PROCESS OPTIMIZATION, FORMULATION AND EVALUATION OF HYDROGEL {GUARGUM-G-POLY (ACRYLAMIDE)} BASED DOXOFYLLINE MICROBEADS

Objective: The objective of the present study was to improve the physical and chemical properties of natural polymers and to reduce the cost of product by graft copolymerization techniques using a natural polymer (Guar gum) and a synthetic polymer {poly (acrylamide)}. The optimized formulation of hydrogel was formulated as microbeads and loaded with Doxofylline and characterized with different parameters.Methods: Graft copolymer of guar gum-g-poly (acrylamide) was prepared by free radical polymerization technique in a specially designed jacked reaction vessel under constant flow of nitrogen. To initiate the reaction, Ceric ammonium nitrate (CAN) was used as reaction initiator. The graft co-polymer was characterised by using FTIR, TGA, and SEM. Polymeric blend beads of the grafted copolymer with sodium alginate were prepared by cross linking with calcium chloride in ionic gelation method and used to deliver a model new generation anti asthmatic drug, Doxofylline. Preparation condition of beads was optimized by considering the percentage entrapment efficiency, particle size, swelling capacity of beads in different PH conditions and their release data.Results: The formation of grafted copolymers is confirmed by FTIR studies and TGA studies showed a comparatively higher thermal stability of grafted copolymer. The pAAm-g-GG/sodium alginate microbeads were almost spherical in shape as indicated by the SEM studies. Swelling index was found to be maximum in Phosphate buffer PH 7.4 and minimum in Phosphate buffer PH 9.2. Release of doxofylline was found to be in a controlled manner with increasing polyacrylamide content in the copolymer and sodium alginate content in microbeads and higher release was observed in PH 7.4 medium than that of PH 1.2. In vitro release kinetics of doxofylline from the polymeric beads followed Higuchi kinetics model.Conclusion: Hydrogel based Doxofylline microbeads were successfully developed by using optimized batches of Guar gum-g-poly (acrylamide) and sodium alginate by free radical ionization technique. All the characterization parameters came under acceptance criteria.  Key words: Hydrogel, Microbeads, Guar gum, Acrylamide, Sodium alginat

FORMULATION DESIGN AND IN VITRO EVALUATION OF BILAYER SUSTAINED RELEASE MATRIX TABLETS OF DOXOFYLLINE

Objective: To develop bilayer matrix tablet of Doxofylline by providing a loading dose followed by the maintenance dose that suppose to enhance the therapeutic efficacy the drug for acute and sustainable asthma.Methods: Both immediate release layer and sustained release layer were prepared by wet granulation methods. Different Pre compression and post compression characterization of the tablet were carried out. Swelling studies were carried out for all the formulation. To optimise the immediate release layer, similarity (f2) and difference factor (f1) were calculated and optimised IR formulation was used for all formulations of bilayer tablet. In-vitro release studies were carried out in USP II paddle type dissolution apparatus for different formulations and release kinetic studies were carried out different kinetic model. FTIR and DSC studies were carried out for pure drug Doxofylline, IR layer and SR layer of optimised formulation to know the physical and chemical compatibility of drug and excipients. Accelerated stability studies were carried out to confirm the stability of dosage forms.Results: Pre compression and post compression parameters satisfied with pharmacopeia specifications. The formulation that contained highest percent of HPMC had highest swelling index. Formulation DBMF6 showed an initial release of 44% of drug within one hour as the loading dose and remaining drug were sustained release up to 12 h. Release kinetic followed Hixon-Crowell kinetic model with drug release mechanism quasi-fickian diffusion. From accelerated stability studies no significant changes in physicochemical properties were noticed.Conclusion: Doxofylline bilayer matrix tablets were successfully developed and can be used as an alternative to the conventional dosage form because it can be therapeutically beneficial for management of asthma.Â

PREPARATION, PHYSICAL CHARACTERIZATION, AND PHARMACOKINETIC STUDY OF DOCETAXEL NANOCRYSTALS

Objective: The main objective of this study was to prepare and evaluate the nanocrystal formulation of docetaxel. Methods: Docetaxel nanocrystals were formulated to improve the water solubility. Docetaxel nanocrystals were prepared by nanoprecipitation method using Tween 80, egg lecithin, and povidone C-12 as stabilizers and poly(lactic-co-glycolic acid) (PLGA) as polymer in acceptable limits. A total of 16 formulations were prepared by changing stabilizer and polymer ratios. The prepared nanocrystals were characterized by particle size, zeta potential, crystalline structure, surface morphology, assay, saturation solubility, and in vitro drug release. Results: Based on particle size, polydispersity index, and zeta potential data, four formulations were optimized. The formulation containing Tween 80 as stabilizer has shown lowest particle size and better drug release than the formulations containing egg lecithin and povidone C-12 as stabilizers. The formulation containing Tween 80 and PLGA has shown still lower sized particles than the Tween 80 alone and exhibited prolonged sustained drug release. The release kinetics of formulations containing Tween 80 and PLGA followed zero-order release kinetics and formulations containing egg lecithin and povidone C-12 followed Higuchi diffusion (non-Fickian). Conclusion: From the study, we concluded that as the type and concentration of stabilizer changed the size and shape of the crystals were also changed and the formulations showed sustained drug release with non-Fickian diffusion

Genome-Wide and Functional Annotation of Human E3 Ubiquitin Ligases Identifies MULAN, a Mitochondrial E3 that Regulates the Organelle's Dynamics and Signaling

Specificity of protein ubiquitylation is conferred by E3 ubiquitin (Ub) ligases. We have annotated ∼617 putative E3s and substrate-recognition subunits of E3 complexes encoded in the human genome. The limited knowledge of the function of members of the large E3 superfamily prompted us to generate genome-wide E3 cDNA and RNAi expression libraries designed for functional screening. An imaging-based screen using these libraries to identify E3s that regulate mitochondrial dynamics uncovered MULAN/FLJ12875, a RING finger protein whose ectopic expression and knockdown both interfered with mitochondrial trafficking and morphology. We found that MULAN is a mitochondrial protein – two transmembrane domains mediate its localization to the organelle's outer membrane. MULAN is oriented such that its E3-active, C-terminal RING finger is exposed to the cytosol, where it has access to other components of the Ub system. Both an intact RING finger and the correct subcellular localization were required for regulation of mitochondrial dynamics, suggesting that MULAN's downstream effectors are proteins that are either integral to, or associated with, mitochondria and that become modified with Ub. Interestingly, MULAN had previously been identified as an activator of NF-κB, thus providing a link between mitochondrial dynamics and mitochondria-to-nucleus signaling. These findings suggest the existence of a new, Ub-mediated mechanism responsible for integration of mitochondria into the cellular environment

CAFET Algorithm Reveals Wnt/PCP Signature in Lung Squamous Cell Carcinoma

We analyzed the gene expression patterns of 138 Non-Small Cell Lung Cancer (NSCLC) samples and developed a new algorithm called Coverage Analysis with Fisher’s Exact Test (CAFET) to identify molecular pathways that are differentially activated in squamous cell carcinoma (SCC) and adenocarcinoma (AC) subtypes. Analysis of the lung cancer samples demonstrated hierarchical clustering according to the histological subtype and revealed a strong enrichment for the Wnt signaling pathway components in the cluster consisting predominantly of SCC samples. The specific gene expression pattern observed correlated with enhanced activation of the Wnt Planar Cell Polarity (PCP) pathway and inhibition of the canonical Wnt signaling branch. Further real time RT-PCR follow-up with additional primary tumor samples and lung cancer cell lines confirmed enrichment of Wnt/PCP pathway associated genes in the SCC subtype. Dysregulation of the canonical Wnt pathway, characterized by increased levels of β-catenin and epigenetic silencing of negative regulators, has been reported in adenocarcinoma of the lung. Our results suggest that SCC and AC utilize different branches of the Wnt pathway during oncogenesis

CXCR4 Expression in Prostate Cancer Progenitor Cells

Tumor progenitor cells represent a population of drug-resistant cells that can survive conventional chemotherapy and lead to tumor relapse. However, little is known of the role of tumor progenitors in prostate cancer metastasis. The studies reported herein show that the CXCR4/CXCL12 axis, a key regulator of tumor dissemination, plays a role in the maintenance of prostate cancer stem-like cells. The CXCL4/CXCR12 pathway is activated in the CD44+/CD133+ prostate progenitor population and affects differentiation potential, cell adhesion, clonal growth and tumorigenicity. Furthermore, prostate tumor xenograft studies in mice showed that a combination of the CXCR4 receptor antagonist AMD3100, which targets prostate cancer stem-like cells, and the conventional chemotherapeutic drug Taxotere, which targets the bulk tumor, is significantly more effective in eradicating tumors as compared to monotherapy

Iodine-catalyzed stereoselective synthesis of allylglycosides, glycosyl cyanides and glycosyl azides

Iodine efficiently catalyzes the glycosidation of glycals with allyltrimethylsilane, trimethylsilyl cyanide and trimethylsilyl azide in dichloromethane at ambient temperature to afford the corresponding 2,3-unsaturated allyl glycosides, glycosyl cyanides and glycosyl azides in excellent yields with high α-selectivity

CLEC5A (MDL-1) is a novel PU.1 transcriptional target during myeloid differentiation

C-type lectin domain family 5, member A (CLEC5A), also known as myeloid DNAX activation protein 12 (DAP12)-associating lectin-1 (MDL-1), is a cell surface receptor strongly associated with the activation and differentiation of myeloid cells. CLEC5A associates with its adaptor protein DAP12 to activate a signaling cascade resulting in activation of downstream kinases in inflammatory responses. Currently, little is known about the transcriptional regulation of CLEC5A. We identified CLEC5A as one of the most highly induced genes in a microarray gene profiling experiment of PU.1 restored myeloid PU.1-null cells. We further report that CLEC5A expression is significantly reduced in several myeloid differentiation models upon PU.1 inhibition during monocyte/macrophage or granulocyte differentiation. In addition, CLEC5A mRNA expression was significantly lower in primary acute myeloid leukemia (AML) patient samples than in macrophages and granulocytes from healthy donors. Moreover, we found activation of a CLEC5A promoter reporter by PU.1 as well as in vivo binding of PU.1 to the CLEC5A promoter. Our findings indicate that CLEC5A expression in monocyte/macrophage and granulocytes is regulated by PU.1