Resveratrol regulates PTEN/Akt pathway through inhibition of MTA1/HDAC unit of the NuRD complex in prostate cancer.

Metastasis associated protein 1 (MTA1) is a component of the nucleosome remodeling and deacetylating (NuRD) complex which mediates gene silencing and is overexpressed in several cancers. We reported earlier that resveratrol, a dietary stilbene found in grapes, can down-regulate MTA1. In the present study, we show that PTEN is inactivated by MTA1 in prostate cancer cells. Further, we show that resveratrol promotes acetylation and reactivation of PTEN via inhibition of the MTA1/HDAC complex, resulting in inhibition of the Akt pathway. In addition, we show that MTA1 knockdown is sufficient to augment acetylation of PTEN indicating a crucial role of MTA1 itself in the regulation of PTEN acetylation contributing to its lipid phosphatase activity. Acetylated PTEN preferentially accumulates in the nucleus where it binds to MTA1. We also show that MTA1 interacts exclusively with PTEN acetylated on Lys¹²5 and Lys¹²8, resulting in diminished p-Akt levels. Finally, using orthotopic prostate cancer xenografts, we demonstrate that both resveratrol treatment and MTA1 knockdown enhance PTEN levels leading to a decreased p-Akt expression and proliferation index. Taken together, our results indicate that MTA1/HDAC unit is a negative regulator of PTEN which facilitates survival pathways and progression of prostate cancer and that resveratrol can reverse this process through its MTA1 inhibitory function.

Serum Zinc Levels in Women with Polycystic Ovarian Syndrome are Lower as Compared to Those without Polycystic Ovarian Syndrome: A Cohort Study.

Background: Zinc is an essential micronutrient, a vital stabiliser and a cofactor in many enzymes such as superoxide dismutase and phospholipase C and also acts as an antioxidant by protecting the sulfhydryl groups of different proteins and enzymes against free radicals. It is unclear if serum zinc levels are correlated with polycystic ovary syndrome (PCOS) and its pathophysiology, although relation between diabetes and insulin resistance has been established. Aims: This study aimed to investigate circulating serum zinc levels in PCOS subjects compared with non-PCOS subjects. Settings and Design: In this cohort study, PCOS subjects were compared with normal subjects aged between 18 and 35. Materials and Methods: All the included subjects underwent measurement of anthropometric parameters, fasting insulin, luteinising hormone, follicle-stimulating hormone, thyroid-stimulating hormone, prolactin, progesterone, oestrogen and serum zinc levels. These values were taken on days 2-5 of the menstrual cycle. Statistical Analysis Used: Univariate analysis and linear regression were performed for serum zinc levels and fasting insulin levels in PCOS subjects and non-PCOS subjects using SPSS (version 21) and Microsoft Excel (2019). Results: Serum zinc levels in the PCOS group were lower than in the control group (P = 0.012). Fasting insulin levels in the PCOS group were higher than in non-PCOS subjects (P = 0.001). We found a negative correlation between zinc and fasting insulin (r = -0.580, P < 0.0001) in the normal group and (r = -0.332, P = 0.019) in the PCOS group. A positive correlation was found between body mass index (BMI) and fasting insulin levels in both the PCOS group (r = 0.227, P = 0.112) and normals (r = 0.612, P < 0.0001). A negative statistically significant correlation between BMI and zinc in both the PCOS group (r = -0.378, P = 0.007) and the non-PCOS group (r = -0.7452, P < 0.0001) was seen. Conclusion: The data suggest that serum zinc levels were found to be lower in PCOS subjects as compared to normal controls and evaluation of these levels may indicate that zinc has a vital role in PCOS pathophysiology.

Trimethoxy-resveratrol and piceatannol administered orally suppress and inhibit tumor formation and growth in prostate cancer xenografts.

BACKGROUND: Resveratrol (Res) is recognized as a promising cancer chemoprevention dietary polyphenol with antioxidative, anti-inflammatory, and anticancer properties. However, the role of its analogues in prostate cancer (PCa) chemoprevention is unknown. METHODS: We synthesized several natural and synthetic analogues of Res and characterized their effects on PCa cells in vitro using a cell proliferation assay. A colony formation assay and in vitro validation of luciferase (Luc) activity was done for LNCaP-Luc cells that were consequently used for in vivo studies. The efficacy of Res, trimethoxy-resveratrol (3M-Res) and piceatannol (PIC) was studied in a subcutaneous (s.c.) model of PCa using oral gavage. Tumor progression was monitored by traditional caliper and bioluminescent imaging. The levels of cytokines in serum were examined by ELISA, and the levels of compounds in serum and tumor tissues were determined by gas chromatography-mass spectrometry. RESULTS: We examined the anti-proliferative activities of Res/analogues in three PCa cell lines. We further compared the chemopreventive effects of oral Res, 3M-Res, and PIC in LNCaP-Luc-xenografts. We found that 2 weeks pretreatment with the compounds diminished cell colonization, reduced tumor volume, and decreased tumor growth in the xenografts. Both 3M-Res and PIC demonstrated higher potency in inhibiting tumor progression compared to Res. Notably, 3M-Res was the most active in inhibiting cell proliferation and suppressing colony formation, and its accumulation in both serum and tumor tissues was the highest. CONCLUSIONS: Our findings offer strong pre-clinical evidence for the utilization of dietary stilbenes, particularly 3M-Res, as novel, potent, effective chemopreventive agents in PCa.

Advancements of Common Gamma-Chain Family Cytokines in Cancer Immunotherapy.

The approval of immunotherapies such as checkpoint inhibitors (CPIs), adoptive cell therapies and cancer vaccines has revolutionized the way cancer treatment is approached. While immunotherapies have improved clinical outcome in a variety of tumor types, some cancers have proven harder to combat using single agents, underscoring the need for multi-targeted immunotherapy approaches. Efficacy of CPIs and cancer vaccines requires patients to have a competent immune system with adequate cell numbers while the efficacy of adoptive cellular therapy is limited by the expansion and persistence of cells after infusion. A promising strategy to overcome these challenges is combination treatment with common gamma-chain cytokines. Gamma-chain cytokines play a critical role in the survival, proliferation, differentiation and function of multiple immune cell types, including CD8 T-cells and NK cells, which are at the center of the anti-tumor response. While the short half-life of recombinant cytokines initially limited their application in the clinic, advancements in protein engineering have led to the development of several next-generation drug candidates with dramatically increased half-life and bioactivity. When combining these cytokines with other immunotherapies, strong evidence of synergy has been observed in preclinical and clinical cancer settings. This promising data has led to the initiation of 70 ongoing clinical trials including IL-2, IL-7, IL-15 and IL-21. This review summarizes the recent advancements of common gamma-chain cytokines and their potential as a cancer immunotherapy.

A tumor suppressor role for EZH2 in diffuse midline glioma pathogenesis.

Pediatric high-grade gliomas, specifically diffuse midline gliomas, account for only 20% of clinical cases but are 100% fatal. A majority of the DMG cases are characterized by the signature K27M mutation in histone H3. The H3K27M mutation opposes the function of enhancer of zeste homolog 2 (EZH2), the methyltransferase enzyme of the polycomb repressor complex 2. However, the role of EZH2 in DMG pathogenesis is unclear. In this study, we demonstrate a tumor suppressor function for EZH2 using Ezh2 loss- and gain-of-function studies in H3WT DMG mouse models. Genetic ablation of Ezh2 increased cell proliferation and tumor grade while expression of an Ezh2 gain-of-function mutation significantly reduced tumor incidence and increased tumor latency. Transcriptomic analysis revealed that Ezh2 deletion upregulates an inflammatory response with upregulation of immunoproteasome genes such as Psmb8, Psmb9, and Psmb10. Ezh2 gain-of-function resulted in enrichment of the oxidative phosphorylation/mitochondrial metabolic pathway namely the isocitrate dehydrogenase Idh1/2/3 genes. Pharmacological inhibition of EZH2 augmented neural progenitor cell proliferation, supporting the tumor suppressive role of EZH2. In vivo 7-day treatment of H3K27M DMG tumor bearing mice with an EZH2 inhibitor, Tazemetostat, did not alter proliferation or significantly impact survival. Together our results suggest that EZH2 has a tumor suppressor function in DMG and warrants caution in clinical translation of EZH2 inhibitors to treat patients with DMG.

The Role of Interleukin-7 in the Formation of Tertiary Lymphoid Structures and Their Prognostic Value in Gastrointestinal Cancers.

Immunotherapies for the treatment of solid tumors continue to develop in preclinical and clinical research settings. Unfortunately, for many patients the tumor fails to respond or becomes resistant to therapies such as checkpoint inhibitors (CPIs) targeting programmed cell death protein-1 (PD-1), programmed death-ligand 1 (PD-L1), and cytotoxic T lymphocyte antigen-4 (CTLA-4). In many cancers, failed response to CPIs can be attributed to poor T cell infiltration, dominant immunosuppression, and exhausted immune responses. In gastrointestinal (GI) cancers T cell infiltration can be dismal, with several reports finding that CD8+ T cells compose less than 2% of all cells within the tumor. Organized aggregates of lymphocytes, antigen-presenting cells, and vessels, together termed tertiary lymphoid structures (TLSs), are hypothesized to be a major source of T cells within solid tumors. The intratumoral formation of these organized immune centers appears to rely on intricate cytokine and chemokine signaling to heterogeneous cell populations such as B and T cells, innate lymphoid cells, fibroblasts, and dendritic cells. In GI cancers, the presence and density of TLSs provide prognostic value for predicting outcome and survival. Further, TLS presence and density associates with favorable responses to CPIs in many cancers. This review highlights the prognostic value of TLSs in GI cancers, the role of the homeostatic cytokine interleukin-7 (IL-7) in TLS formation, and the induction of TLSs in solid tumors by novel therapeutics.

Lysis of aminobisphosphonate-sensitized MCF-7 breast tumor cells by Vγ9Vδ2 T cells.

Aminobisphosphonates are drugs administered for the treatment of bone resorption. They can indirectly activate peripheral γδ T cells and render tumor cells susceptible to lysis by Vγ9Vδ2 T cells. We have investigated the molecules involved in conjugate formation and killing of aminobisphosphonate-treated MCF-7 breast tumor cells by Vγ9Vδ2 T cells. Lysis of aminobisphosphonate (Pamidronate and Zoledronate)-treated MCF-7 tumor cells by Vγ9Vδ2 T cells was assessed by chromium release assays and time-lapse video microscopy. MCF-7 breast cancer cells were chosen as aminobisphosphonates are employed to alleviate bone resorption in this malignancy. Cell cycle profile and expression of MICA, ICAM-I and FasL on aminobisphosphonate-sensitized MCF-7 breast tumor cells was confirmed by flow cytometry. Involvement of γδ TCR and NKG2D in mediating cytotoxicity of aminobisphosphonate-treated MCF-7 breast tumor cells by Vγ9Vδ2 T cells was assessed using blocking antibodies in chromium release assays. MCF-7 tumor cells pretreated with Pamidronate and Zoledronate were efficiently lysed by Vγ9Vδ2 T cells. Pamidronate and Zoledronate treatment of MCF-7 cells induced S phase arrest and did not alter expression of MICA, ICAM-I and FasL. Blocking γδ TCR and NKG2D on Vγ9Vδ2 T cells inhibited lysis of Pamidronate and Zoledronate-treated MCF-7 cells. Inhibiting the perforin-granzyme pathway in Vγ9Vδ2 T cells using concanamycin A reduced their ability to lyse aminobisphosphonate-treated MCF-7 cells. Vγ9Vδ2 T cells form strong conjugates with aminobisphosphonate-treated MCF-7 breast tumor cells. γδ TCR, NKG2D and perforin-granzyme pathway are involved in the lysis of MCF-7 breast tumor cells treated with aminobisphosphonates by Vγ9Vδ2 T cells.

MTA1 drives malignant progression and bone metastasis in prostate cancer.

Prostate cancer often metastasizes to the bone, leading to morbidity and mortality. While metastasis-associated protein 1 (MTA1) is highly overexpressed in metastatic tumors and bone metastatic lesions, its exact role in the development of metastasis is unknown. Here, we report the role of MTA1 in prostate cancer progression and bone metastasis in vitro and in vivo. We found that MTA1 silencing diminished formation of bone metastases and impaired tumor growth in intracardiac and subcutaneous prostate cancer xenografts, respectively. This was attributed to reduced colony formation, invasion, and migration capabilities of MTA1 knockdown cells. Mechanistic studies revealed that MTA1 silencing led to a significant decrease in the expression of cathepsin B (CTSB), a cysteine protease critical for bone metastasis, with an expected increase in the levels of E-cadherin in both cells and xenograft tumors. Moreover, meta-analysis of clinical samples indicated a positive correlation between MTA1 and CTSB. Together, these results demonstrate the critical role of MTA1 as an upstream regulator of CTSB-mediated events associated with cell invasiveness and raise the possibility that targeting MTA1/CTSB signaling in the tumor may prevent the development of bone metastasis in prostate cancer.

MTA1-activated Epi-microRNA-22 regulates E-cadherin and prostate cancer invasiveness.

We have previously shown that metastasis-associated protein 1 (MTA1), a chromatin remodeler, plays an important role in prostate cancer invasiveness, likely through regulation of epithelial-to-mesenchymal transition. Here, we identified miR-22 as an epigenetic-microRNA (Epi-miR) directly induced by MTA1 and predicted to target E-cadherin. Loss-of-function and overexpression studies of MTA1 reinforced its regulatory role in miR-22 expression. MiR-22 directly targets the 3'-untranslated region of E-cadherin, and ectopic overexpression of miR-22 diminishes E-cadherin expression. Overexpression of miR-22 in prostate cancer cells promotes cell invasiveness and migration. Meta-analysis of patient tumor samples indicates a positive correlation between MTA1 and miR-22, supporting their inhibitory effect on E-cadherin expression. Our findings implicate the MTA1/Epi-miR-22/E-cadherin axis as a new epigenetic signaling pathway that promotes tumor invasion in prostate cancer.

Targeting MTA1/HIF-1α signaling by pterostilbene in combination with histone deacetylase inhibitor attenuates prostate cancer progression.

The metastasis-associated protein 1(MTA1)/histone deacetylase (HDAC) unit is a cancer progression-related epigenetic regulator, which is overexpressed in hormone-refractory and metastatic prostate cancer (PCa). In our previous studies, we found a significantly increased MTA1 expression in a prostate-specific Pten-null mouse model. We also demonstrated that stilbenes, namely resveratrol and pterostilbene (Pter), affect MTA1/HDAC signaling, including deacetylation of tumor suppressors p53 and PTEN. In this study, we examined whether inhibition of MTA1/HDAC using combination of Pter and a clinically approved HDAC inhibitor, SAHA (suberoylanilide hydroxamic acid, vorinostat), which also downregulates MTA1, could block prostate tumor progression in vivo. We generated and utilized a luciferase reporter in a prostate-specific Pten-null mouse model (Pb-Cre+ ; Ptenf/f ; Rosa26Luc/+ ) to evaluate the anticancer efficacy of Pter/SAHA combinatorial approach. Our data showed that Pter sensitized tumor cells to SAHA treatment resulting in inhibiting tumor growth and additional decline of tumor progression. These effects were dependent on the reduction of MTA1-associated proangiogenic factors HIF-1α, VEGF, and IL-1ß leading to decreased angiogenesis. In addition, treatment of PCa cell lines in vitro with combined Pter and low dose SAHA resulted in more potent inhibition of MTA1/HIF-1α than by high dose SAHA alone. Our study provides preclinical evidence that Pter/SAHA combination treatment inhibits MTA1/HIF-1α tumor-promoting signaling in PCa. The beneficial outcome of combinatorial strategy using a natural agent and an approved drug for higher efficacy and less toxicity supports further development of MTA1-targeted therapies in PCa.

Dietary pterostilbene is a novel MTA1-targeted chemopreventive and therapeutic agent in prostate cancer.

Overexpression of the epigenetic modifier metastasis-associated protein 1 (MTA1) is associated with aggressive human prostate cancer. The purpose of this study was to determine MTA1- targeted chemopreventive and therapeutic efficacy of pterostilbene, a natural potent analog of resveratrol, in pre-clinical models of prostate cancer. Here, we show that high levels of MTA1 expression in Pten-loss prostate cooperate with key oncogenes, including c-Myc and Akt among others, to promote prostate cancer progression. Loss-of-function studies using human prostate cancer cells indicated direct involvement of MTA1 in inducing inflammation and epithelial-to-mesenchymal transition. Importantly, pharmacological inhibition of MTA1 by pterostilbene resulted in decreased proliferation and angiogenesis and increased apoptosis. This restrained prostatic intraepithelial neoplasia (PIN) formation in prostate-specific Pten heterozygous mice and reduced tumor development and progression in prostate-specific Pten-null mice. Our findings highlight MTA1 as a key upstream regulator of prostate tumorigenesis and cancer progression. More significantly, it offers pre-clinical proof for pterostilbene as a promising lead natural agent for MTA1-targeted chemopreventive and therapeutic strategy to curb prostate cancer.

Resveratrol and pterostilbene epigenetically restore PTEN expression by targeting oncomiRs of the miR-17 family in prostate cancer.

In recent years, not only has the role of miRNAs in cancer become increasingly clear but also their utilization as potential biomarkers and therapeutic targets has gained ground. Although the importance of dietary stilbenes such as resveratrol and pterostilbene as anti-cancer agents is well recognized, our understanding of their miRNA-targeting capabilities is still limited. In our previous study, we reported that resveratrol downregulates PTEN-targeting members of the oncogenic miR-17 family, which are overexpressed in prostate cancer. This study investigates the resveratrol and pterostilbene induced miRNA-mediated regulation of PTEN in prostate cancer. Here, we show that both compounds decrease the levels of endogenous as well as exogenously expressed miR-17, miR-20a and miR-106b thereby upregulating their target PTEN. Using functional luciferase reporter assays, we demonstrate that ectopically expressed miR-17, miR-20a and miR-106b directly target PTEN 3'UTR to reduce its expression, an effect rescued upon treatment with resveratrol and pterostilbene. Moreover, while stable lentiviral expression of miR-17/106a significantly decreased PTEN mRNA and protein levels and conferred survival advantage to the cells, resveratrol and more so pterostilbene was able to dramatically suppress these effects. Further, pterostilbene through downregulation of miR-17-5p and miR-106a-5p expression both in tumors and systemic circulation, rescued PTEN mRNA and protein levels leading to reduced tumor growth in vivo. Our findings implicate dietary stilbenes as an attractive miRNA-mediated chemopreventive and therapeutic strategy, and circulating miRNAs as potential chemopreventive and predictive biomarkers for clinical development in prostate cancer.

Epigenetic potential of resveratrol and analogs in preclinical models of prostate cancer.

Lifestyle, particularly diet, is a risk factor for prostate cancer. Dietary polyphenols such as resveratrol possess anticancer properties and therefore have chemopreventive and therapeutic potential. Resveratrol has pleiotropic effects, exerting its biological activity through multiple pathways and targets, including those associated with cancer. Numerous studies have demonstrated the anticancer effects of resveratrol and, to a lesser extent, its analogs, in tissue culture, while in vivo observations are limited. Here, we provide a concise summary of our results on epigenetic mechanisms of resveratrol and analogs mediated through regulation of chromatin modifier metastasis-associated protein 1 (MTA1) and microRNAs (miRNAs), and highlight the anticancer effects of these compounds in preclinical models of prostate cancer. We suggest that the identified stilbene responsive mechanism-based biomarkers, such as MTA1 and oncogenic miRNAs, may become indicative of treatment efficacy in prostate cancer. Resveratrol analogs with better bioavailability, conferring superior pharmacological potencies and greater anticancer effects, may become stronger candidates for clinical development.

Pterostilbene acts through metastasis-associated protein 1 to inhibit tumor growth, progression and metastasis in prostate cancer.

The development of natural product agents with targeted strategies holds promise for enhanced anticancer therapy with reduced drug-associated side effects. Resveratrol found in red wine, has anticancer activity in various tumor types. We reported earlier on a new molecular target of resveratrol, the metastasis-associated protein 1 (MTA1), which is a part of nucleosome remodeling and deacetylation (NuRD) co-repressor complex that mediates gene silencing. We identified resveratrol as a regulator of MTA1/NuRD complex and re-activator of p53 acetylation in prostate cancer (PCa). In the current study, we addressed whether resveratrol analogues also possess the ability to inhibit MTA1 and to reverse p53 deacetylation. We demonstrated that pterostilbene (PTER), found in blueberries, had greater increase in MTA1-mediated p53 acetylation, confirming superior potency over resveratrol as dietary epigenetic agent. In orthotopic PCa xenografts, resveratrol and PTER significantly inhibited tumor growth, progression, local invasion and spontaneous metastasis. Furthermore, MTA1-knockdown sensitized cells to these agents resulting in additional reduction of tumor progression and metastasis. The reduction was dependent on MTA1 signaling showing increased p53 acetylation, higher apoptotic index and less angiogenesis in vivo in all xenografts treated with the compounds, and particularly with PTER. Altogether, our results indicate MTA1 as a major contributor in prostate tumor malignant progression, and support the use of strategies targeting MTA1. Our strong pre-clinical data indicate PTER as a potent, selective and pharmacologically safe natural product that may be tested in advanced PCa.

Nuclear MTA1 overexpression is associated with aggressive prostate cancer, recurrence and metastasis in African Americans.

Metastasis-associated protein 1 (MTA1), a negative epigenetic modifier, plays a critical role in prostate cancer (PCa) progression. We hypothesized that MTA1 overexpression in primary tumor tissues can predict PCa aggressiveness and metastasis. Immunohistochemical staining of MTA1 was done on archival PCa specimens from University of Mississippi Medical Center and University of Iowa. We found that nuclear MTA1 overexpression was positively correlated with the severity of disease progression reaching its highest levels in metastatic PCa. Nuclear MTA1 overexpression was significantly associated with Gleason > 7 tumors in African Americans but not in Caucasians. It was also a predictor of recurrent disease. We concluded that MTA1 nuclear overexpression may be a prognostic indicator and a future therapeutic target for aggressive PCa in African American men. Our findings may be useful for categorizing African American patients with a higher probability of recurrent disease and metastasis from those who are likely to remain metastasis-free.

Resveratrol and prostate cancer: promising role for microRNAs.

SCOPE: Resveratrol (Res) has anticancer activity in prostate cancer (PCa), which can be attributed to modulation of microRNAs (miRNAs/miRs). miRNAs/miRs are small non-coding RNAs that negatively regulate gene expression. We have analyzed differential miRNA expression in PCa cells treated with Res. METHODS AND RESULTS: Using miRNA microarrays we found that 23 miRNAs were significantly down-regulated and 28 miRNAs were significantly up-regulated after Res treatment. The down-regulated miRs included miR-17-92 and miR-106ab clusters with well recognized oncogenic properties while the up-regulated miRs included several tumor suppressors. Selected miRs were verified by qRT-PCR, including miR-17, miR-20a, miR-20b, miR-106a and miR106b. Since these miRNAs target PTEN (phosphatase and tensin homolog deleted on chromosome 10), we performed Western blot to confirm up-regulation of PTEN in PCa cells. In addition, using TargetScan database, we have identified putative mRNA targets for Res-induced down- and up-regulated miRs. Using a bioinformatics approach, we generated gene networks specifically altered by Res-regulated miRNAs. CONCLUSION: Our results indicate that the dietary compound Res may play an important role in prostate carcinogenesis through modulation of miRNA expression: Res down-regulated oncogenic miRs and up-regulated tumor suppressor miRs in PCa cells. Further in-depth studies are necessary in order to fully recognize the beneficial miRNA-mediated effects of Res in PCa.

gammadelta T cells in cancer immunotherapy: current status and future prospects.

gammadelta T lymphocytes are a distinct T-cell subset that display unique features with respect to T-cell receptor (TCR) gene usage, tissue tropism and antigen recognition. Phosphoantigens contributed by a dysregulated mevalonate pathway or the bacterial nonmevalonate pathway and aminobisphosphonates are capable of activating Vgamma9Vdelta2 T cells. With the aid of synthetic phosphoantigens, large-scale expansion of gammadelta T cells and their adoptive transfer into human hosts is now possible. The present review summarizes triumphs and tribulations of clinical trials using gammadelta T-cell immunotherapy. Adoptive transfer of phosphoantigen-activated gammadelta T cells or coadministration with aminobisphosphonates/cytokines/monoclonal antibodies appear to be promising approaches for cancer immunotherapy. It can be predicted that a comprehensive understanding of the molecular interactions of this unique T-cell subset with other key immune regulators (dendritic cells and regulatory T cells) will provide an impetus to bring this modality of treatment from bench to bedside.