Cutting Edge: LAG3 Dimerization Is Required for TCR/CD3 Interaction and Inhibition of Antitumor Immunity.

Lymphocyte activation gene 3 (LAG3) is an inhibitory receptor that plays a critical role in controlling T cell tolerance and autoimmunity and is a major immunotherapeutic target. LAG3 is expressed on the cell surface as a homodimer but the functional relevance of this is unknown. In this study, we show that the association between the TCR/CD3 complex and a murine LAG3 mutant that cannot dimerize is perturbed in CD8+ T cells. We also show that LAG3 dimerization is required for optimal inhibitory function in a B16-gp100 tumor model. Finally, we demonstrate that a therapeutic LAG3 Ab, C9B7W, which does not block LAG3 interaction with its cognate ligand MHC class II, disrupts LAG3 dimerization and its association with the TCR/CD3 complex. These studies highlight the functional importance of LAG3 dimerization and offer additional approaches to therapeutically target LAG3.

Appraisal of sedimentary alkenones for the quantitative reconstruction of phytoplankton biomass.

Marine primary productivity (PP) is the driving factor in the global marine carbon cycle. Its reconstruction in past climates relies on biogeochemical proxies that are not considered to provide an unequivocal signal. These are often based on the water column flux of biogenic components to sediments (organic carbon, biogenic opal, biomarkers), although other factors than productivity are posited to control the sedimentary contents of the components, and their flux is related to the fraction of export production buried in sediments. Moreover, most flux proxies have not been globally appraised. Here, we assess a proxy to quantify past phytoplankton biomass by correlating the concentration of C37 alkenones in a global suite of core-top sediments with sea surface chlorophyll-a (SSchla) estimates over the last 20 y. SSchla is the central metric to calculate phytoplankton biomass and is directly related to PP. We show that the global spatial distribution of sedimentary alkenones is primarily correlated to SSchla rather than diagenetic factors such as the oxygen concentration in bottom waters, which challenges previous assumptions on the role of preservation on driving concentrations of sedimentary organic compounds. Moreover, our results suggest that the rate of global carbon export to sediments is not regionally constrained, and that alkenones producers play a dominant role in the global export of carbon buried in the seafloor. This study shows the potential of using sedimentary alkenones to estimate past phytoplankton biomass, which in turn can be used to infer past PP in the global ocean.

Formulation and evaluation of sustained release ocular inserts of betaxolol hydrochloride using arabinoxylan from Plantago ovata.

The purpose of the current studies was to develop ocular insert of betaxolol hydrochloride (BXH), using arabinoxylan (AX) as a film former. The inserts were prepared by sandwiching I mg of BXH between two films of AX. Six different formulations of ocular inserts were prepared in such a way that first three formulations contained varying concentrations of AX along with glycerol as plasticizer, whereas, rest of the formulations were added with 0.5mg of sodium alginate, sandwiched between two films of AX along with 1mg of BXH. Chemical compatibilities of the ingredients were assessed by using FTIR. Prepared ocular inserts were subjected to various physicochemical characterizations. The dissolution studies showed that ocular inserts containing sodium alginate with the AX showed sustained release effect better than the formulations with AX alone. Addition of sodium alginate resulted in inhibition of sudden release in initial phase and further sustained the release of drug from ocular inserts. Ocular inserts were pH compatible to the eyes as well as there was no interaction among the drug and excipients, suggesting that the selected excipients were suitable for the development of sustained release ocular inserts of BXH.

Formulation of instant disintegrating buccal films without using disintegrant: An in-vitro study.

The current study was conducted to fabricate Metoclopramide HCL (MCH) and Sumatriptan succinate (SS) instant release buccal films (IRBF) without using any super disintegrant. The solvent casting method was used for the preparation of IRBFs and prepared IRBFs were physicochemically evaluated. Spectrophotometric analysis was done to determine the lambda max followed by the linearity determination of both drugs. Different concentrations such as 100, 125, and 150mg of hydrophilic polymer (HPMC E5) were employed but the concentration of glycerol was variable. Comparatively better results were observed for the formulation with 150mg of HPMC E5 and 30% glycerol. Formulated IRBFs showed good tensile strength with a mean disintegration time of 12.4-28.4 seconds and rapid dissolution with more than 50% drug release within 2 minutes. It was concluded that the chosen combination of polymers was appropriate for the fabrication of MCH and SS buccal strips.

Rosuvastatin, Perindopril and Ezetimibe loaded instant release buccal films: Development and in vitro characterization.

BACKGROUND: Rosuvastatin Calcium and Ezetimibe are used to control cholesterol level while Perindopril Erbumine is used to treat hypertension. Hepatic metabolism reduces the therapeutic effect of these drugs. OBJECTIVE: Instant release buccal films (IRBFs) could possibly be a solution to this issue. The objective of the study was to formulate IRBFs of Rosuvastatin Calcium, Perindopril Erbumine and Ezetimibe using solvent casting technique. METHODS: Polymers used to prepare IRBFs included hydroxypropyl methylcellulose (HPMC E5), PEG 400 (as plasticizer) and Tween 80 (as surfactant). Solvent casting technique was used to fabricate the films, followed by their in-vitro analysis including high performance liquid chromatography (HPLC), X-ray diffraction (XRD), fourier transform infrared evaluation (FTIR), In-vitro dissolution, In-vitro disintegration, stability tests, scanning electron microscopy (SEM), folding fortitude, thickness evaluation, surface pH, tensile strength, weight variation and percentage moisture content. RESULTS: Optical microscopy as well as SEM analysis displayed that the surfaces of IRBFs were smooth with uniform mixing of ingredients. IRBFs disintegrated within 15 seconds while on dissolution they exhibited instant drug release i.e. 100% release in 2 minutes. CONCLUSIONS: The results show promising potential of IRBFs in drug delivery.

Development of thiomer based buccal films for the enhancement of bioavailability: An in-vivo analysis.

Present work was conducted to improve the bioavailability of Tizanidine HCl (TZN) by formulating mucoadhesive buccal films (MBFs) using novel thiolated arabinoxylan (TAX) as film former. MBF's were prepared by solvent casting technique followed by their evaluation for surface morphology and folding endurance. Moreover, pharmacokinetic parameters including Cmax, tmax, t1/2 and AUC were determined after administering standard oral solution (SOS) and MBFs of TZN at a dose of 1mg/kg. Successful thiolation was confirmed by the presence of 4.98 to 7.04 mmol of thiol content per gram of the polymer. Results of in-vivo pharmacokinetics have signified (p=0.0089) the suitability of MBFs as a carrier of drug through buccal route. Results have explored that, t1/2 was increased from 2.51hrs (SOS) to 10 hrs, Cmax from 42.3 ng/ml (SOS) to 105ng/ml and tmax from 2hrs (SOS) to 6h. Conclusively, TAX has exhibited the potential to form MBFs thereby offering sustained release of TZN with improved pharmacokinetic profile.

Development and optimization of flurbiprofen loaded microsponges; an in-vitro evaluation.

Current study was designed with the aim to employ quasi emulsification, and double emulsification techniques for the development of Flurbiprofen (FLB) loaded micro sponges, followed by their physicochemical evaluation. FTIR interpretations exhibited compatibility of ingredients, while crystallographic analysis revealed crystalline nature of pure drug, which was masked upon incorporation into microsponges. Optical microscope and SEM have exposed spherical and spongy surfaces of prepared micro sponges. Micromeritics suggested that the flow properties are excellent and microsponges have remarkable drug entrapment efficiency (98.55±0.08%). In-vitro dissolution studies demonstrated good control over release of FLB until 8th h from the prepared microsponges. However, a difference in cumulated amount of released drug was noticed i.e. EC based formulation has released about 99.3±0.10%, while XG facilitated EC based formulations offered 92.7±2.1% release of the drug. Zeta potential indicated access of negative charge while zeta sizer has described the range of the particle size between 2.6 to 3.5µm. Conclusively the results have advocated the suitability of selected ingredients for incorporation of FLB into microsponges for its sustained delivery.

Antibiotic resistance pattern shown by various pathogens, causing infections in neonates.

This study was schemed to comprehend the latest kaleidoscopic trends of bacterial resistance in neonatal pathogens against all those antibiotics commonly employed as empirical therapy in neonates. The methodological approach included; isolation and subsequent identification of those pathogens having caused bacterial infections in neonates, application of antibiotic sensitivity testing and finally construing the conclusion depicting patterns of antibiotic resistance by various pathogens, isolated from neonatal biological samples. Antibiotic resistance patterns was evident in gram-positive as well as in gram-negative bacteria in all the eight species identified in this study. Even antibiotic drugs which are being commonly relied upon for treating multi-resistant bacterial infections, found to be in effective against many newly emerged resistant bacteria, when used alone. Resistance Antibiotics drugs against which most prominent resistance pattern emerged include; Amikacin sulphate, Linezolid, Piperacillin / Tazobactam, Amoxicillin / Clavulanic acid, Vencomycin, Cefoperazone / Sulbactam, Ceftriaxone sodium, Ciprofloxacin, Cefixime trihydrate and Imipenem. The inferred upshot suggests that antibiotic resistance is emerging fast and ever-changing phenomenon of antibiotic resistance has significantly reduced the therapeutic space to maneuver, particularly, in treating neonatal infections.

Liquid chromatography-mass spectrometry as a general approach for investigating covalent binding of drugs to DNA.

This paper aims at developing a general strategy to study the detection of adducts of drugs with DNA. In particular, ethacrynic acid has been chosen as a model reactive drug that could be able to bind covalently to DNA bases. Such interactions were detected by ultra-high performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS). Principal component analysis (PCA) was applied as an unsupervised method to try to find the potential candidate adduct from MS features. The occurrence of adducts was investigated preliminarily using deoxynucleosides of the guanine, cytosine, adenine, and thymine separately as a way to optimize both separation and detection conditions. Interpretations of MS and MS/MS spectra provided tentative structures of the compounds formed. Conclusions extracted from such simple nucleoside models were further extended to the analysis of DNA adducts. For such a purpose, DNA was incubated in the presence of ethacrynic acid under appropriate experimental conditions and its further enzymatic hydrolysis released the corresponding nucleosides. UHPLC-MS analysis of the resulting test samples under the SRM detection mode confirmed the presence of ethacrynic acid derivatives of nucleosides occurring at very low concentration levels, thus proving the overall performance of the method. Graphical Abstract General approach for investigating drug-DNA adduct formation.

Comparative characterization and cytotoxicity study of TAT-peptide as potential vectors for siRNA and Dicer-substrate siRNA.

Recently, a newly discovered Dicer-substrate siRNA (DsiRNA) demonstrates higher potency in gene silencing than siRNA but both suffer from rapid degradation, poor cellular uptake and chemical instability. Therefore, Tat-peptide was exploited to protect and facilitate their delivery into cells. In this study, Tat-peptide was complexed with siRNA or DsiRNA through simple complexation. The physicochemical properties (particle size, surface charge and morphology) of the complexes formed were then characterized. The ability of Tat-peptide to carry and protect siRNA or DsiRNA was determined by UV-Vis spectrophotometry and serum protection assay, respectively. Cytotoxicity effect of these complexes was assessed in V79 cell line. siRNA-Tat complexes had particle size ranged from 186 ± 17.8 to 375 ± 8.3 nm with surface charge ranged from -9.3 ± 1.0 to +13.5 ± 1.0 mV, depending on the Tat-to-siRNA concentration ratio. As for DsiRNA-Tat complexes, the particle size was smaller than the ones complexed with siRNA, ranging from 176 ± 8.6 to 458 ± 14.7 nm. Their surface charge was in the range of +27.1 ± 3.6 to +38.1 ± 0.9 mV. Both oligonucleotide (ON) species bound strongly to Tat-peptide, forming stable complexes with loading efficiency of more than 86%. These complexes were relatively non cytotoxic as the cell viability of ∼90% was achieved. In conclusion, Tat-peptide has a great potential as siRNA and DsiRNA vector due to the formation of stable complexes with desirable physical characteristics, low toxicity and able to carry high amount of siRNA or DsiRNA.

Thiolated Polymeric Hydrogels for Biomedical Applications: A Review.

Hydrogels are a three-dimensional (3D) network of hydrophilic polymers. The physical and chemical crosslinking of polymeric chains maintains the structure of the hydrogels even when they are swollen in water. They can be modified with thiol by thiol epoxy, thiol-ene, thiol-disulfide, or thiol-one reactions. Their application as a matrix for protein and drug delivery, cellular immobilization, regenerative medicine, and scaffolds for tissue engineering was initiated in the early 21st century. This review focuses on the ingredients, classification techniques, and applications of hydrogels, types of thiolation by different thiol-reducing agents, along with their mechanisms. In this study, different applications for polymers used in thiolated hydrogels, including dextran, gelatin, polyethylene glycol (PEG), cyclodextrins, chitosan, hyaluronic acid, alginate, poloxamer, polygalacturonic acid, pectin, carrageenan gum, arabinoxylan, carboxymethyl cellulose (CMC), gellan gum, and polyvinyl alcohol (PVA) are reviewed.

An Insight into the Role of Artificial Intelligence in the Early Diagnosis of Alzheimer's Disease.

BACKGROUND: The complication of Alzheimer's disease (AD) has made the development of its therapeutic a challenging task. Even after decades of research, we have achieved no more than a few years of symptomatic relief. The inability to diagnose the disease early is the major hurdle behind its treatment. Several studies have aimed to identify potential biomarkers that can be detected in body fluids (CSF, blood, urine, etc.) or assessed by neuroimaging (i.e., PET and MRI). However, the clinical implementation of these biomarkers is incomplete as they cannot be validated. METHODS: This study aimed to overcome the limitation of using artificial intelligence along with technical tools that have been extensively investigated for AD diagnosis. For developing a promising artificial intelligence strategy that can diagnose AD early, it is critical to supervise neuropsychological outcomes and imaging-based readouts with a proper clinical review. CONCLUSION: Profound knowledge, a large data pool, and detailed investigations are required for the successful implementation of this tool. This review will enlighten various aspects of early diagnosis of AD using artificial intelligence.

Synthesis of Thiol-Modified Hemicellulose, Its Biocompatibility, Studies, and Appraisal as a Sustained Release Carrier of Ticagrelor.

Background: Nature has always been considered as the primary source of pharmaceutical ingredients. A variety of hemicelluloses, as well as their modified forms, have been under investigation. Herein, a study was designed to explore the biocompatibility of hemicellulose and its modified form (thiolated hemicellulose) as well as its potential as a pharmaceutical excipient. Method: For thiol modification thiourea was used as the thiol donor, HCl as the catalytic reagent, and methanol was used for washing purposes. Modified polymers were characterized for physicochemical characteristics, including surface morphology, the amorphous or crystalline nature of the particles, modification of polymer by FTIR, and biocompatibilities. For acute oral toxicity study, a single dose of 2 g/kg was administered to albino rats of 200 g average weight (n = 3). Polymers were evaluated as pharmaceutical excipients by preparing compressed tablets of antiplatelet drug (Ticagrelor), followed by various quality control tests, such as swelling index, thickness and diameter, disintegration, and in-vitro drug release. Results: From the results, it was observed that thiol modification has been successfully accomplished as characteristic peaks belonging to -SH group appeared at 2667.7691 cm-1 in FTIR scan. The modified polymer was found safe in the use concentration range, confirming their safe use for in vivo analysis. No significant effect has been observed in the behavior, biological fluid (blood), or on vital organs. Thiolated hemicellulose was found to be an excellent drug retarding polymer as 8 h of dissolution studies showed that 67.08% of the drug has been released. Conclusion: Conclusively, incorporation of thiol moiety made the polymer more mucoadhesive with, and a worthy carrier of, the drug with good biocompatibilities.

Evaluation of Ligustrazine-Based Synthetic Compounds for their Antiproliferative Effects.

BACKGROUND: Ligustrazine and chalcones have been reported previously for various biological activities including anticancer effects. OBJECTIVES: Based on the multitargeted biological activities approach of ligustrazine-based chalcones, in the current study 18 synthetic ligustrazine-containing α, ß-unsaturated carbonyl-based 1, 3- Diphenyl-2-propen-1-one derivatives were evaluated for their inhibitory effects on the growth of five different types of cancer cells. METHODS: All the compounds were evaluated for anticancer effects on various cancer cell lines by propidium iodide fluorescence assay and various other assays were performed for mechanistic studies. RESULTS: A majority of compounds exhibited strong inhibition of cancer cells, especially synthetic compounds 4a and 4b, bearing 1-Pyridin-3-yl-ethanone as a ketone moiety in the main structural backbone were found to be most powerful inhibitors of cancer cell growth. Nine most active compounds among the whole series were selected for further studies related to different cancer targets, including EGFR TK kinases, tubulin polymerization, KAF and BRAFV600E. CONCLUSION: Synthetic derivatives, including 4a-b and 5a-b showed a multitarget approach and strong inhibitory effects on EGFR, FAK and BRAF while three compounds, including 3e bearing methoxy substitution, 4a and 4b with 1-pyridin-3-yl-ethanone moiety showed the inhibition of tubulin polymerization.

Nanoparticles Based Intranasal Delivery of Drug to Treat Alzheimer's Disease: A Recent Update.

Alzheimer Association Report (2019) stated that the 6th primary cause of death in the USA is Alzheimer's Disease (AD), which leads to behaviour and cognitive impairment. Nearly 5.8 million peoples of all ages in the USA have suffered from this disease, including 5.6 million elderly populations. The statistics of the progression of this disease is similar to the global scenario. Still, the treatment of AD is limited to a few conventional oral drugs, which often fail to deliver an adequate amount of the drug in the brain. The reduction in the therapeutic efficacy of an anti-AD drug is due to poor solubility, existence to the blood-brain barrier and low permeability. In this context, nasal drug delivery emerges as a promising route for the delivery of large and small molecular drugs for the treatment of AD. This promising pathway delivers the drug directly into the brain via an olfactory route, which leads to the low systemic side effect, enhanced bioavailability, and higher therapeutic efficacy. However, few setbacks, such as mucociliary clearance and poor drug mucosal permeation, limit its translation from the laboratory to the clinic. The above stated limitation could be overcome by the adaption of nanoparticle as a drug delivery carrier, which may lead to prolong delivery of drugs with better permeability and high efficacy. This review highlights the latest work on the development of promising Nanoparticles (NPs) via the intranasal route for the treatment of AD. Additionally, the current update in this article will draw the attention of the researcher working on these fields and facing challenges in practical applicability.

Fabrication of polyvinyl alcohol based fast dissolving oral strips of sumatriptan succinate and metoclopramide HCL.

Migraine is a throbbing condition, usually associated with nausea and vomiting and requires concurrent administration of anti-migraine along with anti-emetic therapy. The current study was undertaken with an aim to fabricate fast dissolving oral strips (FDOSs) containing Sumatriptan succinate (anti-migraine) and Metoclopramide HCl (anti-emetic) in combination without involving any superdisintegrant. Hydrophilic polymer polyvinyl alcohol (PVA) was used alone with three concentrations of 100, 125, and 150 mg using variable concentrations of glycerol. The solvent casting technique was employed to formulate FDOSs and were evaluated for surface morphology, mechanical properties, surface pH, % moisture content, disintegration time (DT), total dissolving time (TDT), drug contents, and dissolution profile. PVA (150 mg) with 5% glycerol concentration gave best formulation results. FDOSs have exhibited good tensile strength with smooth and uniform surface morphology. DT was ranged from 7.7 to 28 s; while TDT was from 26.4 to 77.6 s. Both polymer and plasticizer concentrations were found to be influencing the characteristics of the strips. Dissolution studies were carried out in distilled water for 15 min and all the formulations have shown released more than 50% drug within first 2 min thereby highlighting the usefulness of FDOSs for the delivery of both drugs in combination significantly. Optimized combination of ingredients was found to be suitable for the formulation of FDOSs for simultaneous delivery of Metoclopramide HCl and Sumatriptan succinate.

Enhancement of Oral Bioavailability of Ibandronate Through Gastroretentive Raft Forming Drug Delivery System: In Vitro and In Vivo Evaluation.

BACKGROUND: Bisphosphonates have very low bioavailability and cause irritation of the esophagus and stomach. This study was planned to improve the oral bioavailability of ibandronate through the formation of a raft in the stomach. Bisphosphonate-induced irritation of the esophagus and stomach is prevented by the formation of a raft. MATERIALS AND METHODS: The nanostructured raft was developed through the use of nanosized citrus pectin (NCP). The particle size of NCP was measured by zeta sizer and SEM. The percentage of NCP and the neutralization profile of raft was studied. The ibandronate, polymers, and the developed formulation were characterized by FTIR, XRD, TGA, and DSC. The release of ibandronate was studied in 0.1 N HCl, 0.5 N HCl, 1 N HCl, and simulated gastric fluid (SGF) and a cell viability study was performed using Caco-2 cells. The PPR5 formulation and Bonish 150 mg tablets were selected as test and reference formulations, respectively, for pharmacokinetic study. Twelve healthy albino rats were taken and divided into two groups using a Latin square crossover design, and the blood samples were collected for 24 hours. RESULTS: The SEM image showed that the particle size of NCP was 159 nm. The raft of PPR5 showed 94% NCP and 45 minutes duration of neutralization. The FTIR and XRD showed chemical stability and a uniform distribution of ibandronate in the raft. The TGA and DSC indicated the thermal stability of formulation. The release of 99.87% ibandronate at 20 minutes was observed in the SGF. The values of C max for the reference and test formulations were 493±0.237 ng/mL and 653±0.097 ng/mL, respectively. The AUC(0-t) of the reference and test formulations was 3708.25±3.418 ng/mL.h and 6899.25±3.467 ng/mL.h, respectively. CONCLUSION: The NCP has been successfully prepared from citrus pectin and has shown effective porous raft formation. The bioavailability of the ibandronate from newly developed PPR5 was higher than the already marketed formulation.

Solid Lipid Nanoparticles of Mycophenolate Mofetil: An Attempt to Control the Release of an Immunosuppressant.

INTRODUCTION: Organ transplantation is a critically important procedure, which requires immune modulation by using immunosuppressants. Development of nanoparticles is an emerging and beneficial engineering process to increase the dissolution rate of poorly soluble immunosuppressants as well as to provide controlled release for better therapeutic outcomes. METHOD: Currently, the nanoprecipitation method was employed to fabricate ß-cyclodextrin (ßCD) facilitated mycophenolate mofetil (MMF)-loaded solid lipid nanoparticles (SLNPs). The prime objectives of the study included, improvement of the dissolution profile of poorly aqueous soluble drug and controlled release from the SLNs to provide steady state drug concentration. Drug release from the prepared SLNs was assessed in two different media, ie, acidic buffer at pH 1.2 and phosphate buffer at pH 7.2 using USP dissolution apparatus for 12 h, followed by the evaluation of drug release mechanism and pattern by applying kinetic models. RESULTS: Justifiably, in acidic medium, the release was found to be 12% more (68%) in comparison to that in basic medium (56%). However, in both dissolution media, drug release was independent of initial concentration (R2>0.95) with non-Fickian type of diffusion mechanism. The outcomes of the study have exhibited that prepared formulations were in nanosized range (80-170 nm) with a net charge of ±23 charge on their surface. They possessed fairly uniform surface with acceptable polydispersity index (0.23±0.09). Scanning electron microscopy (SEM) analysis illustrated that the nanoparticles had uniform particle size and shape. DISCUSSION: The findings show potential applications of the nanoparticles and the method for the development of SLNPs in controlled release of MMF for better therapeutic outcomes. Conclusively, the prepared SLNPs were well designed in nanosized ranges and justifying the once daily controlled release formulation dose of MMF to enhance patient compliance.

Design, mechanism, delivery and therapeutics of canonical and Dicer-substrate siRNA.

Upon the discovery of RNA interference (RNAi), canonical small interfering RNA (siRNA) has been recognized to trigger sequence-specific gene silencing. Despite the benefits of siRNAs as potential new drugs, there are obstacles still to be overcome, including off-target effects and immune stimulation. More recently, Dicer substrate siRNA (DsiRNA) has been introduced as an alternative to siRNA. Similarly, it also is proving to be potent and target-specific, while rendering less immune stimulation. DsiRNA is 25-30 nucleotides in length, and is further cleaved and processed by the Dicer enzyme. As with siRNA, it is crucial to design and develop a stable, safe, and efficient system for the delivery of DsiRNA into the cytoplasm of targeted cells. Several polymeric nanoparticle systems have been well established to load DsiRNA for in vitro and in vivo delivery, thereby overcoming a major hurdle in the therapeutic uses of DsiRNA. The present review focuses on a comparison of siRNA and DsiRNA on the basis of their design, mechanism, in vitro and in vivo delivery, and therapeutics.