Hetalkumar Panchal

DNA sequence analysis by ORF FINDER & GENOMATIX tool: Bioinformatics analysis of some tree species of Leguminosae family

Bioinformatics is rapidly growing field of applied science, here we have done DNA sequence analyses of few legume tree species by bioinformatics tools. In this paper bioinformatics data of some leguminous trees have been explored and brought to one platform. Various analytical bioinformatics tools were used to generate the information for particular species or group of species. DNA sequence analyses have been done by ORF FINDER & GENOMATIX. The results were discussed in context with all available data generated through above methods for leguminous trees. We have done analysis of 23 legume species of Leguminosae family, and it is further classified in three subfamily, 1. Fabaceae (Papilionaceae) 2. Caesalpiniaceae 3. Mimosaeae and made a database which contains all legume species results, but here we have taken only two legume species in two tools to demonstrate our work. We have used DNA sequence from EMBL database.

Phylogenetic analysis of some leguminous trees using CLUSTALW2 bioinformatics tool

A multiple sequence alignment (MSA) is a sequence alignment of three or more biological sequences, generally for Protein. MSA has wide range of applications and to cite few of them such as phylogenetic analysis, protein pattern identification, protein domain identification, prediction of protein structure, structural similarity of amino acids and to get evolutionary similarity. ClustalW2 is a general purpose global multiple sequence alignment program for proteins. It produces biologically meaningful multiple sequence alignments of divergent sequences. The output from ClustalW2 shows the best match for the selected sequences and lines up them in such a way that the identities, similarities and differences can be easily understood. Evolutionary relationships can be seen by creating Cladograms or Phylograms. Firstly, individual weights are assigned to each sequence in a partial alignment in order to down-weight near-duplicate sequences and up-weight the most divergent ones. Secondly, amino acid substitution matrices are varied at different alignment stages according to the divergence of the sequences to be aligned. Thirdly, residue-specific gap penalties and locally reduced gap penalties in hydrophilic regions encourage new gaps in potential loop regions rather than regular secondary structure. Fourthly, positions in early alignments where gaps have been opened receive locally reduced gap penalties to encourage the opening up of new gaps at these positions. In this paper, protein sequences of few legume species from UNIPROT database were taken and focused on MSA for protein sequences for these tree species of family Leguminosae, where ClustalW2 tool have used to generate biological data. The results are discussed with the help of Cladograms and Phylograms for selected tree species.

In silico Structure Modeling and Comparative Analysis of Characterization Properties of Protein Polymers Useful for Protein-Based Nano Particulate Drug Delivery Systems (NPDDS): A Bioinformatics Approach

With an increasing interest in nanoparticulate delivery systems, there is a greater need to identify biomaterials that are biocompatible and safe for human applications. Protein polymers from animal and plant sources are promising materials for designing nanocarriers. Composition of the protein plays an important role for specific drug delivery applications such as drug release, targeting, and stimuli responsive drug release. An important issue in protein polymers is characteristics such as size, charge, and hydrophobicity may play a significant role in phagocytic uptake and initiating a subsequent immune response. This remains to be investigated systematically by analyzing factors that influence nanoparticle characteristics of protein and reduce phagocytic uptake and does not initiate immune response too. Although protein polymers are biodegradable, it is essential to ensure that there must not be premature enzymatic breakdown of the protein nanoparticles in the systemic circulation. Surface modification of the protein nanoparticles can be used to address this issue to propose the necessary modification in the surface of the protein would be great contribution in the nano particulate drug delivery systems (NPPDS). Of the various proteins, gelatin and albumin have been widely studied for drug delivery applications. Plant proteins are yet to be investigated widely for drug delivery applications so there is need to find out the plant proteins capable to act as nanoparticles. The commercial success of albumin-based nanoparticles has created an interest in other proteins. An increased understanding of the physicochemical properties coupled with the developments in rDNA technology will open up new opportunities for protein-based nanoparticulate systems. In the present studies several proteins currently useful for drug delivery system were structurally modeled and has been analyzed to propose the essential characteristics of protein for protein-based NPDDS.

In silico Interactive Study of Periplasmic and Outer Membrane Proteins of Type IV Secreting System (T4ss) in Helicobacter pylori

Helicobacter pylori is gram-negative bacterial pathogen, associated with chronic gastritis and other severe gastro duodenal diseases such as peptic and gastric ulcers, gastric cancer, and mucosa-associated lymphoid tissue (MALT) lymphoma. One of the intriguing virulence factors of H. pylori is cag Pathogenicity Island (cagPAI) which codes for Type IV secreting system. Type IV secreting system is found in many strains of H. pylori. Present work focuses on protein interaction studies to clarify and confirm the interactions and assembly of proteins present in outer membrane and pilus structure of t4ss. During the interaction studies of t4ss we have focused on only CagC, Cag3 and CagM interactions based on steriochemical properties. It is found that the pattern of pilus formation follows the specific order of assembly and the order of assembly is CagC-C dimer formation followed by tetramer formation to form stable pilus assembly of t4ss. The order of interaction at the outer membrane is CagM interacts with CagC-C dimer and then with Cag3 to form sub-assembly of t4ss.This findings of structural interaction can be used to prevent onset of diseases caused by H. pylori by preventing the formation of t4ss, pilus assembly and ultimately the translocation of onco protein CagA in host organism.

In silico investigation and structural characterization of virulent factor and a metallo peptidase present in Helicobacter pylori strain J99

VacA is a high-molecular weight multimeric pore-forming protein encoded by the chromosomal gene vacA of Helicobacter pylori J99 strian. It plays a significant role in the development of gastric cancer in human by inducing the formation of vacuoles. Genomics and proteomics features of an organism have provided a plethora of potential drug targets. The crystal structure of VacA is not available in any structural database; hence a 3D structure is very essential for structural studies and discovery of potential inhibitors against proteins. In this study 3D structure of VacA is modelled a by using Bhageerath: an energy based web enabled computer software suite. According to our study VacA steriochemical validation shows 91.7% residues are in allowed region of Ramachandran plot. Further validation was done by WHAT CHECK to provide evidence that the distribution of the main chain bond lengths and omega bond angles were within limits with Z-score 1.0 and error values are negligible. The modelled protein was submitted to Protein Model Database and can be downloaded with PMDID PM0077963. Further we found that metallo peptidase “M3” cleaves VacA and helps in import mechanism in mitochondria. Structure of metallo peptidase is also not available in any structural database so we modelled and validated its structure. With the help of docking studies we blocked the active site of metallo peptidase by ligand LA3 and 294 with binding energy −5.9 and −5.2 KJ/mol respectively, thus prevented import mechanism of VacA in mitochondria. The inhibitors identified from our study were LA3 and 294 ligands. The investigation concluded that these drugs could be used as the potential inhibitors against the damage of stomach and duodenum, which ultimately reduces the likelihood of ulcer as well as gastric cancer.

Evolutionary studies of some species belonging to leguminosae family based on rbcl gene

Although conifers are of immense ecological and economic value, bioengineering of their chloroplasts remains undeveloped. Understanding chloroplast genomic organization in conifers can facilitate their bioengineering. Members of the conifer II clade (or cupressophytes) are highly diverse in both morphologic features and chloroplast genomic organization. We compared six cupressophyte chloroplast genomes (cpDNAs) that represent four of the five cupressophyte families, including three genomes that are first reported here (Agathis dammara, Calocedrus formosana, and Nageia nagi). The six cupressophyte cpDNAs have lost a pair of large inverted repeats (IRs) and vary greatly in size, organization, and tRNA copies. We demonstrate that cupressophyte cpDNAs have evolved toward reduced size, largely due to shrunken intergenic spacers. In cupressophytes, cpDNA rearrangements are capable of extending intergenic spacers, and synonymous mutations are negatively associated with the size and frequency of rearrangements. The variable cpDNA sizes of cupressophytes may have been shaped by mutational burden and genomic rearrangements. Based on cpDNA organization, our analyses reveal that in gymnosperms, cpDNA rearrangements are phylogenetically informative, which supports the “gnepines” clade. As well, removal of a specific IR influences the minimal rearrangements required for the gnepines and cupressophyte clades, whereby Pinaceae favors removal of IRB but cupressophytes exclusion of IRA. This result strongly suggests that different IR copies have been lost from conifers I and II. Our data helps understand the structural complexity and evolution of cupressophyte cpDNAs.Acute myeloid leukemia (AML) is the second most common form of childhood leukemia and has the worst prognosis of all major childhood cancers. Improving the treatment outcome for patients with AML remains a major clinical challenge. The nucleoside analog, cytarabine (ara-C), has been the mainstay of AML chemotherapy for more than 40 years. However, wide inter-patient variation in treatment response, development of resistance, and severe toxicity remain as major hurdles to effective ara-C chemotherapy. Ara-C is a prodrug that requires activation to ara-CTP by multiple phosphorylation steps. Incorporation of ara-CTP in place of dCTP results in chain termination, thereby blocking DNA and RNA synthesis and causing leukemic cell death. Thus, cellular pathways involved in ara-CTP formation and metabolism as well as in ara-CTP mediated cell death are likely to be significant determinants of ara-C treatment response. Inter-patient variation in relevant pharmacokinetic (PK) and pharmacodynamic (PD) genes may impact the clinical response and toxicity among patients receiving ara-C. We have evaluated genes of importance in ara-C chemotherapy and have found that genetic variation in the ara-C pathway genes had similar prognostic relevance as the well-established factors listed above. We will share our results on ara-C pharmcogenomics and its impact on clinical outcome in AM. Overall our results indicate that understanding of genetic variation in key ara-C metabolic pathway genes might be clinically relevant by providing additional explanation of the variability in clinical response beyond known prognostic factors and might have the potential of being additional prognostic markers of clinical outcome.

De novo RNA seq assembly and annotation of important legume-Vicia sativa L. (SRR403901)

D (Dpp) is the homolog of vertebrate BMP-2 and BMP-4 and functionally interchangeable BMPs. The Bombyx mori (B. mori) and Bombyx mandarina (B. mandarina) Dpp genes share genetic homology with human BMPs and Drosophila Dpp, but few studies have been executed to examine the functions of B. mori and B. mandarina Dpp and its function is not well understood. To date, there was also no reported splicing form of Dpp in silkworm. In this study, we investigated Dpp expression using synthesized cDNA from midgut tissue of B. mandarina by RT-PCR. Interestingly, lower band was discovered with band of full-length Dppc DNA and it was identified as novel splicing form that a part (333 bp) of B. mandarina Dpp was deleted through DNA sequencing analysis. In addition, we found that the delated part in the variant was a portion of proprotein region compared to human BMP-2 and 7 candidate single nucleotide variants (SNVs) were able to affect formation of novel splicing form using variant calling analysis. To the best of our knowledge, this is the first approach to address the novel splicing form of Dpp in B. mandarina which have been not found in B. mori. These results suggest that discovered splicing form of B. mandarina was degenerated in evolution process toward more advanced and domesticated B. mori.Introduction Fusion genes, also known as chimeras, play important roles in tumorigenesis and cancer progression. Then, their role becomes crucial in the areas of biomarkers and therapeutic targets investigation. High-throughput sequencing technologies combined with sophisticated bioinformatics tools might facilitate the discovery of such aberrations. A significant number of bioinformatics algorithms have been developed to detect fusion genes. Detection strategies are quite variegated. In this review, we inspect the strategy of 18 fusion-finder algorithms to understand how these tools call chimeras. Materials and methods In this review, we considered 18 tools which, to the best of our knowledge, are the current state-of-the-art chimera detection tools. Results The considered tools can be classified according to their alignment strategies into four different macro-groups as follows: whole paired-end, paired-end + fragmentation, direct fragmentation and statistical read distribution. The first two techniques require pairedend reads because they exploit encompassing reads during the first alignment phase, while the last two can be applied on both the read formats. Conclusion There is still some work to be done in the area of chimeras detection, especially concerning the definition of common benchmarks and increased specificity.A are powerful reagents for biomedical research, diagnostics and therapeutics. The vast majority of antibody applications have focused on targeting features of the terrestrial proteome. Engineering antibodies to bind RNA directly offers a way to harness powerful technologies to study transcriptomes. We are developing phage display antibody libraries for engineering antibodies that recognize structured RNAs with high affinity and specificity. In one particular application, we are using the antibodies as chaperones for the crystallization and structure determination of RNAs.O meyeriana (O. meyeriana), a GG genome type (2n=24), accumulated plentiful excellent characteristics with respect to tolerance to shade, blast and bacterial blight. However, limited genomic or transcriptomic data of O. meyeriana are currently available. In this study, we present the first comprehensive characterization of the O. meyeriana transcriptome using RNA-seq. We obtained 185,323 contigs with an average length of 1,692 bp and an N50 of 2,391 bp. Through differential expression analysis, we found there were most tissue-specifically expressed genes in roots, and next to stems and leaves. By similarity search against protein databases, 146,450 had at least a significant alignment to existed gene models. Comparison with the Oryza sativa (O. sativa) genome revealed similar rate of alignment across different plant species, however 13% of the O. meyeriana contigs had not been detected as expressed in O. sativa. Furthermore, the enrichment of the plant-pathogen interaction pathway in O. meyeriana showed difference with O. sativa, including the similarity of these aligned candidate genes and the absence of candidate genes involved in the plant-pathogen interaction pathway. In addition, we identified 52 contigs as disease resistance protein which was not existed in O. sativa. Taken together, there are significant differences between O. meyeriana and O. sativa in disease resistance. This information provides a foundation for future investigations into the discovery of a huge amount of novel genes which will facilitate gene mining and provides a basis for comparative studies within the genus Oryza.H mobility group protein A1 (HMGA1) is a non-histone chromosomal protein also known as ‘architectural’ transcription factor that facilitates the assembly of ‘enhanceosome’. Expression of HMGA1 is elevated in a number of human malignancies, however, is practically absent in healthy adults. In the present study, we made an attempt to inhibit hmga1 expression in cancer by using anti-gene strategy. Two triplex forming oligonucleotides (TFOs), TFO-1 and TFO-2 were chosen to target the promoter region of HMGA1 at positions, -284 to -304 and -2800 to -2826 respectively. Stability of DNA triplexes were characterized using UV-Vis spectroscopy, Circular Dichroic spectroscopy, Isothermal titration and Differential scanning calorimetry and was confirmed by gel retardation assay using -32P [ATP]. Expression analysis of HMGA1 was evaluated in HeLa cells using MTT assay, Flow cytometry, Western blot and RT-PCR. Results reveal a higher binding affinity of TFO-1 to HMGA1 as compared to that of TFO-2 which correlates well with the greater efficacy of TFO-1, both in terms of HMGA1 expression and apoptosis. Further, the combination treatment of TFO-1 and adriamycin illustrates an additive effect on HMGA1 expression. Present results indicate that TFO-mediated inhibition of HMGA1 expression is a promising strategy for the development of novel anticancer therapeutics.L plays critical roles in plant development. Depending on the light conditions, plants flexibly elongate or shorten their hypocotyls and leaves. Plant hormones including auxins, gibberellins, and brassinosteroids (BRs) regulate these processes acting alone or in concert. To understand the molecular signature of gene expression under the control of both light and BRs, we performed an Affymetrix genome-wide transcriptome analysis using total RNAs prepared from the four genotypes of Arabidopsis seedlings, Ws-2 wild type, phyB-77, bri1-5, and phyB-77 bri1-5 double. Using the list of genes filtered through the criteria of p 1.5, we made three comparisons: phyB-77 vs. Ws-2; bri1-5vs.Ws-2; and phyB-77 bri1-5vs. Ws-2. Comparisons enabled us to find list of genes under epistatic interactions; bri1-5was epistatic to phyB-77. The genes were further validated through quantitative RT-PCR and transgenic analysis. When At5g53870 was over expressed in the bri1-5 mutant, the dwarf phenotype of bri1-5 was suppressed. Furthermore, we expressed two Peroxidases (PRX2 and PRX73) that were down regulated in phyB-77 but were up regulated in bri1-5 in the antisense orientation; both PRX2 RNAi and PRX73RNAi transgenic plants exhibited elongated hypocotyls and the expression of the target genes was greatly reduced. Our transcriptome analysis followed by qRT-PCR and transgenic expression identified the genes that are critical in light-dependent growth of Arabidopsis.C disease (CD) and ulcerative colitis (UC) are inflammatory bowel diseases (IBD) characterized by chronic and relapsing inflammation of the gastro-intestinal tract (GIT). They cause lifelong suffering, as well as considerable drainage of health care resources. Although their aetiology is still unclear there is a growing body of evidence for a significant microbial factor. Previous research in this area has focused on examining the microbiota composition in stool and in the GIT producing conflicting and inconclusive results. Here, we adapt a new approach and focus on the meta-transcriptome through RNA sequencing of colonic biopsies. Biopsies were collected from inflamed and non-inflamed colonic mucosa from 6 CD and 12 CD patients and sequenced using RNA-Seq using Illumina HiSeq at 15Gb/sample. Raw reads were quality filtered and trimmed using Trimmomatic before aligning to the human genome (hg20) with STAR. The SILVA database along with Bowtie2 was used for identifying and removing rRNA sequences. Remaining reads were aligned using Bowtie2 against a non-redundant gene catalogue constructed from multiple previously published meta-genomic studies of the human GIT. DESeq2 was subsequently used to analyse the count data and identify differentially expressed genes. This led to the finding of microbial genes which are significantly differentially expressed between inflamed and non-inflamed mucosa in bacterial species. Furthermore the count data from these samples show a clear distinction between bacterial gene expression of UC and CD. Thus, our analysis has revealed a clear difference in the gene expression of bacteria in the colon of IBD patients and demonstrated that novel approaches are required in order to understand complex multi-factorial diseases.Materials & Methods: This descriptive analytical study was performed on 100 adult patients (>18 years old) who were affected by cancer and referred to the hospital and the related private centers. The Convenience sampling was performed. Two standard DASS21questionnaires were given to the patients in order to evaluate the rate of their depression; anxiety and religious view standard questionnaire was given them in order to evaluate their religious beliefs. Obtained data were analyzed and investigated using Chi square test and correlation coefficient. Findings among 100 patients, 56 patients were female and 44 patients were male. 23% of the patients were older than 60 years old. 45% were uneducated, 7% elementary educated, 36% guidance school and high school and 12% were under graduated from university. 78% were married, 10% were single and 12% widow. DASS-21 questionnaire indicated rate of depression equal to 36% (24% slight and 12% mean) and rate of anxiety were shown equal to 42% in the under studied patients (14% slight, 18% mean and 10% severe). Also rate of depression, anxiety and religious beliefs of the under studied patients were measured separated from the variables of gender, age, education level and marital statue.R is a common strategy to treat cancer patients. In this case, a high dose (e.g. 80 Gy) of ionizing radiation is often delivered in fractions of 2-3 Gy. However, natural or induced solar radiation can cause deleterious effects to individuals exposed to it at certain conditions of time, dose and distance. The risk/benefit ratio of received ionizing or non-ionizing radiation could be estimated from the expression of key biomarkers involved in the radiation response. The identification and characterization of these biomarkers have been performed from MCF-7 breast cancer cells as well as from biopsies of patients with metastatic breast cancers. SSH and then microarrays has been performed to identify radio-modulated genes. Messenger RNA and total protein have been isolated to performed qPCR and Western blots, respectively. The signalling pathways of new key genes have been unravelled.At present, the exact mechanism and biological basis of aging is unknown. The antagonistic pleiotropy theory and competing model of senescence explain the aging occurrence of some physiological functions of human body from different angles. The single nucleotide polymorphisms (SNPs) of genome suggest the individual’s difference of aging process is associated with SNPs-related individual’s sensitivity to diseases. As the naturally occurring DNA damages and age-associated accumulation of DNA damage as well as decline in gene expression regulation indicate individual life expectancy, whereas the shortening mechanism of telomeric terminals of chromosomes and Hayflick limit of cell division determine human life span.

In silico analysis of different Helicobacter pylori strains and its proteins responsible for gastro-intestinal tract diseases in human

The potential of magnetic nanoparticles (MNP) for targeted therapy, as well as for diagnostic purposes, has been extensively explored both experimentally and clinically. MNP typically used for therapeutic applications are synthetic biodegradable particles in the size range of 50-500 nm with magnetic responsiveness imparted by nanocrystalline iron oxide and the capacity to carry a therapeutic payload either in the bulk of the particle or attached to the MNP surface. MNP formulated in this size range and targeted to steel stents magnetized by a uniform field provide a novel approach for in-stent restenosis treatment that can be readily adapted for clinical use. MNP targeting can address a number of therapeutic challenges facing vascular disease therapy, such as achieving a clinically relevant local effect in the absence of systemic toxicity, extending the drug presence at therapeutically adequate levels at the site of arterial injury with minimal redistribution to non-target tissues, and adjusting the drug dose to the disease status of a treated blood vessel. We recently showed the feasibility of a novel two-source magnetic targeting scheme for guiding therapeutic agents to injured arteries implanted with a magnetizable stent. In this approach a stent is used as the physical platform for targeted delivery due to its ability to focus the magnetic force in its vicinity and attract therapeutic agents configured in magnetically responsive particles to the site of stent implantation. Notably, this targeted delivery scheme is potentially applicable to deep blood vessels and is not restricted to superficial regions, does not require the use of permanently magnetic materials (thus obviating safety concerns), and can be realized using magnetic field intensities and sources that are currently widely used in the clinic (e.g. MRI scanners and magnetic navigation systems). The effectiveness of this strategy was shown in a rat model of carotid stenting, where biodegradable polymer-based magnetic nanoparticles loaded with paclitaxel effectively inhibited neointima formation compared with non-magnetically treated control animals. Targeted delivery of therapeutic agents configured in magnetically responsive nanocarriers using the two-source guidance strategy is a promising experimental direction that has a great potential in vascular disease therapy.An ultrasensitive method for early stage detection of Salmonella is demonstrated using total internal reflection fluorescence (TIRF) microscopy. Salmonella is captured from the milliliters volumes of spiked serum samples using fluorescent magnetic nanoparticles conjugated with anti-Salmonella. The labeled cells are separated from the unbound nanoparticles by simple gravity-based settling in confined volumes, circumventing the use of bulky ultracentrifuges. Thus the fluorescent magnetic nanoparticle conjugates serves both as pathogen preconcentrators and biosensing reagents. The fluorescently-labeled cells are selectively imaged at the bottom of the substrate by evanescent wave illumination using a TIRF microscope. This technique allows rapid visualization of both cultivable and non-cultivable (dead) bacteria unaffected by the antimicrobial properties of fresh blood and the presence of antimicrobial agent in the blood (in case of pretreatment with antibiotics). The current known methods for enteric fever diagnosis using cell culture often have low sensitivity and specificity, and long turnaround times. Thus, the proposed approach could become a promising approach in clinical applications, unlocking many biomedical opportunities.P and siRNAs are likely to be part of future therapeutic arsenals for neurological and oncological diseases. However, effective delivery of these agents represents a major obstacle to their development as pharmaceutical productsproducts. We have developed a non-toxic water-in-oil microemulsion technology, called Aonys®, for the delivery of such agents through buccal and/or rectal mucosa. The mechanism of delivery can be summarized as follows: i) Aonys®-formulated active ingredient is deposited on buccal or rectal mucosa; ii) absorption of the active ingredient through mucosa and combination with apolipoproteins ; iii) secretion into the lymphatic system; iv) systemic circulation of the active ingredient incorporated into VHDL/HDL lipoproteins ; v) cellular uptake via the SR-B1 type lipoprotein receptor. Recent applications of this technology will be described. Peptide 42 (P42) directly interacts with the N17 domain of polyQ-huntingtin implicated in neurodegenerative Huntington’s disease (HD) and inhibits aggregate formation. We evaluated the therapeutic effect of P42 in the R6/2 mice, a model of HD. Presymptomatic treatment of 2-11 week-old animals via buccal and rectal mucosa was performed daily with P42-Aonys® or an empty vector. Several HD-behavioural associated defects (rotarod and foot-clasping time), body weight, and brain section markers were analysed. The data globally show the protective effect of P42 as a presymptomatic treatment. A cDNA construction encoding a mutated (oncogenic) human Cyclin D1 (hCCND1) was transfected in mouse fibroblasts and these cells were transplanted in the flank of nude mice. After tumors reached 0.2 cm diameter, different siRNAs targeting hCCND1 mRNA, scrambled siRNAs, or the empty vector were administered every day for 3 weeks via the rectal mucosa using Aonys®. While tumor size increased regularly with time in control animals (empty vector or scrambled siRNA), the increase was strongly reduced in animals receiving hCCND1-specific siRNAs. Interestingly, after 3 weeks, tumor size increased in animals in which the hCCND1-siRNA treatment was changed to empty vector or scrambled siRNA, and regressed in animals in which the empty vector or scrambled siRNA treatment was changed to hCCND1 siRNA. These data were confirmed by western blot analysis of CCND1 protein in the tumors. These data support the use of Aonys® technology for the in vivo systemic delivery of peptides and non modified siRNAs.Abuse of prescription drugs for nonmedical purposes is a serious worldwide public health issue. Abusers have died when mechanically tampering (e.g. crushing, chewing, grating, or grinding) with medications in an attempt to quickly extract drug from the dosage form. One approach to combat this abuse is to design more resilient dosage forms, often referred to as abuse-deterrent, which need to be evaluated for their deterrence potential through invitro testing. Our objective was to provide a practical way to evaluate the extent of abuse by mechanical manipulation of a dosage form in its solid state. Crushing the dosage form generates particles of different size distribution, which would affect the air-flow property of the crushed particles. The larger the crushed particles are, the less favorable their flow would be. Since abusers have easy access to different household items such as low shear (spoons), or high shear (coffee grinder) mechanical tools, the size distribution of the crushed dosage form may vary. This variation can affect insufflation characteristics and alter the rate and extent of drug absorption. In general, an abuse-deterrent dosage form, if claimed as such, should have very limited flow after being crushed into particles. Flow properties of the crushed dosage form can help the manufacturer evaluate the deterrence potential of their medication. Additionally, this information may help support claims of abuse-deterrence to the FDA for product labeling purposes.Purpose: To evaluate the antiangiogenic effect of individual and mixed paclitaxel (PTX) and rapamycin (RAP) micelles in vitro. Methods: RAP or PTX were conjugated to poly (ethyleneglycol)‐block‐poly(aspartate‐hydrazide) to form PEG‐RAP and PEG‐ PTX and characterized by NMR and GPC. Drug conjugates were assembled into individual and mixed micelles. Physically loaded RAP and PTX micelles were also formed. PTX was loaded into PEG2000‐b‐PLA1800 while RAP or PTX: RAP (1:1) was loaded into PEG4000‐b‐PLA2000. Micelle size was characterized by DLS and loading by RP‐HPLC. The antiangiogenic effect of individual and mixed micelles was evaluated in invitro Human Umbilical Vein Endothelial Cells (HUVEC) through proliferation, tube formation and migration assays. Results: PEG‐RAP and PEG‐PTX were synthesized and characterized by GPC and NMR with individual and mixed micelle sizes between 40‐100nm. PEG‐RAP and PEG‐PTX individual micelles IC50s were 12 nM and 2 nM respectively. Mixed micelles showed synergistic effect in inhibiting the HUVEC cell proliferation. Individual and mixed micelles inhibited HUVEC tube formation and migration with the mixed micelles showing a synergistic effect. Physically loaded PTX and RAP micelles were sized at 36.5 and 33.6 nm respectively. PTX and RAP individual micelles had IC50s of 6.3 nM and 14,051 nM respectively. A synergistic response was observed with PTX: RAP 1:1 micelles. In vitro tube formation was significantly inhibited with different concentrations of PTX and RAP in micelles in comparison to individual micelles. Conclusions: PTX and RAP together, as drug conjugates or physically loaded micelles, demonstrate synergistic antiangiogenic activity.T aim of this work was to develop a gastric retentive multiple dosing pulsatile platform delivery system. Delivery of the APIs ciprofloxacin and verapamil from the multiple platforms was investigated, for potential treatment of infections or cardiovascular disease, respectively. The release rates of the API containing capsules (n=6) were determined using a USP type II apparatus (Hanson SR-PlusTM Dissolution Test Station. Hanson Research Corp. Chatsworth. CA) at a paddle speed of 50 rpni in 900 mL of a 0.1N HC1 buffer solution (pH 1.2) at 37=0.5 °C. The dissolution data obtained were plotted as percent cumulative drug released versus time. The time of 50% drug release of pulse release (T50%) was also calculated by extrapolation on the time axis of each individual release curve. Average and standard deviations for T50% were determined. Ciprofloxacin capsules displayed two T50% pulse release times of 0.3 and 12.8 homs. with the capsule demonstrating immediate floatation. Verapamil capsules showed three T50% pulse release times of 0.3. 8.5. and 17 hours, also with the capsule demonstrating immediate floatation. From this in vitro study, two-pulse release of ciprofloxacin and three-pulse release of verapamil aiming to mimic twice-daily and thricedaily dosing regimens respectively were demonstrated. Implications for the use of the multiple dose platforms are discussed and compared with current multiple-daily and once-daily dosing regimens for both of drugs. By adjusting the time of administration and pulse time to the circadian pattern, the multiple pulse platforms offer a promising direction for chronotherapeutics.S stabilized micelles (SSM) composed of PEGylated phospholipids can be used as carriers for water insoluble small molecules and peptide drugs for targeted delivery to site of action. These micelles are very effective nano-carriers for parenteral applications and promising for transition to the clinics, due to multiple reasons: First, the PEGylated lipid used in SSM composition is already approved in another pharmaceutical product by FDA for human use, so safety concern is minimum. Second, their reproducible and industrial scale preparations are easy, since SSM are formed by self-assembly of PEGylated lipids in aqueous media and they are thermodynamically stable. Third, the final product can be freezedried without any use of cryoand lyo-protectants and stored in dry form for required shelf-life. Fourth, SSM have very low CMC therefore after dilutions in blood, the number of micelles break is not significant. Since monomers in equilibrium with SSM are low, in vivo lipid toxicity should be also low. Sixth, PEG on the surface of the micelle forms a protective layer and avoids the opsonization of the particles and RES uptake. Seventh, PEG can be used to attach active targeting ligand. Last, but not least, the size of SSM (~15nm) is ideal for passive targeting to leaky vasculature by EPR effect, at the same time SSM size is too big to extravasete at the normal vasculature and for renal clearance. My talk will show some data on in-vitro characterization of SSM, and also in-vivo performance of SSM as a drug delivery system on cancer and rheumatoid arthritis animal models.E pain management requires prolonged delivery of the non-narcotic analgesic ketorolac tromethamine (KTM). KTM is having a short biological half life of about 5-7 h and frequent dosing is required in the form of conventional tablets. Keeping the above facts in mind, we have made an attempt to develop a new stomach specific floating in situ gelling system of the drug using pectin as a natural, biodegradable and gelling polymer. A 1%,1.5%, 2% (w/v) sols of pectin containing calcium carbonate and drug were prepared and characterized for rheological behavior, floating lag time (FLT), duration of floating (DOF), physical appearance, in vitro drug release studies and ex vivo studies. Viscosity of the sols was determined at different shear rates and they exhibited shear thinning property of Newtonian systems. In vitro buoyancy of in situ formed gel was determined for the FLT, DOF. FLT of 23 sec and DOF of 24 h were observed with 2 % pectin. The effect of polymer concentration on in vitro drug release from in situ gels was investigated and found that there was a significant decrease in the rate and extent of drug release with increasing concentration. Visual observation of the contents, following administration of 1 ml sols of 2% (w/v) pectin and a marker dye (Aniline blue water soluble dye ) was administered to rat and observed for the presence of gel in rat stomach. The presence of any drug-polymer interaction was studied by FTIR spectroscopy.We evaluated the potential of a histone methylation reversal agent 3-deazaneplanocin A (DZNep) in improving the chemosensitivity of pancreatic cancer to nucleoside analogs (i.e., gemcitabine). DZNep brought delayed but selective cytotoxicity to pancreatic cancer cells without affecting normal human pancreatic ductal epithelial cells. Coexposure of DZNep and gemcitabine induced cytotoxic additivity or synergism in both welland poorly-differentiated pancreatic cell lines. In contrast, DZNep exerted antagonism with gemcitabine against HPDE cells with significant reduction in cytotoxicity compared with the gemcitabine-alone regimen. DZNep marginally depended on purine nucleoside transporters for its cytotoxicity, but the transport dependence was circumvented by acyl derivatization. Drug exposure studies revealed that a short priming with DZNep followed by gemcitabine treatment rather than co-treatment of both agents to produce a maximal chemosensitization response in both gemcitabine-sensitive and gemcitabine-resistant pancreatic cancer cells. DZNep rapidly and reversibly decreased trimethylation of histone H3 lysine 27 but increased trimethylation of lysine 9 in an EZH2and JMJD1A/2C-dependent manner, respectively. However, DZNep potentiation of nucleoside analog chemosensitization was found to be temporally coupled to trimethylation changes in lysine 27 and not lysine 9. Polymeric nanoparticles engineered to chronologically release DZNep followed by gemcitabine produced pronounced chemosensitization and dose-lowering effects. Together, our results identify that an optimized DZNep exposure can presensitize pancreatic cancer cells to anticancer nucleoside analogs and emphasize the promising clinical utilities of histone methylation reversal agents’ and engineered nanoparticle approaches in future pancreatic cancer combination therapies.N have seen a growing interest in drug delivery technologies and the success of the carrier depends on the size, shape and the surface structure and chemistry of the carrier in the context of the delivery pathway. This article develops a map of guildelines for design of nanoparticle based chemotherapy. It reviews the mechanism of delivery in different pathways, physiology and chemistries involved and barriers to transport and delivery of nanocarrier based drugs, specifically for chemotherapeutic drugs. The microenvironment and physiology of a tumor site and its chemical environment is also reviewed, focusing on the impact on delivery. For each of these primary pathways including transdermal, parenteral, mucosal, and oral, nanoparticle size, method of formation and functional characteristics are defined based on the hurdles to transport and mechanisms that aid delivery. Targeting and release mechanisms with these different kinds of nanocarriers are also reviewed. Successful lab and commercially available solutions are compared to these design parameters and a map of nano-carrier design guidelines are developed and presented.N cell lung cancer (NSCLC) is the leading cause of cancer deaths in the United States and the world with a 5 year survival rate of only 15% for all stages combined. The cure rate remains <15% despite some recent advances in chemotherapeutic agents. The size and distribution of NSCLC makes cytoreductive surgery ineffective. Consequently, chemotherapy and/or radiation have been the treatments of choice. Nevertheless, toxicity still remains a major issue with the use of chemotherapy. This toxicity is most often the result of unexpected uptake of these chemotherapeutic agents by normal tissues. The need for increased potency and selectivity in chemotherapy is therefore of high importance in order to more fully harness the benefit of this modality of cancer treatment. We have spent the last few years investigating the use of antibody functionalized nanoparticles as a means of optimizing drug delivery to cancer cells while avoiding normal cells. Nanoparticles of different sizes were functionalized with anti-HER-2 monoclonal antibody (herceptin) by covalent reaction. These nanoparticles were characterized using PCS, SEM, mass spectroscopy and immunofluorescence. Release kinetics of loaded drugs was elucidated. The ability to selectively deliver payload to cancer cells while avoiding normal was assessed in vitro using different lung cancer cells (A549, H460 and calu-3) in comparison to normal cells MRC-5. In vivo distribution of this nanoparticle formulation was assessed using lung cancer mouse models. The results of our findings from this exciting research will be presented at the conference. Sunday A. Shoyele, Med chem 2013, 3:4 http://dx.doi.org/10.4172/2161-0444.S1.009

To study percentage distribution of target genes encoding proteins of different classes in Helicobacter pylori strain J99 and identification of potential therapeutic targets to reduce its proliferation

Helicobacter pylori are one of the most common bacterial pathogens in humans whose seropositivity increases with age and low socio-economic status. Due to presence of its pathogenic-island causes chronic persistent and atrophic gastritis in adults and children that often culminate in development of gastric and duodenal ulcers. Studies indicate that infected individuals have two to sixfold increased risk of developing gastric cancer and mucosal associated lymphoid tissue lymphoma compared to their uninfected counterparts. The complete genome sequences have provided a plethora of potential drug targets. Subtractive study holds the promise of providing a conceptual framework for identification of potential drug targets and providing insights to understand the biological regulatory mechanisms in diseases, which are playing an increasingly important role in searching for novel drug targets from the information contained in genomics. In this paper, we discuss subtractive study approaches for identifying drug targets, with the emphasis on the modelling of target protein and docking of the modelled protein with probable ligand by using computational tools.

古吉拉特邦豆科家庭数据库(GLDB)：存在于印度古吉拉特邦的豆科植物生物信息学数据库

生物数据库在生物信息学中发挥着重要的作用，它们为科学家提供了集中访问各种生物数据的机会。目前，分子数据可用于对许多不同植物物种进行分类、进化和情缘关系分析。我们尝试为豆科家族物种生成初步的生物信息学数据，尽可能在一种平台上提供特定类型的信息。总之，我们做了一个包括在印度的古杰雷特州发现的豆科家族所有信息的数据库，包含每个物种的植物学信息和生物信息学信息，并在一个平台上进行分析。创建这种类型的数据库反映出生物信息学可以在植物数据库中发挥重要作用的当代的跨学科方法。开发生物信息学数据库并将所有这些相关信息放在社会特别是科学界一个广泛的平台上，使之得到进一步延伸。