Synthesis of trehalose-based chemical tools for the study of the mycobacterial membrane

Corynebacteriales including the causative agent of many diseases such as tuberculosis are known to be extremely resistant against external stress as well as to antibiotic treatments which is believed to be related to the singular architecture of their mycomembrane. Over the last decades, both bioorthogonal chemical reporters and fluorescent probes for the metabolic labeling of bacterial cell glycans were developed including several trehalose-based probes to study the dynamics of mycomembrane components. This review presents an exhaustive view on the reported syntheses of trehalose-based probes enabling the study of the mycomembrane biogenesis

2′-Modified thymidines with bioorthogonal cyclopropene or sydnone as building blocks for copper-free postsynthetic functionalization of chemically synthesized oligonucleotides

International audienceThe development of facile methods for conjugating relevant probes, ligands, or delivery agents onto oligonucleotides (ONs) is highly desirable both for fundamental studies in chemical biology and for improving the pharmacology of ONs in medicinal chemistry. Numerous efforts have been focused on the introduction of bioorthogonal groups onto phosphoramidite building blocks, allowing the controlled chemical synthesis of reactive ONs for postsynthetic modifications. Among these building blocks, alkyne, cyclooctynes, trans-cyclooctene, and norbornene have been proved to be compatible with automated solid-phase chemistry. Herein, we present the development of novel 2′-functionalized nucleoside phosphoramidite monomers comprising bioorthogonal methylcyclopropene or sydnone moieties and their introduction for the first time to ON solid-phase synthesis. Traceless ON postsynthetic modifications with reactive complementary probes were successfully achieved through either inverse electron-demand Diels–Alder (iEDDA) reactions or strain-promoted sydnone–alkyne cycloaddition (SPSAC). These results expand the set of bioorthogonal phosphoramidite building blocks to generate ONs for postsynthetic labeling

Sequence-Specific Base Pair Mimics Are Efficient Topoisomerase IB Inhibitors

Topoisomerase IB controls DNA topology by cleaving DNA transiently. This property is used by inhibitors, such as camptothecin, that stabilize, by inhibiting the religation step, the cleavage complex, in which the enzyme is covalently attached to the 3′-phosphate of the cleaved DNA strand. These drugs are used in clinics as antitumor agents. Because three-dimensional structural studies have shown that camptothecin derivatives act as base pair mimics and intercalate between two base pairs in the ternary DNA–topoisomerase–inhibitor complex, we hypothesized that base pairs mimics could act like campthotecin and inhibit the religation reaction after the formation of the topoisomerase I–DNA cleavage complex. We show here that three base pair mimics, nucleobases analogues of the aminophenyl-thiazole family, once targeted specifically to a DNA sequence were potent topoisomerase IB inhibitors. The targeting was achieved through covalent linkage to a sequence-specific DNA ligand, a triplex-forming oligonucleotide, and was necessary to position and keep the nucleobase analogue in the cleavage complex. In the absence of triplex formation, only a weak binding to the DNA and topoisomerase I-mediated DNA cleavage was observed. The three compounds were equally active once conjugated, implying that the intercalation of the nucleobase upon triplex formation is the essential feature for the inhibition activity

Borinic Acids as New Fast-Responsive Triggers for Hydrogen Peroxide Detection.

Detection of hydrogen peroxide (H2O2), which is responsible for numerous damages when overproduced, is crucial for a better understanding of H2O2-mediated signalling in physiological and pathological processes. For this purpose, various“off-on” small fluorescent probes relying on a boronate trigger have been developed. However, they suffer from low kinetics and do not allow forH2O2-detectionwith a short response time. Therefore, more reactive sensors are still awaited. To address this issue, we have successfully developed the first generation of borinic-based fluorescent probes containing a coumarin-scaffold. We determined the in vitrokinetic constants of the probe toward H2O2-promotedoxidation. We measured 1.9x104m-1.s-1as a second order rate constant, which is 10 000 faster than its boronic counterpart (1.8 m-1.s-1). This remarkable reactivity was also effective in a cellular context, rendering the borinic trigger an advantageous new tool for H2O2detection

Identification of Novel Inhibitors of DNA Methylation by Screening of a Chemical Library

In order to discover new inhibitors of the DNA methyltransferase 3A/3L complex, we used a medium-throughput nonradioactive screen on a random collection of 1120 small organic compounds. After a primary hit detection against DNA methylation activity of the murine Dnmt3A/3L catalytic complex, we further evaluated the EC₅₀ of the 12 most potent hits as well as their cytotoxicity on DU145 prostate cancer cultured cells. Interestingly, most of the inhibitors showed low micromolar activities and little cytotoxicity. Dichlone, a small halogenated naphthoquinone, classically used as pesticide and fungicide, showed the lowest EC₅₀ at 460 nM. We briefly assessed the selectivity of a subset of our new inhibitors against hDNMT1 and bacterial Dnmts, including M. SssI and EcoDam, and the protein lysine methyltransferase PKMT G9a and the mode of inhibition. Globally, the tested molecules showed a clear preference for the DNA methyltransferases, but poor selectivity among them. Two molecules including Dichlone efficiently reactivated YFP gene expression in a stable HEK293 cell line by promoter demethylation. Their efficacy was comparable to the DNMT inhibitor of reference 5-azacytidine

Synthesis and Evaluation of Analogues of <i>N</i>‑Phthaloyl‑l‑tryptophan (RG108) as Inhibitors of DNA Methyltransferase 1

DNA methyltransferases (DNMT) are promising drug targets in cancer provided that new, more specific, and chemically stable inhibitors are discovered. Among the non-nucleoside DNMT inhibitors, <i>N</i>-phthaloyl-l-tryptophan <b>1</b> (RG108) was first identified as inhibitor of DNMT1. Here, <b>1</b> analogues were synthesized to understand its interaction with DNMT. The indole, carboxylate, and phthalimide moieties were modified. Homologated and conformationally constrained analogues were prepared. The latter were synthesized from prolino­homotryptophan derivatives through a methodology based amino–zinc–ene–enolate cyclization. All compounds were tested for their ability to inhibit DNMT1 in vitro. Among them, constrained compounds <b>16</b>–<b>18</b> and NPys derivatives <b>10</b>–<b>11</b> were found to be at least 10-fold more potent than the reference compound. The cytotoxicity on the tumor DU145 cell line of the most potent inhibitors was correlated to their inhibitory potency. Finally, docking studies were conducted in order to understand their binding mode. This study provides insights for the design of the next-generation of DNMT inhibitors

Rational Design of Bisubstrate-Type Analogues as Inhibitors of DNA Methyltransferases in Cancer Cells

Aberrant DNA hypermethylation of promoter of tumor suppressor genes is commonly observed in cancer, and its inhibition by small molecules is promising for their reactivation. Here we designed bisubstrate analogues-based inhibitors, by mimicking each substrate, the <i>S</i>-adenosyl-l-methionine and the deoxycytidine, and linking them together. This approach resulted in quinazoline–quinoline derivatives as potent inhibitors of DNMT3A and DNMT1, some showing certain isoform selectivity. We highlighted the importance of (i) the nature and rigidity of the linker between the two moieties for inhibition, as (ii) the presence of the nitrogen on the quinoline group, and (iii) of a hydrophobic group on the quinazoline. The most potent inhibitors induced demethylation of <i>CDKN2A</i> promoter in colon carcinoma HCT116 cells and its reactivation after 7 days of treatment. Furthermore, in a leukemia cell model system, we found a correlation between demethylation of the promoter induced by the treatment, chromatin opening at the promoter, and the reactivation of a reporter gene