Issue 1 2009

miRNA and viral infections in vertebrates

Issue 1 2009 / 7 February 2009 /

For plants and invertebrates, RNA interference is firmly established as an important antiviral mechanism. Even before Fire, Mello, and co-workers described RNA interference (RNAi) in worms in 1998₁ it was becoming clear that plants have an RNA-dependent pathway that protects against viral infections₂. The pathway, then termed post-transcriptional gene silencing (PTGS), helps plants like tobacco recover from initial viral infections and ensures that plants are protected from subsequent infections from the same or similar viral strains₃. Subsequent studies have revealed that plant PTGS and Fire and Mello’s RNAi are identical – the triggers are short RNAs derived from long double-stranded RNAs (dsRNA)₄. Incorporated into the RNA-induced silencing complex (RISC), RISC cleaves transcripts like viral messenger RNAs (mRNAs) with antisense complementary to the short RNAs.

In vertebrates, long dsRNA triggers the protein kinase R (PKR) and interferon responses which give general RNA decay and shutdown of protein synthesis. These responses are potent antiviral defences on their own and it is therefore unclear whether RNAi has the same antiviral role in vertebrates as in plants and invertebrates₅. What is becoming clear, however, is that a different aspect of the RNAi pathway is important in many vertebrate viral infections. (more…)

Tagged with: Jorn Klein, microRNA, Norwegian University of Science and Technology, Pål Sætrom, RNA, RNAi

Article 1: the importance of sample quality for qPCR

Issue 1 2009 / 7 February 2009 /

The fluorescence-based quantitative real-time polymerase chain reaction (qPCR)_1-3, has the ability to detect and measure minute amounts of DNA in a wide range of samples extracted from numerous sources. In combination with reverse transcription (RT), the use of this technology has revolutionised life sciences, agriculture and medical research_4,5. In addition, many diagnostic applications have been developed, including microbial quantification, cancer recurrence risk assessment, gene dosage determination, identification of transgenes in genetically modified foods, and detection of extremely low copy targets for forensic investigations_6-11. The simplicity of assay design and execution, together with sensitivity and specificity have made this the method of choice for nucleic acid quantification that is reflected by the prodigious number of peer-reviewed publications reporting qPCR data. Inevitably, a corresponding number of different protocols, reagent recipes, analysis methods and reporting formats are also being used, which is resulting in the scientific literature being corrupted with publications reporting insignificant and conflicting results. The inexperienced or casual qPCR user is vulnerable to the production of inaccurate data because even assays of extremely poor quality usually yield results that are amenable to statistical manipulation. (more…)

Tagged with: qPCR, Sigma-Aldrich, Stephen Bustin, Tania Nolan

High Content Analysis approach for targeted gene silencing and probing nanoscale cell responses

Issue 1 2009 / 7 February 2009 /

High Content Analysis and Screening technologies (HCA) ushered in a new era in the biomedical research, enabling the scientists to uncover previously unknown disease mechanisms and to introduce innovative approaches to the development of the new generations of therapeutic drugs with a potential to selectively target individual genes, molecules and subcellular organelles. The quest for the novel therapeutic targets starting from thousands of candidate molecules and compounds, narrowing down to a few leads and their subsequent optimisation and practical application, which for many years has been almost an exclusive privilege of the large pharmaceutical companies, is quickly becoming a routine in the academic research environment.

Investigations into vast collections of small molecule chemicals, genome-wide siRNA libraries and large sets of functionalised nanomaterials for their potential biomedical utilisation, for instance, in addition to being prohibitively time-and labour-consuming by conventional methods, all inevitably generate an extensive output of information difficult for routine analysis and correct interpretation within a confined time frame and restricted resourses. HCA technology is perfectly designed to overcome these limitations, permitting for multiplexed quantitative “on the fly” processing of large massives of data, in addition giving an opportunity of detailed retrospective data analysis thereby providing efficient solutions even within the shortfalls of manpower and funding frequently experienced by academic research centres and which have a potential to become more common in the current economic situation worldwide. We provide here an overview of two extremely beneficial HCA application scenarios currently utilised by the researchers at the Institute of Molecular Medicine, Trinity College Dublin, for tackling extensive massives of experimental data in search of siRNA tools targeting inflammation-associated processes, and for screening the safety and intracellular distribution of nanomaterials with promising biomedical application potential. (more…)

Tagged with: Aideen Long, HCA (High Content Analysis), Trinity College Dublin, Yuri Volkov

Cellular Imaging and Analysis 2009

Issue 1 2009 / 7 February 2009 /

Date: 7 – 8 February 2009
Venue: Institute for Molecular Medicine Trinity College Dublin, College Green, Dublin 2, Ireland
Website: www.europeanpharmaceuticalevents.com

Cellular Imaging and Analysis is undoubtedly one of the hottest topics in the field of pharmaceutical research and development. Technical or diagnostic progress made almost hourly, this multidisciplinary set of challenges and opportunities is proving one of the most profoundly effective in the history of science. (more…)

Tagged with: CIA (Cellular Imaging and Analysis), Show Preview

Progress by the Proteomics Standards Initiative

Issue 1 2009 / 7 February 2009 /

There are compelling reasons for regularising the capture and description of proteomics data. Adhering to community-consensus specifications for the annotation of data sets can increase confidence in results and the conclusions drawn upon them, and supports data re-use; working with standard formats and vocabularies can raise efficiency and facilitates sophisticated approaches to data handling and analysis. The Human Proteome Organisation’s Proteomics Standards Initiative (HUPO PSI) is a standards generating body comprising diverse members of the proteomics community and related trades. It develops reporting guidelines, data formats and vocabulary terms with which to describe the components of a proteomics experiment. This article briefly explores the benefits accruing to the use of reporting standards, for academics and for those in a commercial setting; describes HUPO PSI, its products and the status quo with respect to compatible tools and databases; and closes by pulling back to consider multi-domain investigations in the life sciences.

To underpin understanding of the components and outputs of a bioscience investigation it is important to have access to both the data and the available metadata (in essence, this is ‘data about the data’ such as instrument parameters, protocol and operator details, and the provenance of the sample being analysed)_1,2. Focused, appropriate annotation simplifies the assessment of the validity of a data set, supports its comprehension when revisited or presented to others, and facilitates re-use either by the originator or third parties via public databases such as PRIDE3. The re-use of a data set increases the return on the initial investment in the work that generated it, and can reap collateral citations for its originator. Where data are particularly voluminous or complex, as is frequently the case in proteomics, such standard solutions increase in value by facilitating the design and construction of data pipelines and repositories. (more…)

Tagged with: Chris Taylor, European Bioinformatics Institute, Lennart Martens, Proteomics

Biomarkers and the tumour microenvironment

Issue 1 2009 / 7 February 2009 /

The current cost of developing a new medicine for the treatment of human disease has been estimated at $1 to $2 billion (€750-1.5 million1,2). Given progressive increases in the cost of developing new drugs, pharmaceutical companies are facing significant pressure to streamline discovery methods and increase the translational efficiency of their newly discovered compounds. (more…)

Tagged with: Biomarkers, Brett Hall, Johnson & Johnson

The 14th Annual Meeting of the Association of Biomolecular Resource Facilities – Application and Optimisation of Existing and Emerging Biotechnologies

Issue 1 2009 / 7 February 2009 /

Date: 7-10 February 2009
Venue: Memphis Cook Convention Centre, Memphis, Tennessee
Website: www.abrf2009.org

The ABRF 2009 meeting will present the latest developments in life science technologies and the use of these technologies. Leaders in the field will give presentations including scientific sessions and technical workshops on genomics, proteomics, imaging, and other technologies such as next generation sequencing, genotyping, microarrays, real-time PCR, proteomics and mass spectrometry, protein production, optical imaging, bioinformatics and bio-IT. Participants will be able to interact with an array of vendors in the exhibit hall and in exhibitor workshops. (more…)

Tagged with: ABRF, Show Preview

ITC: affinity is not everything

Issue 1 2009 / 7 February 2009 /

During the optimisation of drug candidates, improvements in affinity and selectivity play a critical role. This task is usually accomplished by establishing accurate correlations between the affinity/selectivity of different chemical scaffolds and through chemical modifications to a selected scaffold.

Isothermal titration calorimetry (ITC) is universally acknowledged as the technique that provides the most accurate measurements of binding affinity. It is the gold standard against which other techniques are compared. ITC, however, is not only able to measure binding affinity but also the magnitude of the different thermodynamic forces that determine the binding energy. Since different chemical functionalities contribute differently to the binding forces, the knowledge acquired by ITC also provides precise guidelines for optimisation of drug candidates. (more…)

Tagged with: Ernesto Freire, Johns Hopkins University, Thermal Analysis

Raman spectroscopic techniques for biotechnology and bioprocessing

Issue 1 2009 / 7 February 2009 /

Biotechnological expertise is becoming increasingly important within the pharmaceutical industry, and will play a pivotal role in the monitoring of fermentations, particularly their optimisation within the framework of Process Analytical Technologies (PAT). The ability to harness biological processes for the development of drug therapies, so called ‘biopharmaceuticals’ provides treatments that range from small molecule antibiotics to large recombinant proteins. Typically, synthesis of these drug products is enabled through the exploitation of bacterial, yeast, mammalian or plant cells. One of the earliest examples of protein biopharmaceuticals was the use of recombinant DNA technology to modify ‘Escherichia coli’ for the production of Human Insulin, which was followed by the development of Human Growth Hormone and Human Blood Clotting Factor₁.

More recently, liberalisation of the legislation controlling the development of stem cell technologies, allows for further opportunities in the development of biopharmaceuticals. In support of all these efforts, there is a pressing need to develop rapid and accurate methods for bioprocess monitoring, both for product yield optimisation and quality assurance purposes. Often these methods utilise analytical techniques, such as vibrational spectroscopy to obtain broad chemical profiles of the reaction or biotransformation in real-time, ultimately on-line. These data are ‘holistic’ in nature (since many functional groups are measured) and can be interpreted using multivariate pattern recognition methods in a variety of ways, and a common theme is the calibration of spectral data to product concentration, usually employing the partial least squares regression algorithm_2,3. Although these methodologies are well developed within the traditional chemical and pharmaceutical industries, the additional challenges of dealing with living processes have generated a great deal of interest in the evolution of PAT for biopharmaceuticals. (more…)

Tagged with: Raman Spectroscopy, Roger Jarvis, Royston Goodacre, University of Manchester, Victoria Brewster

PAT Roundtable: trends and issues

Issue 1 2009 / 7 February 2009 /

European Pharmaceutical Review invited four individuals to discuss their views and opinions on current trends and issues surrounding PAT. (more…)

Tagged with: Ametek Process Instruments, ASTM International, FOSS NIRSystems Inc, Martin Warman, Martin Warman Consultancy Ltd, PAT, Pat Picariello, Robert Mattes, Tony Slapikas