Metastasis-associated protein 1 is known to play role in prostate cancer progression and metastasis. Studies of MTA1-mediated prostate cancer tumorigenesis, progression and metastasis have been hampered by a lack of suitable experimental system. Here we describe a new conditional mouse model that shows strong MTA1 expression in the prostate. This model was generated through MTA1 transgene knocked into the mouse ROSA26 (R26) locus using CAG-LoxP-Stop-LoxP (LSL)-2HA-MTA1-T2A-GFP-pA construct (Applied StemCell,Inc, Menlo Park, CA). Briefly, after generation of a genetically modified founder mouse line Rosa26-3attP, resulted MTA1 transgene embryos were injected into foster mice to produce MTA1 founders, designated as R26-LSL-MTA1. To monitor tumor formation and development in live animals by using IVIS, we have crossbred R26-LSL-MTA1 with Pb-Cre⁺;Luc and generated transgenic lines that express MTA1 and Luc upon Cre-mediated removal of the LSL cassette, specifically in the prostate epithelium (Pb-Cre⁺;R26^MTA1;R26^Luc) (MTA1-tg). We confirmed prostate-specific expression of the MTA1 transgene on mRNA and protein levels. We evaluated prostate histology for differences in histology of control normal prostates (Pb-Cre^-; MTA1-tg) and MTA1-tg mice at as early as 13-weeks of age. MTA1 overexpressing prostate showed signs of mPIN characterized by glandular structures composed of atypical proliferating luminal cells that fill out the lumen. We detected differences in cellular proliferation, as determined by increased PCNA and decreased p27 as well as Ki67 immunofluorescent staining in MTA1-tg prostate. Moreover, expression of several oncogenic MTA1-guided molecules was increased in the prostates of MTA1-tg mice. We have previously reported on the role of MTA1-mediated signaling in prostate tumor progression in the prostate-specific Pten heterozygous (Pb-Cre⁺; Pten^+/f) and Pten null (Pb-Cre⁺; Pten^f/f ) mice. To determine the role of MTA1 overexpression in the context of PTEN loss, we generated MTA1-tg; Pten^+/f and MTA1-tg; Pten^f/f mice to evaluate whether MTA1overexpression in the prostate promotes an earlier onset, greater total incidence, accelerated tumor growth rate and metastasis. We predict that MTA1 overexpression will cause higher activation of downstream pathways to promote cancer cell survival, which will result in increased incidence of invasive adenocarcinoma and probably lymph node metastasis. Mice will be sacrificed at different time points for prostate tissue and lymph nodes isolation and subsequent histopathological, immunochemical and molecular evaluation. Citation Format: Avinash Kumar, Swati Dhar, Nasir A. Butt, Pravin R. Phadatare, Kshiti Dholakia, Jogarao Vedula, Virginia Donovan, Anait S. Levenson. A novel MTA1 knock-in mouse model for the mechanistic and therapeutic studies of MTA1-driven prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5107.

The overexpression of metastasis-associated protein 1 (MTA1) in prostate cancer (PCa) contributes to tumor aggressiveness and metastasis. We have reported the inhibition of MTA1 by resveratrol and its potent analog pterostilbene in vitro and in vivo. We have demonstrated that pterostilbene treatment blocks the progression of prostatic intraepithelial neoplasia and adenocarcinoma in mouse models by inhibiting MTA1 expression and signaling. In the current study, we investigated the MTA1 targeted anticancer effects of Gnetin C, a resveratrol dimer, in comparison with resveratrol and pterostilbene. Using DU145 and PC3M PCa cells, we found that Gnetin C downregulates MTA1 more potently than resveratrol and pterostilbene. Further, Gnetin C demonstrated significant MTA1-mediated inhibitory effect on cell viability, colony formation, and migration, while showing a more potent induction of cell death than resveratrol or pterostilbene. In addition, we identified Gnetin C-induced substantial ETS2 (erythroblastosis E26 transformation-specific 2) downregulation, which is not only MTA1-dependent, but is also independent of MTA1 as a possible mechanism for the superior anticancer efficacy of Gnetin C in PCa. Together, these findings underscore the importance of novel potent resveratrol dimer, Gnetin C, as a clinically promising agent for the future development of chemopreventive and possibly combinatorial therapeutic approaches in PCa.

Although the link between diet and cancer is complex, epidemiological data confirm that diet is a risk factor for prostate cancer and indicate a reduced prostate cancer incidence associated with a diet rich in vegetables and fruits. Because of the known protective effect of grape seed extract (GSE) against prostate cancer, we evaluated the effects of grape powder extract (GPE) on cell viability, proliferation, and metastatic capability. Importantly, we explored the possible novel mechanism of GPE through metastasis-associated protein 1 (MTA1) downregulation in prostate cancer, since our previous studies indicated resveratrol (Res)- and pterostilbene (Pter)-induced MTA1-mediated anticancer activities in prostate cancer. We found that GPE inhibited the cell viability and growth of prostate cancer cells only at high 100 μg/mL concentrations. However, at low 1.5–15 μg/mL concentrations, GPE significantly reduced the colony formation and wound healing capabilities of both DU145 and PC3M cells. Moreover, we found that GPE inhibited MTA1 in a dose-dependent manner in these cells, albeit with considerably less potency than Res and Pter. These results indicate that stilbenes such as Res and Pter specifically and potently inhibit MTA1 and MTA1-associated proteins compared to GPE, which contains low concentrations of Res and mainly consists of other flavonoids and anthocyanidins. Our findings support continued interest in GPE as a chemopreventive and anti-cancer agent against prostate cancer but also emphasize the unique and specific properties of stilbenes on MTA1-mediated anticancer effects on prostate cancer.

Overexpression of chromatin modifier protein, metastasis-associated protein 1 (MTA1) in prostate cancer contributes to tumor aggressiveness and metastasis. MTA1 ChiP-Seq analysis identified downstream targets that are transcriptionally regulated by MTA1 and suggested a link between MTA1 and ETS2. The effects of ETS2 in cancer are context-dependent and both oncogenic and tumor suppressive functions have been described. We have shown that there is a positive correlation between MTA1 and ETS2 using loss of function studies and various preclinical models of prostate cancer. Our conclusion that MTA1 functions as a co-activator for regulating ETS2 expression in prostate cancer leading to promotion of prostate cancer progression, prompted us to look for pharmacological inhibitors of MTA1/ETS2 axis. In our previous studies, we reported inhibition of MTA1 by resveratrol and its potent analog pterostilbene in vitro and in vivo. We have demonstrated that pterostilbene treatment blocks the progression of PIN and adenocarcinoma in xenografts and transgenic mouse models by inhibiting MTA1 expression and signaling. Here, we found that Gnetin C, double-resveratrol, has more potent inhibition of MTA1/ETS2 axis than other stilbenes. Gnetin C, a resveratrol dimer, is found abundantly in melinjo plant widely cultivated in Southeast Asia; its seeds and fruits are common ingredients in Indonesian culinary. As resveratrol and pterostilbene, Gnetin C has been reported to possess anti-inflammatory and anticancer activity, however it has not been considered as chemopreventive agent in prostate cancer. Using two prostate cancer cell lines, namely DU145 and PC3M cells, we found that Gnetin C shows significant inhibitory effect on cell proliferation, more potent effects in colony formation and wound healing assays than resveratrol or pterostilbene. Importantly, Gnetin C specifically and strongly downregulates MTA1 and ETS2 expression in prostate cancer cells. Taken together, our findings implicate the potential of Gnetin C in MTA1/ETS2-mediated chemoprevention and therapy in prostate cancer. Citation Format: Urvi Kolhatkar, Kshiti Dholakia, Gabriela Sikorska, Avinash Kumar, Luis A. Martinez, Anait S. Levenson. Gnetin C as a candidate for targeted chemopreventive and therapeutic measures in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 253.