Sirtuin modulators: past, present, and future perspectives.

Sirtuins are NAD+-dependent protein lysine deacylase and mono-ADP ribosylases present in both prokaryotes and eukaryotes. The sirtuin family comprises seven isoforms in mammals, each possessing different subcellular localization and biological functions. Sirtuins have received increasing attention in the past two decades given their pivotal functions in a variety of biological contexts, including cytodifferentiation, transcriptional regulation, cell cycle progression, apoptosis, inflammation, metabolism, neurological and cardiovascular physiology and cancer. Consequently, modulation of sirtuin activity has been regarded as a promising therapeutic option for many pathologies. In this review, we provide an up-to-date overview of sirtuin biology and pharmacology. We examine the main features of the most relevant inhibitors and activators, analyzing their structure-activity relationships, applications in biology, and therapeutic potential.

First-in-Class Inhibitors of the Ribosomal Oxygenase MINA53.

MINA53 is a JmjC domain 2-oxoglutarate-dependent oxygenase that catalyzes ribosomal hydroxylation and is a target of the oncogenic transcription factor c-MYC. Despite its anticancer target potential, no small-molecule MINA53 inhibitors are reported. Using ribosomal substrate fragments, we developed mass spectrometry assays for MINA53 and the related oxygenase NO66. These assays enabled the identification of 2-(aryl)alkylthio-3,4-dihydro-4-oxoypyrimidine-5-carboxylic acids as potent MINA53 inhibitors, with selectivity over NO66 and other JmjC oxygenases. Crystallographic studies with the JmjC demethylase KDM5B revealed active site binding but without direct metal chelation; however, molecular modeling investigations indicated that the inhibitors bind to MINA53 by directly interacting with the iron cofactor. The MINA53 inhibitors manifest evidence for target engagement and selectivity for MINA53 over KDM4-6. The MINA53 inhibitors show antiproliferative activity with solid cancer lines and sensitize cancer cells to conventional chemotherapy, suggesting that further work investigating their potential in combination therapies is warranted.

Recent advances in epigenetic proteolysis targeting chimeras (Epi-PROTACs).

PROteolysis TArgeting Chimeras (PROTACs) are heterobifunctional molecules that trigger the poly-ubiquitination of the protein of interest (POI) inducing its degradation via the recruitment of the ubiquitin-proteasome system, thus suppressing the POI's intracellular levels and indirectly all its functions. Recently, one of the fields where the protein knockdown induced by PROTACs has demonstrated to serve as a promising biochemical tool and to provide new opportunities for drug discovery is the epigenetics (epi-PROTACs). A full inhibition of the functions of all domains of a specific epigenetic POI (e-POI), rather than just the block of its catalytic/single domain activity, is in fact a new more effective modality to hit an e-POI and, in principle, the complex it belongs to, and potentially to treat the related diseases, first cancer. In this review, we will present the most relevant progresses made, especially in the last two years, in the application of PROTACs technology to the three main classes of e-POIs: "writers", "erasers" and "readers". Emphasis will be devoted to the medicinal chemistry aspects of the epi-PROTACs design, preparation, and optimization and to the comparison with small molecule epi-drugs for both epi-targets functional annotation and potential anticancer therapy purposes.

Targeting histone acetylation/deacetylation in parasites: an update (2017-2020).

Histone modifying enzymes have vital roles in the growth and survival of both parasites and humans. Targeting the epigenome can be a new strategy for the treatment of parasitic diseases. Compounds modulating histone acetylation/deacetylation have recently been reported hampering Plasmodium, Schistosoma, Leishmania, and Trypanosoma infections. Beside new histone deacetylase inhibitors, PfGCN5 and bromodomain inhibitors have been recently described to inhibit Plasmodium proliferation. Sm histone deacetylase 8 and SmSIRT2, as well as Leishmania and Trypanosoma sirtuins (SIR2rps), seem to be the most reliable targets to effectively fight the related protozoan infections. The selectivity toward parasite over mammalian cells is still an open question, and significant optimization efforts of epidrugs are still required to improve potency/selectivity and decrease toxicity. Recent reports on the alteration of cellular signaling pathways provoked by parasite infection through changes in the host acetylation/deacetylation status at gene promoters may suggest novel therapeutic strategies to treat these diseases.

Identification of Inhibitors to Trypanosoma cruzi Sirtuins Based on Compounds Developed to Human Enzymes.

Chagas disease is an illness caused by the protozoan parasite Trypanosoma cruzi, affecting more than 7 million people in the world. Benznidazole and nifurtimox are the only drugs available for treatment and in addition to causing several side effects, are only satisfactory in the acute phase of the disease. Sirtuins are NAD+-dependent deacetylases involved in several biological processes, which have become drug target candidates in various disease settings. T. cruzi presents two sirtuins, one cytosolic (TcSir2rp1) and the latter mitochondrial (TcSir2rp3). Here, we characterized the effects of human sirtuin inhibitors against T. cruzi sirtuins as an initial approach to develop specific parasite inhibitors. We found that, of 33 compounds tested, two inhibited TcSir2rp1 (15 and 17), while other five inhibited TcSir2rp3 (8, 12, 13, 30, and 32), indicating that specific inhibitors can be devised for each one of the enzymes. Furthermore, all inhibiting compounds prevented parasite proliferation in cultured mammalian cells. When combining the most effective inhibitors with benznidazole at least two compounds, 17 and 32, demonstrated synergistic effects. Altogether, these results support the importance of exploring T. cruzi sirtuins as drug targets and provide key elements to develop specific inhibitors for these enzymes as potential targets for Chagas disease treatment.

From PARP1 to TNKS2 Inhibition: A Structure-Based Approach.

Tankyrases (TNKSs) have recently gained great consideration as potential targets in Wnt/ß-catenin pathway-dependent solid tumors. Previously, we reported the 2-mercaptoquinazolin-4-one MC2050 as a micromolar PARP1 inhibitor. Here we show how the resolution of the X-ray structure of PARP1 in complex with MC2050, combined with the computational investigation of the structural differences between TNKSs and PARP1/2 active sites, provided the rationale for a structure-based drug design campaign that with a limited synthetic effort led to the discovery of the bis-quinazolinone 5 as a picomolar and selective TNKS2 inhibitor, endowed with antiproliferative effects in a colorectal cancer cell line (DLD-1) where the Wnt pathway is constitutively activated.

Sirtuin modulators: where are we now? A review of patents from 2015 to 2019.

In recent years, sirtuins (SIRTs) gained an increasing consideration because of their multiple key roles in several biological settings such as the regulation of transcription, energetic metabolism, cell cycle progression, and cytodifferentiation, apoptosis, neuro- and cardio-protection, inflammation, cancer onset and progression. Since there is mounting evidence in favor of potential therapeutic applications of SIRT modulators in various age-related disorders, the search about them is quite active. Areas covered: This review includes the patents regarding SIRT modulators released from 2015 to 2019 and provides an overview of the most relevant SIRT modulators.Expert opinion: Despite the knowledge about this family of broad-spectrum protein lysine deacylases has recently massively increased, there are still open questions, first of all, the exact nature of their involvement in various age-related conditions. The search for isoform-specific SIRT activators and inhibitors is still at its infancy, a limited number of patents describing them has been released, and not many clinical trials are ongoing. However, it is extremely likely that the successes obtained in the structural elucidation and structure-based design approaches that very recently have led to potent and specific SIRT modulators will pave the way for the development of further compounds selective for every single isoform.

Tranylcypromine-Based LSD1 Inhibitors: Structure-Activity Relationships, Antiproliferative Effects in Leukemia, and Gene Target Modulation.

LSD1 is a lysine demethylase highly involved in initiation and development of cancer. To design highly effective covalent inhibitors, a strategy is to fill its large catalytic cleft by designing tranylcypromine (TCP) analogs decorated with long, hindered substituents. We prepared three series of TCP analogs, carrying aroyl- and arylacetylamino (1 a-h), Z-amino acylamino (2 a-o), or double-substituted benzamide (3 a-n) residues at the C4 or C3 position of the phenyl ring. Further fragments obtained by chemical manipulation applied on the TCP scaffold (compounds 4 a-i) were also prepared. When tested against LSD1, most of 1 and 3 exhibited IC50 values in the low nanomolar range, with 1 e and 3 a,d,f,g being also the most selective respect to monoamine oxidases. In MV4-11 AML and NB4 APL cells compounds 3 were the most potent, displaying up to sub-micromolar cell growth inhibition against both cell lines (3 a) or against NB4 cells (3 c). The most potent compounds in cellular assays were also able to induce the expression of LSD1 target genes, such as GFI-1b, ITGAM, and KCTD12, as functional read-out for LSD1 inhibition. Mouse and human intrinsic clearance data highlighted the high metabolic stability of compounds 3 a, 3 d and 3 g. Further studies will be performed on the new compounds 3 a and 3 c to assess their anticancer potential in different cancer contexts.