PSP_MCSVM: brainstorming consensus prediction of protein secondary structures using two-stage multiclass support vector machines

Secondary structure prediction is a crucial task for understanding the variety of protein structures and performed biological functions. Prediction of secondary structures for new proteins using their amino acid sequences is of fundamental importance in bioinformatics. We propose a novel technique to predict protein secondary structures based on position-specific scoring matrices (PSSMs) and physico-chemical properties of amino acids. It is a two stage approach involving multiclass support vector machines (SVMs) as classifiers for three different structural conformations, viz., helix, sheet and coil. In the first stage, PSSMs obtained from PSI-BLAST and five specially selected physicochemical properties of amino acids are fed into SVMs as features for sequence-to-structure prediction. Confidence values for forming helix, sheet and coil that are obtained from the first stage SVM are then used in the second stage SVM for performing structure-to-structure prediction. The two-stage cascaded classifiers (PSP_MCSVM) are trained with proteins from RS126 dataset. The classifiers are finally tested on target proteins of critical assessment of protein structure prediction experiment-9 (CASP9). PSP_MCSVM with brainstorming consensus procedure performs better than the prediction servers like Predator, DSC, SIMPA96, for randomly selected proteins from CASP9 targets. The overall performance is found to be comparable with the current state-of-the art. PSP_MCSVM source code, train-test datasets and supplementary files are available freely in public domain at: http://sysbio.icm.edu.pl/secstruct and http://code.google.com/p/cmater-bioinfo

A whole cell pathway screen reveals seven novel chemosensitizers to combat chloroquine resistant malaria

Due to the widespread prevalence of resistant parasites, chloroquine (CQ) was removed from front-line antimalarial chemotherapy in the 1990s despite its initial promise of disease eradication. Since then, resistance-conferring mutations have been identified in transporters such as the PfCRT, that allow for the efflux of CQ from its primary site of action, the parasite digestive vacuole. Chemosensitizing/ chemoreversing compounds interfere with the function of these transporters thereby sensitizing parasites to CQ once again. However, compounds identified thus far have disappointing in vivo efficacy and screening for alternative candidates is required to revive this strategy. In this study, we propose a simple and direct means to rapidly screen for such compounds using a fluorescent-tagged CQ molecule. When this screen was applied to a small library, seven novel chemosensitizers (octoclothepin, methiothepin, metergoline, loperamide, chlorprothixene, L-703,606 and mibefradil) were quickly elucidated, including two which showed greater potency than the classical chemosensitizers verapamil and desipramine

Next generation transcriptomes for next generation genomes using est2assembly

<p>Abstract</p> <p>Background</p> <p>The decreasing costs of capillary-based Sanger sequencing and next generation technologies, such as 454 pyrosequencing, have prompted an explosion of transcriptome projects in non-model species, where even shallow sequencing of transcriptomes can now be used to examine a range of research questions. This rapid growth in data has outstripped the ability of researchers working on non-model species to analyze and mine transcriptome data efficiently.</p> <p>Results</p> <p>Here we present a semi-automated platform '<it>est2assembly</it>' that processes raw sequence data from Sanger or 454 sequencing into a hybrid <it>de-novo </it>assembly, annotates it and produces GMOD compatible output, including a SeqFeature database suitable for GBrowse. Users are able to parameterize assembler variables, judge assembly quality and determine the optimal assembly for their specific needs. We used <it>est2assembly </it>to process <it>Drosophila </it>and <it>Bicyclus </it>public Sanger EST data and then compared them to published 454 data as well as eight new insect transcriptome collections.</p> <p>Conclusions</p> <p>Analysis of such a wide variety of data allows us to understand how these new technologies can assist EST project design. We determine that assembler parameterization is as essential as standardized methods to judge the output of ESTs projects. Further, even shallow sequencing using 454 produces sufficient data to be of wide use to the community. <it>est2assembly </it>is an important tool to assist manual curation for gene models, an important resource in their own right but especially for species which are due to acquire a genome project using Next Generation Sequencing.</p

Cloning and characterization of a 9-lipoxygenase gene induced by pathogen attack from Nicotiana benthamiana for biotechnological application

<p>Abstract</p> <p>Background</p> <p>Plant lipoxygenases (LOXs) have been proposed to form biologically active compounds both during normal developmental stages such as germination or growth as well as during responses to environmental stress such as wounding or pathogen attack. In our previous study, we found that enzyme activity of endogenous 9-LOX in <it>Nicotiana benthamiana </it>was highly induced by agroinfiltration using a tobacco mosaic virus (TMV) based vector system.</p> <p>Results</p> <p>A <it>LOX </it>gene which is expressed after treatment of the viral vectors was isolated from <it>Nicotiana benthamiana</it>. As the encoded LOX has a high amino acid identity to other 9-LOX proteins, the gene was named as <it>Nb-9-LOX</it>. It was heterologously expressed in yeast cells and its enzymatic activity was characterized. The yeast cells expressed large quantities of stable 9-LOX (0.9 U ml^-1cell cultures) which can oxygenate linoleic acid resulting in high yields (18 μmol ml^-1cell cultures) of hydroperoxy fatty acid. The product specificity of Nb-9-LOX was examined by incubation of linoleic acid and Nb-9-LOX in combination with a 13-hydroperoxide lyase from watermelon (Cl-13-HPL) or a 9/13-hydroperoxide lyase from melon (Cm-9/13-HPL) and by LC-MS analysis. The result showed that Nb-9-LOX possesses both 9- and 13-LOX specificity, with high predominance for the 9-LOX function. The combination of recombinant Nb-9-LOX and recombinant Cm-9/13-HPL produced large amounts of C₉-aldehydes (3.3 μmol mg^-1crude protein). The yield of C₉-aldehydes from linoleic acid was 64%.</p> <p>Conclusion</p> <p>The yeast expressed Nb-9-LOX can be used to produce C₉-aldehydes on a large scale in combination with a <it>HPL </it>gene with 9-HPL function, or to effectively produce 9-hydroxy-10(<it>E</it>),12(<it>Z</it>)-octadecadienoic acid in a biocatalytic process in combination with cysteine as a mild reducing agent.</p

A novel method, digital genome scanning detects KRAS gene amplification in gastric cancers: involvement of overexpressed wild-type KRAS in downstream signaling and cancer cell growth

<p>Abstract</p> <p>Background</p> <p>Gastric cancer is the third most common malignancy affecting the general population worldwide. Aberrant activation of KRAS is a key factor in the development of many types of tumor, however, oncogenic mutations of <it>KRAS </it>are infrequent in gastric cancer. We have developed a novel quantitative method of analysis of DNA copy number, termed digital genome scanning (DGS), which is based on the enumeration of short restriction fragments, and does not involve PCR or hybridization. In the current study, we used DGS to survey copy-number alterations in gastric cancer cells.</p> <p>Methods</p> <p>DGS of gastric cancer cell lines was performed using the sequences of 5000 to 15000 restriction fragments. We screened 20 gastric cancer cell lines and 86 primary gastric tumors for <it>KRAS </it>amplification by quantitative PCR, and investigated <it>KRAS </it>amplification at the DNA, mRNA and protein levels by mutational analysis, real-time PCR, immunoblot analysis, GTP-RAS pull-down assay and immunohistochemical analysis. The effect of <it>KRAS </it>knock-down on the activation of p44/42 MAP kinase and AKT and on cell growth were examined by immunoblot and colorimetric assay, respectively.</p> <p>Results</p> <p>DGS analysis of the HSC45 gastric cancer cell line revealed the amplification of a 500-kb region on chromosome 12p12.1, which contains the <it>KRAS </it>gene locus. Amplification of the <it>KRAS </it>locus was detected in 15% (3/20) of gastric cancer cell lines (8–18-fold amplification) and 4.7% (4/86) of primary gastric tumors (8–50-fold amplification). <it>KRAS </it>mutations were identified in two of the three cell lines in which <it>KRAS </it>was amplified, but were not detected in any of the primary tumors. Overexpression of KRAS protein correlated directly with increased <it>KRAS </it>copy number. The level of GTP-bound KRAS was elevated following serum stimulation in cells with amplified wild-type <it>KRAS</it>, but not in cells with amplified mutant <it>KRAS</it>. Knock-down of <it>KRAS </it>in gastric cancer cells that carried amplified wild-type <it>KRAS </it>resulted in the inhibition of cell growth and suppression of p44/42 MAP kinase and AKT activity.</p> <p>Conclusion</p> <p>Our study highlights the utility of DGS for identification of copy-number alterations. Using DGS, we identified <it>KRAS </it>as a gene that is amplified in human gastric cancer. We demonstrated that gene amplification likely forms the molecular basis of overactivation of KRAS in gastric cancer. Additional studies using a larger cohort of gastric cancer specimens are required to determine the diagnostic and therapeutic implications of <it>KRAS </it>amplification and overexpression.</p

Omega-3 polyunsaturated fatty acid supplementation versus placebo on vascular health, glycaemic control, and metabolic parameters in people with type 1 diabetes: a randomised controlled preliminary trial

Background: The role of omega-3 polyunsaturated fatty acids (n-3PUFA), and the potential impact of n-3PUFA supplementation, in the treatment and management of type 1 diabetes (T1D) remains unclear and controversial. Therefore, this study aimed to examine the efficacy of daily high-dose-bolus n-3PUFA supplementation on vascular health, glycaemic control, and metabolic parameters in subjects with T1D. Methods: Twenty-seven adults with T1D were recruited to a 6-month randomised, double-blind, placebo-controlled trial. Subjects received either 3.3 g/day of encapsulated n-3PUFA or encapsulated 3.0 g/day corn oil placebo (PLA) for 6-months, with follow-up at 9-months after 3-month washout. Erythrocyte fatty acid composition was determined via gas chromatography. Endpoints included inflammation-associated endothelial biomarkers (vascular cell adhesion molecule-1 [VCAM-1], intercellular adhesion molecule-1 [ICAM-1], E-selectin, P-selectin, pentraxin-3, vascular endothelial growth factor [VEGF]), and their mediator tumor necrosis factor alpha [TNFα] analysed via immunoassay, vascular structure (carotid intima-media thickness [CIMT]) and function (brachial artery flow mediated dilation [FMD]) determined via ultrasound technique, blood pressure, glycosylated haemoglobin (HbA1c), fasting plasma glucose (FPG), and postprandial metabolism. Results: Twenty subjects completed the trial in full. In the n-3PUFA group, the mean ± SD baseline n-3PUFA index of 4.93 ± 0.94% increased to 7.67 ± 1.86% (P  0.05). Conclusions: This study indicates that daily high-dose-bolus of n-3PUFA supplementation for 6-months does not improve vascular health, glucose homeostasis, or metabolic parameters in subjects with T1D. The findings from this preliminary RCT do not support the use of therapeutic n-3PUFA supplementation in the treatment and management of T1D and its associated complications. Trial Registration ISRCTN, ISRCTN40811115. Registered 27 June 2017, http://www.isrctn.com/ISRCTN40811115

Combinatorial Effect of Non-Steroidal Anti-inflammatory Drugs and NF-κB Inhibitors in Ovarian Cancer Therapy

Several epidemiological studies have correlated the use of non-steroidal anti-inflammatory drugs (NSAID) with reduced risk of ovarian cancer, the most lethal gynecological cancer, diagnosed usually in late stages of the disease. We have previously established that the pro-apoptotic cytokine melanoma differentiation associated gene-7/Interleukin-24 (mda-7/IL-24) is a crucial mediator of NSAID-induced apoptosis in prostate, breast, renal and stomach cancer cells. In this report we evaluated various structurally different NSAIDs for their efficacies to induce apoptosis and mda-7/IL-24 expression in ovarian cancer cells. While several NSAIDs induced apoptosis, Sulindac Sulfide and Diclofenac most potently induced apoptosis and reduced tumor growth. A combination of these agents results in a synergistic effect. Furthermore, mda-7/IL-24 induction by NSAIDs is essential for programmed cell death, since inhibition of mda-7/IL-24 by small interfering RNA abrogates apoptosis. mda-7/IL-24 activation leads to upregulation of growth arrest and DNA damage inducible (GADD) 45 α and γ and JNK activation. The NF-κB family of transcription factors has been implicated in ovarian cancer development. We previously established NF-κB/IκB signaling as an essential step for cell survival in cancer cells and hypothesized that targeting NF-κB could potentiate NSAID-mediated apoptosis induction in ovarian cancer cells. Indeed, combining NSAID treatment with NF-κB inhibitors led to enhanced apoptosis induction. Our results indicate that inhibition of NF-κB in combination with activation of mda-7/IL-24 expression may lead to a new combinatorial therapy for ovarian cancer

Prognostic factors in prostate cancer

Prognostic factors in organ confined prostate cancer will reflect survival after surgical radical prostatectomy. Gleason score, tumour volume, surgical margins and Ki-67 index have the most significant prognosticators. Also the origins from the transitional zone, p53 status in cancer tissue, stage, and aneuploidy have shown prognostic significance. Progression-associated features include Gleason score, stage, and capsular invasion, but PSA is also highly significant. Progression can also be predicted with biological markers (E-cadherin, microvessel density, and aneuploidy) with high level of significance. Other prognostic features of clinical or PSA-associated progression include age, IGF-1, p27, and Ki-67. In patients who were treated with radiotherapy the survival was potentially predictable with age, race and p53, but available research on other markers is limited. The most significant published survival-associated prognosticators of prostate cancer with extension outside prostate are microvessel density and total blood PSA. However, survival can potentially be predicted by other markers like androgen receptor, and Ki-67-positive cell fraction. In advanced prostate cancer nuclear morphometry and Gleason score are the most highly significant progression-associated prognosticators. In conclusion, Gleason score, capsular invasion, blood PSA, stage, and aneuploidy are the best markers of progression in organ confined disease. Other biological markers are less important. In advanced disease Gleason score and nuclear morphometry can be used as predictors of progression. Compound prognostic factors based on combinations of single prognosticators, or on gene expression profiles (tested by DNA arrays) are promising, but clinically relevant data is still lacking

Azithromycin-chloroquine and the intermittent preventive treatment of malaria in pregnancy

In the high malaria-transmission settings of sub-Saharan Africa, malaria in pregnancy is an important cause of maternal, perinatal and neonatal morbidity. Intermittent preventive treatment of malaria in pregnancy (IPTp) with sulphadoxine-pyrimethamine (SP) reduces the incidence of low birth-weight, pre-term delivery, intrauterine growth-retardation and maternal anaemia. However, the public health benefits of IPTp are declining due to SP resistance. The combination of azithromycin and chloroquine is a potential alternative to SP for IPTp. This review summarizes key in vitro and in vivo evidence of azithromycin and chloroquine activity against Plasmodium falciparum and Plasmodium vivax, as well as the anticipated secondary benefits that may result from their combined use in IPTp, including the cure and prevention of many sexually transmitted diseases. Drug costs and the necessity for external financing are discussed along with a range of issues related to drug resistance and surveillance. Several scientific and programmatic questions of interest to policymakers and programme managers are also presented that would need to be addressed before azithromycin-chloroquine could be adopted for use in IPTp