Elevated CSF levels of TACE activity and soluble TNF receptors in subjects with mild cognitive impairment and patients with Alzheimer’s disease.

Expression levels of tumor necrosis factor (TNF) receptors, TNFR1 and TNFR2, are significantly changed in the brains and cerebrospinal fluid (CSF) with Alzheimer’s disease (AD). Moreover, we also found that, in an Alzheimer’s mouse model, genetic deletion of TNF receptor (TNFR1) reduces amyloid plaques and amyloid beta peptides (Aβ) production through β-secretase (BACE1) regulation. TNF-α converting enzyme (TACE/ADAM-17) does not only cleave pro- TNF-α but also TNF receptors, however, whether the TACE activity was changed in the CSF was not clear. In this study, we examined TACE in the CSF in 32 AD patients and 27 age-matched healthy controls (HCs). Interestingly, we found that TACE activity was significantly elevated in the CSF from AD patients compared with HCs. Furthermore, we also assayed the CSF levels of TACE cleaved soluble forms of TNFR1 and TNFR2 in the same patients. We found that AD patients had higher levels of both TACE cleaved soluble TNFR1 (sTNFR1) and TNFR2 (sTNFR2) in the CSF compared to age- and gender-matched healthy controls. Levels of sTNFR1 correlated strongly with the levels of sTNFR2 (rs = 0.567-0.663, p < 0.01). The levels of both sTNFR1 and sTNFR2 significantly correlated with the TACE activity (rs = 0.491-0.557, p < 0.05). To examine if changes in TACE activity and in levels of cleaved soluble TNFRs are an early event in the course of AD, we measured these molecules in the CSF from 47 subjects with mild cognitive impairment (MCI), which is considered as a preclinical stage of AD. Unexpectedly, we found significantly higher levels of TACE activity and soluble TNFRs in the MCI group than that in AD patients. These results suggest that TACE activity and soluble TNF receptors may be potential diagnostic candidate biomarkers in AD and MCI.

for more information on the Roskamp Institute and Alzheimer’s please visit:

http://www.mullanalzheimer.com

http://www.mullanalzheimer.info

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Genetic deletion of TNF receptor suppresses excitatory synaptic transmission via reducing AMPA receptor synaptic localization in cortical neurons.

The distribution of postsynaptic glutamate receptors has been shown to be regulated by proimmunocytokine tumor necrosis factor α (TNF-α) signaling. The role of TNF-α receptor subtypes in mediating glutamate receptor expression, trafficking, and function still remains unclear. Here, we report that TNF receptor subtypes (TNFR1 and TNFR2) differentially modulate α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) clustering and function in cultured cortical neurons. We find that genetic deletion of TNFR1 decreases surface expression and synaptic localization of the AMPAR GluA1 subunit, reduces the frequency of miniature excitatory postsynaptic current (mEPSC), and reduces AMPA-induced maximal whole-cell current. In addition, these results are not observed in TNFR2-deleted neurons. The decreased AMPAR expression and function in TNFR1-deleted cells are not significantly restored by short (2 h) or long (24 h) term exposure to TNF-α. In TNFR2-deleted cells, TNF-α promotes AMPAR trafficking to the synapse and increases mEPSC frequency. In the present study, we find no significant change in the GluN1 subunit of NMDAR clusters, location, and mEPSC. This includes applying or withholding the TNF-α treatment in both TNFR1- and TNFR2-deleted neurons. Our results indicate that TNF receptor subtype 1 but not 2 plays a critical role in modulating AMPAR clustering, suggesting that targeting TNFR1 gene might be a novel approach to preventing neuronal AMPAR-mediated excitotoxicity.-He, P., Liu, Q., Wu, J., Shen, Y. Genetic deletion of TNF receptor suppresses excitatory synaptic transmission via reducing AMPA receptor synaptic localization in cortical neurons.

for more information on the Roskamp Institute and Alzheimer’s please visit:

http://www.mullanalzheimer.com

http://www.mullanalzheimer.info

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Proteomic CNS profile of delayed cognitive impairment in mice exposed to gulf war agents.

Gulf War Illness (GWI) is a chronic multisymptom condition with a central nervous system (CNS) component, for which there is no treatment available. It is now believed that the combined exposure to Gulf War (GW) agents, including pyridostigmine bromide (PB) and pesticides, such as permethrin (PER), was a key contributor to the etiology of GWI. In this study, a proteomic approach was used to characterize the biomolecular disturbances that accompany neurobehavioral and neuropathological changes associated with combined exposure to PB and PER. Mice acutely exposed to PB and PER over 10 days showed an increase in anxiety-like behavior, psychomotor problems and delayed cognitive impairment compared to control mice that received vehicle only. Proteomic analysis showed changes in proteins associated with lipid metabolism and molecular transport in the brains of GW agent-exposed mice compared to controls. Proteins associated with the endocrine and immune systems were also altered, and dysfunction of these systems is a prominent feature of GWI. The presence of astrogliosis in the GW agent-exposed mice compared to control mice further suggests an immune system imbalance, as is observed in GWI. These studies provide a broad perspective of the molecular disturbances driving the late pathology of this complex illness. Evaluation of the potential role of these biological functions in GWI will be useful in identifying molecular pathways that can be targeted for the development of novel therapeutics against GWI.

for more information on the Roskamp Institute and Alzheimer’s please visit:

http://www.mullanalzheimer.com

http://www.mullanalzheimer.info

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Institute targets in on Alzheimer’s

May 7, 2007 – At St. James Hospital in Dublin, Ireland, a nurse is checking the blood pressure of a 62-year-old man. Later, a tech will use a hand-held scanner to measure blood flow in his middle cerebral artery.

The patient has never met Michael Mullan, nor Fiona Crawford. But some 4,150 miles away, they could hardly be happier about his test results.

They show that the man, whose identity is confidential, is showing no adverse reactions to a drug that might be a promising treatment for Alzheimer’s disease.

That is good news for Mullan and Crawford. Directors of the Roskamp Institute, on Whitfield Avenue in southern Manatee County, they are running a clinical trial of Nilvadipine, a drug widely used abroad but not approved in the United States.

The drug is intended to lower blood pressure. But in studies on mice, Roskamp researchers found it also reduced the level of a protein in the brain believed to be at the heart of Alzheimer’s.

The Ireland trial just completed its second phase, a safety study, and the 20 patients taking the drug showed almost no ill effects, paving the way for a broader study.

The clinical trial is the institute’s highest-profile project, but its two dozen scientists also work in fields including drug addiction and head injury.

With even more promising treatments deep in its labs, the little nonprofit academic center is becoming more and more a biotechnology start-up.

Birth of the institute

The institute was born in 1997, but its origins date back another 40 years.

Bob Roskamp was a high school physics teacher. His older brother, diagnosed with schizophrenia, killed himself.

Roskamp tells the story with understatement: “Obviously, it catches your attention in a big way.”

The loss of his brother prompted him to give up teaching and work on opening homes for developmentally disabled adults. He later moved into developing housing communities for seniors.

He began selling his companies — including his Freedom Group to American Retirement Group for $23 million in cash and $14.9 million in stock — leaving him with a question: “What do we do with these surplus dollars?”

The answer is in the logo of the institute bearing his name: “Curing Diseases of the Mind.”

Roskamp and his wife, Diane, are among the pioneers of a style of philanthropy, now more widely followed, of carefully aiming resources at one target and tracking performance.

“Instead of spreading it thinly throughout the world, we would ‘rifle-shot’ it, and have a lot more fun with it,” he said.

Roskamp historically has given the institute about one-third of its operating budget, the rest coming through grants and contracts.

But as federal money has been diverted elsewhere, and the institute’s work has grown more complex, Roskamp has had to pick up more of the tab. He gave just over $1 million in 2005, but $4.6 million this year, he said.

His involvement began with funding a lab at the University of South Florida, then a teaching position eventually filled by Mullan.

In 2003, the institute left the university amid personnel and bureaucratic disputes and moved into the former Bausch & Lomb building in southern Manatee County. Roskamp said university overhead was too expensive, and that scientists can work faster in the independent setting.

“You bring your entrepreneurial hat to this kind of research,” he said. “We’re results-oriented people.”

The scientists appreciate not having to wait for grants and approvals.

“We have the funding from Bob and Diane that allow us to move forward very quickly when we make a new discovery,” Crawford said. “We are really free to focus on the research.”

Paddling mice

Most Alzheimer’s researchers now focus on a protein known as beta-amyloid that forms when a chemical process goes awry. The protein clumps together outside neurons, eventually killing those memory cells.

Mullan and Crawford were part of a team that in 1991 published papers establishing a link between a human gene, on chromosome 21, and early onset of the disease in patients in their 40s and 50s.

A year later, after they came to the United States, they published a paper looking at another genetic link to early-onset Alzheimer’s.

The research made beta-amyloid a prime suspect in Alzheimer’s, and stopping its buildup a potential cure.

Roskamp researchers look for existing drugs and new compounds with that effect. Nilvadipine showed enough promise to test it on lab mice engineered to have Alzheimer’s.

At 10 months, with the disease in full bloom, some mice got the drug while others got a placebo. Later, they scanned the mice’s brains for blood flow.

In the color-coded pictures, the brighter the image, the better the blood flow. The mice getting Nilvadipine scan largely red and orange.

The control animals scan mostly blue-green.

Mullan points to a chart with results of a more traditional test, tracking a mouse’s effort to paddle its way out of a mouse-sized swimming pool.

The mice repeat the test, day after day. Normal mice figure it out and eventually swim a shorter route to the exit.

Mice with Alzheimer’s disease never figure it out, actually taking longer and longer routes.

“But if you give them Nivaldipine, they go from here to here; they do better,” Mullan said, pointing at the figures. “And if you give Nilvadipine to a normal mouse, they do best of all.”

The drug appears to slow or stop the production of beta-amyloid, Mullan said.

As the brain grudgingly yields the mysteries of its architecture, Roskamp’s scientists are exploring how those diseases of the mind may be related by more than just words.

‘Blocking that burst’

In 2002 the institute teamed with the Department of Veterans Affairs to look at memory skills in people who had received traumatic brain injuries.

They found that people who had a gene linked to Alzheimer’s disease recovered less memory function than those without it, even when they had comparable injuries and demographics.

Further research has shown that when the brain suffers such an injury, there is a burst of amyloid production. One theory is that the brain produces the protein as a defense mechanism, but it sometimes overreacts and makes matters worse.

“Blocking that burst appears to be at least one way to try and improve outcomes after head injury,” said Crawford, lead author of the 2002 article.

Last year the Department of Defense approved a $1.5 million grant to the institute to expand that research. It is a particularly urgent matter for the military: Improved body armor is helping soldiers survive explosions — but with little-understood head injuries.

“People can have brain injury even if they didn’t suffer any loss of consciousness, weeks and months of being in the vicinity of an explosive attack,” Crawford said.

For now, that grant is hung up in final reviews.

“We have yet to see dime one,” Roskamp said — but even that $1.5 million pales in comparison with the costs of the Nilvadipine study.

That cuts to the heart of the dilemma scientists face. They might get a moderately effective drug to market quickly, or try to discover the “silver bullet,” which could take many years and exponentially more money.

Roskamp is preparing the foundation to do both.

Just a question of when

To Mullan, the question is not whether there will be treatments for Alzheimer’s.

“It’s the question of when we’re going to get them there,” he said.

It is a question of how effective they will be. Mullan cited three drugs far along in the pipeline, all targeted at reducing amyloid. “They’re not going to be block-busting, perfect drugs by any means, but I think at least two of them will be approved for Alzheimer’s this year.”

Nilvadipine most likely would be like them — useful but not the cure-all. But it offers some advantages.

Developing a new drug from scratch is a much worse bet, Mullan said. Count on 10 to 12 years for development, testing and approvals, with a high failure rate.

“Sometimes they fail spectacularly,” he said, citing Vioxx, a promising drug with unforeseen and deadly side effects.

Nilvadipine, in use for more than a decade, already has a track record for safety. It could help patients while allowing the institute time to find the silver bullet.

By mid-year, the institute hopes to continue the Ireland trial with a double-blind study, meaning neither patients nor doctors know who gets Nilvadipine and who gets a placebo until they conclude. That study, which would involve about 200 people, could cost $20 million, Roskamp said.

Should the drug prove effective, Nilvadipine’s manufacturer could profit — but so would the institute. Because it holds rights to the drug’s newly indicated use, the institute would receive royalty payments, Roskamp said.

The institute has set up a for-profit subsidiary, Roskamp Research LLC, which holds its patents. Roskamp structured it to allow outside backers to support studies, provide the subsidiary with a return on its investment, and guarantee most of the revenue goes to his institute.

“We don’t want to give it to a pharma company, and say here, go make a lot of money off of it,” Roskamp said.

Compounds back in the institute’s labs could provide better returns — and better treatments.

for more information on the Roskamp Institute please visit:

http://www.mullanalzheimer.com

http://www.mullanalzheimer.info

www.rfdn.org

The Roskamp Institute

The Roskamp Institute, a not-for-profit research Institute, is dedicated to finding cures for neuropsychiatric disorders, with the emphasis on Alzheimer’s disease (AD).  Current research at the Roskamp Institute is focused on dissecting the molecular biological pathways implicated in AD pathogenesis in order to develop therapeutic targets specific to AD etiology. Dr. Michael Mullan (Director of the Roskamp Institute) and Fiona Crawford (Associate director of the Roskamp Institute) were part of the original team that discovered a genetic error called the “Swedish mutation” which results in overproduction of  b-amyloid (Ab) by aberrant proteolytic processing of the amyloid precursor protein (APP). This mutation now forms the bases of most mouse model of Alzheimer’s disease. Previously, the Roskamp Institute published an article in a prestigious scientific journal, Nature, showing that Ab plays a normal role in vasoactive mechanisms but also plays a role in vascular abnormalities and neurodegeneration mediated by free radical. Subsequently, Dr. Daniel Paris, a senior scientist at the Roskamp Institute, discovered that the vasoactive effects of Ab are partly mediated via a pro-inflammatory pathway and showed that this effect of Ab on the vasculature can be blocked by inhibiting specific target molecules.  In order to further understand the role of Ab in the vasculature, Dr. Paris investigated the long term effect of Ab on vascular homeostasis. He then discovered that at low doses, Ab promotes angiogenesis, while at high doses, certain forms of Ab peptides are anti-angiogenic. Collectively, these novel findings resulted in new therapeutic prospects for the treatment of Alzheimer’s disease as well as Cancer.

Researchers at the Roskamp Institute also showed that the presence of functional CD40/CD40L signaling is essential for the full development of AD like pathology in transgenic mouse models of AD. In particular, it was demonstrated that accumulation of cerebral Ab is reduced in transgenic mouse models of AD by genetically or pharmacologically reducing the availability of CD40L to CD40. The Roskamp Institute investigators subsequently revealed that loss of functional CD40L diminishes both APP processing to Ab and microglial activation in the brain (Original findings published in journals Science and Nature Neuroscience). CD40L activated pathways in the presence of Ab appear to mediate both of these effects as well as the hyperphosphorylation of murine tau in vivo at epitopes analogous to those which precede tangle formation of human tau. More recently, Dr. Ghania Ait-Ghezala of the Roskamp Institute showed that CD40/CD40L interaction also affects APP via the NF-kB pathway. Using NF-kB inhibitors and SiRNAs to silence diverse elements of the NF-kB pathway, she demonstrated that reduction in levels of both pathological forms of Ab. These results showed that CD40L stimulation may be a key component in AD pathology and that NF-kB pathway may be suitable targets for therapeutic approaches against AD.

Another major focus of research at the Roskamp Institute includes Traumatic Brain Injury (TBI) Program headed by the Associate Director of the Roskamp Institute. Dr. Crawford and her Roskamp Institute team demonstrated an important relationship between apolipoprotein E (APOE) and memory following TBI. She demonstrated that in Veteran’s with TBI, memory performance was significantly worse in individuals who had at least one copy of APOE ε4 allele than those who did not. She had subsequently been funded through the Veteran’s Administration to further study the relationship between different forms of APOE in TBI with the emphasis on finding treatments for this devastating condition.

Drs. Michael Mullan and Fiona Crawford also received funding by the Counterdrug Technology Assessment Center (CTAC) to evaluate the newly emerged genomics and proteomics technology and find biological markers of substance abuse. Recently, Dr. Crawford’s team showed that cocaine treatment of human progenitor neuronal cells results in increased oxidative stress (possibly mediated by inflammatory responses) which precedes cell death. Thus, these findings may have implications for the consequences of cocaine abuse in situations where antioxidant capacity is compromised, as in the aging brain.

As evident here, the Roskamp Institute team has been a pioneer in many area of research in neuropsychiatric diseases and will continue to do so to find novel therapies for these disorders. Currently, a new clinical trial based on the discoveries made at the Roskamp Institute is underway to assess safety and efficacy of nilvadipine in treatment of Alzheimer’s disease. Through the generous support of Diane and Robert Roskamp, the Veteran’s Administration, the National Institutes of Health, CTAC and the Department of Defense, the Roskamp Institute will continue to provide potential avenues for novel therapeutic interventions for neuropsychiatric disorders.

 

for more information on the Roskamp Institute please visit:

http://www.mullanalzheimer.com

http://www.mullanalzheimer.info

www.rfdn.org

Roskamp Institute on Gulf War Illness

The Roskamp Institute located in Sarasota, Florida is the leading research center in the area.  The research focuses on neurological diseases that affect young and elderly men and women of this world. Some of the main focuses are Alzheimer’s disease, addiction, post-traumatic stress disorder (PTSD), multiple scoliosis, Gulf War syndrome and many more. The Director Dr. Michael Mullan leads his team of researchers to exciting findings. Recently the Institute has been given a grant from the VA for their research involving Gulf War Illness. This Illness that inflicts the soldiers that fought during the Gulf War affects their motor skills, memory, stability and other problematic symptoms. How the Illness commenced was through the combination of the neurotoxins and pesticides given to the soldiers. The research done in the Institute led to the finding that shows the toxic mixture affects the long fatty acid chains within the brain. Research continues within the walls of the 42,000 sq. ft research facility hopefully terminating the life altering Illness for our Veterans.

 

for more information on the Roskamp Institute please visit:

http://www.mullanalzheimer.com

http://www.mullanalzheimer.info

www.rfdn.org

Roskamp Institute Scientists Discover Genetic and Biological Consequences of Substance Abuse

A 2004 National Survey on Drug Use and Health reported that about 34 million Americans have had some exposure to cocaine at least once in their lifetime. The addictive properties of this drug is well recognized and results in an increased direct cost in medical care as well as indirect cost due to increased societal burden. The Roskamp Institute, located in Sarasota Florida is currently engaged in research into discovery of biological biomarkers and novel treatments for substance abuse. Michael Mullan MD PhD (Director of the Roskamp Institute), during his medical training in London, worked on addiction and determined that certain addictive disorders had a significant genetic component. It is now well-established that substance dependency has a prominent genetic component and approximately 40-60% of the vulnerability to substances can be sufficiently attributed to these genetic factors. Previously, the Roskamp Institute scientists demonstrated that a polymorphism in gene for mu-opioid receptor (OPRM1 +118A) is a risk for alcohol dependency. This Roskamp Institute team then examined the frequency of OPRM1 +118A carrying genotypes and alleles in several groups of substance-dependent cases compared to individuals with no history of reported substance abuse. These Roskamp Institute scientists found that the OPRM1 +118 polymorphism is a genetic risk factor for substance dependence but not specific to a particular substance (findings published in the journal Molecular Psychiatry).

Dr. Fiona Crawford (Associate director, Roskamp Institute) and Dr. Michael Mullan (Director, Roskamp Institute) received an award from the Counterdrug Technology Assessment Center (CTAC) to evaluate the newly emerged genomics and proteomics technologies and determine their usefulness in finding potential biomarkers and treatments for substance abuse.  Dr. Fiona Crawford along with her Roskamp Institute team using this genomic technology investigated the fundamental transcriptional responses occurring in neurons as a consequence of acute cocaine exposure over a time period. These Roskamp Institute scientists used GeneChip Operating Software from Affymetrix to compare the genomic response in neuronal cells exposed to cocaine compared to cells that were not exposed and showed a time-dependent increase of genes associated with pro-inflammatory and immune responses. These findings suggest that the inflammatory and immune systems maybe involved in modulating response to an acute cocaine exposure (originally published in the Journal of Neurochemistry).

Next, this Roskamp Institute team determined a role of oxidative stress in cocaine exposure in human progenitor neuronal cells. Although, it is evident that cocaine induces oxidative stress in the central nervous system, little is known whether such increase in oxidative stress is also relevant to cell death in cocaine-exposed models. To gain further insight into the role of cocaine-induced oxidative stress, the Roskamp Institute scientists hypothesized that oxidative stress precedes cell death upon cocaine exposure and demonstrated that oxidative stress was significantly increased in neuronal cells treated with cocaine and this phenomenon preceded cell death. Therefore, these findings have implications for cocaine abuse in circumstances where antioxidant system is compromised, as in the aging brain (original findings in the Journal of Neurochemistry International).

The addictive properties of these psychoactive substances is well recognized and results in enormous burden of direct and indirect costs to the US economy. The Roskamp Institute is dedicated to understanding the factors that predispose individuals to substance abuse as well as discover additional molecular targets for therapeutic intervention.

for more information on the Roskamp Institute please visit:

http://www.mullanalzheimer.com

http://www.mullanalzheimer.info

www.rfdn.org