Epigenomic interplay in tumor heterogeneity: Potential of epidrugs as adjunct therapy.

"Heterogeneity" in tumor mass has immense importance in cancer progression and therapy. The impact of tumor heterogeneity is an emerging field and not yet fully explored. Tumor heterogeneity is mainly considered as intra-tumor heterogeneity and inter-tumor heterogeneity based on their origin. Intra-tumor heterogeneity refers to the discrepancy within the same cancer mass while inter-tumor heterogeneity refers to the discrepancy between different patients having the same tumor type. Both of these heterogeneity types lead to variation in the histopathological as well as clinical properties of the cancer mass which drives disease resistance towards therapeutic approaches. Cancer stem cells (CSCs) act as pinnacle progenitors for heterogeneity development along with various other genetic and epigenetic parameters that are regulating this process. In recent times epigenetic factors are one of the most studied parameters that drive oxidative stress pathways essential during cancer progression. These epigenetic changes are modulated by various epidrugs and have an impact on tumor heterogeneity. The present review summarizes various aspects of epigenetic regulation in the tumor microenvironment, oxidative stress, and progression towards tumor heterogeneity that creates complications during cancer treatment. This review also explores the possible role of epidrugs in regulating tumor heterogeneity and personalized therapy against drug resistance.

Nutritive vitamins as epidrugs.

Epigenetic modifications play an important role in disease pathogenesis and therefore are a focus of intense investigation. Epigenetic changes include DNA, RNA, and histone modifications along with expression of non-coding RNAs. Various factors such as environment, diet, and lifestyle can influence the epigenome. Dietary nutrients like vitamins can regulate both physiological and pathological processes through their direct impact on epigenome. Vitamin A acts as a major regulator of above-mentioned epigenetic mechanisms. B group vitamins including biotin, niacin, and pantothenic acid also participate in modulation of various epigenome. Further, vitamin C has shown to modulate both DNA methylation and histone modifications while few reports have also supported its role in miRNA-mediated pathways. Similarly, vitamin D also influences various epigenetic modifications of both DNA and histone by controlling the regulatory mechanisms. Despite the information that vitamins can modulate the epigenome, the detailed mechanisms of vitamin-mediated epigenetic regulations have not been explored fully and hence further detailed studies are required to decipher their role at epigenome level in both normal and disease pathogenesis. The current review summarizes the available literature on the role of vitamins as epigenetic modifier and highlights the key evidences for developing vitamins as potential epidrugs.

Proproliferative function of adaptor protein GRB10 in prostate carcinoma.

Growth factor receptor-binding protein 10 (GRB10) is a well-known adaptor protein and a recently identified substrate of the mammalian target of rapamycin (mTOR). Depletion of GRB10 increases insulin sensitivity and overexpression suppresses PI3K/Akt signaling. Because the major reason for the limited efficacy of PI3K/Akt-targeted therapies in prostate cancer (PCa) is loss of mTOR-regulated feedback suppression, it is therefore important to assess the functional importance and regulation of GRB10 under these conditions. On the basis of these background observations, we explored the status and functional impact of GRB10 in PCa and found maximum expression in phosphatase and tensin homolog (PTEN)-deficient PCa. In human PCa samples, GRB10 inversely correlated with PTEN and positively correlated with pAKT levels. Knockdown of GRB10 in nontumorigenic PTEN null mouse embryonic fibroblasts and tumorigenic PCa cell lines reduced Akt phosphorylation and selectively activated a panel of receptor tyrosine kinases. Similarly, overexpression of GRB10 in PTEN wild-type PCa cell lines accelerated tumorigenesis and induced Akt phosphorylation. In PTEN wild-type PCa, GRB10 overexpression promoted mediated PTEN interaction and degradation. PI3K (but not mTOR) inhibitors reduced GRB10 expression, suggesting primarily PI3K-driven regulation of GRB10. In summary, our results suggest that GRB10 acts as a major downstream effector of PI3K and has tumor-promoting effects in prostate cancer.-Khan, M. I., Al Johani, A., Hamid, A., Ateeq, B., Manzar, N., Adhami, V. M., Lall, R. K., Rath, S., Sechi, M., Siddiqui, I. A., Choudhry, H., Zamzami, M. A., Havighurst, T. C., Huang, W., Ntambi, J. M., Mukhtar, H. Proproliferatve function of adaptor protein GRB10 in prostate carcinoma.

Hypoxia driven glycation: Mechanisms and therapeutic opportunities.

Tumor masses are deprived of oxygen and characterized by enhanced glucose uptake followed by glycolysis. Elevated glucose levels induce non-enzymatic glycosylation or glycation of proteins which leads to accumulation of advanced glycation end products (AGE). These AGE molecules bind to their respective receptors called the receptor for advanced glycation end products (RAGE) and initiate several aberrant signaling pathways leading to onset of diseases such as diabetes, Alzheimer's, atherosclerosis, heart failure and cancer. The role of AGE in cancer progression is being extensively studied in recent years. As cancer cells are hypoxic in nature and adapted to glycolysis, which induces glycation, its effects need to be understood in greater detail. Since AGE-RAGE signaling is involved in cancer progression, inhibition of AGE-RAGE interaction could be a potential therapeutic target. The purpose of this review is to highlight the role of AGE-RAGE interaction in hypoxic cancer cells.

Acetylation-mediated Siah2 stabilization enhances PHD3 degradation in Helicobacter pylori-infected gastric epithelial cancer cells.

Gastric epithelial cells infected with Helicobacter pylori acquire highly invasive and metastatic characteristics. The seven in absentia homolog (Siah)2, an E3 ubiquitin ligase, is one of the major proteins that induces invasiveness of infected gastric epithelial cells. We find that p300-driven acetylation of Siah2 at lysine 139 residue stabilizes the molecule in infected cells, thereby substantially increasing its efficiency to degrade prolyl hydroxylase (PHD)3 in the gastric epithelium. This enhances the accumulation of an oncogenic transcription factor hypoxia-inducible factor 1α (Hif1α) in H. pylori-infected gastric cancer cells in normoxic condition and promotes invasiveness of infected cells. Increased acetylation of Siah2, Hif1α accumulation, and the absence of PHD3 in the infected human gastric metastatic cancer biopsy samples and in invasive murine gastric cancer tissues further confirm that the acetylated Siah2 (ac-Siah2)-Hif1α axis is crucial in promoting gastric cancer invasiveness. This study establishes the importance of a previously unrecognized function of ac-Siah2 in regulating invasiveness of H. pylori-infected gastric epithelial cells.-Kokate, S. B., Dixit, P., Das, L., Rath, S., Roy, A. D., Poirah, I., Chakraborty, D., Rout, N., Singh, S. P., Bhattacharyya, A. Acetylation-mediated Siah2 stabilization enhances PHD3 degradation in Helicobacter pylori-infected gastric epithelial cancer cells.

Targeting epigenome with dietary nutrients in cancer: Current advances and future challenges.

Tumorigenesis and epigenetic are closely linked with each other. Epigenetic changes are potential regulators of gene expression without involving any change in the DNA itself. More interestingly, epigenetic changes are reversible heritable changes which pass through generations. Many dietary bioactive ingredients regulate epigenetic control of cells and influence biochemical processes. Correlation between epigenetic regulation and cancer onset has been well established. Recent studies provide important information on the role of bioactive dietary components in cancer prevention and therapy. Several bioactive components are responsible for modification of the epigenome by affecting DNA methylation, histone modification, micro RNAs (miRNAs) and long non-coding RNAs (lncRNAs). This review summarizes recent advancements in this field and describes the role of many bioactive components in regulating human epigenome and how these modifications can be exploited for prevention and treatment of cancer.

ETS2 and Twist1 promote invasiveness of Helicobacter pylori-infected gastric cancer cells by inducing Siah2.

Helicobacter pylori infection is one of the most potent factors leading to gastric carcinogenesis. The seven in absentia homologue (Siah2) is an E3 ubiquitin ligase which has been implicated in various cancers but its role in H. pylori-mediated gastric carcinogenesis has not been established. We investigated the involvement of Siah2 in gastric cancer metastasis which was assessed by invasiveness and migration of H. pylori-infected gastric epithelial cancer cells. Cultured gastric cancer cells (GCCs) MKN45, AGS and Kato III showed significantly induced expression of Siah2, increased invasiveness and migration after being challenged with the pathogen. Siah2-expressing stable cells showed increased invasiveness and migration after H. pylori infection. Siah2 was transcriptionally activated by E26 transformation-specific sequence 2 (ETS2)- and Twist-related protein 1 (Twist1) induced in H. pylori-infected gastric epithelial cells. These transcription factors dose-dependently enhanced the aggressiveness of infected GCCs. Our data suggested that H. pylori-infected GCCs gained cell motility and invasiveness through Siah2 induction. As gastric cancer biopsy samples also showed highly induced expression of ETS2, Twist1 and Siah2 compared with noncancerous gastric tissue, we surmise that ETS2- and Twist1-mediated Siah2 up-regulation has potential diagnostic and prognostic significance and could be targeted for therapeutic purpose.

Cobalt chloride-mediated protein kinase Cα (PKCα) phosphorylation induces hypoxia-inducible factor 1α (HIF1α) in the nucleus of gastric cancer cell.

Hypoxia promotes cancer progression, and metastasis. The major protein expressed in hypoxic solid cancer is hypoxia-inducible factor 1 (HIF1). We show that enhanced phosphorylation of a conventional protein kinase C isoform, PKCα, at threonine 638 (T(638)) by hypoxia-mimetic cobalt chloride induces HIF1α in nuclei of gastric epithelial cells (GECs). Moreover, phospho-T(638)-PKCα (P-PKCα) interacts with p300-HIF1α complex in the nuclei of hypoxic GECs and PKCα phosphorylation at T(638) enhances transcriptional activity of HIF1α. High P-PKCα expression in neoplastic gastric cancer biopsy samples as compared to nonneoplastic samples suggests that P-PKCα might act as an indicator of gastric cancer progression.

Regulation of Noxa-mediated apoptosis in Helicobacter pylori-infected gastric epithelial cells.

Helicobacter pylori induces the antiapoptotic protein myeloid cell leukemia 1 (Mcl1) in human gastric epithelial cells (GECs). Apoptosis of oncogenic protein Mcl1-expressing cells is mainly regulated by Noxa-mediated degradation of Mcl1. We wanted to elucidate the status of Noxa in H. pylori-infected GECs. For this, various GECs such as AGS, MKN45, and KATO III were either infected with H. pylori or left uninfected. The effect of infection was examined by immunoblotting, immunoprecipitation, chromatin immunoprecipitation assay, in vitro binding assay, flow cytometry, and confocal microscopy. Infected GECs, surgical samples collected from patients with gastric adenocarcinoma as well as biopsy samples from patients infected with H. pylori showed significant up-regulation of both Mcl1 and Noxa compared with noninfected samples. Coexistence of Mcl1 and Noxa was indicative of an impaired Mcl-Noxa interaction. We proved that Noxa was phosphorylated at Ser(13) residue by JNK in infected GECs, which caused cytoplasmic retention of Noxa. JNK inhibition enhanced Mcl1-Noxa interaction in the mitochondrial fraction of infected cells, whereas overexpression of nonphosphorylatable Noxa resulted in enhanced mitochondria-mediated apoptosis in the infected epithelium. Because phosphorylation-dephosphorylation can regulate the apoptotic function of Noxa, this could be a potential target molecule for future treatment approaches for H. pylori-induced gastric cancer.

Arsenic album 30C exhibits crystalline nano structure of arsenic trioxide and modulates innate immune markers in murine macrophage cell lines.

Macrophages are associated with innate immune response and M1-polarized macrophages exhibit pro-inflammatory functions. Nanoparticles of natural or synthetic compounds are potential triggers of innate immunity. As2O3 is the major component of the homeopathic drug, Arsenic album 30C.This has been claimed to have immune-boosting activities, however, has not been validated experimentally. Here we elucidated the underlying mechanism of Ars. alb 30C-mediated immune priming in murine macrophage cell line. Transmission Electron Microscopy (TEM) and X-ray diffraction (XRD) used for the structural analysis of the drug reveals the presence of crystalline As2O3 nanoparticles of cubic structure. Similarly, signatures of M1-macrophage polarization were observed by surface enhanced Raman scattering (SERS) in RAW 264.7 cells with concomitant over expression of M1 cell surface marker, CD80 and transcription factor, NF-κB, respectively. We also observed a significant increase in pro-inflammatory cytokines like iNOS, TNF-α, IL-6, and COX-2 expression with unaltered ROS and apoptosis in drug-treated cells. Enhanced expression of Toll-like receptors 3 and 7 were observed both in transcriptional and translational levels after the drug treatment. In sum, our findings for the first time indicated the presence of crystalline As2O3 cubic nanostructure in Ars. alb 30C which facilitates modulation of innate immunity by activating macrophage polarization.

Methods to Evaluate the Effects of HAT/KAT Inhibition on SIAH2-Driven Reactive Oxygen Species Generation in Helicobacter pylori-Infected Gastric Epithelial Cells.

Helicobacter pylori infection is one of the leading factors that promotes, among other diseases, gastric cancer (GC). Infection of gastric epithelial cells (GECs) by H. pylori enhances the expression as well as acetylation of the E3 ubiquitin ligase SIAH2 which promotes GC progression. The histone acetyltransferase (HAT) activity of p300 catalyzes SIAH2 acetylation following H. pylori infection. Since reactive oxygen species (ROS) generation in H. pylori-infected GECs accelerates GC progression, acetylation-mediated SIAH2 regulation might be a crucial modifier of ROS generation in the infected GECs. Here, we describe a compendium of methods to evaluate the effects of HAT/lysine acetyl transferase (KAT) inhibitors (HAT/KATi) on SIAH2-mediated ROS regulation in H. pylori-infected GECs.

CTK7A, a curcumin derivative, can be a potential candidate for targeting HIF-1α/p300 complex: Evidences from in vitro and computational studies.

Hypoxia-inducible factor-1 (HIF-1) is a transcription factor which plays a critical role in several biochemical pathways, and consists of oxygen-dependent alpha (α) and a constitutively expressed beta (ß) subunit. Under hypoxic conditions, HIF-1α is stabilized and forms a complex with ß subunit and this complex is associated with cancer progression. HIF-1α activity is mainly regulated by its transcriptional co-activator p300 which has histone acetyl-transferase (HAT) activity. p300 HAT activity is very crucial for p300 auto-acetylation and subsequently its interaction with its partner molecule HIF-1α as well as proapoptotic protein p53. p300 is a multi-domain protein and CH1 domain of p300 is the interacting partner of the C-terminal domain (CTD) of HIF-1α as well as p53. Several p300 HAT inhibitors are reported to suppress p300 auto-acetylation which inhibits its interaction with associated partners. We demonstrated that the p300 HAT inhibitor CTK7A down-regulated p300 auto-acetylation, HIF-1α accumulation as well as activity in gastric cancer cell lines. Protein-protein interaction and molecular docking studies revealed a significant decrease in the binding energy of full-length p300 as well as p300-CH1 and HIF-1α-CTD complex in presence of CTK7A. Further, SwissADME, evaluates the drug-likeliness property of CTK7A by analyzing its lipophilicity, size, polarity, solubility, saturation, and flexibility. Our in vitro and in silico data support reduced HIF-1α-p300 interaction in the presence of CTK7A. Hence, CTK7A might be playing a crucial role in down-regulating HIF-1α activity and can be a prospective anticancer drug.

Factors regulating dynamics of angiotensin-converting enzyme-2 (ACE2), the gateway of SARS-CoV-2: Epigenetic modifications and therapeutic interventions by epidrugs.

Angiotensin-converting enzyme-2 (ACE2) is one of the major components of the renin-angiotensin system (RAS) and participates in the physiological functions of the cardiovascular system and lungs. Recent studies identified ACE2 as the receptor for the S-protein of the novel severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and thus acts as the gateway for viral entry into the human body. Virus infection causes an imbalance in the RAS axis and induces acute lungs injury and fibrosis. Various factors regulate ACE2 expression patterns as well as control its epigenetic status at both transcription and translational levels. This review is mainly focused on the impact of environmental toxicants, drugs, endocrine disruptors, and hypoxia as controlling parameters for ACE2 expression and its possible modulation by epigenetic changes which are marked by DNA methylation, histone modifications, and micro-RNAs (miRNAs) profile. Furthermore, we have emphasized on interventions of various phytochemicals and bioactive compounds as epidrugs that regulate ACE2-S-protein interaction and thereby curb viral infection. Since ACE2 is an important component of the RAAS axis and a crucial entry point of SARS-CoV-2, the dynamics of ACE2 expression in response to various extrinsic and intrinsic factors are of contemporary relevance. We have collated updated information on ACE2 expression modulated by epidrugs, and urge to take over further studies on these important physiological regulators to unravel many more systemic linkages related to both metabolic and infectious diseases, in general and SARS-CoV-2 in particular for further development of targeted interventions.

AKT Inhibition Modulates H3K4 Demethylase Levels in PTEN-Null Prostate Cancer.

Hyperactivated AKT kinase due to loss of its negative regulator PTEN influences many aspects of cancer biology, including chromatin. AKT primarily regulates acetyl-CoA production and phosphorylates many histone-modulating enzymes, resulting in their activation or inhibition. Therefore, understanding the therapeutic impact of AKT inhibition on chromatin-related events is essential. Here, we report that AKT inhibition in prostate-specific PTEN knockout mice significantly induces di- and trimethylation of H3K4 with concomitant reduction in H3K9 acetylation. Mechanistically, we observed that AKT inhibition reduces expression of the H3K4 methylation-specific histone demethylases KDM5 family, especially KDM5B expression at transcriptional levels. Furthermore, we observed that AKT negatively regulates miR-137 levels, which transcriptionally represses KDM5B expression. Overexpression of miR-137 significantly reduced KDM5B and increased H3K4 methylation levels but failed to change AKT phosphorylation. Overall, we observed that AKT transcriptionally regulates KDM5B mainly via repression of miR-137. Our data identify a mechanism by which AKT kinase modulates the prostate cancer epigenome through regulating H3K4 methylation. Additional studies on AKT inhibition-mediated induction of H3K4 methylation will help in designing strategies to enhance the therapeutic efficacy of PI3K/AKT inhibitors.

Inhibition of histone/lysine acetyltransferase activity kills CoCl2-treated and hypoxia-exposed gastric cancer cells and reduces their invasiveness.

Hypoxia enhances immortality and metastatic properties of solid tumors. Deregulation of histone acetylation has been associated with several metastatic cancers but its effect on hypoxic responses of cancer cells is not known. This study aimed at understanding the effectiveness of the hydrazinocurcumin, CTK7A, an inhibitor of p300 lysine/histone acetyltransferase (KAT/HAT) activity, in inducing apoptosis of gastric cancer cells (GCCs) exposed to cobalt chloride (CoCl2), a hypoxia-mimetic chemical, or 1% O2. Here, we show that CTK7A-induced hydrogen peroxide (H2O2) generation in CoCl2-exposed and invasive gastric cancer cells (GCCs) leads to p38 MAPK-mediated Noxa expression and thereafter, mitochondrial apoptotic events. Noxa induction in normal immortalized gastric epithelial cells after CTK7A and hypoxia-exposure is remarkably less in comparison to similarly-treated GCCs. Moreover, hypoxia-exposed GCCs, which have acquired invasive properties, become apoptotic after CTK7A treatment to a significantly higher extent than normoxic cells. Thus, we show the potential of CTK7A in sensitizing hypoxic and metastatic GCCs to apoptosis induction.