Vincenzo Carafa

Trials with ‘epigenetic’ drugs: An update

Epigenetic inactivation of pivotal genes involved in correct cell growth is a hallmark of human pathologies, in particular cancer. These epigenetic mechanisms, including crosstalk between DNA methylation, histone modifications and non‐coding RNAs, affect gene expression and are associated with disease progression. In contrast to genetic mutations, epigenetic changes are potentially reversible. Re‐expression of genes epigenetically inactivated can result in the suppression of disease state or sensitization to specific therapies. Small molecules that reverse epigenetic inactivation, so‐called epi‐drugs, are now undergoing clinical trials. Accordingly, the Food and Drug Administration (FDA) and the European Medicines Agency (EMA) for cancer treatment have approved some of these drugs. Here, we focus on the biological features of epigenetic molecules, analyzing the mechanism(s) of action and their current use in clinical practice.

Sirtuins and disease: the road ahead.

Sirtuins represent a promising new class of conserved histone deacetylases, originally identified in yeast. The activity of the sirtuin (SirT) family – made up of seven members (SirT1-7) – is NAD+ dependent. Sirtuins target a wide range of cellular proteins in nucleus, cytoplasm, and mitochondria for post-translational modification by acetylation (SirT1, 2, 3, and 5) or ADP-ribosylation (SirT4 and 6). Sirtuins regulate responses to stress and ensure that damaged DNA is not propagated, thus contrasting the accumulation of mutations. To date, sirtuins have emerged as potential therapeutic targets for treatment of human pathologies such as metabolic, cardiovascular and neurodegenerative diseases, and cancer. SirT1 is the founding member of this class of enzymes and is currently the best known of the group. SirT1 acts in various cellular processes, deacetylating both chromatin and non-histone proteins, and its role in cancer and aging has been extensively studied. SirT1 may play a critical role in tumor initiation and progression as well as drug resistance by blocking senescence and apoptosis, and by promoting cell growth and angiogenesis. Recently, growing interest in sirtuin modulation has led to the discovery and characterization of small molecules able to modify sirtuin activity. The present review highlights SirT mechanism(s) of action and deregulation in cancer, focusing on the therapeutic potential of SirT modulators both in cancer prevention and treatment.

Synthesis of Benzamides Related to Anacardic Acid and Their Histone Acetyltransferase (HAT) Inhibitory Activities

A group of benzamides related to anacardic acid amide CTPB with alkyl chains of defined length were prepared by a five‐step sequence starting from 2,6‐dihydroxybenzoic acid, and their activities were compared with those reported for the HAT inhibitor anacardic acid (AA). The subset of 4‐cyano‐3‐trifluoromethylphenylbenzamides with shorter chains exhibited activities similar to that of AA, as they behaved as human p300 inhibitors, induced a decrease in histone acetylation levels in immortalized HEK cells, and counteracted the action of the HDAC inhibitor SAHA in MCF7 breast cancer cells. Moreover, an analogue with the shortest alkyl chain induced significant apoptosis at 50 μM in U937 leukemia cells.

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.

Antioxidant, antimicrobial and anti-proliferative activities of Solanum tuberosum L. var. Vitelotte

Solanum tuberosum L. var. Vitelotte is a potato variety widely used for human consumption. The pigments responsible for its attractive color belong to the class of anthocyanins. The objectives of this study were to characterize and measure the concentration of anthocyanins in pigmented potatoes and to evaluate their antioxidant and antimicrobial activities and their anti-proliferative effects in solid and hematological cancer cell lines. Anthocyanins exert anti-bacterial activity against different bacterial strains and a slight activity against three fungal strains. The Gram-positive bacterium Staphylococcus aureus and the fungus Rhyzoctonia solani were the most affected microorganisms. Antioxidant activities were evaluated by DPPH and FRAP methods; the extract showed a higher reducing capability than anti-radical activity. Moreover, we found that in different cancer cell models the anthocyanins cause inhibition of proliferation and apoptosis in a dose dependent manner. These biological activities are likely due to the high content of malvidin 3-O-p-coumaroyl-rutinoside-5-O-glucoside and petunidin 3-O-p-coumaroyl-rutinoside-5-O-glucoside.

Structural characterization and comparative modeling of PD-Ls 1–3, type 1 ribosome-inactivating proteins from summer leaves of Phytolacca dioica L.

The amino acid sequence and glycan structure of PD-L1, PD-L2 and PD-L3, type 1 ribosome-inactivating proteins isolated from Phytolacca dioica L. leaves, were determined using a combined approach based on peptide mapping, Edman degradation and ESI-Q-TOF MS in precursor ion discovery mode. The comparative analysis of the 261 amino acid residue sequences showed that PD-L1 and PD-L2 have identical primary structure, as it is the case of PD-L3 and PD-L4. Furthermore, the primary structure of PD-Ls 1-2 and PD-Ls 3-4 have 81.6% identity (85.1% similarity). The ESI-Q-TOF MS analysis confirmed that PD-Ls 1-3 were glycosylated at different sites. In particular, PD-L1 contained three glycidic chains with the well known paucidomannosidic structure (Man)(3) (GlcNAc)(2) (Fuc)(1) (Xyl)(1) linked to Asn10, Asn43 and Asn255. PD-L2 was glycosylated at Asn10 and Asn43, and PD-L3 was glycosylated only at Asn10. PD-L4 was confirmed to be not glycosylated. Despite an overall high structural similarity, the comparative modeling of PD-L1, PD-L2, PD-L3 and PD-L4 has shown potential influences of the glycidic chains on their adenine polynucleotide glycosylase activity on different substrates.

Histone Deacetylase Inhibitors: Recent Insights from Basic to Clinical Knowledge & Patenting of Anti-Cancer Actions

Epigenetic modifications have been causally linked to cancer development and progression, and are potentially reversible by drug treatments. The N-terminal tails of histones contain amino acid residues modifiable by post-translational modifications such as acetylation. Given that HDAC inhibitors induce cancer cell differentiation and death, an increasing number of these compounds has been synthesized in the last ten years. Many HDAC inhibitors are in clinical trials for the treatment of cancer. Two of them, the hydroxamic acid (SAHA) and Romidepsin (FK 228), are approved in the second line treatment of refractory, persistent or relapsed Cutaneous T Cell Lymphoma (CTCL). The growing evidence of the potential benefits of an anti-cancer treatment based on the use of HDAC inhibitors have led to a large number of patent applications all over the world. The aim of this review is to give an overview of the basic current knowledge and molecular mechanisms of HDAC inhibitors and their clinical trials as well as to focus on the recent patent applications existing in the field of HDAC inhibitors and cancer treatment between 2008 and 2010 in USA.

Indole-Derived Psammaplin A Analogues as Epigenetic Modulators with Multiple Inhibitory Activities

A SAR study has been carried out around a modified scaffold of the natural product psammaplin A obtained by replacing the o-bromophenol unit by an indole ring. A series of indole psammaplin A constructs were generated in a short synthetic sequence that starts with the functionalization of the C3 indole position with in situ generated nitrosoacrylate, and this is followed by protection of the β-indole-α-oximinoesters, saponification, condensation with symmetrical diamines, and deprotection. Biochemical and cellular characterization using U937 and MCF-7 cells confirmed that many of these analogues displayed more potent actitivies than the parent natural product. Moreover, in addition to the reported HDAC and DNMT dual epigenetic inhibitory profile of the parent compound, some analogues, notably 4a (UVI5008), also inhibited the NAD(+)-dependent SIRT deacetylase enzymes. The SAR study provides structural insights into the mechanism of action of these multiple epigenetic ligands and paves the way for additional structural exploration to optimize their pharmacological profiles. Because of their multi(epi)target features and their action in ex vivo samples, the indole-based psammaplin A derivatives are attractive molecules for the modulation of epigenetic disorders.

TNF-related apoptosis-inducing ligand: Signalling of a ‘smart’ molecule

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of the tumor necrosis factor super-family and signals via two death receptors, TRAIL-R1 and TRAIL-R2, and two decoy receptors, TRAIL-R3 and TRAIL-R4, differently expressed in normal and cancer cells. TRAIL is mainly studied for its capacity to induce apoptosis preferentially in cancer cells. TRAIL is expressed in a variety of human tissues, in particular in the lymphoid system, suggesting a strong physiological role in the innate immunity. This review will focus on TRAIL gene structure and regulation, protein folding, tissue expression and molecular signalling. Finally, the potential use of TRAIL as anticancer treatment alone or in combination therapy as well as the use of drugs which signal via TRAIL and its receptors will be analyzed.

Identification of Tri‐ and Tetracyclic Pyrimidinediones as Sirtuin Inhibitors

Sirtuins (SIRT1–7) are a family of NAD-dependent deacetylases and/or ADP-ribosyltransferases that modify a broad range of distinct protein substrates with various subcellular localizations. SIRT1 and SIRT2 are mainly localized in the nucleus (SIRT1) or the cytoplasm (SIRT2), SIRT3–5 are typically mitochondrial enzymes, and SIRT6 and SIRT7 are localized in the cell nucleus and nucleolus, respectively. The requirement of NAD as a cofactor for enzymatic activity, along with the ability to deacetylate both histones and a variety of non-histone proteins, suggests that the sirtuins play a crucial role in metabolism and energy-dependent transcription. SIRT1 has been shown to deacetylate more than 30 different acetylated substrates such as H4K16, H3K9, p53, forkhead box class O (FoxO), p300, the DNA repair factor Ku70, peroxisome proliferator-activated receptor g co-activator 1a (PGC-1a), nuclear factor-kB (NF-kB), and many others. Therefore, not surprisingly, SIRT1 is implicated in carcinogenesis, in metabolic and cardiovascular diseases, and in neurodegeneration. In addition to a protective role described for SIRT1 in DNA damage, accumulation of mutations, and genomic instability, SIRT1 has been found to be up-regulated in human lung cancer, prostate cancer, colon carcinoma, and leukemia. Moreover, a deleted-in-breast-cancer (DBC) protein has recently been described as an endogenous inhibitor of SIRT1 to promote p53 acetylation and activation. Cambinol (1; Figure 1), a well-known SIRT inhibitor, is able to inactivate the transcriptional repressor BCL6 in Burkitt’s lymphoma cells through ace-