Cardiac hypertrophy is a thickening of the heart muscle – characterized by increased cell size rather than number – in response to conditions such as high blood pressure and coronary heart disease, which results in a decrease in size of the chambers of the heart, including the left and right ventricles. Since hypertrophy is associated with heart failure, irregular heart rhythms and an increased risk of angina and heart attack, understanding the mechanisms underlying this abnormal thickening is of great importance. Scientists at the Babraham Institute have now identified a new signalling process in the heart which contributes to cardiac hypertrophy. Rhythmical Ca²⁺ increases are fundamental to contraction of the heart muscle, but elevated Ca²⁺ levels also regulate the gene transcription that leads to hypertrophy. The Babraham team found that it is localised increases in Ca²⁺ concentrations in the cell nucleus that activate the genes responsible for hypertrophy. These nuclear Ca²⁺ signals, which are distinct from the Ca²⁺ signals that control the rhythmical contractions of the heart, were found to be generated by opening of the inositol 1,4,5-trisphosphate (IP3) receptor calcium channels (IP3Rs) which surround the nucleus.

These channels open in response to the increased cellular levels of IP3 that are generated on binding of the vasoconstricting peptide, endothelin-1, to receptors on the surface of cardiac myocytes. Nuclear factor of activated T cells (NFAT), which is known to regulate genes involved in pathological hypertrophy, was shown to be activated by these IP3-mediated increases in nuclear Ca²⁺ levels and not by the Ca²⁺ signals associated with contraction. The study confirms the importance of the endothelin pathway in cardiac hypertrophy and provides evidence that modulation of IP3 signalling would be a suitable target for future therapies.

The study is published in the journal Molecular Cell.

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There is no single test that will diagnose Alzheimer’s disease (AD) and a diagnosis of possible or probable AD is currently based on neuropsychological tests together with advanced brain imaging techniques such as functional MRI or PET scans. Simple and reliable diagnostic tests for AD are much needed, and researchers at the University of Georgia have now shown that the levels of two antibodies in the blood correlate with the severity of AD symptoms and may provide just such a test. The team had previously shown that levels of anti-Aβ and anti-RAGE antibodies were significantly higher in AD patients than in healthy individuals and the latest study reveals a direct relationship between the severity of disease and the levels of the two antibodies. Much evidence points to a link between AD and elevated levels of β-amyloid (Aβ) peptides. Binding of Aβ to neuronal membrane receptors for advanced glycation end products (RAGE) is believed to trigger inflammation and contribute to the neurological damage characteristic of AD.

Although it could be years before a diagnostic test based on their work is available for clinical use, the researchers hope that it will, one day, provide a way of identifying people with early AD and those at risk of developing the disease. The study is published in full in The Journals of Gerontology.

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Whereas mammalian fatty acid synthase (FASI) is a multidomain, multifunctional homodimeric protein which carries out all of the enzymatic steps needed for de novo synthesis of long chain fatty acids, bacterial fatty acid synthesis is carried out by a number of discrete enzymes, collectively known as FASII. This difference between FASI and FASII has led to the identification of FASII as a target for antibiotic therapy. The hypothesis is strengthened by the activities of the antiseptic, triclosan, the anti-Mycobacterium tuberculosis agent, isoniazid, and the antifungal antibiotic, cerulenin, which are believed to act primarily by inhibiting steps in the FASII pathway. The natural products, platensimycin and platensin, have also been shown to exhibit broad spectrum Gram-positive antibacterial activity and to inhibit fatty acid biosynthesis.

A recently published letter in the journal Nature, however, suggests that inhibition of FASII as an approach to antimicrobial therapy may be fundamentally flawed. The new study showed that major Gram-positive pathogens, such as streptococci, enterococci and staphylococci, were able to grow in the presence of FASII pathway inhibitors, cerulenin and triclosan, if supplied with exogenous fatty acids at levels that would be present in human serum. Using Streptococcus agalactiae – an opportunistic pathogen that can cause serious meningitis in newborns – as a model, the authors demonstrated that the unsaturated fatty acids, linoleic acid and oleic acid, but not the saturated fatty acids, palmitic acid and stearic acid, were able to overcome the inhibitory effect of cerulenin treatment.

The authors also demonstrated that when S agalactiae is grown in the presence of serum, there is an overall decrease in FASII gene expression. In further experiments, deletion mutants were used to demonstrate that FASII enzymes are dispensable in vivo during S agalactiae infection. Growth of all deletion mutants was severely restricted in standard Todd Hewitt (TH) medium, but all grew comparably with wild type strains in serum or in the presence of added oleic acid or linoleic acid. The mutant strains were also as virulent as wild type strains in animal models, even if the animals were treated with the hypolipidemic agent, fenofibrate. The authors believe that drugs targeting FASII would be ineffective in natural infections unless the bacteria had a requirement for specific fatty acids not present in serum.

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The energy demands of rapidly proliferating cancer cells require high levels of nutrients, including iron. Since cells are sensitive to free iron, they utilise iron storage proteins to protect themselves. Scientists at the German Cancer Research Center (DFKZ) and University Medical Center Mannheim, studying Sézary’s disease, have now shown that manipulation of free intracellular iron can induce apoptosis in T-cell lymphomas.

Sézary’s disease is an aggressive and ultimately fatal type of cutaneous T-cell lymphoma that is resistant to currently available treatments. Apoptosis resistance in leukemias and lymphomas is mediated by aberrant signalling of the NF-_κB pathway. The researchers have demonstrated that cell death of cutaneous T-cell lymphoma cell lines induced by inhibition of the NF-_κB pathway is a result of increased free intracellular iron and reactive oxygen species (ROS). Using T-cells from Sézary patients they show that inhibition of constitutively active NF-_κB causes down-regulation of ferritin heavy chain (FHC) that leads to an increase of free intracellular iron, which, in turn, induces massive generation of ROS. The involvement of FHC was confirmed by direct down-regulation using siRNA.

Importantly, T cells isolated from healthy donors do not display down-regulation of FHC and, therefore, do not show an increase in iron and cell death upon NF-_κB inhibition.

The work, published in the journal Cancer Research, suggests FHC as a novel target for therapeutic intervention in lymphoma.

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Drug-induced liver injury (DILI) is the most frequent cause of acute liver failure in the US and a major obstacle in the development of new medicines. It is therefore not surprising that prediction of liver toxicity is of considerable interest to healthcare professionals and drug companies alike.

Now The Hamner Institutes for Health Sciences and Entelos, Inc. have announced a partnership to create a computer model of liver function in virtual patients to improve understanding of how drugs can sometimes damage the liver.

The partnership supports the FDA’s Critical Path Initiative, which aims to reduce the time taken to develop and approve safe and effective medicines, and two FDA scientists will join the scientific advisory board for the project.

Entelos will use its clinically validated PhysioLab biosimulation platform to build a mathematical model of liver function using information from a variety of sources, incorporating The Hamner’s expertise in liver injury and systems biology. Additional important information will be supplied through Hamner research programs employing novel liver-derived cell models and special metabolism studies made possible by a new Hamner metabolomics laboratory. The objective of the collaboration is to develop a virtual liver that will account for the effects of genetic variations and other factors, such as patient sex, age, behavioural characteristics and environmental influences. In parallel, virtual rodents will be developed that will provide an improved means by which to evaluate preclinical drug effects and mechanisms of liver injury across species.

If successful, the platform will have many potential uses, including guiding the development of new diagnostic tests and new ways to test drug safety in the laboratory.

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Two viruses from the Henipavirus genus, Nipah (NiV) and Hendra (HeV), are recently emerged zoonitic (transmissible from animals to humans) paramyxoviruses that cause encephalitis in humans. HeV, previously known as equine morbillivirus, emerged as the causative agent of an outbreak of fatal respiratory disease in horses and man in Australia in 1994. NiV emerged in 1998/1999 in Malaysia and Singapore causing fatal encephalitis in humans. Human fatality rates can be as high as 75%.

Researchers at Weill Medical College of Cornell University, Australian Animal Health Laboratory, University of Tennessee Health Science Center and Rockefeller University have now developed a high-throughput assay that is able to identify inhibitors that target several stages of the viral life cycle. Their initial screen showed that chloroquine, approved for malaria treatment, inhibited infection with live HeV and NiV at a concentration of 1µM in vitro (IC₅₀=2µM), lower than the plasma concentrations present in humans receiving chloroquine treatment for malaria.

The scientists speculate that the mechanism of action of chloroquine is likely to be inhibition of cathepsin L, a host enzyme essential for processing of the viral fusion glycoprotein and maturation of newly budding virions. In the absence of this processing step, virions are not infectious. Chloroquine has previously been shown to suppress the activity of cathepsin L.

The authors of the current study, published online ahead of print in the Journal of Virology, suggest that the established safety profile and broad experience with chloroquine in humans should provide an option for treating individuals infected by these deadly viruses.

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High affinity binding of nicotine to subtype α4β2 nicotinic acetylcholine receptors in the brain is thought to be critical for nicotine addiction, but the reason for the higher affinity of nicotine at these receptors than at muscle receptors has been difficult to explain, especially given the similarity between the two types of receptors. If nicotine activated muscle receptors as strongly as it does brain receptors, smoking would cause unbearable, and possibly fatal, muscle contractions. Dougherty and colleagues, writing in the journal Nature, have now described a strong cation-π interaction between nicotine and a tryptophan residue, TrpB, in the brain α4β2 receptor. This key interaction is not formed between nicotine and muscle receptors even though the residues around the binding site, including the key tryptophan, are identical in brain and muscle receptors. A point mutation close to TrpB that differentiates α4β2 and muscle receptors appears to affect the exact shape of the binding sites and allows nicotine to interact more strongly with TrpB in the brain receptor. The new insight into differences in binding affinities may help in the development of drugs that target specific nicotinic acetylcholine receptors and, since more than 25% of all tryptophans are believed to be involved in cation-π interactions, may also have relevance for other drug classes. Energies for cation-π interactions, which are non-covalent interactions between the face of an electron-rich π-system and an adjacent cation, are comparable to those for hydrogen bonds.

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Historically, most drugs were discovered either by identifying the active ingredient from traditional medicines or by observing the pharmacological effect of compounds in living animals and it wasn’t until the 1960s that an understanding of the relationship between chemical structure and biological activity began to develop. Since then, attempts have been made to link discrete molecular targets (usually proteins) to particular diseases and to identify small molecules which will interfere with the function of these targets.

Contemporary drug discovery is dominated by this molecular target-based paradigm but, with the advent of high throughput cell-based assays, scientists are now also able to test compounds for a desirable activity in whole cells. A disadvantage of this approach is that the precise protein target is not always easy to identify but, writing in the journal PNAS, Ong et. al. have now described the use of quantitative proteomics (SILAC, stable isotope labelling with amino acids in cell culture) together with affinity enrichment to identify the protein targets.
SILAC is a technique based on mass-spectrometry which allows detection of differences in protein abundance between samples of cells. The growth medium of one cell population contains normal essential amino acids whilst that of the other cell population contains arginine or lysine labelled with stable heavy isotopes (¹³C or ¹⁵N). The growing cells incorporate these amino acids into their proteins, reaching full incorporation after 5 population doublings and producing a characteristic mass shift (6 Da with ¹³C₆-Arg or 8 Da with ¹³C₆¹⁵N₂-Lys). If the cells grown in the presence of the ‘heavy’ amino acids are also treated with a test compound, any difference in the ratio of peak intensities in the mass spectrum for protein pairs between treated and untreated cells indicates that the protein is, directly or indirectly, a target of the test compound. The authors describe the application of the method to the identification of targets for kinase inhibitors and immunophilin binders.

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Chagas’ disease is a parasitic disease caused by the protozoan Trypanosoma cruzi that affects an estimated 16-18 million people in Central and South America. Transmission of T. cruzi is most commonly by insect vectors but the disease may also be spread through blood transfusion and organ transplantation, through contaminated food, and from mother to child. In the first weeks or months after infection, symptoms are generally mild and the disease is often not diagnosed or treated. Over the course of years, however, chronic infection can damage the nervous system, heart and digestive system, leading to death. Only two drugs, with limited efficacy and significant toxicity, are currently available to treat the acute phase of the infection and none is available for treatment of the chronic phase of the disease. A report in the journal PLoS Neglected Tropical Diseases now describes three classes of heteroaromatic compounds which effectively inhibit replication of the parasite in vitro and show low toxicity towards host cells.

One of the compounds, CX1, showed strong tryptanocidal activity, which may be especially relevant for the development of drugs which are active during the chronic stage of the disease when the parasites are intracellular and not actively replicating. The compounds also showed high activity against Leishmania major and Leishmania amazonensi which cause leishmaniasis.

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Approved in 1995, glatiramer acetate (Copaxone®, copolymer-1) is a disease-modifying drug that has been demonstrated to reduce the relapse rate and progression of disability in relapsing-remitting multiple sclerosis (RRMS) patients. The compound is a mixture of synthetic peptides (50-90 amino acids) composed of alanine, glutamic acid, lysine and tyrosine. Originally developed to mimic myelin basic protein, a major component of the neuronal myelin sheath, it was intended for use as an inducer of experimental autoimmune encephalitis (EAE). The unexpected inhibition of EAE that was observed with glatiramer acetate led to clinical trials and subsequent approval for RRMS.

The efficacy of glatiramer acetate has been ascribed to an effect on the adaptive immune response, shifting towards a Th2 polarisation of myelin-specific T-cells. Further studies have demonstrated an immunomodulatory effect on monocytes, macrophages and dendritic cells. However, the full mechanistic picture is still unclear.

Collaborating scientists from the University of Geneva, Technische Universität München and University of California, San Francisco, have now demonstrated an effect of glatiramer acetate on the IL-1 system. Their research has shown that treatment with the polymer increases blood levels of secreted IL-1 receptor antagonist (sIL-1Ra), a natural inhibitor of IL-1β, both in RRMS patients and in EAE mice. In the same subjects, levels of IL-1β were undetectable. Additional in vitro experiments with T-cell contact-activated monocytes, a model relevant to chronic inflammation, showed that glatiramer acetate strongly reduced expression of IL-1β, whilst enhancing expression of sIL-1Ra. This is in contrast to effects in monocytes subjected to acute inflammatory conditions (stimulation with LPS), where glatiramer acetate increased production of both sIL-1Ra and IL-1β. The authors conclude that the effects on the IL-1 system in chronic inflammatory conditions contribute to the therapeutic effects of glatiramer acetate in RRMS.

The study is published in the online early edition of the journal PNAS.

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