An immunoinformatic approach for developing a multi-epitope subunit vaccine against Monkeypox virus.

An in-silico approach was implemented to develop a multi-epitope subunit vaccine construct against the recent outbreak of the Monkeypox virus. The contribution of 10 different antigenic proteins based on their antigenicity led to the selection of 10 HTL, 9 CTL, and 6 BCL epitopes. The construct was further investigated for its allergenicity, antigenicity, and physio-chemical properties using servers such as AllerTOP and Allergen FP, VaxiJen and ANTIGENPro, and ProtParam respectively. The secondary structure of the vaccine was predicted using the SOPMA server followed by I-TASSER for the 3D structure. After refinement and validation of structural stability of the modelled vaccine, a molecular docking assay was implemented to study the interaction of the known TLR4 receptor with that of the constructed vaccine using the ClusPro server. The docked vaccine and TLR4 receptor were studied using the molecular dynamics (MD) simulation to validate the stability of the complex. After codon optimization the cDNA was constructed and in-silico cloning of the vaccine construct was carried out. The vaccine was also subjected to computational immune assay which predicted a powerful immune response against the Monkeypox virus validating that the developed multi-epitope vaccine construct can be a potent vaccine candidate. Supplementary Information: The online version contains supplementary material available at 10.1007/s40203-024-00220-5.

Deciphering the molecular choreography of Janus kinase 2 inhibition via Gaussian accelerated molecular dynamics simulations: a dynamic odyssey.

The Janus kinases (JAK) are crucial targets in drug development for several diseases. However, accounting for the impact of possible structural rearrangements on the binding of different kinase inhibitors is complicated by the extensive conformational variability of their catalytic kinase domain (KD). The dynamic KD contains mainly four prominent mobile structural motifs: the phosphate-binding loop (P-loop), the αC-helix within the N-lobe, the Asp-Phe-Gly (DFG) motif, and the activation loop (A-loop) within the C-lobe. These distinct structural orientations imply a complex signal transmission path for regulating the A-loop's flexibility and conformational preference for optimal JAK function. Nevertheless, the precise dynamical features of the JAK induced by different types of inhibitors still remain elusive. We performed comparative, microsecond-long, Gaussian accelerated molecular dynamics simulations in triplicate of three phosphorylated JAK2 systems: the KD alone, type-I ATP-competitive inhibitor (CI) bound KD in the catalytically active DFG-in conformation, and the type-II inhibitor (AI) bound KD in the catalytically inactive DFG-out conformation. Our results indicate significant conformational variations observed in the A-loop and αC helix motions upon inhibitor binding. Our studies also reveal that the DFG-out inactive conformation is characterized by the closed A-loop rearrangement, open catalytic cleft of N and C-lobe, the outward movement of the αC helix, and open P-loop states. Moreover, the outward positioning of the αC helix impacts the hallmark salt bridge formation between Lys882 and Glu898 in an inactive conformation. Finally, we compared their ligand binding poses and free energy by the MM/PBSA approach. The free energy calculations suggested that the AI's binding affinity is higher than CI against JAK2 due to an increased favorable contribution from the total non-polar interactions and the involvement of the αC helix. Overall, our study provides the structural and energetic insights crucial for developing more promising type I/II JAK2 inhibitors for treating JAK-related diseases.

Microsecond dynamics of H10N7 influenza neuraminidase reveals the plasticity of loop regions and drug resistance due to the R292K mutation.

At the beginning of the last century, multiple pandemics caused by influenza (flu) viruses severely impacted public health. Despite the development of vaccinations and antiviral medications to prevent and control impending flu outbreaks, unforeseen novel strains and continuously evolving old strains continue to represent a serious threat to human life. Therefore, the recently identified H10N7, for which not much data is available for rational structure-based drug design, needs to be further explored. Here, we investigated the structural dynamics of neuraminidase N7 upon binding of inhibitors, and the drug resistance mechanisms against the oseltamivir (OTV) and laninamivir (LNV) antivirals due to the crucial R292K mutation on the N7 using the computational microscope, molecular dynamics (MD) simulations. In this study, each system underwent long 2 × 1 µs MD simulations to answer the conformational changes and drug resistance mechanisms. These long time-scale dynamics simulations and free energy landscapes demonstrated that the mutant systems showed a high degree of conformational variation compared to their wildtype (WT) counterparts, and the LNV-bound mutant exhibited an extended 150-loop conformation. Further, the molecular mechanics Poisson-Boltzmann surface area (MM/PBSA) calculation and MM/GBSA free energy decomposition were used to characterize the binding of OTV and LNV with WT, and R292K mutated N7, revealing the R292K mutation as drug-resistant, facilitated by a decline in binding interaction and a reduction in the dehydration penalty. Due to the broader binding pocket cavity of the smaller K292 mutant residue relative to the wildtype, the drug carboxylate to K292 hydrogen bonding was lost, and the area surrounding the K292 residue was more accessible to water molecules. This implies that drug resistance could be reduced by strengthening the hydrogen bond contacts between N7 inhibitors and altered N7, creating inhibitors that can form a hydrogen bond to the mutant K292, or preserving the closed cavity conformations.

Computational insights into VacA toxin inhibition: harnessing FDA-approved antibiotics against Helicobacter pylori.

Cancer is a condition in which a few of the body's cells grow beyond its control and spread to other outward regions. Globally, gastric cancer (GC) is third most common cause of cancer-related mortality and the fourth most common kind of cancer. Persistent infection of VacA-positive Helicobacter pylori (H. pylori) modulates cellular physiology and leads to GC. About ∼70% of H. pylori are positive for vacuolating cytotoxin-A (VacA), and it infects ∼80-90% of world populations. Herein, for first time, we repurposed FDA-approved gram-negative antibiotics, which are feasible alternatives to existing regimens and may be used in combinatorial treatment against VacA-positive H. pylori. Out of 110 FDA-approved antibiotics, we retrieved 92 structures, which were screened against the VacA protein. Moreover, we determined that the top eight hit antibiotics viz; cefpiramide, cefiderocol, eravacycline, doxycycline, ceftriaxone, enoxacin, tedizolid, and cefamandole show binding free energies of -9.1, -8.9, -8.1, -8.0, -7.9, -7.8, -7.8 and -7.8 Kcal/mol, respectively, with VacA protein. Finally, we performed 100 ns duplicate MD simulations on the top eight selected antibiotics showing strong VacA binding. Subsequently, five antibiotics, including cefiderocol, cefpiramide, doxycycline, enoxacin, and tedizolid show stable ligand protein distance and good binding affinity revealed by the MM-PBSA scheme. Among the five antibiotics cefiderocol act as the most potent inhibitor (-28.33 kcal/mol). Furthermore, we also identified the hotspot residue like Asn-506, Tyr-529, and Phe-483 which control the interaction. Concisely, we identified antibiotics that can be repurposed against VacA of H. pylori and explored their molecular mechanism of interaction with VacA.Communicated by Ramaswamy H. Sarma.

Molecular level investigation for identifying potential inhibitors against thymidylate kinase of monkeypox through in silico approaches.

The need for more advanced and effective monkeypox (Mpox) treatments has become evident with numerous Mpox virus (MPXV) outbreaks. Over the years, interest has increased in developing targeted medicines that are efficient, safe, and precise while avoiding adverse effects. Here, we screened 32409 compounds against thymidylate kinase (TMPK), an emerging target for Mpox treatment. We studied their pharmacological characteristics and analyzed those through all-atom molecular dynamics simulations followed by molecular mechanics Poisson Boltzmann surface area (MM-PBSA) based free energy calculations. According to our findings, the leads CID40777874 and CID28960001 had the highest binding affinities towards TMPK with ΔGbind of -8.04 and -5.58 kcal/mol, respectively, which outperformed our control drug cidofovir (ΔGbind = -2.92 kcal/mol) in terms of binding favourability. Additionally, we observed crucial TMPK dynamics brought on by ligand-binding and identified key residues such as Phe68 and Tyr101 as the critical points of the protein-ligand interaction. The DCCM analysis revealed the role of ligand binding in stabilizing TMPK's binding region, as indicated by residual correlation motions. Moreover, the PSN analysis revealed that the interaction with ligand induces changes in residual network properties, enhancing the stability of complexes. We successfully identified novel compounds that may serve as potential building blocks for constructing contemporary antivirals against MPXV and highlighted the molecular mechanisms underlying their binding with TMPK. Overall, our findings will play a significant role in advancing the development of new therapies against Mpox and facilitating a comprehensive understanding of their interaction patterns.Communicated by Ramaswamy H. Sarma.

Development of chalcone-like derivatives and their biological and mechanistic investigations as novel influenza nuclear export inhibitors.

Concerning the emergence of resistance to current anti-influenza drugs, our previous phenotypic-based screening study identified the compound A9 as a promising lead compound. This chalcone analog, containing a 2,6-dimethoxyphenyl moiety, exhibited significant inhibitory activity against oseltamivir-resistant strains (H1N1 pdm09), with an EC50 value of 1.34 µM. However, it also displayed notable cytotoxicity, with a CC50 value of 41.46 µM. Therefore, compound A9 was selected as a prototype structure for further structural optimization in this study. Initially, it was confirmed that the substituting the α,ß-unsaturated ketone with pent-1,4-diene-3-one as a linker group significantly reduced the cytotoxicity of the final compounds. Subsequently, the penta-1,4-dien-3-one group was utilized as a privileged fragment for further structural optimization. Following two subsequent rounds of optimizations, we identified compound IIB-2, which contains a 2,6-dimethoxyphenyl- and 1,4-pentadiene-3-one moieties. This compound exhibited inhibitory effects on oseltamivir-resistant strains comparable to its precursor (compound A9), while demonstrating reduced toxicity (CC50 > 100 µM). Furthermore, we investigated its mechanism of action against anti-influenza virus through immunofluorescence, Western blot, and surface plasmon resonance (SPR) experiments. The results revealed that compound IIB-2 can impede virus proliferation by blocking the export of influenza virus nucleoprotein. Thusly, our findings further emphasize influenza nuclear export as a viable target for designing novel chalcone-like derivatives with potential inhibitory properties that could be explored in future lead optimization studies.

Plant derived active compounds of ayurvedic neurological formulation, Saraswatharishta as a potential dual leucine zipper kinase inhibitor: an in-silico study.

Recent findings have highlighted the essential role of dual leucine zipper kinase (DLK) in neuronal degeneration. Saraswatharishta (SWRT), an ayurvedic formulation utilized in traditional Indian medicine, has demonstrated effectiveness in addressing neurodegenerative diseases. Herein, we aim to delve into the atomistic details of the mode of action of phytochemicals present in SWRT against DLK. Our screening process encompassed over 500 distinct phytochemicals derived from the main ingredients of the SWRT formulation. Through a comparative analysis of docking scores and relative poses, we successfully identified four novel compounds, which underwent further investigation via 2 × 500 ns long molecular dynamics (MD) simulations. Among the top four compounds, CID16066851 sourced from the Acorus calamus displayed the most stable complex with DLK. The molecular mechanics Poisson - Boltzmann surface area (MM-PBSA) calculations highlighted the significance of electrostatic and van der Waals interactions in the binding recognition process. Additionally, we identified key residues, namely Phe192, Leu243, Val139, and Leu141, as hotspots that predominantly govern the DLK-inhibitor interaction. Notably, the leading compounds are sourced from the Acorus calamus, Syzygium aromaticum, Zingiber officinale, and Anethum sowa plants present in the SWRT formulation. Overall, the findings of our study hold promise for future drug development endeavors combating neurodegenerative conditions.Communicated by Ramaswamy H. Sarma.

Prediction of Rab5B inhibitors through integrative in silico techniques.

Rab5B is a small monomeric G protein that regulates early endocytosis and controls signaling pathways related to cell growth, survival, and apoptosis. Dysregulation of Rab5B protein expression has been linked to the development of several cancers such as leukemia, lymphoma, kidney, prostate, ovarian, breast cancer, etc. Our research shows the first attempt to identify inhibitors that can target Rab5B GTPase. In this study, we performed molecular docking using Autodock Vina 1.5.6 and identified eight molecules with docking scores ranging from -9.8 to -10.6 kcal/mol. Thereafter, we examined the pharmacological characteristics of these compounds, and selected compounds were further analyzed for their conformational dynamics and thermodynamic stability using molecular dynamics simulations and molecular mechanics Poisson-Boltzmann surface area (MM-PBSA)-based free energy calculations. Notably, our findings revealed that strychnine had the highest binding affinity to Rab5B followed by anonaine, helioxanthin, and taiwanin E, with a ΔGbind value of -21.43, -17.11, -15.11, and -14.09 kcal/mol respectively. The binding free energy calculations showed that Van der Waals interactions are the primary contributor to the binding between Rab5B and the inhibitor. The interaction between the inhibitor and Rab5B was shown to be controlled by certain hot spot residues, including Phe45, Tyr48, Ala64, and Ala30. Overall, we believe that these findings could facilitate the exploration and development of potential hits against Rab5B, subject to optimization and further research. Rab5B inhibitory binding affinity of natural plants active compounds.

Advanced Magnetic Resonance Imaging in the Evaluation of a Chronic Non-healing ulcer - A Case Report of Rare Primary Cutaneous Leiomyosarcoma.

INTRODUCTION: Primary cutaneous leiomyosarcoma of the skin is extremely uncommon superficial soft tissue sarcoma. The uniqueness of this case is the exceedingly rarity of the tumor and its potential to be clinically misdiagnosed as squamous cell carcinoma. We report an interesting and rare case of PCL with unusual clinical presentation. CASE REPORT: Our case documents the radiological approach and importance of newer magnetic resonance imaging sequences in helping the diagnosis of a rare case of primary en plaque cutaneous leiomyosarcoma of lower extremity in a 60-year-old male, farmer by occupation with superficial ulceration and local invasion presenting as chronic non-healing ulcerative lesion. CONCLUSION: Contrast-enhanced MRI when coupled with advanced imaging sequence such as magnetic resonance spectroscopy and diffusion-weighted imaging further increases malignant probability of the lesion and guides the surgeon for a definite treatment plan.