Synthesis, Biological Evaluation, and Structure–Activity Relationships of a Novel Class of Apurinic/Apyrimidinic Endonuclease 1 Inhibitors

APE1 is an essential protein that operates in the base excision repair (BER) pathway and is responsible for ≥95% of the total apurinic/apyrimidinic (AP) endonuclease activity in human cells. BER is a major pathway that copes with DNA damage induced by several anticancer agents, including ionizing radiation and temozolomide. Overexpression of APE1 and enhanced AP endonuclease activity have been linked to increased resistance of tumor cells to treatment with monofunctional alkylators, implicating inhibition of APE1 as a valid strategy for cancer therapy. We report herein the results of a focused medicinal chemistry effort around a novel APE1 inhibitor, <i>N</i>-(3-(benzo­[<i>d</i>]­thiazol-2-yl)-6-isopropyl-4,5,6,7-tetrahydrothieno­[2,3-<i>c</i>]­pyridin-2-yl)­acetamide (<b>3</b>). Compound <b>3</b> and related analogues exhibit single-digit micromolar activity against the purified APE1 enzyme and comparable activity in HeLa whole cell extract assays and potentiate the cytotoxicity of the alkylating agents methylmethane sulfonate and temozolomide. Moreover, this class of compounds possesses a generally favorable in vitro ADME profile, along with good exposure levels in plasma and brain following intraperitoneal dosing (30 mg/kg body weight) in mice

MOESM1 of Rilpivirine analogs potently inhibit drug-resistant HIV-1 mutants

Additional file 1: Table S1–S4. The IC50 values (nM) of RPV and the RPV analogs were determined against the NNRTI resistant mutants were measured by using a single round infection assay, n = 4. The concentrations (nM) are measured by reductions in luciferase reporter activity followed by the standard deviations. In parenthesis is the fold-change that describes the difference between the IC50 value of the NNRTI resistant mutant relative to WT

Venglustat Inhibits Protein N‑Terminal Methyltransferase 1 in a Substrate-Competitive Manner

Venglustat is a known allosteric inhibitor for ceramide glycosyltransferase, investigated in diseases caused by lysosomal dysfunction. Here, we identified venglustat as a potent inhibitor (IC50 = 0.42 μM) of protein N-terminal methyltransferase 1 (NTMT1) by screening 58,130 compounds. Furthermore, venglustat exhibited selectivity for NTMT1 over 36 other methyltransferases. The crystal structure of NTMT1-venglustat and inhibition mechanism revealed that venglustat competitively binds at the peptide substrate site. Meanwhile, venglustat potently inhibited protein N-terminal methylation levels in cells (IC50 = 0.5 μM). Preliminary structure–activity relationships indicated that the quinuclidine and fluorophenyl parts of venglustat are important for NTMT1 inhibition. In summary, we confirmed that venglustat is a bona fide NTMT1 inhibitor, which would advance the study on the biological roles of NTMT1. Additionally, this is the first disclosure of NTMT1 as a new molecular target of venglustat, which would cast light on its mechanism of action to guide the clinical investigations

Venglustat Inhibits Protein N‑Terminal Methyltransferase 1 in a Substrate-Competitive Manner

Venglustat is a known allosteric inhibitor for ceramide glycosyltransferase, investigated in diseases caused by lysosomal dysfunction. Here, we identified venglustat as a potent inhibitor (IC50 = 0.42 μM) of protein N-terminal methyltransferase 1 (NTMT1) by screening 58,130 compounds. Furthermore, venglustat exhibited selectivity for NTMT1 over 36 other methyltransferases. The crystal structure of NTMT1-venglustat and inhibition mechanism revealed that venglustat competitively binds at the peptide substrate site. Meanwhile, venglustat potently inhibited protein N-terminal methylation levels in cells (IC50 = 0.5 μM). Preliminary structure–activity relationships indicated that the quinuclidine and fluorophenyl parts of venglustat are important for NTMT1 inhibition. In summary, we confirmed that venglustat is a bona fide NTMT1 inhibitor, which would advance the study on the biological roles of NTMT1. Additionally, this is the first disclosure of NTMT1 as a new molecular target of venglustat, which would cast light on its mechanism of action to guide the clinical investigations

Venglustat Inhibits Protein N‑Terminal Methyltransferase 1 in a Substrate-Competitive Manner

Venglustat is a known allosteric inhibitor for ceramide glycosyltransferase, investigated in diseases caused by lysosomal dysfunction. Here, we identified venglustat as a potent inhibitor (IC50 = 0.42 μM) of protein N-terminal methyltransferase 1 (NTMT1) by screening 58,130 compounds. Furthermore, venglustat exhibited selectivity for NTMT1 over 36 other methyltransferases. The crystal structure of NTMT1-venglustat and inhibition mechanism revealed that venglustat competitively binds at the peptide substrate site. Meanwhile, venglustat potently inhibited protein N-terminal methylation levels in cells (IC50 = 0.5 μM). Preliminary structure–activity relationships indicated that the quinuclidine and fluorophenyl parts of venglustat are important for NTMT1 inhibition. In summary, we confirmed that venglustat is a bona fide NTMT1 inhibitor, which would advance the study on the biological roles of NTMT1. Additionally, this is the first disclosure of NTMT1 as a new molecular target of venglustat, which would cast light on its mechanism of action to guide the clinical investigations

Parallel Chemistry Approach to Identify Novel Nuclear Receptor Ligands Based on the GW0742 Scaffold

We describe the parallel synthesis of novel analogs of GW0742, a peroxisome proliferator-activated receptor δ (PPARδ) agonist. For that purpose, modified reaction conditions were applied, such as a solid-phase palladium-catalyzed Suzuki coupling. In addition, tetrazole-based compounds were generated as a bioisostere for carboxylic acid-containing ligand GW0742. The new compounds were investigated for their ability to activate PPARδ mediated transcription and their cross-reactivity with the vitamin D receptor (VDR), another member of the nuclear receptor superfamily. We identified many potent PPARδ agonists that were less toxic than GW0742, where ∼65 of the compounds synthesized exhibited partial PPARδ activity (23–98%) with EC₅₀ values ranging from 0.007–18.2 μM. Some ligands, such as compound <b>32</b>, were more potent inhibitors of VDR-mediated transcription with significantly reduced PPARδ activity than GW0742, however, none of the ligands were completely selective for VDR inhibition over PPARδ activation of transcription

4‑(3-Chloro-5-(trifluoromethyl)pyridin-2-yl)‑<i>N</i>‑(4-methoxypyridin-2-yl)piperazine-1-carbothioamide (ML267), a Potent Inhibitor of Bacterial Phosphopantetheinyl Transferase That Attenuates Secondary Metabolism and Thwarts Bacterial Growth

4′-Phosphopantetheinyl transferases (PPTases) catalyze a post-translational modification essential to bacterial cell viability and virulence. We present the discovery and medicinal chemistry optimization of 2-pyridinyl-<i>N</i>-(4-aryl)­piperazine-1-carbothioamides, which exhibit submicromolar inhibition of bacterial Sfp-PPTase with no activity toward the human orthologue. Moreover, compounds within this class possess antibacterial activity in the absence of a rapid cytotoxic response in human cells. An advanced analogue of this series, ML267 (<b>55</b>), was found to attenuate production of an Sfp-PPTase-dependent metabolite when applied to Bacillus subtilis at sublethal doses. Additional testing revealed antibacterial activity against methicillin-resistant Staphylococcus aureus, and chemical genetic studies implicated efflux as a mechanism for resistance in Escherichia coli. Additionally, we highlight the in vitro absorption, distribution, metabolism, and excretion and in vivo pharmacokinetic profiles of compound <b>55</b> to further demonstrate the potential utility of this small-molecule inhibitor

Determination of IC₅₀ values for ketoconazole and ketaminazole with CaCYP51 and HsCYP51.

<p>CYP51 reconstitution assays (0.5-ml total volume) containing 1 µM CaCYP51 (A) or 0.3 µM HsCYP51 (B) were performed as detailed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065928#s2" target="_blank">Materials and Methods</a>. Ketoconazole (solid circles) and ketaminazole (hollow circles) concentrations were varied from 0 to 4 µM for CaCYP51 and up to 190 µM for HsCYP51 with the DMSO concentration kept constant at 1% (vol/vol). Mean values from two replicates are shown along with associated standard deviation bars. Relative velocities of 1.0 were equivalent to 1.04 and 2.69 nmoles 14α-demetylated lanosterol produced per minute per nmole CYP51 (min⁻¹) for CaCYP51 and HsCYP51, respectively.</p

Buffer conditions for IC₅₀ assays, with constant substrate concentration and varying inhibitor concentration^a.

a<p>The UV-based manual inhibition data (3 replicates) were fit as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065928#s2" target="_blank">Materials and Methods</a> section.</p

Representative analogues evaluated for pseudoperoxidase activity and IC₅₀ potency (µM), with errors in brackets.

<p>The UV-based manual inhibition data (3 replicates) were fit as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065928#s2" target="_blank">Materials and Methods</a> section.</p