Constitutive and Treatment-Induced CXCL8-Signalling Selectively Modulates the Efficacy of Anti-Metabolite Therapeutics in Metastatic Prostate Cancer

Background: The current study was undertaken to characterize the effect of anti-metabolites on inducing CXCL8 signaling and determining whether the constitutive and/or drug-induced CXCL8 signaling in metastatic prostate cancer (CaP) cells modulates their sensitivity to this class of agent.

Methods: The response of metastatic CaP cells to 5-Fluorouracil (5-FU), Pemetrexed or Tomudex was determined using cell count assays, flow cytometry and PARP cleavage analysis. Quantitative-PCR, ELISA and immunoblots were employed to determine effects of drugs or CXCL8 administration on target gene/protein expression.

Results: Administration of 5-FU but not pemetrexed potentiated CXCL8 secretion and increased CXCR1 and CXCR2 gene expression in metastatic PC3 cells. Consistent with this, the inhibition of CXCL8 signaling using a CXCR2 antagonist, AZ10397767, increased the cytotoxicity of 5-FU by 4-fold (P,0.001), and increased 5-FU-induced apoptosis in PC3 cells (P,0.01). In contrast, while administration of AZ10397767 had no effect on the sensitivity of pemetrexed, the CXCR2 antagonist exerted the greatest effect in increasing the sensitivity of PC3 cells to Tomudex, a directed thymidylate synthase (TS) inhibitor. Subsequent experiments confirmed that administration of recombinant human CXCL8 increased TS expression, a response mediated in part by the CXCR2 receptor. Moreover, siRNA-mediated knockdown of the CXCL8-target gene Bcl-2 increased the sensitivity of PC3 cells to 5-FU.

Conclusions: CXCL8 signaling provides a selective resistance of metastatic prostate cancer cells to specific anti-metabolites by promoting a target-associated resistance, in addition to underpinning an evasion of treatment-induced apoptosis. © 2012 Wilson et al.

Optimizing the balance between false positive and false negative error probabilities of confirmatory methods for the detection of veterinary drug residues

GC-MS data on veterinary drug residues in bovine urine are used for controlling the illegal practice of fattening cattle. According to current detection criteria, peak patterns of preferably four ions should agree within 10 or 20% from a corresponding standard pattern. These criteria are rigid, rather arbitrary and do not match daily practice. A new model, based on multivariate modeling of log peak abundance ratios, provides a theoretical basis for the identification of analytes and optimizes the balance between the avoidance of false positives and false negatives. The performance of the model is demonstrated on data provided by five laboratories, each supplying GC-MS measurements on the detection of clenbuterol, dienestrol and 19 beta-nortestosterone in urine. The proposed model shows a better performance than confirmation by using the current criteria and provides a statistical basis for inspection criteria in terms of error probabilities.

Stability during cooking of anthelmintic veterinary drug residues in beef

Anthelmintic drugs are widely used for treatment of parasitic worms in livestock, but little is known about the stability of their residues in food under conventional cooking conditions. As part of the European Commissionfunded research project ProSafeBeef, cattle were medicated with commercially available anthelmintic preparations, comprising 11 active ingredients (corresponding to 21 marker residues). Incurred meat and liver were cooked by roasting (40 min at 190°C) or shallow frying (muscle 8-12 min, liver 14-19 min) in a domestic kitchen. Raw and cooked tissues and expressed juices were analysed using a novel multi-residue dispersive solid-phase extraction method (QuEChERS) coupled with ultra-performance liquid chromatography-tandem mass spectrometry. After correction for sample weight changes during cooking, no major losses were observed for residues of oxyclozanide, clorsulon, closantel, ivermectin, albendazole, mebendazole or fenbendazole. However, significant losses were observed for nitroxynil (78% in fried muscle, 96% in roast muscle), levamisole (11% in fried muscle, 42% in fried liver), rafoxanide (17% in fried muscle, 18% in roast muscle) and triclabendazole (23% in fried liver, 47% in roast muscle). Migration of residues from muscle into expressed cooking juices varied between drugs, constituting 0% to 17% (levamisole) of total residues remaining after cooking. With the exception of nitroxynil, residues of anthelmintic drugs were generally resistant to degradation during roasting and shallow frying. Conventional cooking cannot, therefore, be considered a safeguard against ingestion of residues of anthelmintic veterinary drugs in beef. © 2011 Taylor & Francis.

The effects of climate change on ovine parasitic gastroenteritis determined using veterinary surveillance and meteorological data for Northern Ireland over the period 1999-2009

While the influence of temperature and moisture on the free-living stages of gastrointestinal nematodes have been described in detail, and evidence for global climate change is mounting, there have been only a few attempts to relate altered incidence or seasonal patterns of disease to climate change. Studies of this type have been completed for England Scotland and Wales, but not for Northern Ireland (NI). Here we present an analysis of veterinary diagnostic data that relates three categories of gastrointestinal nematode infection in sheep to historical meteorological data for NI. The infections are: trichostrongylosis/teladorsagiosis (Teladorsagia/Trichostrongylus), strongyloidosis and nematodirosis. This study aims to provide a baseline for future climate change analyses and to provide basic information for the development of nematode control programmes. After identifying and evaluating possible sources of bias, climate change was found to be the most likely explanation for the observed patterns of change in parasite epidemiology, although other hypotheses could not be refuted. Seasonal rates of diagnosis showed a uniform year-round distribution for Teladorsagia and Trichostrongylus infections, suggesting consistent levels of larval survival throughout the year and extension of the traditionally expected seasonal transmission windows. Nematodirosis showed a higher level of autumn than Spring infection, suggesting that suitable conditions for egg and larval development occurred after the Spring infection period. Differences between regions within the Province were shown for strongyloidosis, with peaks of infection falling in the period September-November. For all three-infection categories (trichostrongylosis/teladorsagiosis, strongyloidosis and nematodirosis), significant differences in the rates of diagnosis, and in the seasonality of disease, were identified between regions. (C) 2012 Elsevier B.V. All rights reserved.

Connectivity Mapping for Candidate Therapeutics Identification Using Next Generation Sequencing RNA-Seq Data

The advent of next generation sequencing technologies (NGS) has expanded the area of genomic research, offering high coverage and increased sensitivity over older microarray platforms. Although the current cost of next generation sequencing is still exceeding that of microarray approaches, the rapid advances in NGS will likely make it the platform of choice for future research in differential gene expression. Connectivity mapping is a procedure for examining the connections among diseases, genes and drugs by differential gene expression initially based on microarray technology, with which a large collection of compound-induced reference gene expression profiles have been accumulated. In this work, we aim to test the feasibility of incorporating NGS RNA-Seq data into the current connectivity mapping framework by utilizing the microarray based reference profiles and the construction of a differentially expressed gene signature from a NGS dataset. This would allow for the establishment of connections between the NGS gene signature and those microarray reference profiles, alleviating the associated incurring cost of re-creating drug profiles with NGS technology. We examined the connectivity mapping approach on a publicly available NGS dataset with androgen stimulation of LNCaP cells in order to extract candidate compounds that could inhibit the proliferative phenotype of LNCaP cells and to elucidate their potential in a laboratory setting. In addition, we also analyzed an independent microarray dataset of similar experimental settings. We found a high level of concordance between the top compounds identified using the gene signatures from the two datasets. The nicotine derivative cotinine was returned as the top candidate among the overlapping compounds with potential to suppress this proliferative phenotype. Subsequent lab experiments validated this connectivity mapping hit, showing that cotinine inhibits cell proliferation in an androgen dependent manner. Thus the results in this study suggest a promising prospect of integrating NGS data with connectivity mapping. © 2013 McArt et al.

A Trip from a Tube to a Chip Applied Micro and Nanotechnology in Biotechnology, Veterinary and Life Sciences

More than 200 known diseases are transmitted via foods or food products. In the United States, food-borne diseases are responsible for 76 million cases of illness, 32,500 cases of hospitalisation and 5000 cases of death yearly. The ongoing increase in worldwide trade in livestock, food, and food products in combination with increase in human mobility (business- and leisure travel, emigration etc.) will increase the risk of emergence and spreading of such pathogens. There is therefore an urgent need for development of rapid, efficient and reliable methods for detection and identification of such pathogens.

Microchipfabrication has had a major impact on electronics and is expected to have an equally pronounced effect on life sciences. By combining micro-fluidics with micromechanics, micro-optics, and microelectronics, systems can be realized to perform complete chemical or biochemical analyses. These socalled ’Lab-on-a-Chip’ will completely change the face of laboratories in the future where smaller, fully automated devices will be able to perform assays faster, more accurately, and at a lower cost than equipment of today. A general introduction of food safety and applied micro-nanotechnology in life sciences will be given. In addition, examples of DNA micro arrays, micro fabricated integrated PCR chips and total integrated lab-on-achip systems from different National and EU research projects being carried out at the Laboratory of Applied Micro- Nanotechnology (LAMINATE) group at the National Veterinary Institute (DTU-Vet) Technical University of Denmark and the BioLabchip group at the Department of Micro and Nanotechnology (DTU-Nanotech), Technical University of Denmark (DTU), Ikerlan-IK4 (Spain) and other 16 partners from different European countries will be presented.

Potential impacts of climate change on veterinary medicinal residues in livestock produce: An island of Ireland perspective

Residues of veterinary medicines are a food safety issue regulated by European legislation. The occurrence of animal diseases necessitating application of veterinary medicines is significantly affected by global and local climate changes. This review assesses potential impacts of climate change on residues in food produced on the island of Ireland. Use of various classes of veterinary drugs in light of predicted local climate change is reviewed with particular emphasis on anthelmintic drugs and consideration is given to residues accumulating in the environment. Veterinary medicine use is predicted to increase as disease burdens increase due to varied climate effects. Locally relevant mitigation and adaptation strategies are suggested to ensure climate change does not adversely affect food safety via increasing drug residues.

Radiosensitising Nanoparticles as Novel Cancer Therapeutics — Pipe Dream or Realistic Prospect?

The field of high atomic number nanoparticle radiosensitising agents is reviewed. After a brief discussion of the new mode of physicochemical action implied by irradiation of high atomic number nanoparticles embedded in biological systems, a series of exemplars are discussed. Silver-, gadolinium- and gold-based nanoparticles are discussed in order of increasing atomic number with functionalisation strategies being outlined. In vitro and in vivo evidence for radio-enhancement and the mechanisms attributed to the increased biological effect are discussed.

RALA-mediated delivery of FKBPL nucleic acid therapeutics

Aims: RALA is a novel 30 mer bioinspired amphipathic peptide that is showing promise for gene delivery. Here, we used RALA to deliver the FK506-binding protein like – FKBPL gene (pFKBPL) – a novel member of the immunophilin protein family. FKBPL is a secreted protein, with overexpression shown to inhibit angiogenesis, tumor growth and stemness, through a variety of intra- and extracellular signaling mechanisms. We also elucidated proangiogenic activity and stemness after utilizing RALA to deliver siRNA (siFKBPL). Materials & methods: The RALA/pFKBPL and RALA/siFKBPL nanoparticles were characterized in terms of size, charge, stability and toxicity. Overexpression and knockdown of FKBPL was assessed in vitro and in vivo. Results: RALA delivered both pFKBPL and siFKBPL with less cytotoxicity than commercially available counterparts. In vivo, RALA/pFKBPL delivery retarded tumor growth, and prolonged survival with an associated decrease in angiogenesis, while RALA/siFKBPL had no effect on tumor growth rate or survival, but resulted in an increase in angiogenesis and stemness. Conclusion: RALA is an effective delivery system for both FKBPL DNA and RNAi and highlights an alternative therapeutic approach to harnessing FKBPL's antiangiogenic and antistemness activity.

QUADrATiC: scalable gene expression connectivity mapping for repurposing FDA-approved therapeutics

Background: Gene expression connectivity mapping has proven to be a powerful and flexible tool for research. Its application has been shown in a broad range of research topics, most commonly as a means of identifying potential small molecule compounds, which may be further investigated as candidates for repurposing to treat diseases. The public release of voluminous data from the Library of Integrated Cellular Signatures (LINCS) programme further enhanced the utilities and potentials of gene expression connectivity mapping in biomedicine. Results: We describe QUADrATiC (http://go.qub.ac.uk/QUADrATiC), a user-friendly tool for the exploration of gene expression connectivity on the subset of the LINCS data set corresponding to FDA-approved small molecule compounds. It enables the identification of compounds for repurposing therapeutic potentials. The software is designed to cope with the increased volume of data over existing tools, by taking advantage of multicore computing architectures to provide a scalable solution, which may be installed and operated on a range of computers, from laptops to servers. This scalability is provided by the use of the modern concurrent programming paradigm provided by the Akka framework. The QUADrATiC Graphical User Interface (GUI) has been developed using advanced Javascript frameworks, providing novel visualization capabilities for further analysis of connections. There is also a web services interface, allowing integration with other programs or scripts.Conclusions: QUADrATiC has been shown to provide an improvement over existing connectivity map software, in terms of scope (based on the LINCS data set), applicability (using FDA-approved compounds), usability and speed. It offers potential to biological researchers to analyze transcriptional data and generate potential therapeutics for focussed study in the lab. QUADrATiC represents a step change in the process of investigating gene expression connectivity and provides more biologically-relevant results than previous alternative solutions.

A gene-signature progression approach to identifying candidate small-molecule cancer therapeutics with connectivity mapping

Gene expression connectivity mapping has gained much popularity recently with a number of successful applications in biomedical research testifying its utility and promise. Previously methodological research in connectivity mapping mainly focused on two of the key components in the framework, namely, the reference gene expression profiles and the connectivity mapping algorithms. The other key component in this framework, the query gene signature, has been left to users to construct without much consensus on how this should be done, albeit it has been an issue most relevant to end users. As a key input to the connectivity mapping process, gene signature is crucially important in returning biologically meaningful and relevant results. This paper intends to formulate a standardized procedure for constructing high quality gene signatures from a user’s perspective.