Identification of Phytochemicals from Arabian Peninsula Medicinal Plants as Strong Binders to SARS-CoV-2 Proteases (3CLPro and PLPro) by Molecular Docking and Dynamic Simulation Studies.

We provide promising computational (in silico) data on phytochemicals (compounds 1-10) from Arabian Peninsula medicinal plants as strong binders, targeting 3-chymotrypsin-like protease (3CLPro) and papain-like proteases (PLPro) of SARS-CoV-2. Compounds 1-10 followed the Lipinski rules of five (RO5) and ADMET analysis, exhibiting drug-like characters. Non-covalent (reversible) docking of compounds 1-10 demonstrated their binding with the catalytic dyad (CYS145 and HIS41) of 3CLPro and catalytic triad (CYS111, HIS272, and ASP286) of PLPro. Moreover, the implementation of the covalent (irreversible) docking protocol revealed that only compounds 7, 8, and 9 possess covalent warheads, which allowed the formation of the covalent bond with the catalytic dyad (CYS145) in 3CLPro and the catalytic triad (CYS111) in PLPro. Root-mean-square deviation (RMSD), root-mean-square fluctuation (RMSF), and radius of gyration (Rg) analysis from molecular dynamic (MD) simulations revealed that complexation between ligands (compounds 7, 8, and 9) and 3CLPro and PLPro was stable, and there was less deviation of ligands. Overall, the in silico data on the inherent properties of the above phytochemicals unravel the fact that they can act as reversible inhibitors for 3CLPro and PLPro. Moreover, compounds 7, 8, and 9 also showed their novel properties to inhibit dual targets by irreversible inhibition, indicating their effectiveness for possibly developing future drugs against SARS-CoV-2. Nonetheless, to confirm the theoretical findings here, the effectiveness of the above compounds as inhibitors of 3CLPro and PLPro warrants future investigations using suitable in vitro and in vivo tests.

Covalent Inhibitors from Saudi Medicinal Plants Target RNA-Dependent RNA Polymerase (RdRp) of SARS-CoV-2.

COVID-19, a disease caused by SARS-CoV-2, has caused a huge loss of human life, and the number of deaths is still continuing. Despite the lack of repurposed drugs and vaccines, the search for potential small molecules to inhibit SARS-CoV-2 is in demand. Hence, we relied on the drug-like characters of ten phytochemicals (compounds 1-10) that were previously isolated and purified by our research team from Saudi medicinal plants. We computationally evaluated the inhibition of RNA-dependent RNA polymerase (RdRp) by compounds 1-10. Non-covalent (reversible) docking of compounds 1-10 with RdRp led to the formation of a hydrogen bond with template primer nucleotides (A and U) and key amino acid residues (ASP623, LYS545, ARG555, ASN691, SER682, and ARG553) in its active pocket. Covalent (irreversible) docking revealed that compounds 7, 8, and 9 exhibited their irreversible nature of binding with CYS813, a crucial amino acid in the palm domain of RdRP. Molecular dynamic (MD) simulation analysis by RMSD, RMSF, and Rg parameters affirmed that RdRP complexes with compounds 7, 8, and 9 were stable and showed less deviation. Our data provide novel information on compounds 7, 8, and 9 that demonstrated their non-nucleoside and irreversible interaction capabilities to inhibit RdRp and shed new scaffolds as antivirals against SARS-CoV-2.

Marine Natural Compound (Neviotin A) Displays Anticancer Efficacy by Triggering Transcriptomic Alterations and Cell Death in MCF-7 Cells.

We investigated the anticancer mechanism of a chloroform extract of marine sponge (Haliclona fascigera) (sample C) in human breast adenocarcinoma (MCF-7) cells. Viability analysis using MTT and neutral red uptake (NRU) assays showed that sample C exposure decreased the proliferation of cells. Flow cytometric data exhibited reactive oxygen species (ROS), nitric oxide (NO), dysfunction of mitochondrial potential, and apoptosis in sample C-treated MCF-7 cells. A qPCR array of sample C-treated MCF-7 cells showed crosstalk between different pathways of apoptosis, especially BIRC5, BCL2L2, and TNFRSF1A genes. Immunofluorescence analysis affirmed the localization of p53, bax, bcl2, MAPKPK2, PARP-1, and caspase-3 proteins in exposed cells. Bioassay-guided fractionation of sample C revealed Neviotin A as the most active compound triggering maximum cell death in MCF-7, indicating its pharmacological potency for the development of a drug for the treatment of human breast cancer.

Novel anti­hepatitis B virus flavonoids sakuranetin and velutin from Rhus retinorrhoea.

Drug­resistance in hepatitis B virus (HBV), especially due to prolonged treatment with nucleoside analogs, such as lamivudine (LAM), remains a clinical challenge. Alternatively, several plant products and isolated phytochemicals have been used as promising anti­HBV therapeutics with no sign of resistance. Among all known Rhus species, R. coriaria, R. succedanea and R. tripartite have been widely studied for their anti­HBV efficacy, however, the effects of R. retinorrhoea have not been previously investigated. The current study reported the isolation of two flavonoids, namely sakuranetin (SEK) and velutin (VEL), from the dichloromethane fraction of R. retinorrhoea aerial parts using chromatography and spectral analyses. The two flavonoids (6.25­50 µg/ml) were pre­tested for non­hepatocytotoxicity using an MTT assay and their dose­ and time­dependent inhibitory activities against HBV [hepatitis B surface antigen (HBsAg) and hepatitis B 'e' antigen (HBeAg)] in cultured HepG2.2.15 cells were assessed by ELISA. SEK and VEL at the selected doses (12.5 µg/ml) significantly inhibited HBsAg by ~58.8 and ~56.4%, respectively, and HBeAg by ~55.5 and ~52.4%, respectively, on day 5. The reference drugs LAM and quercetin (anti­HBV flavonoids), suppressed the production of HBsAg/HBeAg by ~86.4/~64 and ~84.5/~62%, respectively. Furthermore, molecular docking of the flavonoids with HBV polymerase and capsid proteins revealed the formation of stable complexes with good docking energies, thus supporting their structure­based antiviral mechanism. In conclusion, the present study was the first to demonstrate the anti­HBV therapeutic activities of SEK and VEL isolated from R. retinorrhoea.

Identification of potent anti-immunogenic agents through virtual screening, 3D-QSAR studies, and in vitro experiments.

A wealth of literature has highlighted the discovery of various immune modulators, frequently used in clinical practice, yet associated with numerous drawbacks. In light of this pharmacological deficiency, medical scientists are motivated to develop new immune modulators with minimized adverse effects yet retaining the improved therapeutic potential. T-cell differentiation and growth are central to human defense and are regulated by interleukin-2 (IL-2), an immune-modulatory cytokine. However, scientific investigation is hindered due to its flat binding site and widespread hotspot residues. In this regard, a prompt and logical investigation guided by integrated computational techniques was undertaken to unravel new and potential leads against IL-2. In particular, the combination of score-based and pharmacophore-based virtual screening approaches were employed, reducing the data from millions of small molecules to a manageable number. Subsequent docking and 3D-QSAR prediction via CoMFA further helped remove false positives from the data. The reliability of the model was assessed via standard metrics, which explain the model's fitness and the robustness of the model in predicting the activity of new compounds. The extensive virtual screening herein led to the identification of a total of 24 leads with potential anti-IL-2 activity. Furthermore, the theoretical findings were corroborated with in vitro testing, further endorsing the anti-inflammatory potential of the identified leads.

Compounds Related to Saudin and Three New Series of Diterpenoids from Clutia lanceolata.

Clutia lanceolata is a medicinal plant native to Ethiopia and sub-Saharan Africa and to the Arabian Peninsula. It is used traditionally in Saudi Arabia for the treatment of diabetes. Previous phytochemical analysis of this species has been limited to the identification of methylthiocoumarins. Further work has led to isolation of 19 new diterpenoids in three structural classes. Their structures were established by HRMS and by a range of NMR techniques (1H, 13C, COSY, NOESY, HSQC, HMBC), with confirmation for some examples by X-ray crystallography. NOESY and 1H-1H NMR coupling constants gave the relative stereochemical configurations and conformational information, with absolute configurations being established through X-ray crystallography. One example closely related to the known hypoglycemic compound saudin (found in C. richardiana and also in C. lanceolata) and one with a different core tetracycle were found to enhance strongly the glucose-triggered release of insulin from murine pancreatic islets. Biosynthetic proposals for the three groups of new diterpenoids by alternative cyclization of a common precursor are put forward. Lanceolide P (16) is proposed as a lead compound for further development for the treatment of diabetes.

Artificial Intelligence for Antimicrobial Resistance Prediction: Challenges and Opportunities towards Practical Implementation.

Antimicrobial resistance (AMR) is emerging as a potential threat to many lives worldwide. It is very important to understand and apply effective strategies to counter the impact of AMR and its mutation from a medical treatment point of view. The intersection of artificial intelligence (AI), especially deep learning/machine learning, has led to a new direction in antimicrobial identification. Furthermore, presently, the availability of huge amounts of data from multiple sources has made it more effective to use these artificial intelligence techniques to identify interesting insights into AMR genes such as new genes, mutations, drug identification, conditions favorable to spread, and so on. Therefore, this paper presents a review of state-of-the-art challenges and opportunities. These include interesting input features posing challenges in use, state-of-the-art deep-learning/machine-learning models for robustness and high accuracy, challenges, and prospects to apply these techniques for practical purposes. The paper concludes with the encouragement to apply AI to the AMR sector with the intention of practical diagnosis and treatment, since presently most studies are at early stages with minimal application in the practice of diagnosis and treatment of disease.

Structural basis for the mutation-induced dysfunction of the human IL-15/IL-15α receptor complex.

In silico strategies offer a reliable, fast, and inexpensive, way compared to the clumsy in vitro approaches to boost understanding of the effect of amino acid substitution on the structure and consequently the associated function of proteins. In the present work, we report an atomistic-based, reliable in silico structural and energetic framework of the interactions between the receptor-binding domain of the Interleukin-15 (IL-15) protein and its receptor Interleukin-15α (IL-15α), consequently, providing qualitative and quantitative details of the key molecular determinants in ligand/receptor recognition. Molecular dynamics simulations were used to investigate the dynamic behavior of the specific binding between IL-15 and IL-15α followed by estimation of the free energies via molecular mechanics/generalized Born surface area (MM/GBSA). In particular, residues Y26, E46, E53, and E89 of the IL-15 protein receptor-binding domain are identified as main hot spots, shaping and governing the stability of the assembly. These results can be used for the development of neutralizing antibodies and the effective structure-based design of protein-protein interaction inhibitors against the so-called orphan disease, vitiligo.

Noncontact assessment for fatigue based on heart rate variability using IR-UWB radar.

Physical fatigue can be assessed using heart rate variability (HRV). We measured HRV at rest and in a fatigued state using impulse-radio ultra wideband (IR-UWB) radar in a noncontact fashion and compared the measurements with those obtained using electrocardiography (ECG) to assess the reliability and validity of the radar measurements. HRV was measured in 15 subjects using radar and ECG simultaneously before (rest for 10 min before exercise) and after a 20-min exercise session (fatigue level 1 for 0-9 min; fatigue level 2 for 10-19 min; recovery for ≥ 20 min after exercise). HRV was analysed in the frequency domain, including the low-frequency component (LF), high-frequency component (HF) and LF/HF ratio. The LF/HF ratio measured using radar highly agreed with that measured using ECG during rest (ICC = 0.807), fatigue-1 (ICC = 0.712), fatigue-2 (ICC = 0.741) and recovery (ICC = 0.764) in analyses using intraclass correlation coefficients (ICCs). The change pattern in the LH/HF ratios during the experiment was similar between radar and ECG. The subject's body fat percentage was linearly associated with the time to recovery from physical fatigue (R2 = 0.96, p < 0.001). Our results demonstrated that fatigue and rest states can be distinguished accurately based on HRV measurements using IR-UWB radar in a noncontact fashion.

Insilico structure based drug design approach to find potential hits in ventilator-associated pneumonia caused by Pseudomonas aeruginosa.

Tetraacyldisaccharide 4'-kinase (LpxK) is the prime enzyme responsible for the biosynthesis of lipid A. LpxK is a key antibacterial drug target, but it is less exploitation in Pseudomonas aeruginosa and other bacterial species limits its therapeutic use. Pseudomonas aeruginosa is responsible for severe infections like pneumonia and urinary tract infections. The precautionary measures of Pseudomonas aeruginosa infections are decisive as it results in extensive drug resistance, systemic bacteremia, and ventilator-associated pneumonia. The current rational approach highlights exploiting the use of computer-aided drug design approaches to counter Pseudomonas aeruginosa specific LpxK. The various approaches used were exploring the metabolic pathway database (Metacyc), drug target validation using DEG, protein modeling, ligand docking, e-pharmacophore assisted virtual screening, physicochemical and Toxicity profile prediction studies, and molecular simulations in spotting out novel potential hits compounds. The virtual hits which have highly ranked in the study were STOCK4S-16119, STOCK1S -60869, STOCK6S -43621, STOCK6S -3328, and STOCKS-39892 which can act as a scaffold for the establishment of new hits against LpxK and can result in control of Pseudomonas aeruginosa infectivity.

The anti-oxidative, anti-cell proliferative and anti-microbial efficacies of cold-adapted Crepis flexuosa: HPTLC and GC/MS analyses.

The genus Crepis constitutes cold-adapted plant spp., of these some are traditionally used in folk medicine against inflammation or fungal infections without scientific validations. Here, we report the biological activities of Crepis flexuosa total ethanol-extract (CF-EtOH) and its hexane (CF-Hex), ethyl acetate (CF-EtOA), butanol (CF-ButOH), and aqueous (CF-Aqua) fractions. Our in vitro DPPH and ABTS radical-scavenging assays showed CF-EtOH, CF-ButOH and CF-Aqua with maximal, CF-EtOA with moderate, and CF-Hex with mild anti-oxidant activities. When tested on human cancer cell lines, high cytotoxicity was demonstrated by CF-EtOH (IC50: 42.45 µg/ml) and CF-Aqua (IC50: 46.37 µg/ml) on HepG2, followed by CF-Hex (IC50: 63.24 µg/ml) and CF-ButOH (IC50: 65.32 µg/ml) on MCF7 cells. The human primary cell line (HUVEC) had comparatively lower cytotoxicity for the tested samples. Moreover, when assessed for anti-microbial efficacy, CF-ButOH and CF-Aqua exhibited the strongest activity (MIC: 156.25 µg/ml) against S. aureus, E. faecalis and C. albicans. Further, while the developed RP-HPTLC identified the bioactive flavonoid luteolin-7-O-glucoside (17.58 mg/g), GS/MS analysis revealed sixteen compounds in C. flexuosa extract. In conclusion, we for the first time show the promising anti-oxidative, anti-cell proliferative and anti-microbial efficacies of C. flexuosa. This warrants further phytochemical and bio-efficacy studies towards isolations and identifications of active principles.

Exploring the Role of Antioxidants to Combat Oxidative Stress in Malaria Parasites.

BACKGROUND: Malaria, a global challenge, is a parasitic disease caused by Plasmodium species. Approximately 229 million cases of malaria were reported in 2019. Major incidences occur in various continents, including African and Eastern Mediterranean Continents and South-East Asia. INTRODUCTION: Despite the overall decline in global incidence from 2010 to 2018, the rate of decline has been almost constant since 2014. The morbidity and mortality have been accelerated due to reactive oxygen species (ROS) caused by oxidative stress generated by the parasite responsible for the destruction of host metabolism and cell nutrients. METHODS: The excessive release of free radicals is associated with the infection in the animal or human body by the parasites. This may be related to a reduction in nutrients required for the generation of antioxidants and the destruction of cells by parasite activity. Therefore, an intensive literature search has been carried out to find the natural antioxidants used to neutralize the free radicals generated during malarial infection. RESULTS: The natural antioxidants may be useful as an adjuvant treatment along with the antimalarial chemotherapeutics to reduce the death rate and enhance the success rate of malaria treatment. CONCLUSION: In this manuscript, an attempt has been made to provide significant insight into the antioxidant activities of herbal extracts against malaria parasites.

Structure-Based Discovery of Potent Staphylococcus aureus Thymidylate Kinase Inhibitors by Virtual Screening.

INTRODUCTION: Multidrug-resistant bacteria are rapidly increasing worldwide, increasing antibiotic resistance. The exploitation, misuse, overuse, and decrease of the therapeutic potential of currently available antibiotics have resulted in the development of resistance against bacteria. As the most common bacterial pathogen in humans, Staphylococcus aureus can cause many adverse health effects. In fighting multidrug-resistant Staphylococcus aureus, scientists have identified an extremely relevant target - SaTMPK. SaTMPK is essential for DNA synthesis, which, in turn, is necessary for the replication and cell division of bacteria. OBJECTIVE: To perform multi-stage screening using the ZINC database, followed by molecular docking, ADMET profiling, molecular dynamics simulations, and energy calculations. METHODS: Based on the similar pharmacophoric characteristics of existing SaTMPK crystal structures, a model of interaction-based pharmacophores was developed. We then performed molecular docking studies on the positive hits obtained from the pharmacophore screening. Compounds that exhibited good molecular interactions within the SaTMPK binding sites were further evaluated using in-silico ADMET profiling. RESULTS: In a multi-stage screening campaign, three compounds were shortlisted that exhibited physicochemical characteristics suitable for human administration. CONCLUSION: The findings from this study should contribute to in vitro and in vivo studies for clinical applications.

Structure-Based Virtual Screening to Identify Negative Allosteric Modulators of NMDA.

BACKGROUND: NMDA (N-methyl-D-aspartate) receptor is one of the ionotropic receptor subtypes of glutamate, the most abundant excitatory neurotransmitter in the human brain. Besides physiological roles in learning and memory, neuronal plasticity and somatosensory function NMDAR overstimulation are also implicated in a pathophysiological mechanism of 'excitotoxicity.' In this study, an allosteric site has been focused on to design inhibitors of the most abundant form of this receptor of utility in many acute (stroke, traumatic brain injury) and chronic neurodegenerative diseases such as Parkinson's disease, Huntington's, Alzheimer's, and others. METHODS: In order to target this specific site at the interdimer interface of the ligand-binding domain of GluN2A-containing NMDA-Rs, blood-brain barrier-permeable potentially therapeutic compounds, as opposed to only pharmacological tools currently available, were sought. Pharmacophorebased virtual screening, docking, computational ADME prediction techniques, and MD simulation studies were used. RESULTS: Proceeding through the in-silico methodology, the study was successful at reaching 5 compounds from ChEMBL Database, which were predicted to be potential NMDA inhibitor drugs. CONCLUSION: The products of the study are compounds that have been validated through pharmacophore and score-based screening and MD simulation techniques to be allosterically inhibiting NMDA receptors and with favorable pharmacokinetic profiles. They are likely to be therapeutic agents ready for in-vitro and in-vivo testing.

An updated patent review on drugs for the treatment of tuberculosis (2018-present).

INTRODUCTION: Tuberculosis (TB) caused by Mycobacterium tuberculosis (M.tb) has been a global challenge as 1.4 million deaths were reported in 2019, which included deaths attributed to HIV-TB co-infection. It is curable by the prescribed Directly Observed Treatment Short (DOTS) course, but the situation becomes critical and alarming due to multi-drug resistant (MDR) and extensively drug-resistant (XDR) tuberculosis. Hence there has been an urgent need to develop novel M.tb chemotherapeutics to overcome this situation. AREAS COVERED: This review provides an overview and update on recent developments on the novel therapeutics for the treatment of TB from the important published and granted patents (2018-present). EXPERT OPINION: The discovery of potent chemotherapeutics with reduced toxicity to combat M.tb particularly MDR and XDR-TB is a major challenge in antitubercular drug development. The missing of any doses during the DOTS treatment and poor immunity particularly in HIV patients has been a major cause for the development of drug resistance. Hence the major focus has to be on novel targets with their inhibitors and novel molecules both of natural and synthetic origins along with repurposed drugs for the complete eradication of tuberculosis.

Probing the mechanism of peptide binding to REV response element RNA of HIV-1; MD simulations and free energy calculations.

Ribonucleic acid (RNA) of HIV-1 contains a 350 nucleotide, highly structured, cis-acting element called RRE (REV-response-element RNA), essential for virus replication. REV is a natural peptide that binds to RRE and transports it from the nucleus to cytoplasm where it is expressed into a new virus. The synthetic peptide known as RSG-1.2 also binds the RRE element and competes with REV. The purpose of study is to rationally design novel peptides such as RSG peptide with improved binding affinity to prevent the transport of HIV-1 RNA and so replication of virus. Herein, we performed MD simulation and free energy calculations to evaluate the interactions and binding free energies of REV (PDB ID: 4PMI) and RSGs peptides (PDB IDs: 1G70 and 1I9F) with RRE. The protein-RNA interactions were analyzed using the MM-PBSA method. Results suggest that REV has more binding free energy -188.41 kcal/mol than two RSG peptides with total binding free energy -120.97 and -141.46 kcal/mol. The ARG and ASN were found to be important residues of REV. In the RRE sequence, the nucleotides 62-67 and 78-84 were found to be important contributors in binding free energy. This study play a major role in elaboration of binding REV and RSG1-2 with RRE element and pave the way for further synthesis of peptide that can bind with RRE element and can be selected as therapeutic agent for HIV.Communicated by Ramaswamy H. Sarma.

Mechanistic insights into the inhibitory activity of FDA approved ivermectin against SARS-CoV-2: old drug with new implications.

The novel corona virus (Covid-19) has become a great challenge worldwide since 2019, as no drug has been reported yet. Different clinical trials are still under way. Among them is Ivermectin (IVM), an FDA approved drug which was recently reported as a successful candidate to reduce SARS-CoV-2 viral load by inhibiting Importin-α1 (IMP-α1) protein which subsequently affects nuclear transport of viral proteins but its basic binding mode and inhibitory mechanism is unknown. Therefore, we aimed to explore the inhibitory mechanism and binding mode of IVM with IMP-α1 via different computational methods. Initially, comparative docking of IVM was performed against two different binding sites (Nuclear Localization Signal (NLS) major and minor sites) of IMP-α1 to predict the probable binding mode of IVM. Then, classical MD simulation was performed (IVM/NLS-Major site and IVM/NLS-Minor site), to predict its comparative stability dynamics and probable inhibitory mechanism. The stability dynamics and biophysical analysis of both sites highlighted the stable binding of IVM within NLS-Minor site by establishing and maintaining more hydrophobic contacts with crucial residues, required for IMP-α1 inhibition which were not observed in NLS-major site. Altogether, these results recommended the worth of IVM as a possible drug to limit the SARS-CoV-2 viral load and consequently reduces its progression.Communicated by Ramaswamy H. Sarma.

Novel Anti-Hepatitis B Virus Activity of Euphorbia schimperi and Its Quercetin and Kaempferol Derivatives.

Natural or plant products, because of their structural diversity, are a potential source for identifying new anti-hepatitis B virus (HBV) agents. Here, we report the anti-HBV activity of Euphorbia schimperi and its quercetin (QRC) and kaempferol derivatives. The anti-HBV-active methanol fraction of E. schimperi was subjected to chromatographic techniques, leading to isolation of three flavonols, following their structure determination by 1H and 13C NMR spectroscopies. Their cytotoxicity and anti-HBV potential were assessed using HBV reporter HepG2.2.15 cells, and their modes of action were delineated by molecular docking. The isolated compounds identified as quercetin-3-O-glucuronide (Q3G), quercetin-3-O-rhamnoside (Q3R), and kaempferol-3-O-glucuronide (K3G) were non-cytotoxic to HepG2.2.15 cells. The viral HBsAg/HBeAg production on day 5 was significantly inhibited by K3G (∼70.2/∼73.4%), Q3G (∼67.8/∼72.1%), and Q3R (∼63.2%/∼68.2%) as compared to QRC (∼70.3/∼74.8%) and lamivudine (∼76.5/∼84.5%) used as standards. The observed in vitro anti-HBV potential was strongly supported by in silico analysis, which suggested their structure-based activity via interfering with viral Pol/RT and core proteins. In conclusion, this is the first report on the anti-HBV activity of E. schimperi-derived quercitrin-3-O-glucuronide, quercitrin-3-O-rhamnoside, and kaempferol-3-O-glucuronide, most likely through interfering with HBV proteins.