Energy Restriction-mimetic Agents Induce Apoptosis in Prostate Cancer Cells in Part through Epigenetic Activation of KLF6 Tumor Suppressor Gene Expression

Abstract

Although energy restriction has been recognized as an important target for cancer prevention, the mechanism by which energy restriction-mimetic agents (ERMAs) mediate apoptosis remains unclear. By using a novel thiazolidinedione-derived ERMA, CG-12 (Wei, S., Kulp, S. K., and Chen, C. S. (2010) J. Biol. Chem. 285, 9780 -9791), vis-à-vis 2-deoxyglucose and glucose deprivation, we obtain evidence that epigenetic activation of the tumor suppressor gene Kruppel-like factor 6 (KLF6) plays a role in ERMA-induced apoptosis in LNCaP prostate cancer cells. KLF6 regulates the expression of many proapoptotic genes, and shRNA-mediated KLF6 knockdown abrogated the ability of ERMAs to induce apoptosis. Chromatin immunoprecipitation analysis indicates that this KLF6 transcriptional activation was associated with increased histone H3 acetylation and histone H3 lysine 4 trimethylation occupancy at the promoter region. Several lines of evidence demonstrate that the enhancing effect of ERMAs on these active histone marks was mediated through transcriptional repression of histone deacetylases and H3 lysine 4 demethylases by down-regulating Sp1 expression. First, putative Sp1-binding elements are present in the promoters of the affected histone-modifying enzymes, and luciferase reporter assays indicate that site-directed mutagenesis of these Sp1 binding sites significantly diminished the promoter activities. Second, shRNA-mediated knockdown of Sp1 mimicked the repressive effect of energy restriction on these histone-modifying enzymes. Third, ectopic Sp1 expression protected cells from the repressive effect of CG-12 on these histone-modifying enzymes, thereby abolishing the activation of KLF6 expression. Together, these findings underscore the intricate relationship between energy restriction and epigenetic regulation of tumor suppressor gene expression, which has therapeutic relevance to foster novel strategies for prostate cancer therapy.A hallmark feature of tumorigenesis is the shift of cellular metabolism from oxidative phosphorylation to aerobic glycolysis, the so-called Warburg effect, which provides a growth advantage to cancer cells in the microenvironment (1, 2). Recent evidence indicates that the high rate of glycolysis in tumor cells is attributable to the dysregulation of multiple oncogenic signaling pathways (1), including those mediated by hypoxia-inducible factor 1 (3), Akt (4), c-Myc (5), and p53 (6). This glycolytic shift is considered to be a fundamental property of neoplasia, and the high rate of glucose uptake in glycolytic tumor cells, including those of lung, breast, liver, and colon, is the basis for imaging tumors by [18 F]2-fluoro-2-deoxyglucose positron emission tomography (7). Moreover, in light of the in vivo efficacy of dietary caloric restriction and natural productbased energy restriction-mimetic agents (ERMAs) 2 such as 2-deoxyglucose (2-DG) and resveratrol in suppressing carcinogenesis in various animal models, targeting aerobic glycolysis represents a therapeutically relevant strategy for cancer prev...

Development of a Novel Class of Glucose Transporter Inhibitors

“…Compound 30 -induced ER stress was manifested by increased expression of the two ER stress markers, glucose-regulated protein (GRP)78 and growth arrest- and DNA damage-inducible gene (GADD)153. Moreover, reminiscent of the demonstrated effect of compound 4 on the epigenetic activation of KLF6, 38 compound 30 increased the expression of this tumor suppressor protein in a dose-dependent manner.…”

“…Equally important, compound 30 shared the reported activities of compound 4 , 2-DG, and glucose starvation in eliciting energy restriction-associated cellular responses in LNCaP cells, including β-TrCP-facilitated protein degradation, adenosine monophosphate-activated protein kinase (AMPK) activation, and endoplasmic reticulum (ER) stress. 30, 38 Western blot analysis indicates that compound 30 dose dependently increased β-TrCP expression, leading to the downregulated expression of its substrates cyclin D1 and Sp1, as well as the Sp1 target AR (Fig. 4C).…”

Activation of silenced tumor suppressor genes in prostate cancer cells by a novel energy restriction‐mimetic agent

“…The suppression of energy metabolism by CG‐12 leads to an intricate signaling network mediated by silent information regulator 1, AMP‐activated protein kinase, and oxidative stress, the interplay among which culminates in autophagy and apoptosis in cancer cells. More recently, we demonstrated an epigenetic effect of CG‐12 in cancer cells involving histone acetylation and H3 lysine 4 methylation, leading to the transcriptional activation of Kruppel‐like factor 6 ( KLF6 ) and a series of proapoptotic genes 9. In this study, we report the unique ability of CG‐5, a structurally optimized CG‐12 derivative (Fig.…”

OSU-CG5, a novel energy restriction mimetic agent, targets human colorectal cancer cells in vitro

“…These responses include β-TrCP-facilitated protein degradation, ER stress and adenosine monophosphate-activated protein kinase (AMPK) activation [9,17] . OSU-CG5 and resveratrol treatment led to a dose-dependent increase in β-TrCP expression, which led to the downregulation of its substrates, cyclin D1 and Sp1 (Figure 4).…”