Emerging epigenetics, metabolomics and oxidative stress in cancer chemoprevention: the potential of dietary phytochemicals
Creators: name: Wang, Lujing
Publisher: name: No Publisher Supplied
Year: 2022

description: Cancer chemoprevention is defined as the strategy to block or slow the onset of premalignant tumors using relatively nontoxic chemical substance. Recently, accumulating experimental evidence has suggested that epigenetic alterations are involved in cancer development. The scope of epigenetics lies on the molecular interface between genetics and environmental factors; external factors switch genes on and off by influencing how cells read the genes, therefore, to regulate the transcriptomic profiles of organisms. DNA methylation, histone covalent modification and remodeling as well as miRNA-mediated gene silencing represents the major mechanisms that play important roles in epigenetic control of gene expression. Increasing evidence supporting redox imbalance and aberrant reactive oxygen species (ROS) are closely linked to the oncogenesis of various cancers. ROS-induced oxidative stress regulates multiple redox signaling pathways that ultimately impact on cellular metabolic rewiring. Interestingly, epigenetic modifications such as DNA methylation and histone acetylation are sensitive to cellular metabolic status. Strong molecular link between metabolic reprogramming and epigenetic modifications through key metabolic intermediates, such as nicotinamide adenine dinucleotide (NAD), α-ketoglutarate (aKG), S-adenosyl methionine (SAM), and Acetyl-CoA (AcCoA), which are co-factors for the epigenetic enzymes and work as hubs between epigenetic processes and oxidative stress responses has been reported.This thesis focused on elucidating the underlying intricate biological connectivity between metabolomic, epigenomic and transcriptomic regulation in blocking pro-tumorigenic signaling and elicit cancer-protective effects by dietary phytochemicals including Fucoxanthin (FX), Butyrate (B), Ursolic acid (UA) and Curcumin. The Nuclear Factor Erythroid-2 like 2 (NRF2)-ARE (antioxidant response element) signaling axis plays a critical role in many phytochemical-mediated cellular defense against oxidative and chemical stresses via induction of cellular defense and antioxidant enzymes. Normal healthy cells often maintain a low basal Nrf2 expression/activation, while cancer cells hold high intrinsic Nrf2 activity. Activation of Nrf2 protects normal cells from oxidative stress and cell damage, while activation of Nrf2 in cancer cells enhances drug resistance and cancer cell survival. Inhibition of Nrf2, on the other hand, sensitizes cancer cells to chemotherapeutic agents. Specifically, studies have shown that the higher the presence of the phase II detoxifying/antioxidant enzymes including heme oxygenase 1 (HO-1), NAD(P)H quinone oxidoreductase 1 (NQO-1), NADPH, and superoxide dismutase (SOD) in the normal healthy tissue/cells, the less susceptible to cancers. NRF2 and its interaction with AREs increase the transcription of these enzymes in normal condition. Previous in vivo studies have shown that NRF2 knockout (KO) mice exhibit significantly lower levels of cellular defense in various tissues, with an increased risk of developing carcinogen-induced cancers. Oppositely, cancerous and otherwise diseased cells often exhibit dysfunctional NRF2 regulation and overexpression of NRF2 in the nucleus of cells. This overabundance of NRF2 in the nucleus leads to increased expression of antioxidant response and cell survival genes often leading to drug insensitivity or resistance. This has become a common area of research for many groups and several small molecule inhibitors of NRF2 have been developed and implicated in overcoming multidrug resistance in cancer.Our group utilized multi-omics approaches to evaluate the role of Nrf2 and the impact of FX on tumor promoter 12-O-tetradecanolyphorbal-13-acetate (TPA)-induced normal skin cell JB6 transformation. FX blocked TPA-induced ROS and oxidized glutathione (GSSG)/reduced glutathione (GSH), an oxidative stress index, in Nrf2 wild-type (WT) but not Nrf2-Knockdown (KD) cells. Both Nrf2 KD and TPA altered cellular metabolisms and metabolites which are tightly coupled to epigenetic machinery. The suppressive effects of FX on TPA-enhanced SAM (S-adenosyl methionine)/SAH (S-adenosylhomocysteine), a biosensor of the cellular metabolic state to influence the activity of methyltransferase enzymes, was abrogated by Nrf2 KD indicating Nrf2 plays a critical role in FX-mediated metabolic rewiring and its potential consequences on epigenetic reprogramming. FX/Nrf2’s redox signaling drives metabolic rewiring causing epigenetic and transcriptomic reprogramming potentially contributing to the protection of TPA-induced JB6 cellular transformation skin cancer model. Besides the FX/Nrf2-mediated cancer protective effects in normal healthy cells, we also found that butyrate (B), a short-chain fatty acid produced from dietary fiber, increased NRF2 negative regulator Kelch-like ECH-associated protein 1 (KEAP1) expression (KEAP1-NRF2 signaling pathway) through inhibiting it’s promoter CpG methylation which further result in the NRF2 inhibition in colorectal cancer (CRC) HCT116 cells. Associative analysis of DEGs (differentially expressed genes) from RNA-seq and DMRs (differentially methylated regions) from CpG methyl-seq identified the tumor suppressor gene ATP binding cassette A1 (ABCA1) and tumor promote gene Early growth response protein 3 (EGR3) were correlated with their promoters’ CpG methylation indicating B regulates cancer markers through modulating their promoter methylation. B activated the mitochondrial tricarboxylic acid (TCA) cycle while inhibited the methionine metabolism which are both tightly coupled to the epigenetic machinery. B also regulated the epigenetic enzymes/genes including DNMT1, HAT1, KDM1A, KDM1B and TET1. Altogether, B’s regulation of metabolites coupled to the epigenetic enzymes illustrates the underlying biological connectivity between metabolomics and epigenomics. And B regulates KEAP1-NRF2 signaling, drives metabolic rewiring, CpG methylomic and transcriptomic reprogramming contributing to the overall antitumor effect in CRC cell model. In addition to the regulation effects of FX and B via NRF2 signaling pathways, UA as a natural pentacyclic triterpenoid carboxylic acid phytochemical also known to possess antioxidant, anti-inflammatory, and cancer-preventive/anti-cancer effects. We found UA protects against Pten (phosphatase and tensin homologue deleted on chromosome 10; one of the most frequently mutated/deleted tumor suppressor genes which is mutated in 30-63% of primary prostate cancer (PCa)), KO-induced tumorigenesis at different stages of PCa. Epigenomic CpG methyl-seq revealed UA attenuated Pten KO-induced differentially methylated regions (DMRs) profiles. Transcriptomic RNA-seq showed UA abrogated Pten KO-induced differentially expressed genes (DEGs) of PCa related oncogenes’ Has3, Cfh and Msx1 overexpression indicating UA plays a crucial role in Pten KO-mediated gene regulation and its potential consequences on cancer prevention. Pathway analysis revealed UA elicits stronger protective effect on Pten KO-induced inflammatory- and cancer-associated signaling pathways modulation at early stage (12-week age) compared to late stage (20-week age) of PCa. Associative analysis of DEGs and DMRs identified the mRNA expression of tumor suppressor genes BDH2, and oncogenes Ephas, Isg15 and Nos2 were correlated with the promoter CpG methylation status in the early-stage comparison groups indicating UA could regulate the tumor promoter/suppressor genes through modulating their promoter methylation at early stage of prostate tumorigenesis. Metabolomic study showed UA attenuated Pten KO-regulated cancer-associated metabolism like purine metabolism/metabolites correlating with RNAseq findings, glycolysis/gluconeogenesis metabolism, as well as epigenetic-related metabolites pyruvate and lactate indicating UA plays a critical role in Pten KO-mediated metabolic and epigenetic reprogramming and its consequences on cancer development. Given that oxidative stress and inflammation-mediated epigenetic, metabolomic and transcriptomic reactions are important (micro) environmental factors in malignancy transformation, understanding the role of redox and inflammation as well as the metabolic metabolisms in epigenetic regulation could bring novel insights in cancer prevention. In summary, the phytochemicals used in current studies including FX, B and UA exert the antioxidant and cancer prevention effects via rewiring the metabolomic and reprogramming epigenomic and transcriptomic profiles in different cancer cell and animal models, and Nrf2 play roles in these regulations via regulating the redox hemostasis. The detailed mechanisms of these corresponding regulation effects were investigated in the following specific chapters.