Blog Archives

Natural Antiviral Proteins Discovered

Viruses are economical with their own genetic information, relying on requisition of host cellular proteins to complete their lifecycle. In a study to identify cellular proteins required by the H1N1 influenza virus, collaborators from Howard Hughes Medical Institute, Harvard Medical School, Massachusetts General Hospital, Yale Medical School, and the Wellcome Trust Sanger Institute have now made an unexpected discovery.

The research team set up large arrays of cultured human cells and used siRNA molecules to block expression of individual genes. They then examined the effect of the knockdowns on H1N1 activity by measuring changes in the presence of viral protein on the surface on infected cells. The work identified more than 120 genes which were required to be expressed for H1N1 infectivity but, surprisingly, also identified a class of genes that permitted greater influenza replication when they were blocked.

The proteins encoded by these genes were members of the interferon-inducible transmembrane (IFITM) family, IFITM-1, IFITM-2 and IFITM-3. These IFITMs were known to be produced at low levels in most cells, and at higher levels in cells exposed to immune-stimulating interferon, but their functions were poorly understood. Knockdown of IFITM-3, in particular, increased replication of H1N1 by 5 to 10-fold. Conversely, by over-expressing IFITM-3 the team were able to completely inhibit H1N1 replication.

To the researchers’ surprise, increased expression of IFITMs also blocked the replication of completely different viruses, including strains of West Nile and Dengue viruses. Although the proteins are not effective against all viruses, the discovery could lead to new antiviral therapeutics. Further work is required to elucidate the precise mechanisms by which the IFITMs exert their effects.

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


Alzheimer’s Disease: Marijuana Maybe Good, Gingko Biloba Not So Good?

There are no medicines that can prevent Alzheimer’s disease and current drugs ease symptoms but do little to stop progression of the disease. There is consequently much current interest in whether lifestyle choices or herbal remedies can prevent or alter the course of Alzheimer’s disease. Two recent reports describe the effect of marijuana-like compounds in elderly rats and extracts of Gingko Biloba in elderly humans.

cannabis sativaPrevious work has suggested that people who regularly smoked marijuana in the 1960s and 1970s are less likely to develop Alzheimer’s disease in later life. Now scientists at Ohio State University have described details of how constituents of marijuana can combat inflammation and possibly stimulate neurogenesis. The team had found that treatment with WIN-55212-2, a compound with similar pharmacological properties to tetrahydrocannabinol (THC), made a small improvement to the ability of elderly rats to perform memory tests. They then went on to look at how the drug was working, and showed that it lowered inflammation in the hippocampus by acting on the TRPV1 receptor. The team also showed that the action of WIN-55212-2 on CB1 and CB2 receptors led to the generation of new brain cells. The group are continuing to investigate which receptors are most important for reducing inflammation and stimulating neurogenesis. This knowledge could lead to the discovery of new drugs to reduce inflammation and increase the production of new neurons before memory loss becomes apparent.

WIN55212-2 is a full agonist at the CB1 receptor and is not approved as a drug since it elicits cannabis-like effects in humans.

Ginkgo bilobaA second report discusses the results of the first randomized, double-blind trial to assess the effectiveness of Ginkgo biloba in reducing the incidence of dementia. Ginkgo is an antioxidant that helps to protect cells in vitro from oxidative damage. Some studies have suggested that ginkgo may also protect against the toxic effects of β-amyloid. Although previous clinical trials of ginkgo leaf extracts have had mixed results, many older people continue to use the herb in the hope of warding off the onset of Alzheimer’s disease. The new study, however, has shown that ginkgo gave no benefit in reducing all-cause dementia or Alzheimer’s dementia. The study involved more than 3000 men and women aged 75 or older who were randomised to placebo or treatment groups. The treated group received 120mg ginkgo twice a day for an average of 6 years. At the end of the study, dementia developed in 246 people taking ginkgo compared with 277 people in the placebo group. The authors note that an effect may have been seen had the study carried on for longer since it is known that there is a significant time lag between initial brain changes and the manifestation of clinical dementia.


New Targets to Combat HIV Infection

HIV capsidThe human immunodeficiency virus (HIV) uses host cell surface receptors such as CD4, CCR5 and CXCR4 to gain entry into cells. Recently, monoclonal antibodies and small molecules that block these receptors have joined the armoury of drugs used to combat HIV infection. A new report in the journal Cell describes a genome-wide analysis to search for other virus-host interactions that are important for the early steps of HIV infection.

More than 40 host factors that regulate capsid uncoating and reverse transcription were identified along with 15 cellular factors that facilitate entry of the HIV preintegration complex into the cell nucleus and integration of proviral DNA.

Cytoskeletal proteins and microtubules were found to play roles in early HIV replication, and proteins involved in DNA-damage repair were found to influence the initiation of reverse transcription and the accumulation of HIV-1 DNA products prior to integration. Other cellular systems discovered to play roles in the early stages of HIV infection include nucleic acid binding proteins and the ubiquitin-proteasome pathway. Protein phosphorylation and dephosphorylation events were found to have a possible role in the regulation of HIV-1 reverse transcription, and prostaglandins were shown to have a potential role in HIV-1 nuclear import. Several cellular factors were also found to be important for HIV-1 DNA integration.

The study revealed totally new pathways in HIV infection, including involvement of Notch signalling in reverse transcription. A full understanding of the role of each of these host factors in cell types infected by HIV will be essential to assess their contribution to disease progression. However, given the success of compounds which block viral entry receptors, modulation of the activity of some of the newly discovered pathways could provide novel strategies for the treatment of HIV/AIDS.


New Results for PTC124 in Cystic Fibrosis

Cystic fibrosis (CF) is a hereditary disease characterised by the production of thick sticky mucus which results in frequent lung infections. CF is caused by any one of a number of mutations in a gene called the cystic fibrosis transmembrane conductance regulator (CFTR) which encodes a protein that transports chloride ions across cell membranes. In about 10% of patients worldwide, and more than 50% of patients in Israel, CF is caused by nonsense mutations in the messenger RNA for CFTR. Premature stop codons prevent production of functional full-length protein: patients with nonsense-mutation CF produce very little functional CFTR and often have a severe form of CF.

New Phase II results published in The Lancet show that an orally bioavailable small molecule demonstrates activity in nonsense-mutation CF. PTC124 was designed to induce ribosomes to selectively read through premature stop codons to produce functional CFTR. The data show that treatment with PTC124 results in statistically significant improvements in the chloride channel function of patients.

PTC124
Nonsense mutations account for a significant number of cases of most inherited diseases and PTC124 may have the potential to treat diseases other than CF.


Calpain Inhibitors in Models of Alzheimer’s Disease

Calpains are calcium-activated cysteine proteases which, when abnormally activated, can initiate degradation of proteins essential for neuronal survival. A report in Journal of Clinical Investigation describes the effect of the selective calpain inhibitor, BDA-410, in a transgenic mouse model of Alzheimer’s Disease.
BDA-410
Calpain inhibition by BDA-410 improved spatial-working memory and associative fear memory in APP/PS1 mice. The authors put forward a hypothesis in which amyloid peptides trigger a cascade leading to calpain activation and, ultimately, synaptic dysfunction and cognitive abnormalities. BDA-410 did not alter levels of amyloid oligomers or plaque load but restored normal phosphorylation levels of transcription factor CREB and normal distribution of synaptic protein, synapsin I. These results suggest that calpain inhibition might provide a new strategy for alleviating memory loss in Alzheimer’s Disease.

BDA-410 has previously been shown to be a potent inhibitor of cysteine proteases of the malaria parasite, Plasmodium falciparum.


Huntington’s Drug Approved in US

tetrabenazineHuntington’s Disease is a hereditary neurological disorder caused by a fault in the gene that produces a protein called Huntingtin.

The function of Huntingtin is not clear, but the presence of abnormal protein increases the rate of neuronal cell death and gives rise to physical, cognitive and psychiatric symptoms. Personality or mood changes may be the earliest signs of the disease, followed by uncontrollable jerky movements (chorea) and problems with memory. Although there is no cure for Huntington’s Disease, tetrabenazine (Xenazine®) has been shown to substantially reduce chorea. The drug is currently licensed for the treatment of Huntington’s Disease in a number of other countries, and has now been approved for use in the US.


Cell Receptor for HCMV Entry Identified

Human Cytomegalovirus (HCMV) is a member of the Herpes virus family and infects 50-80% of adults in the US. After infection, the virus remains latent in the body for the rest of the person’s life. Most people who are infected with HCMV after birth remain asymptomatic unless their immune system is compromised by drugs (transplant recipients), HIV infection or old age. In such individuals, HCMV infections can become life-threatening.

The route by which the virus enters cells is not well understood, but a report in the journal Nature now shows that the HCMV glycoprotein B binds to cell-surface platelet-derived growth factor receptor-α(PDGFR-α), which acts as an entry receptor for HCMV. PDGFR-α is a receptor tyrosine kinase, and both laboratory and clinical strains of HCMV trigger autophosphorylation of the receptor, resulting in activation of downstream signalling pathways. Cells in which PDGFR-α was genetically deleted were non-permissive to HCMV entry, viral gene expression or infectious virus production. An HCMV glycoprotein B neutralizing antibody, an antibody to PDGRF-α and Gleevec, an inhibitor of the receptor kinase activity, also completely blocked viral entry. These data suggest that PDGFR-α is critical for HCMV infection and that the receptor provides a novel target for anti-HCMV therapies.
Gleevec
PDGFRs also play an important role in angiogenesis and have been linked to the pathogenesis of some tumors. Gleevec is marketed by Novartis for treating certain types of cancer.


MS Drug Could Also Fight Viral Infections

A drug that is being developed to treat multiple sclerosis (MS) by damping down the immune system may also have the potential to treat viral infections. A study published in the journal, Nature, showed that mice treated with the drug FTY-720 (fingolimod) were able to clear a viral meningitis infection that persisted in untreated mice.

It is, at first sight, surprising that a drug developed to suppress the immune response in MS patients can help to fight a viral infection.
Fingolimod
FTY720-P is an agonist of the sphingosine-1-phosphate receptor and causes lymphopenia by preventing egress of lymphocytes from the lymph nodes. The new finding builds on the observation that more easily cleared strains of the virus also cause lymphocytes to become sequestered in lymph nodes. The reason why trapping circulating lymphocytes allows a more robust response to infection is not clear, but may be linked to the fact the lymph nodes are where the immune response is primed. Clearance of the virus does not occur when CD4 T cells are absent at the time of treatment, indicating that the drug is not exerting direct antiviral effects. Some viruses, including HIV, replicate at high levels in lymph nodes and the team plan to test the effect of FTY720 on infection with other viruses.

FTY720 is currently in Phase III clinical trials to test its safety and efficacy as a disease modifying therapy for relapsing-remitting MS.


Novel Mechanism of γ-Secretase Modulation

It is widely believed that reducing amyloid plaque formation will provide a disease-modifying treatment for Alzheimer’s Disease, either halting or slowing cognitive decline. Amyloid peptides, which aggregate to form neurotoxic plaques, are formed by proteolysis of a precursor protein by the action of two enzymes, β-secretase and γ-secretase. Inhibition of either of these proteases should prevent the formation of amyloid peptides, and much effort has been devoted to the identification of inhibitors. γ-Secretase, particularly, is a promiscuous enzyme and hydrolyses a number of other substrates, including Notch. The Notch signalling pathway is important in many cellular processes and so modulators rather than inhibitors of γ-secretase are preferred.

Amyloid pathway

Relatively early in the search for modulators of γ-secretase, the surprising discovery was made that certain nonsteroidal anti-inflammatory drugs (NSAIDs), including ibuprofen, effectively suppress amyloid peptide production while sparing processing of Notch and other γ-secretase substrates. Further studies identified tarenflurbil (R-flurbiprofen, Flurizan, MPC-7869) as a γ-secretase modulator with the potential to treat Alzheimer’s Disease. A recent report in Nature (Kukar et. al., Nature 2008, 453, 925-929) has now shed new light on how such compounds prevent amyloid formation.

tarenflurbil

Using photoprobes, the group were able to show that flurbiprofen does not bind to the γ-secretase protein complex but, instead, binds to the amyloid precursor protein.

Disappointingly, Myriad Genetics have recently discontinued development of Flurizan since it failed in a phase III clinical trial in Alzheimer’s patients.

The study by Kukar et. al. does, however, suggest that it may be possible to identify other small molecules that reduce the formation of amyloid peptides by binding to the precursor protein rather than to the γ-secretase complex. It remains to be seen whether similar ‘substrate-binding’ inhibitors of other proteases can be identified.


Fitness without Effort?

AICARResearchers at the Salk Institute have shown that agonists of both AMP-activated protein kinase (AMPK ) and a peroxisome proliferator-activated receptor (PPAR) can mimic some of the beneficial effects of exercise in mice. In a treadmill running test, the PPAR-β/δ agonist, GW 1516 (GW 501516), acted synergistically with exercise to increase running endurance after 4 weeks. The AMPK agonist, AICAR, surprisingly enhanced running endurance even in sedentary mice, also after 4 weeks dosing. PPAR-δ and AMPK agonists have the potential to treat diseases such as diabetes, where exercise has been shown to be beneficial and to offer protection against obesity, but also have the more controversial potential to increase endurance in athletes.
GW1516
Like exercise, AICAR and GW1516 trigger a variety of changes that contribute improved endurance and the ability of muscle cells to burn fat. A phase II clinical trial of GW1516 for the potential treatment of dyslipidemia has been completed.