic modification makes it a potential therapeutic target. To date, DNA methyltransferase inhibitors (DNMTi) have not demonstrated clinical efficacy in prostate cancer, with one of the major obstacles being the inability to monitor drug activity during the trial. Given the high frequency and specificity of GSTP1 DNA methylation in prostate cancer, we investigated whether GSTP1 is a useful marker of DNMTi treatment efficacy. LNCaP prostate cancer cells were treated with 5-aza-2′-deoxycytidine (5-aza-CdR) either with a single high dose (5–20 µM), every alternate day (0.1–10 µM) or daily (0.005–2.5 µM). A daily treatment regimen with 5-aza-CdR was optimal, with significant suppression of cell proliferation achieved with doses of 0.05 µM or greater (p<0.0001) and induction of cell death from 0.5 µM (p<0.0001). In contrast, treatment with a single high dose of 20 µM 5-aza-CdR inhibited cell proliferation but was not able to induce cell death. Demethylation of GSTP1 was observed with doses of 5-aza-CdR that induced significant suppression of cell proliferation (≥0.05 µM). Re-expression of the GSTP1 protein was observed only at doses of 5-aza-CdR (≥0.5 µM) associated with induction of cell death. Treatment of LNCaP cells with a more stable DNMTi, Zebularine required at least a 100-fold higher dose (≥50 µM) to inhibit proliferation and was less potent in inducing cell death, which corresponded to a lack of GSTP1 protein re-expression. We have shown that GSTP1 DNA methylation and protein expression status is correlated with DNMTi treatment response in prostate cancer cells. Since GSTP1 is methylated in nearly all prostate cancers, our results warrant its testing as a marker of epigenetic therapy response in future clinical trials. We conclude that the DNA methylation and protein expression status of GSTP1 are good indicators of DNMTi efficacy

Dysregulated fibronectin trafficking by Hsp90 inhibition restricts prostate cancer cell invasion

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.The molecular chaperone Hsp90 is overexpressed in prostate cancer (PCa) and is responsible for the folding, stabilization and maturation of multiple oncoproteins, which are implicated in PCa progression. Compared to first-in-class Hsp90 inhibitors such as 17-allylamino-demethoxygeldanamycin (17-AAG) that were clinically ineffective, second generation inhibitor AUY922 has greater solubility and efficacy. Here, transcriptomic and proteomic analyses of patient-derived PCa explants identified cytoskeletal organization as highly enriched with AUY922 treatment. Validation in PCa cell lines revealed that AUY922 caused marked alterations to cell morphology, and suppressed cell motility and invasion compared to vehicle or 17-AAG, concomitant with dysregulation of key extracellular matrix proteins such as fibronectin (FN1). Interestingly, while the expression of FN1 was increased by AUY922, FN1 secretion was significantly decreased. This resulted in cytosolic accumulation of FN1 protein within late endosomes, suggesting that AUY922 disrupts vesicular secretory trafficking pathways. Depletion of FN1 by siRNA knockdown markedly reduced the invasive capacity of PCa cells, phenocopying AUY922. These results highlight a novel mechanism of action for AUY922 beyond its established effects on cellular mitosis and survival and, furthermore, identifies extracellular matrix cargo delivery as a potential therapeutic target for the treatment of aggressive PCa

A patient-derived explant (PDE) model of hormone-dependent cancer.

Breast and prostate cancer research to date has largely been predicated on the use of cell lines in vitro or in vivo. These limitations have led to the development of more clinically relevant models, such as organoids or murine xenografts that utilize patient-derived material; however, issues related to low take rate, long duration of establishment, and the associated costs constrain use of these models. This study demonstrates that ex vivo culture of freshly resected breast and prostate tumor specimens obtained from surgery, termed patient-derived explants (PDEs), provides a high-throughput and cost-effective model that retains the native tissue architecture, microenvironment, cell viability, and key oncogenic drivers. The PDE model provides a unique approach for direct evaluation of drug responses on an individual patient's tumor, which is amenable to analysis using contemporary genomic technologies. The ability to rapidly evaluate drug efficacy in patient-derived material has high potential to facilitate implementation of personalized medicine approaches.Cancer Research UK and ERC

A ZEB1-miR-375-YAP1 pathway regulates epithelial plasticity in prostate cancer

MicroRNA-375 (miR-375) is frequently elevated in prostate tumors and cell-free fractions of patient blood, but its role in genesis and progression of prostate cancer is poorly understood. In this study, we demonstrated that miR-375 is inversely correlated with epithelial–mesenchymal transition signatures (EMT) in clinical samples and can drive mesenchymal–epithelial transition (MET) in model systems. Indeed, miR-375 potently inhibited invasion and migration of multiple prostate cancer lines. The transcription factor YAP1 was found to be a direct target of miR-375 in prostate cancer. Knockdown of YAP1 phenocopied miR-375 overexpression, and overexpression of YAP1 rescued anti-invasive effects mediated by miR-375. Furthermore, transcription of the miR-375 gene was shown to be directly repressed by the EMT transcription factor, ZEB1. Analysis of multiple patient cohorts provided evidence for this ZEB1-miR-375-YAP1 regulatory circuit in clinical samples. Despite its anti-invasive and anti-EMT capacities, plasma miR-375 was found to be correlated with circulating tumor cells in men with metastatic disease. Collectively, this study provides new insight into the function of miR-375 in prostate cancer, and more broadly identifies a novel pathway controlling epithelial plasticity and tumor cell invasion in this disease

Androgen receptor inhibits estrogen receptor-alpha activity and Is prognostic in breast cancer

There is emerging evidence that the balance between estrogen receptor-α (ERα) and androgen receptor (AR) signaling is a critical determinant of growth in the normal and malignant breast. In this study, we assessed AR status in a cohort of 215 invasive ductal breast carcinomas. AR and ERα were coexpressed in the majority (80-90%) of breast tumor cells. Kaplan-Meier product limit analysis and multivariate Cox regression showed that AR is an independent prognostic factor in ERα-positive disease, with a low level of AR (less than median of 75% positive cells) conferring a 4.6-fold increased risk of cancer-related death (P = 0.002). Consistent with a role for AR in breast cancer outcome, AR potently inhibited ERα transactivation activity and 17β-estradiol–stimulated growth of breast cancer cells. Transfection of MDA-MB-231 breast cancer cells with either functionally impaired AR variants or the DNA-binding domain of the AR indicated that the latter is both necessary and sufficient for inhibition of ERα signaling. Consistent with molecular modeling, electrophoretic mobility shift assays showed binding of the AR to an estrogen-responsive element (ERE). Evidence for a functional interaction of the AR with an ERE in vivo was provided by chromatin immunoprecipitation data, revealing recruitment of the AR to the progesterone receptor promoter in T-47D breast cancer cells. We conclude that, by binding to a subset of EREs, the AR can prevent activation of target genes that mediate the stimulatory effects of 17β-estradiol on breast cancer cells

Human DECR1 is an androgen-repressed survival factor that regulates PUFA oxidation to Protect prostate tumor cells from ferroptosis

Fatty acid β-oxidation (FAO) is the main bioenergetic pathway in human prostate cancer (PCa) and a promising novel therapeutic vulnerability. Here we demonstrate therapeutic efficacy of targeting FAO in clinical prostate tumors cultured ex vivo, and identify DECR1, encoding the rate-limiting enzyme for oxidation of polyunsaturated fatty acids (PUFAs), as robustly overexpressed in PCa tissues and associated with shorter relapse-free survival. DECR1 is a negatively-regulated androgen receptor (AR) target gene and, therefore, may promote PCa cell survival and resistance to AR targeting therapeutics. DECR1 knockdown selectively inhibited β-oxidation of PUFAs, inhibited proliferation and migration of PCa cells, including treatment resistant lines, and suppressed tumor cell proliferation and metastasis in mouse xenograft models. Mechanistically, targeting of DECR1 caused cellular accumulation of PUFAs, enhanced mitochondrial oxidative stress and lipid peroxidation, and induced ferroptosis. These findings implicate PUFA oxidation via DECR1 as an unexplored facet of FAO that promotes survival of PCa cells.status: publishe