Sirtuin functions and modulation: from chemistry to the clinic

Sirtuins are NAD+ -dependent histone deacetylases regulating important metabolic pathways in prokaryotes and eukaryotes and are involved in many biological processes such as cell survival, senescence, proliferation, apoptosis, DNA repair, cell metabolism, and caloric restriction. The seven members of this family of enzymes are considered potential targets for the treatment of human pathologies including neurodegenerative diseases, cardiovascular diseases, and cancer. Furthermore, recent interest focusing on sirtuin modulators as epigenetic players in the regulation of fundamental biological pathways has prompted increased efforts to discover new small molecules able to modify sirtuin activity. Here, we review the role, mechanism of action, and biological function of the seven sirtuins, as well as their inhibitors and activators

Simplification of the tetracyclic SIRT1-selective inhibitor MC2141: Coumarin- and pyrimidine-based SIRT1/2 inhibitors with different selectivity profile

n/

Anticancer activities of anthocyanin extract from genotyped Solanum tuberosum L. "Vitelotte"

n/

Epi-Regulation of Cell Death in Cancer

How do organisms regulate the correct balance between the production of “new” cells and the elimination of the “old” ones, remains an important biology issue under investigation. Cell(s) death represents a fundamental process involved in organism development and cell homeostasis, whose alteration is considered one hallmark of cancer and lead to drug resistance and consequently treatment failure. The recent re-classification of cell death has identified new molecular programs in which several proteins have a pivotal role. Several studies have highlighted a direct link between epigenetic modifications and cell death mechanisms. Different epi-modifications have been described, capable of regulating diverse key players implicated in cell death, leading to uncontrolled proliferation of cancer cells. Scientific efforts are focused on the understanding the epigenetic regulation of cell death mechanisms by developing tools and/or new epi-molecules able to overcome cell death resistance. The development of new epi-molecular tools can overcome cell death deregulation thus potentially improving the sensitivity to the anti-tumor therapies. This chapter focuses on the main epigenetic deregulations in cell death mechanisms in cancer

Benzodeazaoxaflavins as Sirtuin Inhibitors with Antiproliferative Properties in Cancer Stem Cells

n/

CBX2 shapes chromatin accessibility promoting AML via p38 MAPK signaling pathway

Abstract Background: The dynamic epigenome and proteins specialized in the interpretation of epigenetic marks critically contribute to leukemic pathogenesis but also offer alternative therapeutic avenues. Targeting newly discovered chromatin readers involved in leukemogenesis may thus provide new anticancer strategies. Accumulating evidence suggests that the PRC1 complex member CBX2 is overexpressed in solid tumors and promotes cancer cell survival. However, its role in leukemia is still unclear. Methods: We exploited reverse genetic approaches to investigate the role of CBX2 in human leukemic cell lines and ex vivo samples. We also analyzed phenotypic effects following CBX2 silencing using cellular and molecular assays and related functional mechanisms by ATAC-seq and RNA-seq. We then performed bioinformatic analysis of ChIPseq data to explore the influence of histone modifications in CBX2-mediated open chromatin sites. Lastly, we used molecular assays to determine the contribution of CBX2-regulated pathways to leukemic phenotype. Results: We found CBX2 overexpressed in leukemia both in vitro and ex vivo samples compared to CD34+ cells. Decreased CBX2 RNA levels prompted a robust reduction in cell proliferation and induction of apoptosis. Similarly, sensitivity to CBX2 silencing was observed in primary acute myeloid leukemia samples. CBX2 suppression increased genome-wide chromatin accessibility followed by alteration of leukemic cell transcriptional programs, resulting in enrichment of cell death pathways and downregulation of survival genes. Intriguingly, CBX2 silencing induced epigenetic reprogramming at p38 MAPK-associated regulatory sites with consequent deregulation of gene expression. Conclusions: Our results identify CBX2 as a crucial player in leukemia progression and highlight a potential druggable CBX2-p38 MAPK network in AML

Epigenetic Silencing of Peroxisome Proliferator-Activated Receptor γ Is a Biomarker for Colorectal Cancer Progression and Adverse Patients' Outcome

The relationship between peroxisome proliferator-activated receptor γ (PPARG) expression and epigenetic changes occurring in colorectal-cancer pathogenesis is largely unknown. We investigated whether PPARG is epigenetically regulated in colorectal cancer (CRC) progression. PPARG expression was assessed in CRC tissues and paired normal mucosa by western blot and immunohistochemistry and related to patients' clinicopathological parameters and survival. PPARG promoter methylation was analyzed by methylation-specific-PCR and bisulphite sequencing. PPARG expression and promoter methylation were similarly examined also in CRC derived cell lines. Chromatin immunoprecipitation in basal conditions and after epigenetic treatment was performed along with knocking-down experiments of putative regulatory factors. Gene expression was monitored by immunoblotting and functional assays of cell proliferation and invasiveness. Methylation on a specific region of the promoter is strongly correlated with PPARG lack of expression in 30% of primary CRCs and with patients' poor prognosis. Remarkably, the same methylation pattern is found in PPARG-negative CRC cell lines. Epigenetic treatment with 5′-aza-2′-deoxycytidine can revert this condition and, in combination with trichostatin A, dramatically re-activates gene transcription and receptor activity. Transcriptional silencing is due to the recruitment of MeCP2, HDAC1 and EZH2 that impart repressive chromatin signatures determining an increased cell proliferative and invasive potential, features that can experimentally be reverted. Our findings provide a novel mechanistic insight into epigenetic silencing of PPARG in CRC that may be relevant as a prognostic marker of tumor progression

SIRT1 pharmacological activation rescues vascular dysfunction and prevents thrombosis in MTHFR deficiency

Beyond well-assessed risk factors, cardiovascular events could be also associated with the presence of epigenetic and genetic alterations, such as the methylenetetrahydrofolate-reductase (MTHFR) C677T polymorphism. This gene variant is related to increased circulating levels of homocysteine (Hcy) and cardiovascular risk. However, heterozygous carriers have an augmented risk of cardiovascular accidents independently from normal Hcy levels, suggesting the presence of additional deregulated processes in MTHFR C677T carriers. Here, we hypothesize that targeting Sirtuin 1 (SIRT1) could be an alternative mechanism to control the cardiovascular risk associated to MTHFR deficiency condition. Flow Mediated Dilatation (FMD) and light transmission aggregometry assay were performed in subjects carrying MTHFR C677T allele after administration of resveratrol, the most powerful natural clinical usable compound that owns SIRT1 activating properties. MTHFR C677T carriers with normal Hcy levels revealed endothelial dysfunction and enhanced platelet aggregation associated with SIRT1 downregulation. SIRT1 activity stimulation by resveratrol intake was able to override these abnormalities without affecting Hcy levels. Impaired endothelial function, bleeding time, and wire-induced thrombus formation were rescued in a heterozygous Mthfr-deficient (Mthfr+/-) mouse model after resveratrol treatment. Using a cell-based high-throughput multiplexed screening (HTS) assay, a novel selective synthetic SIRT1 activator, namely ISIDE11, was identified. Ex vivo and in vivo treatment of Mthfr+/- mice with ISIDE11 rescues endothelial vasorelaxation and reduces wire-induced thrombus formation, effects that were abolished by SIRT1 inhibitor. Moreover, platelets from MTHFR C677T allele carriers treated with ISIDE11 showed normalization of their typical hyper-reactivity. These results candidate SIRT1 activation as a new therapeutic strategy to contain cardio and cerebrovascular events in MTHFR carriers

Dual Tumor Suppressor and Tumor Promoter Action of Sirtuins in Determining Malignant Phenotype

Sirtuins (SIRTs), class III histone deacetylases, are differentially expressed in several human cancers, where they display both oncogenic and tumor-suppressive properties depending on cellular context and experimental conditions. SIRTs are involved in many important biological processes and play a critical role in cancer initiation, promotion, and progression. A growing body of evidence indicates the involvement of SIRTs in regulating three important tumor processes: epithelial-to-mesenchymal transition (EMT), invasion, and metastasis. Many SIRTs are responsible for cellular metabolic reprogramming and drug resistance by inactivating cell death pathways and promoting uncontrolled proliferation. In this review, we summarize current knowledge on the role of SIRTs in cancer and discuss their puzzling dual function as tumor suppressors and tumor promoters, important for the future development of novel tailored SIRT-based cancer therapies

CHARACTERIZATION OF NEW RIPKs FAMILY MEMBERS: HIGHLIGHTS ON THE BIOLOGICAL ROLE OF RIPK2 AND RIPK4 IN CANCER

Receptor-Interacting Protein Kinases (RIPKs) are a seven-member family of Ser/Thr protein kinases involved in host defense, inflammatory phenomena as well as in cell death. To date their role in tumorigenesis is still unclear. Little scientific evidence explains the direct involvement of RIPK2 and RIPK4 in pathogenesis, especially in cancer. Several cell death regulation strategies have been studied in order to improve anticancer therapies and, among these, the study of the RIP kinase family is particularly interesting, which offers a therapeutic alternative in cases of resistance to apoptotic processes. It is known that RIPK2, through its CARD functional domain, is able to trigger the activation of NF-kB or the MAP kinase pathway, playing a fundamental role in the immune response and inflammation while RIPK4 appears to participate both physically and functionally in several TRAF-dependent pathways leading to the activation of NF-kB pathway. Methods: Western blot, RT-qPCR, Immunoprecipitation, Transfection. Results. Since both kinases are differentially expressed in different cancer types and the molecular mechanisms involved are poorly characterized, one of the main objectives of the project is to characterize the oncogenic role of these proteins by studying key molecular interactors and activated protein complexes. For this purpose, different cancer cell models exhibiting variable expression levels were chosen. Jurkat and Raji tumoral cell lines, in which RIPK2 is respectively poor and high expressed, as well as HL-60 and PANC-1 tumoral cell lines for RIPK4 investigation will be considered. To identify molecular players, MS/MS analysis coupled immunoprecipitation experiments were performed. Waiting for these results, molecular screening experiments have been started in order to identify molecules that modulate the expression of these kinases, that are structural scaffolds useful for the synthesis of new molecules. Conclusions. Further studies will be needed to better understand the biological activity of the investigated proteins and to understand the possible cross talk between different family members