In vitro antifungal activity analysis of Streptomyces sp. strain 196 against Candida albicans and Aspergillus flavus.

Numerous bioactive compounds have been reported to be produced by the members of the genus Streptomyces. During our previous studies, Streptomyces sp. strain 196 was tested for its antimicrobial activity, and bioactive compounds produced by this strain were characterized LC-MS and 1H NMR. To examine the antifungal potential of strain 196 is the goal of the current investigation. Present investigation is focused on exploring antifungal activity of extract of strain 196 (196EA) on membrane disruption potential against two fungi Candida albicans ATCC 90028 and Aspergillus flavus ITCC 5599. Results revealed that the MIC value is higher for A. flavus than for C. albicans which is 450 µg/mL and 250 µg/mL, respectively. Disc diffusion and spot assay also correspond to the values of the MIC for their respective pathogen. In growth curve analysis, lag and log phase are significantly affected by the extract of strain 196. The effects of extract from strain 196 on plasma membrane disruption of Candida albicans and Aspergillus flavus were analyzed in terms of ergosterol quantification assay, cellular leakage, proton efflux measurement (PM-ATPase), plasma membrane integrity assay (PI), and DNA damage assay (DAPI). Results shown that the extract of strain 196 has the potential to inhibit the cell membrane of the both pathogenic fungi which was further confirmed with the help of scanning electron microscopic (SEM) studies.

Multi-targeted effects of D-carvone against Non-Small Cell Lung Cancer (NSCLC): A network pharmacology-based study.

Non-small cell lung cancer (NSCLC) is a complex malignancy with a high degree of heterogeneity, representing approximately 85% of all lung cancer cases. The treatment landscape for NSCLC has been revolutionised by incorporating targeted and immunotherapies; however, novel therapeutic modalities are consistently needed to enhance the treatment outcomes. Indeed, alternative anti-cancer therapies involving natural products have drawn the attention of clinicians and scientists owing to their remarkable chemopreventive potential, often displaying minimal toxicity. D-carvone (CN) is one such natural product that has exhibited numerous promising therapeutic benefits, yet its efficacy against NSCLC remains enigmatic. In the present study, network pharmacological studies and molecular docking in conjunction with in-vitro validation were used to elucidate the underlying mechanism of action of CN comprehensively. Different databases revealed a total of 77 putative anti-NSCLC targets of CN. The identified core targets were utilised to construct a "Compound- Target- Disease" network by Cytoscape (v3.9.0). Further analysis identified 5 core/ hub targets of CN including JAK2, ERK1, ESR1, GSK3B and HSP90AA1. Molecular docking indicated a strong binding interaction of the compound with these core targets. Also, Gene Ontology and KEGG analysis validated the involvement of multiple biological processes. Additionally, CN significantly inhibited cell proliferation, clonogenicity, and wound healing potential while promoting apoptosis in a dose-dependent manner in H1299 and A549 cell lines as examined by flow cytometry, morphological assessment, and western blotting. In conclusion, this study delineates the therapeutic effects of CN on NSCLC, thus highlighting CN as a putative drug candidate for further analysis.

Natural products from Streptomyces spp. as potential inhibitors of the major factors (holoRdRp and nsp13) for SARS-CoV-2 replication: an in silico approach.

The COVID-19 pandemic caused unprecedented damage to humanity, and while vaccines have been developed, they are not fully effective against the SARS-CoV-2 virus. Limited targeted drugs, such as Remdesivir and Paxlovid, are available against the virus. Hence, there is an urgent need to explore and develop new drugs to combat COVID-19. This study focuses on exploring microbial natural products from soil-isolated bacteria Streptomyces sp. strain 196 and RI.24 as a potential source of new targeted drugs against SARS-CoV-2. Molecular docking studies were performed on holoRdRp and nsp13, two key factors responsible for virus replication factor. Our in silico studies, K-252-C aglycone indolocarbazole alkaloid (K252C) and daunorubicin were found to have better binding affinities than the respective control drugs, with K252C exhibiting binding energy of - 9.1 kcal/mol with holoRdRp and - 9.2 kcal/mol with nsp13, and daunorubicin showing binding energy at - 8.1 kcal/mol with holoRdRp and - 9.3 kcal/mol with nsp13. ADMET analysis, MD simulation, and MM/GBSA studies indicated that K252C and daunorubicin have the potential to be developed as targeted drugs against SARS-CoV-2. The study concludes that K252C and daunorubicin are potential lead compounds that might suppress the inhibition of SARS-CoV-2 replication among the tested microbial compounds and could be developed as targeted drugs against COVID-19. In the future, further in vitro studies are required to validate these findings.

In-silico and in-vitro evaluation of antifungal bioactive compounds from Streptomyces sp. strain 130 against Aspergillus flavus.

Streptomyces spp. are considered excellent reservoirs of natural bioactive compounds. The study evaluated the bioactive potential of secondary metabolites from Streptomyces sp. strain 130 through PKS-I and NRPS gene-clusters screening. GC-MS analysis was done for metabolic profiling of bioactive compounds from strain 130 in the next set of experiments. Identified antifungal compounds underwent ADMET analyses to screen their toxicity. All compounds' molecular docking was done with the structural gene products of the aflatoxin biosynthetic pathway of Aspergillus flavus. MD simulations were utilized to evaluate the stability of protein-ligand complexes under physiological conditions. Based on the in-silico studies, compound 2,4-di-tert butyl-phenol (DTBP) was selected for in-vitro studies against Aspergillus flavus. Simultaneously, bioactive compounds were extracted from strain 130 in two different solvents (ethyl-acetate and methanol) and used for similar assays. The MIC value of DTBP was found to be 314 µg/mL, whereas in ethyl-acetate extract and methanol-extract, it was 250 and 350 µg/mL, respectively. A mycelium growth assay was done to analyze the effect of compounds/extracts on the mycelium formation of Aspergillus flavus. In agar diffusion assay, zone of inhibitions in DTBP, ethyl-acetate extract, and methanol extract were observed with diameters of 11.3, 13.3, and 7.6 mm, respectively. In the growth curve assay, treated samples have delayed the growth of fungi, which signified that the compounds have a fungistatic nature. Spot assay has determined the fungal sensitivity to a sub-minimum inhibitory concentration of antifungal compounds. The study's results suggested that DTBP can be exploited for antifungal-drug development.Communicated by Ramaswamy H. Sarma.

Monoterpenes as potential antifungal molecules against Candida cell membranes: in-vitro and in-silico studies.

Azoles are the frequently used antifungal drugs that target the enzyme lanosterol 14 α-demethylase (erg11p). This enzyme plays a vital role in ergosterol biosynthesis and hence maintainenance of cell membrane fluidity and integrity. The emergence of resistance to azoles and their fungistatic nature against several fungal pathogens is the major challenge to combat invasive candidiasis. Therefore, there is an urgent need to discover new antifungals with better efficacy. This study targets erg11 protein using in silico approach and identifies the monoterpene compounds (α-terpineol, carveol, and terpinene-4-ol) based on docking score and ligand interaction analysis. Further dynamic behavior of best-docked compounds with erg11p was analyzed by various parameters of MD simulation. The binding free energy of selected compounds towards the definitive pocket was also calculated. To further investigate the antifungal activity of selected compounds, in vitro studies were conducted on C. albicans. Studies thus suggest that the proposed the mechanism of antifungal action of test compounds involves targeting the ergosterol biosynthetic pathway. The compounds were explored for their effect on the disruption of membrane integrity by studying ERG11gene expression analysis, scanning electron microscopy, PI uptake (fluorescence microscopy,) and H+-extrusion. The results suggest that the selected monoterpenes are safer natural antifungals that disrupt membrane integrity by inhibiting ergosterol biosynthesis and other membrane associated structures.Communicated by Ramaswamy H. Sarma.

In Vitro and In Vivo Investigation of a Dual-Targeted Nanoemulsion Gel for the Amelioration of Psoriasis.

Psoriasis, due to its unique pathological manifestations and the limited success of existing therapeutic modalities, demands dedicated domain research. Our group has developed nanotherapeutics consisting of bioactives such as Thymoquinone (TQ) and Fulvic acid (FA), which have been successfully incorporated into a Nanoemulsion gel (NEG), taking kalonji oil as oil phase. The composition is aimed at ameliorating psoriasis with better therapeutic outcomes. TQ is a natural bio-active that has been linked to anti-psoriatic actions. FA has anti-inflammatory actions due to its free radical and oxidant-scavenging activity. Our previous publication reports the formulation development of the NEG, where we overcame the pharmaco-technical limitations of combining the above two natural bioactives. In vitro evaluation of the optimized NEG was carried out, which showed an enhanced dissolution rate and skin permeation of TQ. This work furthers the pharmaceutical progression of dual-targeted synergistic NEG to treat psoriasis. A suitable animal model, BALB/c mice, has been used to conduct the in vivo studies, which revealed the effective anti-psoriatic action of TQ. Molecular docking studies corroborated the results and revealed a good binding affinity for both the targets of TNF-α (Tumor necrosis factor) and IL-6 (Interlukin-6). Tissue uptake by Confocal laser scanning microscopy (CLSM), a skin interaction study of the gel formulation, and an antioxidant free radical scavenging assay (1-1 Diphenyl-2-picrylhydrazyl DPPH) were also carried out. It was concluded that the NEG may be effective in treating psoriasis with minimal side effects.

Integrated network pharmacology and experimental analysis unveil multi-targeted effect of 18α- glycyrrhetinic acid against non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) is one of the most malignant types of cancer with soaring incidence rates worldwide, attributed to its heterogeneity and complex etiology. Evidently, alternative anti-cancer therapies comprising traditional medicines and natural products have gained attention for their ability to act as chemopreventive agents with minimal toxicities, either alone or in combination. Accumulating studies have substantiated the inevitability of network pharmacology studies for effectively mapping molecular targets of natural products against multifaceted diseases, including cancer. The 18α-Glycyrrhetinic acid (18α-GA), a triterpenoid found in licorice plants, has shown promising medicinal properties, although, its mechanism of action against NSCLC yet remains elusive. The present study was conducted to explore the anti- NSCLC potential of 18α-GA, employing integrative network pharmacology, molecular docking, and experimental research. Initially, network analysis revealed 181 common targets of 18α-GA in NSCLC as shown in the "compound-target- disease" network employing Cytoscape 3.8.2. Further analyses identified EGFR, AKT1, PI3KR1, MAPK1, IGF1, and SRC as the most crucial hub targets of 18α-GA against NSCLC. Moreover, molecular docking simulations and functional enrichment analyses indicated the involvement of multiple signaling pathways in suppressing NSCLC. Subsequent in-vitro studies verified the antiproliferative effect of 18α-GA on two NSCLC cancer cell lines, H1299 and A549. Mechanistically, 18α-GA arrested cell cycle at the G1 phase, induced apoptosis, decreased migratory potential, and protein expression levels of EGFR-PI3K/AKT, as examined by flow cytometry, morphological assessment, RT-PCR, and western blot. In conclusion, this study delineates the therapeutic potential and underlying mechanism(s) of 18α-GA as a putative novel drug against NSCLC. However, further studies are warranted to elucidate the complete molecular mechanism(s) using animal models of NSCLC.