Hepatocellular carcinoma (HCC) is a deadly tumour whose causative agents are generally well known, but whose pathogenesis remains poorly understood. Nevertheless, key genetic alterations are emerging from a heterogeneous molecular landscape, providing information on the tumorigenic process from initiation to progression. Among these molecular alterations, those that affect epigenetic processes are increasingly recognised as contributing to carcinogenesis from preneoplastic stages. The epigenetic machinery regulates gene expression through intertwined and partially characterised circuits involving chromatin remodelers, covalent DNA and histone modifications, and dedicated proteins reading these modifications. In this review, we summarise recent findings on HCC epigenetics, focusing mainly on changes in DNA and histone modifications and their carcinogenic implications. We also discuss the potential drugs that target epigenetic mechanisms for HCC treatment, either alone or in combination with current therapies, including immunotherapies.

Artemisinin and its derivatives (ARTs) were reported to display heme-dependent antitumor activity. On the other hand, histone deacetylase inhibitors (HDACi) were known to be able to promote heme synthesis in erythroid cells. Nevertheless, the effect of HDACi on heme homeostasis in non-erythrocytes remains unknown. We envisioned that the combination of HDACi and artesunate (ARS) might have synergistic antitumor activity through modulating heme synthesis. In vitro studies revealed that combination of ARS and HDACi exerted synergistic tumor inhibition by inducing cell death. Moreover, this combination exhibited more effective antitumor activity than either ARS or HDACi monotherapy in xenograft models without apparent toxicity. Importantly, mechanistic studies revealed that HDACi coordinated with ARS to increase 5-aminolevulinate synthase (ALAS1) expression, and subsequent heme production, leading to enhanced cytotoxicity of ARS. Notably, knocking down ALAS1 significantly blunted the synergistic effect of ARS and HDACi on tumor inhibition, indicating a critical role of ALAS1 upregulation in mediating ARS cytotoxicity. Collectively, our study revealed the mechanism of synergistic antitumor action of ARS and HDACi. This finding indicates that modulation of heme synthesis pathway by the combination based on ARTs and other heme synthesis modulators represents a promising therapeutic approach to solid tumors.

Esophageal cancers are highly aggressive tumors with poor prognosis despite some recent advances in surgical and radiochemotherapy treatment options. This study addressed the feasibility of drugs targeting epigenetic modifiers in esophageal squamous cell carcinoma (ESCC) and esophageal adenocarcinoma (EAC) cells. We tested inhibition of histone deacetylases (HDACs) by SAHA, MS-275, and FK228, inhibition of DNA methyltransferases by Azacytidine (AZA) and Decitabine (DAC), and the effect of combination treatment using both types of drugs. The drug targets, HDAC1/2/3 and DNMT1, were expressed in normal esophageal epithelium and tumor cells of ESCC or EAC tissue specimens, as well as in non-neoplastic esophageal epithelial (Het-1A), ESCC (OE21, Kyse-270, Kyse-410), and EAC (OE33, SK-GT-4) cell lines. In vitro, HDAC activity, histone acetylation, and p21 expression were similarly affected in non-neoplastic, ESCC, and EAC cell lines post inhibitor treatment. Combined MS-275/AZA treatment, however, selectively targeted esophageal cancer cell lines by inducing DNA damage, cell viability loss, and apoptosis, and by decreasing cell migration. Non-neoplastic Het-1A cells were protected against HDACi (MS-275)/AZA treatment. RNA transcriptome analyses post MS-275 and/or AZA treatment identified novel regulated candidate genes (up: BCL6, Hes2; down: FAIM, MLKL), which were specifically associated with the treatment responses of esophageal cancer cells. In summary, combined HDACi/AZA treatment is efficient and selective for the targeting of esophageal cancer cells, despite similar target expression of normal and esophageal cancer epithelium, in vitro and in human esophageal carcinomas. The precise mechanisms of action of treatment responses involve novel candidate genes regulated by HDACi/AZA in esophageal cancer cells. Together, targeting of epigenetic modifiers in esophageal cancers may represent a potential future therapeutic approach.

Histone deacetylases (HDAC) play a crucial role in transcriptional regulation and are often deregulated in many cancers. However, global HDAC enzymatic activity has never been investigated in Chronic Lymphocytic Leukemia (CLL). We measured HDAC activity in protein extracts from CD19+ B-cells purified from 114 CLL patients with a median follow-up of 91 months (range: 11-376). HDAC activity was equivalent in CLL and normal B-cells but higher in patients who died during the study than in living patients (152.1 vs. 65.04 pmol; P = 0.0060). Furthermore, HDAC activity correlated with treatment-free survival (TFS; P = 0.0156) and overall survival (OS; P < 0.0001): patients with low HDAC activity (n = 75) had a median TFS and OS of 101 and > 376 months, respectively, whereas patients with high HDAC activity (n = 39) had a median TFS and OS of 47 and 137 months, respectively. Multivariate analyses indicated that HDAC activity is an independent predictor of OS (hazard ratio = 7.68; P = 0.0017). Finally, HDAC activity increased after B-cell receptor stimulation using IgM, suggesting a role for microenvironment stimuli (n = 10; P = 0.0371). In conclusion, high HDAC activity in CLL B-cells is associated with shorter TFS and OS and is an independent marker of OS, refining the use of other prognostic factors. This work provides a biological base for the use of HDAC inhibitors in CLL treatment.