Image: Flickr - bibendum84 New research has shown that compounds that affect cellular energy status could also be used to treat hepatitis C virus (HCV) infections. Metformin, which is used to treat type II diabetes, and 5-amino-1-β-D-ribofuranosyl-1H-imidazole-4-carboxamide (AICAR), which has been shown to mimic the beneficial effects of exercise in mice, stimulate AMP-activated protein kinase (AMPK). AMPK is a key sensor of cellular energy status and regulates both lipid and glucose metabolism to maintain cellular energy balance and protect against metabolic stress. An increase in the AMP/ATP ratio initiates an AMPK-mediated switch from activities that consume ATP, such as lipid production, to activities that produce ATP, such as lipid and glucose oxidation.
Infection with viruses might be expected to activate AMPK because of the energy demands put on the cell by viral replication, but research led by scientists at the University of Leeds has shown instead that HCV switches off AMPK so that the cell continues to produce the lipids needed to provide new viral particles with a protective outer coat. When the team treated HCV-infected cells with metformin or AICAR, AMPK activity was restored and viral replication was inhibited.
The team plan to carry out a small scale clinical trial to investigate the effects of AMPK activators in HCV infection and hope that such drugs may provide much-needed new treatments for HCV.
Image: Adapted from Flickr - Horia Varlan Cholesterol is essential for all animal life but high levels of cholesterol – when associated with low density lipoprotein (LDL) – are linked to an increased risk of atherosclerosis, heart disease and stroke. Circulating cholesterol can also be transported by high density lipoprotein (HDL); HDL is believed to be able to remove cholesterol from atheroma within arteries and cholesterol associated with HDL is considered to be beneficial for cardiovascular health. Cholesterol levels are determined by dietary intake, de novo synthesis and secretion by the liver: cholesterol absorption blockers and HMG-CoA reductase inhibitors (statins), which block cholesterol synthesis, are used clinically to reduce cholesterol levels.
A study led by researchers at the University of Cincinnati has now identified a new potential target for the control of cholesterol. The study, carried out in mice, found that circulation of cholesterol is regulated in the brain by the ‘hunger hormone’, ghrelin, which inhibits the melanocortin 4 receptor (MC4R) in the hypothalamus and is important for the regulation of food intake and energy expenditure. Increased levels of ghrelin were associated with increased levels of circulating HDL cholesterol, which the authors attribute to a reduction in the uptake of cholesterol by the liver. Genetically deleting or chemically blocking MCR4 in the CNS also led to increased levels of HDL cholesterol, suggesting that MCR4 is key to central control of cholesterol.
More studies are need to determine whether the effects observed in mice can be applied to humans but the finding that a neural circuit may be directly involved in the control of cholesterol metabolism by the liver could provide a target for new treatments to control cholesterol.
Image: Flickr - Alaskan Dude The cytoskeleton plays a key role in regulating many cellular functions; it maintains cell shape, protects the cell, enables cellular motion, and has important roles in proliferation and differentiation. Metastasising cancer cells exploit the cytoskeleton to produce protrusions that allow them to invade surrounding tissue and enter the blood system from where they can spread to distant tissues and seed new tumours.
The protrusions, known as pseudopodia, are highly specialised ‘feet’ that the cell uses to pull itself forward across the underlying surface. A team led by researchers at the University of California, San Diego has now identified a previously unknown kinase – termed pseudopodium-enriched atypical kinase one or PEAK1 – that regulates the cytoskeleton and plays a central role in the formation of pseudopodia. Preliminary studies in mice suggest that PEAK1 is important during tumour growth and the team also showed that PEAK1 levels are increased in primary and metastatic samples from human colon cancer patients. Whether PEAK1 is capable of transforming non-tumour cells into cancer cells has not yet been determined but the fact that PEAK1 has kinase activity suggests that it may be possible to design specific inhibitors which could help to elucidate its role in both normal and cancer cells. PEAK1, which is a 190-kDa non-receptor tyrosine kinase, could serve as a clinical biomarker that predicts whether a cancer is likely to metastasise and could also be a target for future cancer treatments.
Although the recent sporadic outbreaks of influenza A virus H5N1 and of a new variant of H1N1 in 2009 were less serious than initially feared, public health responses gave an indication of the potential for pandemic influenza A to wreak havoc amongst human populations. Timely development of vaccines should help to contain future outbreaks, but effective antiviral medicines will also be needed. Circulating strains of influenza A virus with resistance to existing neuraminidase inhibitors have already been discovered, and new molecular targets would provide additional protection in the event of a fresh outbreak.
Image: Wikimedia Commons - Armondo
Nucleozin Researchers led by a team at the University of Hong Kong have now identified a compound, nucleozin, which can aggregate the viral nucleoprotein and prevent its transport into the nucleus. The nucleoprotein plays critical roles in viral RNA replication and genome assembly, and nucleozin was shown to block replication of H1N1, H3N2, and H5N1 viruses in cell culture experiments and also to protect mice from lethal challenge with highly pathogenic avian influenza virus A H5N1.
The study, which is published in Nature Biotechnology, shows that the nucleoprotein is a viable drug target and could lead to the development of new treatments to control the impact of future influenza A outbreaks. Potential binding sites for nucleozin on the influenza nucleoprotein were also predicted using molecular docking models.
Ming dynasty acupuncture chart - Wikimedia Commons The origins of acupuncture are lost in antiquity and, today, the effectiveness of such treatment remains controversial. Despite the sometimes extravagant claims of proponents, there is little scientific basis to explain how acupuncture works and many researchers believe that the effects are attributable to a placebo effect. In a study published in Nature Neuroscience, researchers led by a team at the University of Rochester Medical Center have now suggested a physiological mechanism that may explain the analgesic effects of acupuncture.
The team found that adenosine was released during acupuncture in mice and that the analgesic effect of acupuncture could be replicated by direct injection of an adenosine A1 receptor agonist, 2-chloro-N(6)-cyclopentyladenosine (CCPA). Adenosine, which is released in response to injury or inflammation, has known pain-relieving properties and acupuncture was found to be ineffective in A1 receptor knock-out mice. 2′-Deoxycoformycin (dCF), a potent inhibitor of adenosine deaminase, was found to boost the effects of acupuncture, increasing the accumulation of adenosine in tissue as well as the duration of analgesia.
It will be interesting to see whether similar effects are observed in human subjects and also whether adenosine is also released during ‘sham’ acupuncture treatment in which needles are pressed against the skin without puncturing it – lack of differentiation from sham treatment is one of the main reasons that detractors cite for the placebo effect of acupuncture.
Crystal structure of inactivated form of CDPK1 from toxoplasma gondii - PDB ID 3KU2 Apicomplexan parasites such as Toxoplasma gondii and Plasmodium species can cause serious diseases in humans and domestic animals. Because the parasites are eukaryotes and share many metabolic pathways with their hosts, it has proved difficult to develop safe and effective treatments but researchers at Washington University School of Medicine in St. Louis have now identified an essential kinase in T. gondii which is unlike human kinases and more closely resembles those found in plants. In a study published in Nature, the team used conditional suppression to show that T. gondii calcium-dependent protein kinase 1 (TgCDPK1) is essential for survival of the parasite. The enzyme controls the ability of T. gondii parasites to secrete microneme proteins which allow the parasites to control their movement and move in and out of host cells.
3-MB-PPI It should be possible to exploit the differences between the parasite kinase and human kinases to develop potent and selective inhibitors of the parasite enzyme and the team have already identified compounds that block CDPK1 signalling without affecting human cells. Pyrazolopyrimidine-derived compounds such as 3-MB-PPI were found to specifically inhibit TgCDPK1 and disrupt the parasite’s life cycle at stages dependent on microneme secretion. The disruption was dependent on TgCDPK1 inhibition since parasites expressing a mutant kinase not sensitive to the inhibitors.
Calcium-dependent protein kinases have a kinase domain similar to that of calmodulin-dependent kinase, regulated by a calcium-binding domain in the C terminus. X-ray structures of TgCDPK1, published in Nature Structural and Molecular Biology, showed that, in the auto-inhibited (apo) form, the C-terminal activation domain resembles a calmodulin protein with an unexpected long helix in the N terminus that inhibits the kinase domain in the same manner as calmodulin-dependent kinase II. Calcium binding triggers reorganization of the C-terminal activation domain into a highly intricate fold, leading to its relocation around the base of the kinase domain to a site remote from the substrate binding site. This large conformational change constitutes a distinct mechanism in calcium signal-transduction pathways.
CDPK1 may play a similar role in Plasmodium species which cause malaria, but the researchers predict that it may be harder to selectively inhibit the Plasmodium enzymes.
Image: Flickr - Esther Kirby Programmed cell death (apoptosis) is essential to maintain homeostasis within living organisms and is controlled by a variety of intra- and extra-cellular signals. Activation of the death receptor CD95 (also known as Fas or Apo-1) by its physiological ligand, CD95 ligand (Fas ligand), leads to apoptosis in many tissues and is especially important in the immune system.
Resistance to apoptosis is also key to the survival of malignant cells but the role of CD95 in cancer progression is complex: although the down-regulation of CD95 that is frequently observed in tumour cells could contribute to their survival, complete loss of CD95 is rare in human cancers. On the other hand, many cancer cells express large quantities of CD95 and are highly sensitive to CD95-mediated apoptosis in vitro. Cancer patients often have high levels of CD95 ligand, suggesting that CD95 could perhaps promote the growth of tumours through non-apoptotic activities. Scientists from the University of Chicago and Northwestern University Feinberg School of Medicine have further investigated the role of CD95 in several human cancer cell lines and in mouse models of liver and ovarian cancer. Cancer cells – regardless of their sensitivity to CD95-mediated apotosis – were found to produce CD95 ligand and to depend on constitutive activity of CD95 for optimal growth. In the mouse models, deletion of CD95 reduced both the incidence and size of tumours. In further studies, the tumour promoting activity of CD95 was shown to be mediated by pathways involving JNK and Jun but not caspase-8.
The study, which is published in Nature, suggests that, paradoxically, reducing rather than enhancing activity of the death receptor CD95 may be an effective way to control the growth and proliferation of cancer cells. Further research is needed to understand the switch between signalling ‘die’ and ‘grow’, but eventually soluble CD95 or antibodies against CD95 ligand could find a role in the treatment of cancer.
Scanning electron micrograph showing asbestos fibres; Source – Wikimedia Commons MicroRNAs (miRNAs) are small (21-23 nucleotides), single-stranded RNA molecules that function as regulators of gene expression. The human genome encodes several hundred miRNAs and abnormal expression of these has been associated with cancer progression. We have previously reported on miRNA involvement in cholesterol regulation, amyotrophic lateral sclerosis and liver cancer. Now a collaboration between Rosetta Genomics, NYU Langone Medical Center and Vanderbilt School of Medicine has identified the potential utility of miR-33 for development of therapies targeting malignant mesothelioma (MM).
MM is a rare cancer that has been associated with exposure to asbestos dust (although a small proportion of patients have never been exposed). Taking anywhere between 20 and 50 years for symptoms to develop, the cancer affects the mesothelium, a thin layer of tissue surrounding the internal organs. The most common form of MM is pleural, involving the lining of the lungs, but it may also affect other tissues such as the peritoneum. Current treatment options, including surgery and chemotherapy, are limited and often confounded by late diagnosis because of the absence of symptoms.
This latest study found that MM cell lines derived from patients with aggressive disease failed to express miR-31, a microRNA that has also been linked to suppression of breast cancer metastasis. Functional studies, where miR-31 was reintroduced to the cells, showed that the microRNA could inhibit proliferation, migration, invasion, and clonogenicity of mesothelioma cells. miR-31 suppressed expression of a number of factors associated with maintenance of DNA replication and cell cycle progression, including the pro-survival phosphatase PPP6C. The mRNA for PPP6C, which contains three miR-31-binding sites in its 3′-UTR, was down-regulated when miR-31 was present and up-regulated in clinical MM specimens compared to matched normal tissues.
Whilst the study, published in the Journal of Biological Chemistry, reveals a key role for miR-31 in MM, considerable challenges remain to exploit this finding for therapy of the disease.
Hypusine The eukaryotic translation initiation factor eIF5A, which exists in two isoforms, was originally thought to be involved in formation of the first peptide bond during mRNA translation, but more recent work has implicated it as a translation elongation factor. In mammalian cells it has variously been associated with mediation of proliferation, apoptosis and inflammatory responses, although its mechanisms of action have remained vague. It has also been identified as a cofactor of the Rev trans-activator protein of HIV-1. eIF5A is unique in that it is the only known protein to contain the amino acid hypusine, formed posttranslationally via the sequential action of deoxyhypusine synthase (DHS) and deoxyhypusine hydroxylase (DOHH) acting at a specific lysine residue.
GC7 Based on the role of eIF5A in inflammation, a multi-institutional research team led by scientists at Indiana University School of Medicine has explored involvement of the protein in pancreatic islet dysfunction during the development of diabetes. In a low-dose streptozotocin mouse model of diabetes the team found that depletion of eIF5A (using siRNA) protected the mice from pancreatic β-cell loss and hyperglycemia. The depletion of eIF5A resulted in impaired translation of inducible nitric oxide synthase (iNOS)-encoding mRNA within islet cells. Further studies using an inhibitor of DHS, N1-guanyl-1,7-diaminoheptane (GC7), demonstrated a requirement for hypusination in the action of eIF5A.
The study, published in the Journal of Clinical Investigation, demonstrates a role for eIF5A in inflammation-induced damage to islet cells. Since the negative effects of eIF5A depend on hypusination, DHS may represent a viable therapeutic target for diabetes. Further work will be necessary to establish the role of this pathway in development and progression of the human disease.
Photomicrograph showing a mature sporangium of a Mucor species mould; Image: Wikimedia commons; source – CDC Mucormycosis is a potentially fatal infection of the sinuses, brain, or lungs, which is most commonly caused by the fungus Rhizopus oryzae. Even if the disease is successfully treated with antifungal agents together with surgery to remove necrotic tissue, survivors are typically left with considerable disfigurement. The condition is seen most often in people with diabetic ketoacidosis (DKA) who have elevated serum glucose and iron levels and a team of researchers at the University of California Los Angeles has now discovered why these individuals are more susceptible to infection. In mucormycosis, fungal invasion of blood vessels results in the formation of blood clots and destruction of local tissue and the team set out to identify the endothelial cell receptor that the fungus uses, and also whether iron and glucose play a role in regulating the expression of the receptor.
In human endothelial cells, glucose-regulated protein 78 (GRP78) was found to play a key role in endocytosis of R. oryzae and subsequent damage; enhanced expression of the protein in the presence of high concentrations of glucose, especially when iron levels are also elevated, offers an explanation of the increased susceptibility of individuals with DKA to mucormycosis. Mice with DKA, which have elevated levels of glucose and available iron, and which are also susceptible to mucormycosis, showed increased expression of GRP78 in sinus, lungs, and brain compared with normal mice. In further studies, treatment of DKA mice with GRP78-specific immune serum was shown to protect them from mucormycosis.
The study, which is published in the Journal of Clinical investigation, provides a new understanding of the pathogenesis of R. oryzae and may lead to new treatments for potentially lethal mucormycosis.
