Intratympanic injections of emulsion-like dispersions to augment cinnarizine amount in a healthy rabbit inner ear model.

Aim: To investigate eutectic liquid-based emulsion-like dispersions for intratympanic injections to augment cinnarizine permeability across round window membrane in a healthy rabbit inner ear model. Methods: Two-tier systematic optimization was used to get the injection formula. The drug concentrations in perilymph and plasma were analyzed via. Ultra-performance liquid chromatography-tandem mass spectrometry method after 30-, 60-, 90- and 120-min post intratympanic injection time points in rabbits. Results: A shear-thinning behavior, immediate drug release (∼98.80%, 10 min) and higher cell viability (>97.86%, 24 h) were observed in dispersions. The cinnarizine level of 8168.57 ± 1236.79 ng/ml was observed in perilymph at 30 min post intratympanic injection in rabbits. Conclusion: The emulsion-like dispersions can augment drug permeability through round window membrane.

[Box: see text].

Ambient particulate matter (PM2.5) exposure contributes to neurodegeneration through the microbiome-gut-brain axis: Therapeutic role of melatonin.

Exposure to ambient particulate matter (PM2.5) has been shown to disturb the gut microbiome homeostasis and cause initiation of neuroinflammation and neurodegeneration via gut-brain bi-directional axis. Polyaromatic hydrocarbons (PAHs), which are carcinogenic and mutagenic, are important organic constituents of PM2.5 that could be involved in the microbiome-gut-brain axis-mediated neurodegeneration. Melatonin (ML) has been shown to modulate the microbiome and curb inflammation in the gut and brain. However, no studies have been reported for its effect on PM2.5-induced neuroinflammation. In the current study, it was observed that treatment with ML at 100 µM significantly inhibits microglial activation (HMC-3 cells) and colonic inflammation (CCD-841 cells) by the conditioned media from PM2.5 exposed BEAS2B cells. Further, melatonin treatment at a dose of 50 mg/kg to C57BL/6 mice exposed to PM2.5 (at a dose of 60 µg/animal) for 90 days significantly alleviated the neuroinflammation and neurodegeneration caused by PAHs in PM2.5 by modulating olfactory-brain and microbiome-gut-brain axis.

Bio-nanoconjugates of lithocholic acid/IR 780 for ROS-mediated apoptosis and optoacoustic imaging applications in breast cancer.

A new lithocholic acid/IR 780 conjugate (LIC) was designed and synthesized for theranostic applications in triple-negative breast cancer. Lithocholic acid is an antitumor biomacromolecule and acts via multiple molecular targets. IR 780 iodide is a fluorescent NIR organic dye researched as a photothermal agent in cancer therapy. A combined conjugate, LIC can have wide applications as a Photothermal/chemotherapeutic and imaging agent in cancer therapy. LIC was characterized and evaluated for its photothermal cytotoxic effect in breast cancer cell lines. Further, to improve the bioavailability of the LIC, a polymeric (PLGA) nanosystem was developed and characterized. The resultant lithocholic acid/IR 780 polymeric nanoconjugates (LIPNCs) were well taken up by the cells and are evident by the inherent red fluorescence of LIC. The LIPNCs also exhibited commendable heat generation when exposed to NIR light (808 nm). The in-vitro anti-cancer studies of LIPNCs also revealed a significant NIR light-based photothermal efficacy (cytotoxic dose 0.75 µM) when compared to the free conjugate (LIC) or the parent moieties. Further cell-based fluorescent and molecular assays showed that LIPNCs induced ROS-mediated apoptotic cell death concurrently being physiologically biocompatible. In-vitro photoacoustic imaging of the LICs exhibited signals comparable to free IR780 dye. Future in vivo studies with LIPNCs or LIC may prove beneficial for developing a promising translational system for its wide application in image-guided cancer theranostics.

Analytical Quality-by-Design-Based Systematic Optimization of RP-HPLC Method Conditions to Analyze Simultaneously Chloroquine Phosphate and Flavopiridol in Stress-Induced Combined Drug Solution and Pharmaceutical Emulsions.

A design of experiments (DoE)-driven RP-HPLC method conditions was employed to analyze simultaneously chloroquine (CQ) phosphate and flavopiridol (FLAP) in emulsions and solution. After subjecting the various critical method attributes to preliminary risk assessment and screening by Pareto-chart-based fractional factorial design, the 17 runs were produced in Box-Behnken design for optimization. Analysis of variance, lack of fit, prediction equations, 3D response surface plots and contour plots were used to evaluate the critical analytical attributes such as retention time, tailing factor and theoretical plate count. The optimized RP-HPLC method conditions include 262 nm as detection wavelength, 37°C temperature for column, 20-µl injection volume, 1-ml/min flow rate and mobile phase mixture [70:30 ratio of 0.4% triethylamine in methanol&sodium phosphate buffer (11 mM, pH 3.0)]. The studied validation parameters were found within the ICH-prescribed limits. Exposing the combined drug solution at oxidative stress condition resulted to diminish the FLAP recovery value (53.39 ± 0.86) and arrival of an extra chromatographic peak. However, the % drug entrapment efficiency values of 96.22 ± 2.47 and 85.86 ± 3.66, respectively, were noticed for CQ phosphate and FLAP in emulsions. Thus, DoE-driven approach could be helpful for systematically optimizing RP-HPLC method conditions.

Phyto-metabolomics of phlogacanthus thyrsiformis by using LC-ESI-QTOF-MS/MS and GC/QTOF-MS: Evaluation of antioxidant and enzyme inhibition potential of extracts.

Phlogacanthus thyrsiformis (P. thyrsiformis) is a non-conventional edible plant that has been used as a vegetable and as a traditional medicine to treat various diseases. This non-conventional edible plant is widespread in India, Yunnan-Chinese provinces, and Vietnam. Despite this potential claim, the phytochemical investigation of plants remains incomplete. In the present study, the chemical profile analysis of P. thyrsiformis leaves extracts was performed using LC-ESI-QTOF-MS/MS and GC/QTOF-MS and assessed for its antioxidants and enzyme inhibitory potential. A cursory examination of the phytometabolites of different solvents extracts revealed chloroform extract has different metabolites, including phenols, terpenes, glycosides and alkaloids. 113 and 138 metabolites, including primary and secondary metabolites, were identified using LC-ESI-QTOF-MS/MS and GC/QTOF-MS, respectively. Total phenolic content was observed highest in chloroform extract (60.59 ± 1.25 mg GAE/g). Maximum antioxidant activity was found to be in chloroform extract, which was identified by DPPH (0.88 ± 0.29 mg/mL), ABTS (0.54 ± 0.1 mg/mL) and FRAP assay (26.46 ± 1.65 mg AA/g extract). Ethyl acetate extract have the highest α-amylase (0.09 ± 0.11 mg/mL) and α-glucosidase (0.088 ± 0.13 mg/mL) enzyme inhibition potential. For all the detected molecules, docking and molecular dynamics studies were performed. To the best of our knowledge, this is the first report that discusses LC-ESI-QTOF-MS/MS and GC/QTOF-MS-based metabolites profiling of P. thyrsiformis that can help to treat various diseases.

Extraction of small molecule from human plasma by prototyping 3D printed sorbent through extruded filament for LC-MS/MS analysis.

Analytical sample preparation techniques are regarded as crucial steps for analyzing compounds from different biological matrices. The development of new extraction techniques is a modern trend in the bioanalytical sciences. 3D printed techniques have emerged as a valuable technology for prototyping devices in customized shapes for a cost-effective way to advance analytical sample preparation techniques. The present study aims to fabricate customized filaments through the hot-melt extrusion (HME) technique followed by fused deposition modeling mediated 3D printing process for rapid prototyping of 3D printed sorbents to extract a sample from human plasma. Thus, we fabricated our own indigenous filament using poly (vinyl alcohol), Eudragit® RSPO, and tri-ethyl citrate through HME to prototype the fabricated filament into a 3D printed sorbent for the extraction of small molecules. The 3D sorbent was applied to extract hydrocortisone from human plasma and analyzed using a validated LC-MS/MS method. The extraction procedure was optimized, and the parameters influencing the sorbent extraction were systematically investigated. The extraction recovery of hydrocortisone was found to be >82% at low, medium, and high quality control samples, with a relative standard deviation of <2%. The intra-and inter-day precisions for hydrocortisone ranged from 1.0% to 12% and 2.0%-10.0%, respectively, whereas the intra-and inter-day accuracy for hydrocortisone ranged from 93.0% to 111.0% and 92.0% to 110.0%, respectively. The newly customizable size and shape of the 3D printed sorbent opens new possibilities for extracting small molecules from human plasma.

Transporter-mediated drug-drug interactions: advancement in models, analytical tools, and regulatory perspective.

Drug-drug interactions mediated by transporters are a serious clinical concern hence a tremendous amount of work has been done on the characterization of the transporter-mediated proteins in humans and animals. The underlying mechanism for the transporter-mediated drug-drug interaction is the induction or inhibition of the transporter which is involved in the cellular uptake and efflux of drugs. Transporter of the brain, liver, kidney, and intestine are major determinants that alter the absorption, distribution, metabolism, excretion profile of drugs, and considerably influence the pharmacokinetic profile of drugs. As a consequence, transporter proteins may affect the therapeutic activity and safety of drugs. However, mounting evidence suggests that many drugs change the activity and/or expression of the transporter protein. Accordingly, evaluation of drug interaction during the drug development process is an integral part of risk assessment and regulatory requirements. Therefore, this review will highlight the clinical significance of the transporter, their role in disease, possible cause underlying the drug-drug interactions using analytical tools, and update on the regulatory requirement. The recent in-silico approaches which emphasize the advancement in the discovery of drug-drug interactions are also highlighted in this review. Besides, we discuss several endogenous biomarkers that have shown to act as substrates for many transporters, which could be potent determinants to find the drug-drug interactions mediated by transporters. Transporter-mediated drug-drug interactions are taken into consideration in the drug approval process therefore we also provided the extrapolated decision trees from in-vitro to in-vivo, which may trigger the follow-up to clinical studies.

Geometry encoded functional programming of tumor homing peptides for targeted drug delivery.

Poly-peptide molecules have shown promising applications in drug delivery and tumor targeting. A series of tumor homing peptides were designed by exhaustively sampling low energy geometrical basins of amino acids at specific sites of a peptide molecule to induce a conformational lock. This peptide library was pruned to a limited set of eight molecules, employing electrostatic interactions, docking, and molecular dynamics simulations. These designed and optimized peptides were synthesized and tested on various cell lines, including breast cancer (MDA-MB-231), cervical cancer (HeLa), osteosarcoma (U2-OS), and non-cancerous mammary epithelial cells (MCF-10A) using confocal microscopy and flow cytometry. Peptides show differential uptake in cancerous MDA-MB-231, HeLa, U2-OS, and non-cancerous MCF-10A cells. Confocal imaging verified their ability to penetrate even in 3D tumorospheres of MDA-MB-231 cells. Further, experiments of mitochondrial membrane potential depolarization and Caspase-3 activation confirmed that their cytotoxic effects are by apoptosis. Homing ability of the designed peptides in in vivo system and fluorescence imaging with clinical samples of human origin have further confirmed that the in vitro studies are qualitatively identical and quantitatively comparable in their ability to selectively recognize tumor cells. Overall, we present a roadmap for the functional programming of peptide-based homing and penetrating molecules that can perform selective tumor targeting.

Synthesis, Characterizations, and Use of O-Stearoyl Mannose Ligand-Engineered Lipid Nanoarchitectonics for Alveolar Macrophage Targeting.

The main challenging aspect in the management of tuberculosis (TB) diseases is effective alveolar macrophages targeting. Macrophage mannose receptor plays a predominant role in stimulating immune systems by TB pathogen. Our earlier in silico computational studies revealed that O-stearoyl mannose (OSM) possesses a higher affinity with macrophage mannose receptors. Therefore, keeping this in view, we developed OSM with the association of stearic acid and d-mannose as initial reactants by the esterification process. Preliminary confirmation of reaction was assessed with thin-layer chromatography experimentation, whereas further confirmation followed by in vitro characterization with several analytical experimental tools such as fourier transform near-infrared, differential scanning calorimetry, and electrospray ionization-assisted mass spectrometry confirms the formation of the OSM. This synthesized and well-characterized OSM as a ligand was further incubated with surface-engineered lipid nanoarchitectonics to achieve OSM ligand-engineered lipid nanoarchitectonics and earlier explored for its safety study through hemolysis assay and potential in vitro triggering efficiency in human alveolar macrophages (THP-1 cells) to validate its active targeting efficiency. Graphical Abstract [Figure: see text].