Synthesis, Structural Investigations, DNA/BSA Interactions, Molecular Docking Studies, and Anticancer Activity of a New 1,4-Disubstituted 1,2,3-Triazole Derivative.

We report herein a new 1,2,3-triazole derivative, namely, 4-((1-(3,4-dichlorophenyl)-1H-1,2,3-triazol-4-yl)methoxy)-2-hydroxybenzaldehyde, which was synthesized by copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC). The structure of the compound was analyzed using Fourier transform infrared spectroscopy (FTIR), 1H NMR, 13C NMR, UV-vis, and elemental analyses. Moreover, X-ray crystallography studies demonstrated that the compound adapted a monoclinic crystal system with the P21/c space group. The dominant interactions formed in the crystal packing were found to be hydrogen bonding and van der Waals interactions according to Hirshfeld surface (HS) analysis. The volume of the crystal voids and the percentage of free spaces in the unit cell were calculated as 152.10 Å3 and 9.80%, respectively. The evaluation of energy frameworks showed that stabilization of the compound was dominated by dispersion energy contributions. Both in vitro and in silico investigations on the DNA/bovine serum albumin (BSA) binding activity of the compound showed that the CT-DNA binding activity of the compound was mediated via intercalation and BSA binding activity was mediated via both polar and hydrophobic interactions. The anticancer activity of the compound was also tested by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay using human cell lines including MDA-MB-231, LNCaP, Caco-2, and HEK-293. The compound exhibited more cytotoxic activity than cisplatin and etoposide on Caco-2 cancer cell lines with an IC50 value of 16.63 ± 0.27 µM after 48 h. Annexin V suggests the induction of cell death by apoptosis. Compound 3 significantly increased the loss of mitochondrial membrane potential (MMP) levels in Caco-2 cells, and the reactive oxygen species (ROS) assay proved that compound 3 could induce apoptosis by ROS generation.

Mechanistic insight into impact of phosphorylation on the enzymatic steps of farnesyltransferase.

Farnesyltransferase (FTase) is a heterodimeric enzyme, which catalyzes covalent attachment of the farnesyl group to target proteins, thus coordinating their trafficking in the cell. FTase has been demonstrated to be highly expressed in cancer and neurological diseases; hence considered as a hot target for therapeutic purposes. However, due to the nonspecific inhibition, there has been only one inhibitor that could be translated into the clinic. Importantly, it has been shown that phosphorylation of the α-subunit of FTase increases the activity of the enzyme in certain diseases. As such, understanding the impact of phosphorylation on dynamics of FTase provides a basis for targeting a specific state of the enzyme that emerges under pathological conditions. To this end, we performed 18 µs molecular dynamics (MD) simulations using complexes of (non)-phosphorylated FTase that are representatives of the farnesylation reaction. We demonstrated that phosphorylation modulated the catalytic site by rearranging interactions between farnesyl pyrophosphate (FPP)/peptide substrate, catalytic Zn2+ ion/coordinating residues and hot-spot residues at the interface of the subunits, all of which led to the stabilization of the substrate and facilitation of the release of the product, thus collectively expediting the reaction rate. Importantly, we also identified a likely allosteric pocket on the phosphorylated FTase, which might be used for specific targeting of the enzyme. To the best of our knowledge, this is the first study that systematically examines the impact of phosphorylation on the enzymatic reaction steps, hence opens up new avenues for drug discovery studies that focus on targeting phosphorylated FTase.

Inhibition of SARS-CoV-2 main protease: a repurposing study that targets the dimer interface of the protein.

Coronavirus disease-2019 (COVID-19) was firstly reported in Wuhan, China, towards the end of 2019, and emerged as a pandemic. The spread and lethality rates of the COVID-19 have ignited studies that focus on the development of therapeutics for either treatment or prophylaxis purposes. In parallel, drug repurposing studies have also come into prominence. Herein, we aimed at having a holistic understanding of conformational and dynamical changes induced by an experimentally characterized inhibitor on main protease (Mpro) which would enable the discovery of novel inhibitors. To this end, we performed molecular dynamics simulations using crystal structures of apo and α-ketoamide 13b-bound Mpro homodimer. Analysis of trajectories pertaining to apo Mpro revealed a new target site, which is located at the homodimer interface, next to the catalytic dyad. Thereafter, we performed ensemble-based virtual screening by exploiting the ZINC and DrugBank databases and identified three candidate molecules, namely eluxadoline, diosmin, and ZINC02948810 that could invoke local and global conformational rearrangements which were also elicited by α-ketoamide 13b on the catalytic dyad of Mpro. Furthermore, ZINC23881687 stably interacted with catalytically important residues Glu166 and Ser1 and the target site throughout the simulation. However, it gave positive binding energy, presumably, due to displaying higher flexibility that might dominate the entropic term, which is not included in the MM-PBSA method. Finally, ZINC20425029, whose mode of action was different, modulated dynamical properties of catalytically important residue, Ala285. As such, this study presents valuable findings that might be used in the development of novel therapeutics against Mpro.Communicated by Ramaswamy H. Sarma.

Dimethyl sulfoxide reduces the stability but enhances catalytic activity of the main SARS-CoV-2 protease 3CLpro.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for coronavirus disease 2019 (COVID-19), one of the most challenging global pandemics of the modern era. Potential treatment strategies against COVID-19 are yet to be devised. It is crucial that antivirals that interfere with the SARS-CoV-2 life cycle be identified and developed. 3-Chymotrypsin-like protease (3CLpro) is an attractive antiviral drug target against SARS-CoV-2, and coronaviruses in general, because of its role in the processing of viral polyproteins. Inhibitors of 3CLpro activity are screened in enzyme assays before further development of the most promising leads. Dimethyl sulfoxide (DMSO) is a common additive used in such assays and enhances the solubility of assay components. However, it may also potentially affect the stability and efficiency of 3CLpro but, to date, this effect had not been analyzed in detail. Here, we investigated the effect of DMSO on 3CLpro-catalyzed reaction. While DMSO (5%-20%) decreased the optimum temperature of catalysis and thermodynamic stability of 3CLpro, it only marginally affected the kinetic stability of the enzyme. Increasing the DMSO concentration up to 20% improved the catalytic efficiency and peptide-binding affinity of 3CLpro. At such high DMSO concentration, the solubility and stability of peptide substrate were improved because of reduced aggregation. In conclusion, we recommend 20% DMSO as the minimum concentration to be used in screens of 3CLpro inhibitors as lead compounds for the development of antiviral drugs against COVID-19.

Design and synthesis of novel cylopentapyrazoles bearing 1,2,3-thiadiazole moiety as potent antifungal agents.

In drug-resistant phytopathogenic fungi, there has been extensive research on microbiological and antifungal drug development. In this study, a novel series of cylopentapyrazole bearing a 1,2,3-thiadiazole ring 2a-e were designed and synthesized according to the principle of combination of bioactive structures. Thus, we have employed a [3 + 2] cycloaddition with 4-methyl-[1,2,3] thiadiazole-5-carboxylic acid hydrazones 1a-e and cyclopentadiene ring. Novel synthesized compounds were identified with IR, 1H and 13C NMR, mass spectrometry and elemental analysis then, antifungal activities were assayed. Based on our study, a combination of the compounds 1a and 2b possess remarkable antifungal activity against Botrytis cinerea AHU 9424 with 100% inhibition. EC50 values were calculated by studying different doses in combinations with high inhibition rates. The combination of 1a + 2b has an EC50 value at 6.37 and 13.85 µg/ml concentrations against B. cinerea and F. culmorum, respectively. The combination of compound 1a + 2b, having a cylopentapyrazole ring on the 1,2,3-thiadiazole backbone, shows promising fungicidal activity and deserves further development. Additionally, the homology model of the CYP51 enzyme that belongs toFusarium moniliformewas generated using CYP51B (PDB ID: 6CR2), and molecular docking was performed using this homology model for each compound. The results of this study clearly indicate that these novel compounds can be identified as promising lead compounds and potential fungicidal agents in future.