Scientists have identified a new target for the diagnosis and treatment of age-related macular degeneration (AMD). AMD most commonly affects people over the age of fifty and is the leading cause of vision loss in those over sixty. The rarer, but more serious form of the disease, wet AMD, is caused by abnormal blood vessel growth under the macula (centre of the retina) and can develop very quickly. Although the condition is painless, the new blood vessels leak blood and fluid which lift the macula and destroy sharp central vision. The condition can be treated with laser surgery, photodynamic therapy or anti-vascular endothelial growth factor (anti-VEGF) injections into the eye, although none of these treatments provides a complete cure and loss of vision may continue despite treatment.
The new study, published online on Jun 14th in Nature, shows that blocking the eosinophil/mast cell chemokine receptor, CCR3, can reduce the abnormal blood vessel growth that leads to AMD. The researchers detected CCR3 protein in eye tissue from people with AMD but not in eye tissue from people of the same age who did not have the disease. In studies in mice, blocking CCR3, either by genetic engineering or using antibodies, reduced the number of abnormal blood vessels. In the mice, targeting CCR3 was shown to be more effective than targeting VEGF (70% vs 60%), suggesting that CCR3 blockers could also provide an effective treatment for patients with AMD. The team injected anti-CCR3 antibodies attached to semiconductor nanocrystals into mice and, using conventional ocular angiography techniques, were able to visualise the abnormal blood vessels even before they had penetrated the retina. Since CCR3 was detected at an early stage in the development of the disease, the researchers hope that the new imaging technology could be used diagnostically, and that early detection will provide better opportunities to prevent structural damage and preserve vision.
Huntington’s disease (HD) is an inherited neurological disorder that causes a wide range of symptoms including involuntary movements, clumsiness, lack of concentration, memory lapses, mood swings, and depression. Although the disease mechanism is not fully understood, it is known that sufferers have a defect on the short arm of chromosome 4 and produce abnormal versions of the protein, huntingtin. One of the puzzles surrounding the progression of HD has been that, although abnormal huntingtin proteins accumulate in cells everywhere in the body, they predominantly kills cells in the corpus striatum, the part of the brain that controls movement.
Writing in the journal Science, scientists at John Hopkins School of Medicine now report that the answer may lie with the small G protein, Rhes (Ras homologue enriched in striatum), which is found almost exclusively in the corpus striatum. In cell culture experiments using both mouse and human cells, Rhes was found to bind much more tightly to mutant huntingtin than to normal protein. When added singly to cell cultures, neither Rhes nor mutant huntingtin had any effect on survival but, when they were added together, half of the cells died within 48 hours.
Abnormal huntingtin proteins aggregate and form clumps, but there are fewer of these clumps in the corpus striatum of HD patients than in other brain regions or elsewhere in the body, suggesting that clumping of the protein may actually protect the cells. The team found that, in their cell culture experiments, adding Rhes to cells with abnormal huntingtin led to fewer clumps, although the cells died. The results suggest clumping of abnormal huntingtin may prevent it from causing cell death and that Rhes might be responsible for preventing abnormal protein from clumping. Rhes was also shown to promote sumoylation of mutant huntingtin. The team are currently exploring whether removing Rhes from mice with HD will prevent cell death, and hope that it may eventually be possible to design drugs which will specifically target Rhes to treat HD.
For the most part, cancer therapy has been aimed at exploiting pathways that are present in cancer cells and not in normal cells but two studies published in the May 29th issue of the journal Cell suggest potential for an alternative approach. Blocking the activity of oncogenic protein kinases – either with antibodies or small molecules – has become an important field of interest in cancer research but, despite the prevalence of RAS mutations in human tumours, inhibition of oncogenic RAS has not been realised as a therapeutic strategy. As an alternative to directly targeting RAS, US researchers have now identified ‘normal’ genes that are needed for cell survival in the presence of mutant, but not wild-type, KRAS.
To identify genes that are essential for survival only in the context of mutant KRAS, the researchers used a short hairpin RNA (shRNA) library to carry out high-throughput loss-of-function RNA interference (RNAi) screens in cancer cell lines as well as in normal cells. Dozens of potential drug targets were identified including serine/threonine kinase 33 (STK33) and mitotic polo-like kinase 1 (PLK1).
Patients with KRAS tumours are more likely to survive if they also have reduced expression of genes in the PLK1 pathway, suggesting that PLK1 inhibitors may have the potential to prolong survival. Although not required by normal cells, STK33 was found to be essential for the survival of cancer cells, irrespective of their tissue of origin, again suggesting therapeutic potential for inhibitors.
As well as identifying new targets for which it may be possible to develop therapeutically useful inhibitors, the two studies demonstrate the potential of RNAi screens to discover functional dependencies between oncogenes and normal genes in cancer cells. Targeting proteins which are essential for the survival of cancer cells, but not normal cells, could lead to a substantial therapeutic window, especially if only partial knock-down is needed to kill cancer cells.
The neurodegenerative disorder, Huntington’s Disease (HD, Huntington’s Chorea), is caused by mutations in the gene for the protein Huntingtin (Htt). Mutant Htt (mHtt) results when the number of trinucleotide repeats, in this case the CAG sequence encoding glutamine, exceeds a threshold value. Typically, HD affects patients when the number of repeats is greater than 35.
Although the mechanisms by which mHtt causes the disease are poorly understood, proteolysis of the mutant protein is key and neurotoxicity is attributed to the cleaved N-terminal fragments of mHtt. Scientists at the California Institute of Technology and the Wallenberg Neuroscience Center, Lund, Sweden have now reported that the pro-inflammatory kinase, IKKβ , can regulate mHtt proteolysis in response to neuronal DNA damage.
The team demonstrated that DNA damage in neurons induced by etoposide or γ-irradiation results in cleavage of both wild-type and mutant Htt and that the proteolysis requires IKKβ. Elevation of IKKα, inhibition of IKKβ expression or inhibition of IKKβ catalytic activity all suppressed proteolysis and increased neuronal resistance to DNA damage.
Since elevated neuronal DNA damage is observed in HD patients and animal models of HD, inhibition of IKKβ activity may represent a treatment option for the disease. The study is published in the journal PLoS one.
Asthma involves an immune response to inhaled allergens and chemical irritants, but the limited efficacy of existing treatments aimed at modifying this response suggest that additional physiological mechanisms may be at work in the disease process. In a report published in the May 19th online Early Edition of PNAS, US researchers have now found that the ion channel, TRPA1, plays a key role in allergic asthma. TRPA1 is a ‘chemosensor’ that is activated by mustard oil as well as by a number of endogenous and exogenous stimuli known to be triggers of asthmatic inflammation. The team found that, compared with wild type mice, animals that did not express TRPA1 showed fewer asthma symptoms, with reduced inflammation, airway mucus and bronchoconstriction. Although the exact role of TRPA1 in asthmatic inflammation is not yet understood, the ion channel is known to be activated by allergens such as cigarette smoke that can trigger asthma attacks. TRPA1 is found in airway nerves and the researchers believe that blocking TRPA1 may prevent infiltration of the lung by the inflammatory cells responsible for asthma symptoms such as wheezing and overproduction of mucus. In further studies, the team went on to show that treatment with the TRPA1 antagonist, HC-030031, reduced the symptoms of allergic asthma in mice. TRPA1 antagonists have previously been shown to reduce chronic inflammatory and neuropathic pain. The discovery of a role for TRPA1 as a neuronal mediator of allergic airway inflammation could lead to new treatments for allergic asthma and Hydra Biosciences, whose scientists contributed to the study, hope to start human clinical trials with a novel TRPA1 inhibitor within 12 months.
A number of human studies have linked lack of sleep to weight gain – decreased insulin sensitivity and glucose tolerance as well as disruption of the natural balance between the appetite hormones grehlin and leptin have been put forward to explain this link between disrupted sleep and weight gain. In studies in rodents, researchers at Merck have now shown that T-type calcium channels regulate both sleep and body weight maintenance. Mice lacking CaV3.1 T-type calcium channels were known to have altered sleep/wake patterns and the new study showed that these mice are also resistant to weight gain induced by a high fat diet.
Writing in the Journal of Clinical Investigation, the researchers report that the knock-out mice gained significantly less weight and had less body fat than their wild-type littermates when fed a high fat diet. The resistance to weight gain of the knock-out mice could not be fully explained by reduced food intake, an overall increase in activity or increased metabolic rate. In further studies, a selective T-type channel antagonist, TTA-A2, was shown to prevent, and even reverse, weight gain induced by a high fat diet, and also to improve body composition to greater extent than the widely used appetite suppressant, fenfluramine. TTA-A2, when dosed either prior to the sleep phase or during the wake phase, was found to promote sleep – a result which was unexpected since the knock-out mice have increased wake time compared with wild type animals. Although the reasons for the observed differences between pharmacological antagonism and genetic knock-out remain to be fully explained, the study highlights the potential for antagonism of T-type calcium channels as a novel weight loss strategy.
The authors suggest that the benefits of T-type calcium channel antagonists may be the result of better alignment of feeding patterns and the circadian rhythm and that sleep or circadian treatments may be of particular benefit for people struggling to lose weight or maintain weight loss because of poor diet.
Two recent reports in the British Journal of Cancer highlight the importance of chemokines in the spread and progression of cancer.
Using a mouse xenograft model, researchers from the University of Nebraska Medical Center showed that CXCR1 and CXCR2 receptors are important in the progression of melanoma, the most aggressive form of skin cancer. The study, which is published in the May 12th issue of the journal, investigated the effect of overexpression of CXCR1 or CXCR2 in two different human melanoma cell lines – one non-tumourigenic and the other with low tumourigenicity. Both cell lines showed low endogenous expression of the receptors. Both CXCR1- or CXCR2-overexpressing melanoma cells showed enhanced proliferation, chemotaxis and invasiveness in vitro. In vivo, overexpression induced tumourigenicity in the non- tumourigenic cell line and significantly enhanced tumour growth in the low-tumourigenic cell line. Tumours generated from CXCR1- or CXCR2-overexpressing melanoma cells showed significantly increased tumour cell proliferation and microvessel density as well as reduced apoptosis, indicating an essential role for the receptors in growth, survival, motility and invasion of human melanoma.
The second study, published in the 12th May advance online edition of the journal, investigated the effect of CXCR3 receptors on the metastasis of colorectal cancer (CRC). Liver and lung metastases are the main cause of death from colorectal cancer and the study looked at the ability of CXCR3-expressing CRC cell lines from humans or mice to induce metastases in immunodeficient and immunocompetent mice, respectively. In vitro, activation of CXCR3 on both human and mouse CRC cells by its cognate ligands induced migratory and growth responses which could be blocked by the CXCR3 receptor antagonist, AMG487. In vivo, preventative or curative treatment with AMG487 markedly inhibited implantation and growth of both human and mouse CRC cells in the lung, but had no effect on metastases affecting the liver.
The axons of nerve cells are sheathed by an insulating layer of myelin which is made up of about 80% lipid and 20% protein. Demyelination, leading to impaired or lost conduction of signals along the nerve, is a hallmark of multiple sclerosis (MS). In relapsing-remitting MS myelin can be replaced but, after repeated attacks, the repair system becomes less efficient. Researchers at the University of Medicine and Dentistry of New Jersey have now identified a key pathway which regulates the production of new oligodendrocytes – the myelin-producing cells of the CNS – and the production of myelin. They found that activation of the mammalian target of rapamycin (mTOR) is essential for oligodendrocyte differentiation at the late progenitor to immature oligodendrocyte transition. The effects were found to be mediated via two distinct signalling complexes, mTORC1 and mTORC2. mTORC2 was found to control myelin gene expression at the mRNA level whereas mTORC1 influenced expression of myelin basic protein via an alternative mechanism.
Although it remains to be determined whether stimulation of the mTOR pathway or removal of some inhibitory mechanism would be most appropriate, allowing the pathway to function normally could provide new treatments for MS and other demyelinating diseases.
The study is published in the May 13th online edition of the Journal of Neuroscience.
Meningitis – infection of the cerebrospinal fluid and protective membranes surrounding the brain and spinal cord – can be caused by infection with either viruses or bacteria. Viral meningitis is typically relatively mild and self-limiting whereas bacterial meningitis is much more serious and can result in severe brain damage or even death. Bacterial meningitis in children is almost exclusively caused by infection with one of three bacterial strains: Streptococcus pneumoniae, Neisseria meningitidis, or Haemophilus influenza. Exactly how these bacteria are able to breach the blood-brain barrier and cause infection was not understood, but researchers from the University of Nottingham and St. Jude Children’s Research Hospital have now discovered that all three pathogens use the same receptor on human cerebrovascular endothelial cells to begin the process of crossing the barrier. Bacteria need to have some way of fixing their position and becoming established and use attachment molecules – known as adhesins – to achieve this. Some bacteria take the attachment process a stage further and use the adhesins as a first step towards gaining entry into host cells. It was known that the three bacteria responsible for most cases of meningitis share the same strategy for the second step of crossing endothelial cells and the new study has shown that they also use the same host cell receptor, the laminin receptor, for initial attachment. Other infectious agents known to use the laminin receptor to gain access to the CNS include prions and some neurotropic viruses, although the interaction of the laminin receptor with bacteria appears to be somewhat different to that of other pathogens.
The study suggests that blocking the interaction between bacterial adhesins and the laminin receptor might offer broad protection against bacterial meningitis and may lead to better treatments and prevention strategies. The study is published in full in the Journal of Clinical Investigation.
Researchers from University of Rochester Medical Center have shown that eliminating the gene for cyclophilin A completely protects mice from developing abdominal aortic aneurysms, a late stage complication of atherosclerosis. An aortic aneurysm is a thin, weakened section of the aorta which can rupture, leading to massive internal blood loss and death. Aneurysms occur most frequently in the abdominal section of the aorta and cause around 15,000 deaths a year, most in older men. In abdominal aortic aneurysm, angiotensin II is known to stimulate oxidative stress in blood vessels leading to increased activity of matrix metalloproteinases which, in turn, degrade the matrix structure of the vessel wall. Increased activity of matrix metalloproteinases also plays a role in atherosclerosis, allowing smooth muscle cells from the blood vessel walls to contribute to the development of plaques. In both abdominal aortic aneurysm and atherosclerosis, angiotensin II also contributes to local inflammation by recruiting immune cells to the blood vessel wall.
Using genetically modified mice, cyclophilin A was found to promote all three events involved in angiotensin II mediated damage to blood vessels – oxidative stress, matrix degradation and inflammation. Cyclophilin A is highly expressed in vascular smooth muscle cells and studies showed that both intracellular and extracellular cyclophilin A are required for generation of reactive oxygen species and activation of matrix metalloproteinases. The team is hoping to develop anti-cyclophilin A drugs that will reduce the processes underlying cardiovascular diseases such as abdominal aortic aneurysm, atherosclerosis and hypertension. The study is published in full in the journal Nature Medicine.