PROTAC Beyond Cancer- Exploring the New Therapeutic Potential of Proteolysis Targeting Chimeras.

In the realm of oncology, the transformative impact of PROTAC (PROteolysis TAget-ing Chimeras) technology has been particularly pronounced since its introduction in the 21st cen-tury. Initially conceived for cancer treatment, PROTACs have evolved beyond their primary scope, attracting increasing interest in addressing a diverse array of medical conditions. This ex-panded focus includes not only oncological disorders but also viral infections, bacterial ailments, immune dysregulation, neurodegenerative conditions, and metabolic disorders. This comprehensive review explores the broadening landscape of PROTAC application, high-lighting ongoing developments and innovations aimed at deploying these molecules across a spectrum of diseases. Careful consideration of the design challenges associated with PROTACs reveals that, when appropriately addressed, these compounds present significant advantages over traditional therapeutic approaches, positioning them as promising alternatives. To evaluate the efficacy of PROTAC molecules, a diverse array of assays is employed, ranging from High-Throughput Imaging (HTI) assays to Cell Painting assays, CRBN engagement assays, Fluorescence Polarization assays, amplified luminescent proximity homogeneous assays, Time-resolved fluorescence energy transfer assays, and Isothermal Titration Calorimetry assays. These assessments collectively contribute to a nuanced understanding of PROTAC performance. Looking ahead, the trajectory of PROTAC technology suggests its potential recognition as a ver-satile therapeutic strategy for an expansive range of medical conditions. Ongoing progress in this field sets the stage for PROTACs to emerge as valuable tools in the multifaceted landscape of medical treatments.

SARS-CoV-2 inhibitory potential of fish oil-derived 2-pyrone compounds by acquiring linoleic acid binding site on the spike protein.

Traditional medicines have reportedly treated SARS-CoV-2 infection. Substantial evidence shows that fish oil supplements promote human immune function, suggesting they may lessen susceptibility to SARS-CoV-2 infection and suppress viral replication by inducing interferon. Fish oil was subjected to partition chromatography and separated into two compounds (EP01 and DH01). Isolated compounds were purified and characterized using UV, FTIR, NMR, and mass spectrometry to confirm their identity. Molecular docking was studied on the SARS CoV-2 variants of concern; SARS CoV-2 WT (PDB: 6VXX), SARS CoV-2 Alpha variant (PDB: 7LWS), SARS CoV-2 Delta variant (PDB: 7TOU), SARS CoV-2 Gamma variant (PDB: 7V78), SARS CoV-2 Kappa variant (PDB: 7VX9), and SARS CoV-2 Omicron variant (PDB: 7QO7) and TMPRSS2 (PDB: 7Y0E). Further selected protein-ligand complexes were subjected to 100 ns MD simulations to predict their biological potential in the SARS-CoV-2 treatment. In-vitro biological studies were carried out to support in-silico findings. Isolated compounds EP01 and DH01 were identified as 5-Tridecyltetrahydro-2H-pyran-2-one and 5-Heptadecyltetrahydro-2H-pyran-2-one, respectively. The compound EP01 significantly reduced (93.24 %) the viral RNA copy number with an IC50 of ~8.661 µM. EP01 proved to be a potent antiviral by in-vitro method against the SARS-CoV-2 clinical isolate, making it a promising antiviral candidate, with a single dose capable of preventing viral replication.

Quest for Anti-SARS-CoV-2 antiviral therapeutics: in-silico and in-vitro analysis of edible mushroom- Cordyceps militaris.

BACKGROUND: The emergence and evolution of SARS-CoV-2 resulted a severe threat to public health globally. Due to the lack of an effective vaccine with durable immunity, the disease transited into the endemic phase, necessitating potent antiviral therapy including a scientific basis for current traditional herbal medicine. OBJECTIVE: This study aimed to conduct a pharmacoinformatic analysis of selected chemical ingredients and in-vitro evaluation of Cordyceps militaris extract against SARS-CoV-2. MATERIALS AND METHODS: C. militaris, the widely used fungus in conventional herbal medicine, was subjected to computational investigation using molecular docking, molecular dynamic simulation and network pharmacology analysis followed by the in-vitro assay for evaluating its anti-SARS-CoV-2 potential. RESULTS: The molecular docking analysis of C. militaris revealed the Cordycepin's highest affinity (-9.71 kcal/mol) than other molecules, i.e., Cicadapeptin-I, Cicadapeptin-II, Cordycerebroside-B, and N-Acetyl galactosamine to the receptor binding domain of the SARS-CoV-2 spike protein. C. militaris aqueous extract could reduce the SARS-CoV-2 viral copy numbers by 50.24% using crude extract at 100 µg/mL concentration. CONCLUSION: These findings suggest that C. militaris has promising anti-SARS-CoV-2 activity and may be explored as traditional medicine for managing the COVID-19 surge in the endemic phase.

Identification of novel hit molecules targeting M. tuberculosis polyketide synthase 13 by combining generative AI and physics-based methods.

In this work we investigated the Pks13-TE domain, which plays a critical role in the viability of the mycobacteria. In this report, we have used a series of AI and Physics-based tools to identify Pks13-TE inhibitors. The Reinvent 4, pKCSM, KDeep, and SwissADME are AI-ML-based tools. AutoDock Vina, PLANTS, MDS, and MM-GBSA are physics-based methods. A combination of these methods yields powerful support in the drug discovery cycle. Known inhibitors of Pks13-TE were collected, curated, and used as input for the AI-based tools, and Mol2Mol molecular optimisation methods generated novel inhibitors. These ligands were filtered based on physics-based methods like molecular docking and molecular dynamics using multiple tools for consensus generation. Rigorous analysis was performed on the selected compounds to reduce the chemical space while retaining the most promising compounds. The molecule interactions, stability of the protein-ligand complexes and the comparable binding energies with the native ligand were essential factors for narrowing the ligands set. The filtered ligands from docking, MDS, and binding energy colocations were further tested for their ADMET properties since they are among the essential criteria for this series of molecules. It was found that ligands Mt1 to Mt6 have excellent predicted pharmacokinetic, pharmacodynamic and toxicity profiles and good synthesisability.

Identification of Mycobacterium tuberculosis transcriptional repressor EthR inhibitors: Shape-based search and machine learning studies.

Tuberculosis has been a challenge to the world since prehistoric times, and with the advent of drug-resistant strains, it has become more challenging to treat this infection. Ethionamide (ETH), a second-line drug, acts as a prodrug and targets mycolic acid synthesis by targeting the enoyl-acyl carrier protein reductase (InhA) enzyme. Mycobacterium tuberculosis (Mtb) EthR is an ethA gene repressor required to activate prodrug ETH. Recent studies suggest targeting the EthR could lead to newer drug molecules that would help better activate the ETH or complement this process. In this report, we have attempted and successfully identified three new molecules from the drug repurposing library that can target EthR protein and function as ETH boosters. These molecules were obtained after rigorous filtering of the database for their physicochemical, toxicological properties and safety. The molecular docking, molecular dynamics simulations and binding energy studies yielded three compounds, Ethyl (2-amino-4-((4-fluorobenzyl)amino)phenyl)carbamate) (L1), 2-((2,2-Difluorobenzo [d] [1,3]dioxol-5-yl)amino)-2-oxoethyl (E)-3-(5-bromofuran-2-yl)acrylate (L2), and N-(2,3-Dihydrobenzo [b] [1,4]dioxin-6-yl)-4-(2-((4-fluorophenyl)amino)-2-oxoethoxy)-3-methoxy benzamide (L3) are potential EthR inhibitors. We applied machine learning methods to evaluate these molecules for toxicity and synthesisability, suggesting safety and ease of synthesis for these molecules. These molecules are known for other pharmacological activities and can be repurposed faster as adjuvant therapy for tuberculosis.

Machine learning assisted methods for the identification of low toxicity inhibitors of Enoyl-Acyl Carrier Protein Reductase (InhA).

Tuberculosis (TB) is one of the life-threatening infectious diseases with prehistoric origins and occurs in almost all habitable parts of the world. TB mainly affects the lungs, and its etiological agent is Mycobacterium tuberculosis (Mtb). In 2022, more than 10 million people were infected worldwide, and 1.3 million were children. The current study considered the in-silico and machine learning (ML) approaches to explore the potential anti-TB molecules from the SelleckChem database against Enoyl-Acyl Carrier Protein Reductase (InhA). Initially, the entire database of ∼ 119000 molecules was sorted out through drug-likeness. Further, the molecular docking study was conducted to reduce the chemical space. The standard TB drug molecule's binding energy was considered a threshold, and molecules found with lower affinity were removed for further analyses. Finally, the molecules were checked for the pharmacokinetic and toxicity studies, and compounds found to have acceptable pharmacokinetic parameters and were non-toxic were considered as final promising molecules for InhA. The above approach further evaluated five molecules for ML-based toxicity and synthetic accessibility assessment. Not a single molecule was found toxic and each of them was revealed as easy to synthesise. The complex between InhA and proposed and standard molecules was considered for molecular dynamics simulation. Several statistical parameters showed the stability between InhA and the proposed molecule. The high binding affinity was also found for each of the molecules towards InhA using the MM-GBSA approach. Hence, the above approaches and findings exposed the potentiality of the proposed molecules against InhA.

Current biomarkers and treatment strategies in Alzheimer disease: An overview and future perspectives.

Alzheimer's disease (AD), a progressive degenerative disorder first identified by Alois Alzheimer in 1907, poses a significant public health challenge. Despite its prevalence and impact, there is currently no definitive ante mortem diagnosis for AD pathogenesis. By 2050, the United States may face a staggering 13.8 million AD patients. This review provides a concise summary of current AD biomarkers, available treatments, and potential future therapeutic approaches. The review begins by outlining existing drug targets and mechanisms in AD, along with a discussion of current treatment options. We explore various approaches targeting Amyloid ß (Aß), Tau Protein aggregation, Tau Kinases, Glycogen Synthase kinase-3ß, CDK-5 inhibitors, Heat Shock Proteins (HSP), oxidative stress, inflammation, metals, Apolipoprotein E (ApoE) modulators, and Notch signaling. Additionally, we examine the historical use of Estradiol (E2) as an AD therapy, as well as the outcomes of Randomized Controlled Trials (RCTs) that evaluated antioxidants (e.g., vitamin E) and omega-3 polyunsaturated fatty acids as alternative treatment options. Notably, positive effects of docosahexaenoic acid nutriment in older adults with cognitive impairment or AD are highlighted. Furthermore, this review offers insights into ongoing clinical trials and potential therapies, shedding light on the dynamic research landscape in AD treatment.

Nutritional Composition, Mineral Profiling, In Vitro Antioxidant, Antibacterial and Enzyme Inhibitory Properties of Selected Indian Guava Cultivars Leaf Extract.

Psidium guajava L. is a small evergreen tree known for its magnificent medicinal and nutritional value. This study aimed to evaluate the nutritional profile and in vitro pharmacological potentialities of the different leaf extracts of four cultivars of Psidium guajava namely Surka chitti, Allahabad safeda, Karela, and Lucknow-49. The standard procedures of the Association of Official Analytical Chemists (AOAC) were followed to carry out the nutritional analysis and all of the cultivars recorded the presence of elements at a nominal range. The highest presence of phenols (125.77 mg GAE/g) and flavonoids (92.38 mg QE/g) in the methanolic leaf extract of the Karela cultivar was recorded. A wide range of minerals such as sodium, phosphorus, magnesium, zinc, and boron were recorded with a higher percentage in the Karela cultivar of Psidium guajava. In the enzyme inhibitory assays, Allahabad safeda showed potential inhibition with an IC50 of 113.31 ± 1.07, 98.2 ± 0.66 and 95.73 ± 0.39 µg/mL in α-amylase, α-glucosidase, and tyrosinase inhibition assays, respectively. The strong antioxidant effect was established by Lucknow-49 (IC50 of 74.43 ± 1.86 µg/mL) and Allahabad safeda (IC50 of 78.93 ± 0.46 µg/mL) for ABTS and DPPH assays, respectively. The ethyl acetate and methanolic leaf extracts of the Allahabad safeda cultivar showed better inhibition against Pseudomonas aeruginosa with an MIC of 14.84 and 28.69 µg/mL, respectively. A decent mean zone of inhibition was recorded in methanolic leaf extract that ranged from 21-25 mm in diameter against the tested bacterial strains (Proteus vulgaris, Bacillus subtilis, and P. aeruginosa). This is the first scientific report on the comparative and comprehensive analysis of indigenous guava cultivars to evidently shortlist the elite cultivars with enriched dietary nutrition and biological activities.

Rutin from Begonia roxburghii modulates iNOS and Sep A activity in treatment of Shigella flexneri induced diarrhoea in rats: An in vitro, in vivo and computational analysis.

In developing countries, diarrhoea is a major issue of concern, where consistent use of antibiotics has resulted in several side effects along with development of resistance among pathogens against these antibiotics. Since natural products are becoming the treatment of choice, therefore present investigation involves mechanistic evaluation of antidiarrhoeal potential of Begonia roxburghii and its marker rutin against Shigella flexneri (SF) induced diarrhoea in rats following in vitro, in vivo and in silico protocols. The roots of the plant are used as vegetable in the North East India and are also used traditionally in treating diarrhoea. Phytochemically standardized ethanolic extract of B. roxburghii (EBR) roots and its marker rutin were first subjected to in vitro antibacterial evaluation against SF. Diarrhoea was induced in rats using suspension of SF and various diarrhoeagenic parameters were examined after first, third and fifth day of treatment at 100, 200 and 300 mg/kg, p.o. with EBR and 50 mg/kg, p.o. with rutin respectively. Additionally, density of SF in stools, stool water content, haematological and biochemical parameters, cytokine profiling, ion concentration, histopathology and Na+/K+-ATPase activity were also performed. Molecular docking and dynamics simulation studies of ligand rutin was studied against secreted extracellular protein A (Sep A, PDB: 5J44) from SF and Inducible nitric oxide synthase (iNOS, PDB: 1DD7) followed by network pharmacology. EBR and rutin demonstrated a potent antibacterial activity against SF and also showed significant recovery from diarrhoea (EBR: 81.29 ± 0.91% and rutin: 75.27 ± 0.89%) in rats after five days of treatment. EBR and rutin also showed significant decline in SF density in stools, decreased cytokine expression, potential antioxidant activity, cellular proliferative nature and recovered ion loss due to enhanced Na+/K+-ATPase activity, which was also supported by histopathology. Rutin showed a very high docking score of -11.61 and -9.98 kcal/mol against iNOS and Sep A respectively and their stable complex was also confirmed through dynamics, while network pharmacology suggested that, rutin is quite capable of modulating the pathways of iNOS and Sep A. Thus, we may presume that rutin played a key role in the observed antidiarrhoeal activity of B. roxburghii against SF induced diarrhoea.

A Review on Five and Six-Membered Heterocyclic Compounds Targeting the Penicillin-Binding Protein 2 (PBP2A) of Methicillin-Resistant Staphylococcus aureus (MRSA).

Staphylococcus aureus is a common human pathogen. Methicillin-resistant Staphylococcus aureus (MRSA) infections pose significant and challenging therapeutic difficulties. MRSA often acquires the non-native gene PBP2a, which results in reduced susceptibility to ß-lactam antibiotics, thus conferring resistance. PBP2a has a lower affinity for methicillin, allowing bacteria to maintain peptidoglycan biosynthesis, a core component of the bacterial cell wall. Consequently, even in the presence of methicillin or other antibiotics, bacteria can develop resistance. Due to genes responsible for resistance, S. aureus becomes MRSA. The fundamental premise of this resistance mechanism is well-understood. Given the therapeutic concerns posed by resistant microorganisms, there is a legitimate demand for novel antibiotics. This review primarily focuses on PBP2a scaffolds and the various screening approaches used to identify PBP2a inhibitors. The following classes of compounds and their biological activities are discussed: Penicillin, Cephalosporins, Pyrazole-Benzimidazole-based derivatives, Oxadiazole-containing derivatives, non-ß-lactam allosteric inhibitors, 4-(3H)-Quinazolinones, Pyrrolylated chalcone, Bis-2-Oxoazetidinyl macrocycles (ß-lactam antibiotics with 1,3-Bridges), Macrocycle-embedded ß-lactams as novel inhibitors, Pyridine-Coupled Pyrimidinones, novel Naphthalimide corbelled aminothiazoximes, non-covalent inhibitors, Investigational-ß-lactam antibiotics, Carbapenem, novel Benzoxazole derivatives, Pyrazolylpyridine analogues, and other miscellaneous classes of scaffolds for PBP2a. Additionally, we discuss the penicillin-binding protein, a crucial target in the MRSA cell wall. Various aspects of PBP2a, bacterial cell walls, peptidoglycans, different crystal structures of PBP2a, synthetic routes for PBP2a inhibitors, and future perspectives on MRSA inhibitors are also explored.

Process Optimization for the Bioinspired Synthesis of Gold Nanoparticles Using Cordyceps militaris, Its Characterization, and Assessment of Enhanced Therapeutic Efficacy.

The promising therapeutic implications of nanoparticles have spurred their development for biomedical applications. An eco-friendly methodology synthesizes gold nanoparticles using Cordyceps militaris, an edible mushroom (Cord-Au-NPs), using a quality-by-design approach (central composite design). UV-visible spectroscopy analysis revealed an absorption peak at 540-550 nm, thus confirming the synthesis of gold nanoparticles. Cord-Au-NPs have a crystalline structure, as evidenced by the diffraction peaks. The zeta potential value of -19.42 mV signifies the stability of Cord-Au-NPs. XRD study shows gold facets and EDX analysis revealed a strong peak of spherical nanoparticles in the gold region with a mean particle size of 7.18 nm and a polydispersity index of 0.096. The obtained peaks are closely associated with phenolic groups, lipids, and proteins, as examined by FTIR, suggesting that they function as the reducing agent. Cord-Au-NPs exhibited dose-dependent antioxidant, antidiabetic, and antibacterial activity. The method is eco-friendly, nontoxic, less time-consuming, and does not use synthetic materials, leading to higher capabilities in biomedical applications.

Isolation and Characterization of Anti-Inflammatory Compounds from Ficus microcarpa L.f. Stem Bark.

The anti-inflammatory effect of the ethyl acetate extract of F. microcarpa bark (EAFMB) was investigated in acute and chronic (21 days) inflammation induced in Wistar albino rats. EAFMB (200 mg/kg b.w.) exhibited comparable anti-inflammatory effects to the reference drug, with a reduction of 59.48% at 4 h in acute inflammation and 83.96% on day 21 in chronic inflammation. Bioassay-guided fractionation using DPPH radical scavenging activity led to isolating and identifying three compounds from EAFMB: oleanolic acid, catechin, and p-hydroxycinnamic acid. All these compounds demonstrated the concentration-dependent inhibition of COX enzymes and the protection of egg albumin from heat-induced denaturation. Catechin exhibited the highest COX inhibition (COX-1 and COX-2 IC50 = 9.02 and 50.38 µM, respectively) and anti-denaturation effect (IC50 = 27.13 µg/mL) compared to oleanolic acid and p-hydroxycinnamic acid. These isolated compounds are likely responsible for the anti-inflammatory activities of F. microcarpa bark.

Analysing the impact of eriosematin E from Eriosema chinense Vogel. against different diarrhoeagenic pathovars of Escherichia coli using in silico and in vitro approach.

Since diarrhoea is reportedly the third largest cause of fatality among kids, therefore it is considered to be one of the major areas of concerns among developing nations. The main causative agents of diarrhoea include Escherichia coli, Vibrio cholera, and Shigella spp where E. coli shares the maximum contribution. The roots of the plant Eriosema chinense Vogel. (Fabaceae) are traditionally used by the native tribes of Meghalaya, India to treat diarrhoea. From previous reports, the plant and its marker eriosematin E have been reported to have antidiarrhoeal potential against pathogenic and nonpathogenic diarrhoea. Therefore, the objective of the current investigation was to use in silico studies to determine the efficacy of eriosematin E against different diarrhoeagenic strains of E. coli. Six different pathovars of E. coli i.e. enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enterohaemorrhagic E. coli (EHEC), enteroaggregative E. coli (EAEC), uropathogenic E. coli (UPEC) and enteroinvasive E. coli (EIEC) were subjected to docking simulation studies utilizing Glide module of Schrodinger Maestro 2018-1 MM Share Version. Based on the obtained binding energy and balance between H-bonding, hydrophobic, and salt bridge interactions eriosematin E was found to be most effective against EPEC followed by EAEC and ETEC, while UPEC and EHEC were moderately affected. The molecular dynamics studies suggested a higher affinity of eriosematin E towards heat-labile enterotoxin b-pentamer from ETEC. The in vitro antibacterial studies against the universal strain S. aureus 12981 and E. coli 10418 revealed the effectiveness of eriosematin E showing MIC values of ≥256 µg/mL.Communicated by Ramaswamy H. Sarma.

Unlocking the potential of PROTACs: A comprehensive review of protein degradation strategies in disease therapy.

The technology known asPROTACs (PROteolysisTArgeting Chimeras) is a method of protein degradation. Utilising bifunctional small molecules, the ubiquitin-proteosome system (UPS) is used to induce the ubiquitination and degradation of target proteins. In addition to being novel chemical knockdown agents for biological studies that are catalytic, reversible, and rapid, PROTACs used in the treatment for disorders like cancer, immunological disorders, viral diseases, and neurological disorders. The protein degradation field has advanced quickly over the last two years, with a significant rise in research articles on the subject as well as a quick rise in smallmolecule degraders that are currently in or will soon enter the clinical stage. Other new degrading technologies, in addition to PROTAC and molecular glue technology, are also emerging rapidly. In this review article, we mainly focuses on various PROTAC molecules designed with special emphasis on targeted cellular pathways for different diseases i.e., cancer, Viral diseases Immune disorders, Neurodegenerative diseases, etc. We discussed about new technologies based on PROTACs such as Antibody PROTAC, Aptamers, Dual target, Folate caged, TF PROTAC, etc. Also, we listed out the PROTACs which are in clinical trials.

Fabrication of chitosan nanocomposites loaded with biosynthetic metallic nanoparticles and their therapeutic investigation.

The present research demonstrates the formation of zinc oxide nanoparticles facilitated by Cissus quadrangularis (CQ-ZnONPs) and subsequent synthesis of chitosan-conjugated nanocomposites (CQ-CS/ZnONCs) along with their biological assessment. The biosynthesized nanoparticles and nanocomposites were physicochemically characterized and therapeutically assessed for their antioxidant, antibacterial, and antidiabetic potential. The formation of CQ-ZnONPs and CQ-CS/ZnONCs was preliminarily validated by the change in color and subsequently by UV-visible spectroscopic analysis. The crystalline peaks associated with the CQ-ZnONPs in CQ-CS/ZnONCs were established by XRD analysis. Morphological evaluation of CQ-ZnONPs and CQ-CS/ZnONCs was carried out through FE-SEM and HRTEM studies. The particle size of the CQ-ZnONPs and CQ-CS/ZnONCs was 243.3 nm and 176.6 nm, with a PDI of 0.188 and 0.199, respectively. Nanoparticles and nanocomposites expressed Zeta potential of -15.7 mV and -16.2 mV, respectively. The CQ-ZnONPs and CQ-CS/ZnONCs showed good radical effectiveness with various in-vitro assays. The formulated nanoparticles and nanocomposites displayed significant antibacterial activity against the selected bacterial pathogens. CQ-CS/ZnONCs presented noteworthy α-amylase and α-glucosidase inhibitory effects compared to CQ-ZnONPs with IC50 of 73.66 ± 1.21 µg/mL and 87.59 ± 1.29 µg/mL, respectively. Moreover, the synthesized CQ-CS/ZnONCs demonstrated 98.92 ± 0.39% and 99.58 ± 0.16% wound contraction (at 7 and 14 mg, respectively), significantly (p < 0.05) higher than the standard and CQ-ZnONPs. Thus, the CQ-ZnONPs and CQ-CS/ZnONCs could effectively develop promising drug delivery systems to inhibit pathogens and chronic tissue repair.

TLK1-mediated RAD54 phosphorylation spatio-temporally regulates Homologous Recombination Repair.

Environmental agents like ionizing radiation (IR) and chemotherapeutic drugs can cause severe damage to the DNA, often in the form of double-strand breaks (DSBs). Remaining unrepaired, DSBs can lead to chromosomal rearrangements, and cell death. One major error-free pathway to repair DSBs is homologous recombination repair (HRR). Tousled-like kinase 1 (TLK1), a Ser/Thr kinase that regulates the DNA damage checkpoint, has been found to interact with RAD54, a central DNA translocase in HRR. To determine how TLK1 regulates RAD54, we inhibited or depleted TLK1 and tested how this impacts HRR in human cells using a ISce-I-GR-DsRed fused reporter endonuclease. Our results show that TLK1 phosphorylates RAD54 at three threonines (T41, T59 and T700), two of which are located within its N-terminal domain (NTD) and one is located within its C-terminal domain (CTD). Phosphorylation at both T41 and T59 supports HRR and protects cells from DNA DSB damage. In contrast, phosphorylation of T700 leads to impaired HRR and engenders no protection to cells from cytotoxicity and rather results in repair delay. Further, our work enlightens the effect of RAD54-T700 (RAD54-CTD) phosphorylation by TLK1 in mammalian system and reveals a new site of interaction with RAD51.

Targeting dormant phenotype acquired mycobacteria using natural products by exploring its important targets: In vitro and in silico studies.

The dormant phenotype of Mycobacterium tuberculosis that develops during infection poses a major challenge in disease treatment, since these bacilli show tolerance to front-line drugs. An in vitro hypoxia dormancy model was established, which produced phenotypically dormant Mycobacterium smegmatis after prolonged incubation under conditions of low oxygen, low pH, and nutrient limitation. Bacilli in this model displayed the classical dormancy characters, including loss of acid fastness, altered morphology, and, most importantly, tolerance to front-line drugs. The dormant form of M. smegmatis was treated with drugs and phytomolecules. Three phytomolecules exhibited activity against dormant bacilli, as shown by lack of regrowth in solid and liquid media. Further investigation of dormancy-active hits was carried out using in silico approaches to understand the druggable targets of these phytomolecules in dormant bacilli. For this study, molecular docking, molecular dynamics simulations (MDS), and molecular mechanics-generalized born solvent accessibility (MM-GBSA)-based binding energy (ΔGbind) calculations were performed. Five different targets, namely, isocitrate lyase (ICL), GMP synthase, LuxR, DosR, and serine/threonine protein kinase (STPK), from M. smegmatis and M. tuberculosis were studied in details. DosR and STPK were found to be the common targets in both the species that were more prone to the phytomolecules. The standard DosR inhibitor, HC104A, showed a lower dock score and binding energy of -4.27 and -34.50 kcal/mol, respectively, compared to the natural products under study. The phytomolecule, icariin, showed better docking score (dock score = -5.92 kcal/mol with and binding energy ΔGbind= -52.96 kcal/mol) with DosR compared to known DosR inhibitor, HC104A (dock score = -4.27 kcal/mol and binding energy ΔGbind = -34.50 kcal/mol). Similarly, the known STPK inhibitor MRCT67127 showed a lower dock score and binding energy of -4.25 and -29.43 kcal/mol, respectively, compared to the phytomolecule, icariin (dock score = -5.74 kcal/mol and ΔGbind= -43.41 kcal/mol). These compounds might ultimately lead to new therapeutics or may be useful as adjuvants to the first-line drugs to reduce the lengthy anti-TB therapy in the future.

Ethnological validation of Ashwagandha (Withania somnifera L. Dunal) ghrita as 'Vajikarana Rasayana': In-silico, in-vitro and in-vivoapproach.

ETHNOPHARMACOLOGICAL RELEVANCE: Processing cow ghee (clarified butterfat) with therapeutic herbs, i.e. ghrita, is recognized for augmenting the therapeutic efficacy of plant materials. Ashwagandha ghrita (AG) is an effective Ayurvedic formulation consisting of Indian ginseng, i.e., Withania somnifera (L.) Dunal, the main constituent used to treat infertility, weakness, gynaecological disorders, and general debility. OBJECTIVES: The present investigation was undertaken to corroborate the ethnopharmacological claim of AG as 'Vajikarana Rasayana' for its aphrodisiac potential using bioinformatics (in-silico) and experimental (in-vitro and in-vivo) approaches. METHODS: AG was formulated as per the methods reported in Ayurved sarsangraha. AG was further subjected to HPLC, GCMS analysis, and biological (acute toxicity and aphrodisiac) assessment per the standard procedures. Thirty-eight bioactives of Indian ginseng were subjected to computational studies (molecular docking and network pharmacology) to confirm the plausible mechanism. RESULTS: AG was found to be safe up to 2000 mg/kg body wt., and it showed dose-dependent upsurge (p < 0.01 and p < 0.05, wherever necessary) in mount and intromission frequency, genital grooming, and anogenital sniffing at 150 and 300 mg/kg body weight suggesting aphrodisiac activity. In-vitro studies demonstrated significant relaxation of the Corpus Cavernosal Smooth Muscle at all concentrations in a dose-dependent manner. Furthermore, the results of molecular modelling studies were in agreement with the biological activity and showed interaction with phosphodiesterase-5 as a possible target. CONCLUSION: AG exhibited an aphrodisiac effect and substantiated the traditional claim of Indian ginseng-based ghrita formulation as 'Vajikarana Rasayana'.