An in-silico analysis of ivermectin interaction with potential SARS-CoV-2 targets and host nuclear importin α.

Ivermectin (IVM) is a broad-spectrum antiparasitic agent, having inhibitory potential against wide range of viral infections. It has also been found to hamper SARS-CoV-2 replication in vitro, and its precise mechanism of action against SARS-CoV-2 is yet to be understood. IVM is known to interact with host importin (IMP)α directly and averts interaction with IMPß1, leading to the prevention of nuclear localization signal (NLS) recognition. Therefore, the current study seeks to employ molecular docking, molecular mechanics generalized Born surface area (MM-GBSA) analysis and molecular dynamics simulation studies for decrypting the binding mode, key interacting residues as well as mechanistic insights on IVM interaction with 15 potential drug targets associated with COVID-19 as well as IMPα. Among all COVID-19 targets, the non-structural protein 9 (Nsp9) exhibited the strongest affinity to IVM showing -5.30 kcal/mol and -84.85 kcal/mol binding energies estimated by AutoDock Vina and MM-GBSA, respectively. However, moderate affinity was accounted for IMPα amounting -6.9 kcal/mol and -66.04 kcal/mol. Stability of the protein-ligand complexes of Nsp9-IVM and IMPα-IVM was ascertained by 100 ns trajectory of all-atom molecular dynamics simulation. Structural conformation of protein in complex with docked IVM exhibited stable root mean square deviation while root mean square fluctuations were also found to be consistent. In silico exploration of the potential targets and their interaction profile with IVM can assist experimental studies as well as designing of COVID-19 drugs. Communicated by Ramaswamy H. Sarma.

Association of Fluorescent Protein Pairs and Its Significant Impact on Fluorescence and Energy Transfer.

Fluorescent proteins (FPs) are commonly used in pairs to monitor dynamic biomolecular events through changes in proximity via distance dependent processes such as Förster resonance energy transfer (FRET). The impact of FP association is assessed by predicting dimerization sites in silico and stabilizing the dimers by bio-orthogonal covalent linkages. In each tested case dimerization changes inherent fluorescence, including FRET. GFP homodimers demonstrate synergistic behavior with the dimer being brighter than the sum of the monomers. The homodimer structure reveals the chromophores are close with favorable transition dipole alignments and a highly solvated interface. Heterodimerization (GFP with Venus) results in a complex with ≈87% FRET efficiency, significantly below the 99.7% efficiency predicted. A similar efficiency is observed when the wild-type FPs are fused to a naturally occurring protein-protein interface system. GFP complexation with mCherry results in loss of mCherry fluorescence. Thus, simple assumptions used when monitoring interactions between proteins via FP FRET may not always hold true, especially under conditions whereby the protein-protein interactions promote FP interaction.

Zerumbone Induces Apoptosis in Breast Cancer Cells by Targeting αvß3 Integrin upon Co-Administration with TP5-iRGD Peptide.

Cell-penetrating peptides (CPPs) are highly promising tools to deliver therapeutic molecules into tumours. αVß3 integrins are cell-matrix adhesion receptors, and are considered as an attractive target for anticancer therapies owing to their roles in the process of metastasis and angiogenesis. Therefore, this study aims to assess the effect of co-administration of zerumbone (ZER) and ZERencapsulated in hydroxypropyl-ß-cyclodextrin with TP5-iRGD peptide towards cell cytotoxicity, apoptosis induction, and proliferation of normal and cancerous breast cells utilizing in vitro assays, as well as to study the molecular docking of ZER in complex with TP5-iRGD peptide. Cell viability assay findings indicated that ZER and ZERencapsulated in hydroxypropyl-ß-cyclodextrin (ZER-HPßCD) inhibited the growth of estrogen receptor positivebreast cancer cells (ER+ MCF-7) at 72 h treatment with an inhibitory concentration (IC)50 of 7.51 ± 0.2 and 5.08 ± 0.2 µg/mL, respectively, and inhibited the growth of triple negative breast cancer cells (MDA-MB-231) with an IC50 of 14.96 ± 1.52 µg/mL and 12.18 ± 0.7 µg/mL, respectively. On the other hand, TP5-iRGD peptide showed no significant cytotoxicity on both cancer and normal cells. Interestingly, co-administration of TP5-iRGD peptide in MCF-7 cells reduced the IC50 of ZER from 7.51 ± 0.2 µg/mL to 3.13 ± 0.7 µg/mL and reduced the IC50 of ZER-HPßCD from 5.08 ± 0.2 µg/mL to 0.49 ± 0.004 µg/mL, indicating that the co-administration enhances the potency and increases the efficacy of ZER and ZER-HPßCD compounds. Acridine orange (AO)/propidium iodide (PI) staining under fluorescence microscopy showed evidence of early apoptosis after 72 h from the co-administration of ZER or ZER-HPßCD with TP5-iRGD peptide in MCF-7 breast cancer cells. The findings of the computational modelling experiment provide novel insights into the ZER interaction with integrin αvß3 in the presence of TP5-iRGD, and this could explain why ZER has better antitumor activities when co-administered with TP5-iRGD peptide.

Novel Approaches for Estimating Female Sex Worker Population Size in Conflict-Affected South Sudan.

BACKGROUND: Limited data exist describing the population size of female sex workers (FSW) in South Sudan. A population size estimation exercise among FSW was undertaken in Juba and Nimule during the Eagle Survey. OBJECTIVE: The study aimed to estimate the number of FSW in Juba and Nimule to inform resource allocation and service provision for FSW. METHODS: We utilized service and unique object multipliers, and 3-source capture-recapture methods in conjunction with a respondent-driven sampling (RDS) survey to estimate the number of FSW in Juba and Nimule. For service multiplier, the number of FSW testing for HIV in 2015 (Juba) and 2016 (Nimule) was obtained from the LINKAGES program targeting FSW. Survey participants were asked whether they had been tested for HIV by LINKAGES during the relevant period. A total of 2 separate unique object distributions were conducted in Juba and Nimule. In Nimule, these were combined to produce a 3-source capture-recapture estimate. The exercise involved distribution of key chains and bangles to FSW, documentation of the number of those who received unique objects, and questions during RDS survey to assess whether participants received unique objects. RESULTS: In Juba, the service multiplier method yielded an estimate of 5800 (95% CI 4927-6673) FSW. The unique object estimate (key chain and RDS participation) yielded 5306 (95% CI 4673-5939). Another estimate using RDS participation and receipt of a bangle yielded a much lower estimate of 1863 (95% CI 1776-1951), as did a 2-source estimate of key chain and bangle (2120, 95% CI 2028-2211). A 3-source capture-recapture estimate could not be produced because aggregate rather than individual level data were collected during the third capture. The multiplier estimate using key chain and RDS participation was taken as the final population estimate for FSW in Juba, which constitutes more than 6% of the female population aged 15 to 64 years. In Nimule, the service multiplier method yielded an estimate of 9384 (95% CI 8511-10,257). The 2-source estimates for key chain and RDS yielded 6973 (95% CI 4759-9186); bangles and RDS yielded a higher estimate of 13,104 (95% CI 7101-19,106); key chains and bangles yielded a lower estimate of 1322 (95% CI 1223-1420). The 3-source capture-recapture method using Bayesian nonparametric latent-class model-based estimate yielded a population of 2694 (95% CI 1689-6945), and this was selected as the final estimate for Nimule, which constitutes nearly 40% of female population aged 15 to 64 years. CONCLUSIONS: The service and unique object multiplier, and 3-source capture-recapture methods were successfully used to estimate the number of FSW in Nimule, whereas service and unique object multiplier methods were successfully used in Juba. These methods yielded higher than previously estimated FSW population sizes. These estimates will inform resource allocation and advocacy efforts to support services for FSW.

Zerumbone binding to estrogen receptors: an in-silico investigation.

Breast cancer is the most frequent malignancy among females worldwide. Estrogen receptor (ER) mediate important pathophysiological signaling pathways induced by estrogens, and is regarded as a promising target for the treatment of breast cancer. Zerumbone (2,6,9,9-tetramethylcycloundeca-2,6,10-trien-1-one; ZER), a chemical constituent present in the Zingiber zerumbet is known to exhibit anti-breast cancer activity by modulating several proteins to induce apoptosis. Medicinal chemists usually exploit lead compounds of natural origin to develop molecules with improved pharmacological properties. Current study is intended to utilize molecular modeling techniques to investigate the interaction of ZER with estrogen receptors. AutoDock was used to predict the binding modes of ZER and target receptors. Stability of the ZER-ER complex was verified by molecular dynamics simulation using Desmond software. Docked ZER was further optimized by density functional theory (DFT) using Gaussian09 program. Analysis of docked conformations in terms of binding energy disclosed estrogen receptor-ß (ERß) as more promising than estrogen receptor-α (ERα). Evaluation of MD trajectories of ZER bound to both ERα and ERß showed appreciable stability with minimum Cα-atom root mean square deviation shifts. DFT based global reactivity descriptors such as electron affinity, hardness, chemical potential, electronegativity and electrophilicity index, calculated from the energies of highest occupied and lowest unoccupied molecular orbitals underscored the electronic features governing viability of the ZER for interaction with the target receptors. In conclusion, these findings can be exploited to design and develop novel anticancer agents based on the lead compound, ZER.

NSAIDs as potential treatment option for preventing amyloid ß toxicity in Alzheimer's disease: an investigation by docking, molecular dynamics, and DFT studies.

Aggregation of amyloid beta (Aß) protein considered as one of contributors in development of Alzheimer's disease (AD). Several investigations have identified the importance of non-steroidal anti-inflammatory drugs (NSAIDs) as Aß aggregation inhibitors. Here, we have examined the binding interactions of 24 NSAIDs belonging to eight different classes, with Aß fibrils by exploiting docking and molecular dynamics studies. Minimum energy conformation of the docked NSAIDs were further optimized by density functional theory (DFT) employing Becke's three-parameter hybrid model, Lee-Yang-Parr (B3LYP) correlation functional method. DFT-based global reactivity descriptors, such as electron affinity, hardness, softness, chemical potential, electronegativity, and electrophilicity index were calculated to inspect the expediency of these descriptors for understanding the reactive nature and sites of the molecules. Few selected NSAID-Aß fibrils complexes were subjected to molecular dynamics simulation to illustrate the stability of these complexes and the most prominent interactions during the simulated trajectory. All of the NSAIDs exhibited potential activity against Aß fibrils in terms of predicted binding affinity. Sulindac was found to be the most active compound underscoring the contribution of indene methylene substitution, whereas acetaminophen was observed as least active NSAID. General structural requirements for interaction of NSAIDs with Aß fibril include: aryl/heteroaryl aromatic moiety connected through a linker of 1-2 atoms to a distal aromatic group. Considering these structural requirements and electronic features, new potent agents can be designed and developed as potential Aß fibril inhibitors for the treatment of AD.

Novel Aryl Substituted Pyrazoles as Small Molecule Inhibitors of Cytochrome P450 CYP121A1: Synthesis and Antimycobacterial Evaluation.

Three series of biarylpyrazole imidazole and triazoles are described, which vary in the linker between the biaryl pyrazole and imidazole/triazole group. The imidazole and triazole series with the short -CH2- linker displayed promising antimycobacterial activity, with the imidazole-CH2- series (7) showing low MIC values (6.25-25 µg/mL), which was also influenced by lipophilicity. Extending the linker to -C(O)NH(CH2)2- resulted in a loss of antimycobacterial activity. The binding affinity of the compounds with CYP121A1 was determined by UV-visible optical titrations with KD values of 2.63, 35.6, and 290 µM, respectively, for the tightest binding compounds 7e, 8b, and 13d from their respective series. Both binding affinity assays and docking studies of the CYP121A1 inhibitors suggest type II indirect binding through interstitial water molecules, with key binding residues Thr77, Val78, Val82, Val83, Met86, Ser237, Gln385, and Arg386, comparable with the binding interactions observed with fluconazole and the natural substrate dicyclotyrosine.

Analysis of the binding sites of vitamin D 1α-hydroxylase (CYP27B1) and vitamin D 24-hydroxylase (CYP24A1) for the design of selective CYP24A1 inhibitors: Homology modelling, molecular dynamics simulations and identification of key binding requirements.

A homology model of human CYP27B1 was built using MOE and was further optimised by molecular dynamics simulations of the hCYP27B1 homology model and a hCYP27B1-SDZ-88357 complex. Docking results from the hCYP27B1-SDZ-88357 complex showed amino acids Arg107, Asn387 and Asp320 have an important role in binding interaction, with Asp320 part of the important acid-alcohol pair situated in the I-helix with the conserved sequence (A/G) GX (E/D) (T/S), which assumes an essential role in the binding of an oxygen molecule for catalysis. Additional docking experiments with selective hCYP27B1 or hCYP24A1 inhibitors using both the hCYP27B1 model and a triple mutant hCYP24A1 model provided further support for the importance of H-bonding interactions with the three identified active site amino acids. To confirm the role of Arg107, Asn387 and Asp320 in the active site of hCYP27B1 compounds were designed that would form H-bonding interactions, as determined from docking experiments with the hCYP27B1 model. Subsequent synthesis and CYP24A1 and CYP27B1 enzyme assays of the designed compounds 1a and 1b showed a∼5-fold selectivity for CYP27B1 confirming the importance of Asp320 in particular and also Asn387 and Arg107 as important amino acids for CYP27B1 inhibitory activity.

Synthesis, molecular modelling and CYP24A1 inhibitory activity of novel of (E)-N-(2-(1H-imidazol-1-yl)-2-(phenylethyl)-3/4-styrylbenzamides.

CYP24A1 (25-hydroxyvitamin D-24-hydroxylase) is a useful enzyme target for a range of medical conditions including cancer, cardiovascular and autoimmune disease, which show elevated CYP24A1 levels and corresponding reduction of calcitriol (the biologically active form of vitamin D). A series of (E)-N-(2-(1H-imidazol-1-yl)-2-(phenylethyl)-3/4-styrylbenzamides have been synthesised using an efficient synthetic route and shown to be potent inhibitors of CYP24A1 (IC50 0.11-0.35µM) compared with the standard ketoconazole. Molecular modelling using our CYP24A1 homology model showed the inhibitors to fill the hydrophobic binding site, forming key transition metal interaction between the imidazole nitrogen and the haem Fe3+ and multiple interactions with the active site amino acid residues.