Discerning computational, in vitro and in vivo investigations of self-assembling empagliflozin polymeric micelles in type-2 diabetes.

BACKGROUND: Empagliflozin (EMPA) is an SGLT2 inhibitor, a new class of anti-diabetic medication, indicated for treating type-2 diabetes. Its low permeability, poor solubility and bioavailability limits its use in management of diabetes. The study was aimed to formulate EMPA loaded polymeric micelles (PMs) to overcome these obstacles in oral absorption. METHODOLOGY: In silico studies-molecular docking, molecular dynamic simulation (MDS), and quantum chemical calculation were employed to study the interaction of EMPA with different polymers. EMPA loaded TPGS polymeric micelles (EMPA-TPGS-PMs) were formulated by direct dissolution method and characterized in terms of surface morphology, entrapment, particle size, in vitro drug release, and in vitro cytotoxicity (HEK293 cells). In vivo pharmacokinetic and pharmacodynamic studies were also performed. RESULTS: The results suggested a good interaction between TPGS and EMPA with lowest binding energy compared to other polymers. Further MDS results and DFT calculations validated the stable binding of the complex hence TPGS was selected for further wet lab experiments. The EMPA-TPGS complex displayed lower value of Total energy (T.E.) than its individual components, indicating the overall stability of the complex while, the energy band gap (∆E) value lied between the two individual molecules, signifying the better electron transfer between HOMO and LUMO of the complex. Based on the solubility, entrapment and cytotoxicity studies, 5% TPGS was selected for formulating drug loaded micelles. EMPA-TPGS5-PMs presented a size of 9.008 ± 1.25 nm, Polydispersity index (PDI) of 0.254 ± 0.100, a controlled release behaviour upto 24 h. SEM and AFM images of the nanoformulation suggested spherical particles whereas, DSC, and PXRD studies confirmed the loss of crystallinity of EMPA. A 3.12-folds higher AUC and a greater reduction in blood glucose levels was exhibited by EMPA-TPGS5-PMs in comparison to EMPA-SUSP in mice model. CONCLUSION: EMPA-TPGS-PMs has exhibited better bio absorption and therapeutic effectiveness in diabetes treatment. This improved performance would open the possibility of dose reduction, reduced dosing frequency & dose-related side effects, improving pharmaco-economics and thereby improved overall compliance to the patient. However, this translation from bench to bedside would necessitate studies in higher animals and human volunteers.

Molecular modelling studies and identification of novel phytochemical inhibitor of DLL3.

Prostate cancer has been recently considered the most diagnosed cancer in male. DLL3 is overexpressed in CRPC-NE but not in localised prostate cancer or BPH. There are no effective treatments for neuroendocrine differentiated prostate cancer due to a lack of understanding of DLL3 structure and function. The structure of DLL3 is not yet determined using any experimental techniques. Hence, the structure-based drug discovery approach against prostate cancer has not shown great success. In present study, molecular modelling techniques were employed to generate three-dimensional structure of DLL3 and performed its thorough structural analysis. Further, all-atom molecular dynamics simulation was performed to obtain energetically favourable conformation. Further, we used a virtual screening using a library of >13800 phytochemicals from the IMPPAT database and other literature to select the best possible phytochemical inhibitor for DLL3 and identified the top five compounds. Relative binding affinity was calculated using the MM-PBSA approach. ADMET properties of the screened compounds reveal the toxic effect of Gnemonol C. We believe these studied physicochemical properties, functional domain identification, and binding site identification would be very useful to gain more structural and functional insights of DLL3; also, it can be used to infer their pharmacodynamics properties of DLL3 which was recently reported as an important prostate cancer target. The current study also proposes that Ergosterol Peroxide, Dioslupecin A, Mulberrofuran K, and Caracurine V have strong affinities and could serve as plausible inhibitors against DLL3. We believe this study would further help develop better drug candidates against neuroendocrine prostate cancer.Communicated by Ramaswamy H. Sarma.

Friedelin, a novel inhibitor of CYP17A1 in prostate cancer from Cassia tora.

In prostate cancer (PC), drugs targeting CYP17A1 have shown great success in regulating PC progression. However, successful drug molecules show adverse side effects and therapeutic resistance in PC. Therefore, we proposed to discover the potent phytochemical-based inhibitor against CYP17A1 using virtual screening. In this study, a phytochemicals library of ∼13800 molecules was selected to screen the best possible inhibitors against CYP17A1. A molecular modelling approach investigated detailed intermolecular interactions, their structural stability, and binding affinity. Further, in vitro and in vivo studies were performed to confirm the anticancer activity of identified potential inhibitor against CYP17A1. Friedelin from Cassia tora (CT) is identified as the best possible inhibitor from the screened library. MD simulation study reveals stable binding of Friedelin to conserved binding pocket of CYP17A1 with higher binding affinity than studied control, that is, Orteronel. Friedelin was tested on hormone-sensitive (22Rv1) and insensitive (DU145) cell lines and the IC50 value was found to be 72.025 and 81.766 µg/ml, respectively. CT extract showed a 25.28% IC50 value against 22Rv1, ∼92.6% increase in late Apoptosis/Necrosis, and three folds decrease in early apoptosis in treated cells compared to untreated cells. Further, animal studies show a marked decrease in prostate weight by 39.6% and prostate index by 36.5%, along with a reduction in serum PSA level by 71.7% and testosterone level by 92.4% compared to the testosterone group, which was further validated with histopathological studies. Thus, we propose Friedelin and CT extract as potential leads, which could be taken further for drug development in PC.[Figure: see text]Communicated by Ramaswamy H. Sarma.

Isolation and characterization of Lactococcus garvieae from the fish gut for in vitro fermentation with carbohydrates from agro-industrial waste.

This study focused on agro-industrial waste such as fruit peels by extracting prebiotics as a carbon source for lactic acid bacteria (LAB). Four strains of LAB were selected from Oreochromis niloticus (B2 and B3) and Nemipterus japonicas (R4 and R5), and identified as Lactococcus garvieae through 16S rRNA gene sequencing. The analysis of probiotic characteristics revealed that all four strains were able to tolerate sodium chloride (up to 7 %), bile salt (up to 3 %), and broad range of pH (2-9). Further, analysis of polysaccharide contents in the agro-industrial waste materials such as peels of pineapple, orange, lemon, sugarcane, pomegranate, and sweet lemon revealed that the concentration ranged from 3.91-163.85 mg/g. It was observed that orange peels (20.38-140.99 mg/g), sweet lemon peels (22.03-161.93 mg/g), and pomegranate peels (38.19-163.85 mg/g) yielded maximum indigestible polysaccharide. Evaluation of synbiotic combination of probiotic and prebiotic revealed that L. garvieae strains had better fermentation efficiency with orange, sweet lemon, and pineapple compared to lemon, sugarcane, and pomegranate. In nutshell, different types of agro-industrial waste evaluated in this research were found to be a cheap and fermentable carbon sources for LAB. Further study should be conducted to analyze this symbiotic combination as feed supplements for fish in aquaculture as well as various fermentation industries.

Molecular identification and biocontrol activity of sugarcane rhizosphere bacteria against red rot pathogen Colletotrichum falcatum.

A total of 226 sugarcane rhizosphere-associated bacterial strains from the six different cultivars were screened against three pathogenic strains of C. falcatum (cfNAV, cfCHA, and cf8436) for the suppression of red rot disease. On the basis of mycelial growth inhibition in dual culture assay, 26 bacteria were selected for further characterization of morphology, biochemical activity, plant-growth-promoting (PGP) activity, antifungal potential and molecular identity by 16S rRNA gene sequence. On the basis of the 16S rRNA gene sequencing, it was found that the isolates belonged to proteobacteria (13), Firmicutes (10), and Bacteroides (3). The antagonistic bacteria tested for PGP traits revealed that 10 strains were able to solubilize tricalcium phosphate, 11 strains were able to produce siderophore, and 14 strains were able to grow in the N-free medium. The quantitative estimation of indole-3-acetic acid production was ranged from 21.58 to 66.31 µg/mL. On the basis of PGP and biocontrol traits, five strains Ochrobactrum intermedium (TRD14), Acinetobacter sp. (PK9), Bacillus sp. (RSC29 and KR91) and Escherichia sp. (VRE34) were further chosen for pot trial under greenhouse conditions on highly susceptible variety CoC671. The results showed that the pathogen-inoculated sugarcane plants were able to germinate but died within one month. However, the CoC671 inoculated with selected biocontrol strains found protected from disease and an increase in plant growth parameters on par with carbendazim fungicides. This study proves that the isolates identified in this study could be used as an alternative to chemical fungicides to control red rot pathogen of sugarcane plants.