Development of Urea-Bond-Containing Michael Acceptors as Antitrypanosomal Agents Targeting Rhodesain.

Human African Trypanosomiasis (HAT) is a neglected tropical disease widespread in sub-Saharan Africa. Rhodesain, a cysteine protease of Trypanosoma brucei rhodesiense, has been identified as a valid target for the development of anti-HAT agents. Herein, we report a series of urea-bond-containing Michael acceptors, which were demonstrated to be potent rhodesain inhibitors with K i values ranging from 0.15 to 2.51 nM, and five of them showed comparable k 2nd values to that of K11777, a potent antitrypanosomal agent. Moreover, most of the urea derivatives exhibited single-digit micromolar activity against the protozoa, and the presence of substituents at the P3 position appears to be essential for the antitrypanosomal effect. Replacement of Phe with Leu at the P2 site kept unchanged the inhibitory properties. Compound 7 (SPR7) showed the best compromise in terms of rhodesain inhibition, selectivity, and antiparasitic activity, thus representing a new lead compound for future SAR studies.

Development of Reduced Peptide Bond Pseudopeptide Michael Acceptors for the Treatment of Human African Trypanosomiasis.

Human African Trypanosomiasis (HAT) is an endemic protozoan disease widespread in the sub-Saharan region that is caused by T. b. gambiense and T. b. rhodesiense. The development of molecules targeting rhodesain, the main cysteine protease of T. b. rhodesiense, has led to a panel of inhibitors endowed with micro/sub-micromolar activity towards the protozoa. However, whilst impressive binding affinity against rhodesain has been observed, the limited selectivity towards the target still remains a hard challenge for the development of antitrypanosomal agents. In this paper, we report the synthesis, biological evaluation, as well as docking studies of a series of reduced peptide bond pseudopeptide Michael acceptors (SPR10-SPR19) as potential anti-HAT agents. The new molecules show Ki values in the low-micro/sub-micromolar range against rhodesain, coupled with k2nd values between 1314 and 6950 M-1 min-1. With a few exceptions, an appreciable selectivity over human cathepsin L was observed. In in vitro assays against T. b. brucei cultures, SPR16 and SPR18 exhibited single-digit micromolar activity against the protozoa, comparable to those reported for very potent rhodesain inhibitors, while no significant cytotoxicity up to 70 µM towards mammalian cells was observed. The discrepancy between rhodesain inhibition and the antitrypanosomal effect could suggest additional mechanisms of action. The biological characterization of peptide inhibitor SPR34 highlights the essential role played by the reduced bond for the antitrypanosomal effect. Overall, this series of molecules could represent the starting point for further investigations of reduced peptide bond-containing analogs as potential anti-HAT agents.

Development of novel dipeptide nitriles as inhibitors of rhodesain of Trypanosoma brucei rhodesiense.

In this paper, we developed a new series of dipeptide nitriles that were demonstrated to be reversible rhodesain inhibitors at nanomolar level, with EC50 values against cultured T. b. brucei in the micromolar range. We also proved that our dipeptide nitriles directly bind to the active site of rhodesain acting as competitive inhibitors. Within the most interesting compounds, the dipeptide nitrile 2b showed the highest binding affinity towards rhodesain (Ki = 16 nM) coupled with a good antiparasitic activity (EC50 = 14.1 µM). Moreover, for the dipeptide nitrile 3e, which showed a Ki = 122 nM towards the trypanosomal protease, we obtained the highest antiparasitic activity (EC50 = 8.8 µM). Thus, given the obtained results both compounds could certainly represent new lead compounds for the discovery of new drugs to treat Human African Trypanosomiasis.

Ensemble-based high-throughput virtual screening of natural ligands using the Super Natural-II database against cell-wall protein dTDP-4-dehydrorhamnose reductase (RmlD) in Mycobacterium tuberculosis.

The disease Tuberculosis (TB) is caused by a bacterium called Mycobacterium tuberculosis (Mtb). The bacterial cell-wall consists of peptidoglycan layer maintains the cellular integrity and cell viability. The main problem resides in the cell cycle of Mycobacterium tuberculosis in its quiescent form which is not targeted by any drugs hence there is an immediate need for new antibiotics to target the cell wall. The current study deals with the dTDP-4-dehydrorahmnose reductase (RmlD) which is the final enzyme in the series of cell-wall proteins of Mtb. The RmlD is a part of Carbohydrate biosynthesis has been considered as a good drug target for the novel class of antibiotics. Our study begins with the protein structure prediction, Homology studies were conducted using the Phyre2 web server. The structure is then refined and subjected to molecular dynamics simulations for 50 ns using GROMACS. The clustering analysis has been carried out and generated 41 clusters with 2 Å as the cut-off. Blind docking virtual screening was performed against RmlD protein using the Super Natural-II database with AutoDock4.0. its results helped to screen top ligands based on best binding energies. In both dockings, there are some common residues in which the ligands are interacting and forming the Hydrogen bonds such as Asp-105, Val-158, Thr-160, Gly-161, Arg-224, Arg-256. The ligand-567 giving the best results by being in the top-3 of all the clusters in both blind docking as well as the active-site docking. Hence ligand-567 can be a potential inhibitor of RmlD which can further inhibit the cell-wall synthesis of Mycobacterium tuberculosis.Communicated by Ramaswamy H. Sarma.

Translational suppression of SARS-COV-2 ORF8 protein mRNA as a Viable therapeutic target against COVID-19: Computational studies on potential roles of isolated compounds from Clerodendrum volubile leaves.

The open reading frame 8 (ORF8) protein of SARS-CoV-2 has been implicated in the onset of cytokine storms, which are responsible for the pathophysiology of COVID-19 infection. The present study investigated the potential of isolated compounds from Clerodendrum volubile leaves to stall oxidative bursts in vitro and interact with ORF8 mRNA segments of the SARS-CoV-2 whole genome using computational tools. Five compounds, namely, harpagide, 1-(3-methyl-2-butenoxy)-4-(1-propenyl)benzene, ajugoside, iridoid glycoside and erucic acid, were isolated from C. volubile leaves, and their structures were elucidated using conventional spectroscopy tools. Iridoid glycoside is being reported for the first time and is thus regarded as a new compound. The ORF8 mRNA sequences of the translation initiation sites (TIS) and translation termination sites (TTSs) encoding ORF8 amino acids were retrieved from the full genome of SARS-CoV-2. Molecular docking studies revealed strong molecular interactions of the isolated compounds with the TIS and TTS of ORF8 mRNA. Harpagide showed the strongest binding affinity for TIS, while erucic acid was the strongest for TTS. The immunomodulatory potentials of the isolated compounds were investigated on neutrophil phagocytic respiratory bursts using luminol-amplified chemiluminescence technique. The compounds significantly inhibited oxidative burst, with 1-(3-methyl-2-butenoxy)-4-(1-propenyl)benzene having the best activity. Ajugoside and erucic acid showed significant inhibitory activity on T-cell proliferation. These results indicate the potential of C. volubile compounds as immunomodulators and can be utilized to curb cytokine storms implicated in COVID-19 infection. These potentials are further corroborated by the strong interactions of the compounds with the TIS and TTS of ORF8 mRNA from the SARS-CoV-2 whole genome.

Tetrahydroquinazole-based secondary sulphonamides as carbonic anhydrase inhibitors: synthesis, biological evaluation against isoforms I, II, IV, and IX, and computational studies.

A library of variously decorated N-phenyl secondary sulphonamides featuring the bicyclic tetrahydroquinazole scaffold was synthesised and biologically evaluated for their inhibitory activity against human carbonic anhydrase (hCA) I, II, IV, and IX. Of note, several compounds were identified showing submicromolar potency and excellent selectivity for the tumour-related hCA IX isoform. Structure-activity relationship data attained for various substitutions were rationalised by molecular modelling studies in terms of both inhibitory activity and selectivity.

Carbonic anhydrase activation profile of indole-based derivatives.

Carbonic Anhydrase Activators (CAAs) could represent a novel approach for the treatment of Alzheimer's disease, ageing, and other conditions that require remedial achievement of spatial learning and memory therapy. Within a research project aimed at developing novel CAAs selective for certain isoforms, three series of indole-based derivatives were investigated. Enzyme activation assay on human CA I, II, VA, and VII isoforms revealed several effective micromolar activators, with promising selectivity profiles towards the brain-associated cytosolic isoform hCA VII. Molecular modelling studies suggested a theoretical model of the complex between hCA VII and the new activators and provide a possible explanation for their modulating as well as selectivity properties. Preliminary biological evaluations demonstrated that one of the most potent CAA 7 is not cytotoxic and is able to increase the release of the brain-derived neurotrophic factor (BDNF) from human microglial cells, highlighting its possible application in the treatment of CNS-related disorders.

L-leucine stimulation of glucose uptake and utilization involves modulation of glucose - lipid metabolic switch and improved bioenergetic homeostasis in isolated rat psoas muscle ex vivo.

The antidiabetic effect of l-leucine has been attributed to its modulatory effect on glucose uptake and lipid metabolism in muscles. However, there is a dearth on its effect on glucose metabolism in muscles. Thus, the present study investigated the effect of l-leucine - stimulated glucose uptake on glucose metabolism, dysregulated lipid metabolic pathways, redox and bioenergetic homeostasis, and proteolysis in isolated psoas muscle from Sprague Dawley male rats. Isolated psoas muscles were incubated with l-leucine (30-240 µg/mL) in the presence of 11.1 mMol glucose at 37 ˚C for 2 h. Muscles incubated in only glucose served as the control, while muscles not incubated in l-leucine and/or glucose served as the normal control. Metformin (6.04 mM) was used as the standard antidiabetic drug. Incubation with l-leucine caused a significant increase in muscle glucose uptake, with an elevation of glutathione levels, superoxide dismutase, catalase, E-NTPDase and 5'nucleotidase activities. It also led to the depletion of malondialdehyde and nitric oxide levels, ATPase, chymotrypsin, acetylcholinesterase, glycogen phosphorylase, glucose-6-phosphatase, fructose-1,6-bisphosphatase and lipase activities. There was an alteration in lipid metabolites, with concomitant activation of glycerolipid metabolism, fatty acid metabolism, and fatty acid elongation in mitochondria in the glucose-incubated muscle (negative control). Incubation with l-leucine reversed these alterations, and concomitantly deactivated the pathways. These results indicate that l-leucine-enhanced muscle glucose uptake involves improved redox and bioenergetic homeostasis, with concomitant suppressed proteolytic, glycogenolytic and gluconeogenetic activities, while modulating glucose - lipid metabolic switch.

Inhibition studies on carbonic anhydrase isoforms I, II, IV and IX with N-arylsubstituted secondary sulfonamides featuring a bicyclic tetrahydroindazole scaffold.

Carbonic Anhydrases (CAs) are pharmaceutically relevant targets for the treatment of several disease conditions. The ubiquitous localization of these enzymes and the high homology shared by the different isoforms represent substantial impediments for the discovery of potential drugs devoid of off-target side effects. As a consequence, substantial efforts are still needed to allow for the full realization of the pharmacological potential of CA modulators. In this contribution, starting from our previous studies, we describe the synthesis of a set of new bicyclic tetrahydroindazoles featuring a secondary sulfonamide. Biological evaluation of the inhibitory activity against the hCA I, II, IV, and IX isoforms allowed drawing a structure-activity relationship profile that was rationalized through theoretical studies. This allowed dissecting the new molecules into the single portions influencing the zinc chelation properties and the selectivity profile thereby offering a new platform for the discovery of new isotype selective CA inhibitors.

Landscape of ROD9 Island: Functional annotations and biological network of hypothetical proteins in Salmonella enterica.

Salmonella, an Enterobacteria is a therapeutically important pathogen for the host. The advancement of genome sequencing of S. enterica serovar Enteritidis have identified a distinct ROD9 pathogenic island, imparting virulence. The occurrence of 17 ROD9 hypothetical proteins, necessitates subsequent bioinformatics approach for structural and functional aspects of protein-protein relations or networks in different pathogenic phenotypes express. A collective analysis using predictive bioinformatics tools that includes NCBI-BLASTp and BLAST2GO annotated the motif patterns and functional significance. The VFDB identified 10 virulence proteins at both genomic and metagenomic level. Phylogenetic analysis revealed a divergent and convergent relationship between 17 ROD9 and 41 SP-1 proteins. Here, combining a comprehensive approach from sequence based, motif recognitions, domain identification, virulence ability to structural modelling provides a precise function to ROD9 proteins biological network, for which no experimental information is available.

Genetics of PCOS: A systematic bioinformatics approach to unveil the proteins responsible for PCOS.

Polycystic ovary syndrome (PCOS) is a hormonal imbalance in women, which causes problems during menstrual cycle and in pregnancy that sometimes results in fatality. Though the genetics of PCOS is not fully understood, early diagnosis and treatment can prevent long-term effects. In this study, we have studied the proteins involved in PCOS and the structural aspects of the proteins that are taken into consideration using computational tools. The proteins involved are modeled using Modeller 9v14 and Ab-initio programs. All the 43 proteins responsible for PCOS were subjected to phylogenetic analysis to identify the relatedness of the proteins. Further, microarray data analysis of PCOS datasets was analyzed that was downloaded from GEO datasets to find the significant protein-coding genes responsible for PCOS, which is an addition to the reported protein-coding genes. Various statistical analyses were done using R programming to get an insight into the structural aspects of PCOS that can be used as drug targets to treat PCOS and other related reproductive diseases.