FUNDING PERIOND APRIL 1, 2002 - MARCH 31, 2004

Yuzhi Chen, Ph.D.
McLean Hospital
Belmont, Massachusetts
Project: APP-BP1, the Cell Cycle and Alzheimer's Disease

There is increasing evidence that neurons in Alzheimer's disease (AD) enter the cell cycle (a period of DNA replication leading up to cell division), but because they are locked into a non-dividing state, the attempt to divide causes them to die. Work from many investigators suggests that the amyloid precursor protein (APP) is a signaling molecule whose function may be to regulate cell death. Dr. Chen has found a cell cycle protein called APP-BP1 that binds to APP, and its over-expression in neurons leads to cell death. The present work is based on the hypothesis that AD results from a gradual dysfunction of the APP interaction with APP-BP1, and this dysfunction leads to neuronal death by the abnormal signaling through APP. Dr. Chen will examine the properties of APP and APP-BP1 to identify the protein domains and their function; he will determine where these proteins are located within the cells; he will over-express the proteins in neurons in culture to determine their function; and he will determine if other proteins bind to APP-BP1 and play a role in AD. The data obtained from these studies will help to understand the normal function of APP and AP-BP1 in the brain and identify how these proteins may malfunction in AD resulting in cell death.

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Yoon Cho, Ph.D.
University of Bordeaux I
Talence Cedex, France
Project: Behavioral and in vivo Electrophysiological Characterization of APP Transgenic Mice

Although amyloid beta (Aß) deposits in the brain constitute a neuropathological hallmark of Alzheimer's disease, the role these plaques play in the onset and progression of the disease is unknown and controversial. Transgenic technologies provide powerful tools for studying this phenomenon, and a number of mouse models have been created that develop amyloid plaques similar to those in patients with Alzheimer's disease. Current evidence suggests that the transgenic mouse models exhibit behavioral and cognitive abnormalities that might underlie deficits in learning and memory that are early diagnostic features of the disease. However, there has been no direct evaluation to determine the effects of amyloid accumulation in the brain on cellular functioning in live animals. This proposal uses state-of-the-art electrophysiological recording and behavioral techniques to examine the physiology of behavior in brain areas such as the hippocampus, a region that is critical for certain forms of memory and also susceptible to amyloid plaque deposition. The goal of this study is to provide a comprehensive characterization of the effects of amyloid deposition on behavior and on perturbations of normal cellular function in life that produce memory deficits. In addition, this approach should provide a means to assess the time of onset at which the first subtle signs of memory loss appear as the disease slowly progresses.

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Mony de Leon, Ed.D.
New York University
New York, New York
Project: The Preclinical Diagnosis of AD

The accurate preclinical diagnosis of late-onset Alzheimer's disease is a necessary goal. Studies have shown that damage to the hippocampus and entorhinal cortex, the major memory processing center of the brain, occur early in the course of AD. Imaging studies have shown that reduction in the size of the hippocampus can assist in predicting the decline from mild cognitive impairment (MCI) to AD. In addition, the tau protein and amyloid beta peptide levels can also serve as early diagnostic markers of AD. These markers are potentially useful for the identification of early AD pathology; however, there remains considerable uncertainty regarding their predictive value or their capacity to monitor the course of AD. Dr. de Leon is conducting a longitudinal study of normal and MCI individuals to demonstrate that combining information from imaging studies and biochemical markers improves both sensitivity and specificity for the prediction of cognitive decline and helps characterize early brain changes observed in AD. His ultimate goal is to identify different markers and characterize their relationships in order to improve the early diagnosis of AD.

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Sylvain Dore, Ph.D.
The Johns Hopkins University
Baltimore, Maryland
Project: Role of Heme Oxygenase 2 Interactions in AD Models

The onset and progression of Alzheimer's disease (AD) can be quite varied. To date, it is very difficult to predict an individual's risk or even to detect the preliminary signs of the condition before dementia appears. Dr. Dore has been studying the neuronal form of a protein called heme oxygenase (HO2), which may play a pivotal role in AD. It is already believed that oxidative stress plays a role in AD, and that certain types of highly reactive oxygen species (ROS) cause cellular damage. The control of the production and excretion of these molecules is necessary to prevent cellular damage and is maintained by antioxidants. HO2 has antioxidant properties and also binds to amyloid precursor protein (APP). Furthermore, an essential component for HO2 activity is cytochrome P450 reductase (CPR) and CPR is a key element in generating ROS. Dr. Dore hypothesizes that CPR, in conjunction with HO2, regulates the generation of ROS and is important in AD. The objective of this research is to characterize the interactions between APP and HO2 using cell culture systems and transgenic mouse models. New mouse models lacking HO2 will be bred, and it is expected that significantly more plaques will be apparent in mice with HO2 deletions compared to normal mice. Dr. Dore will also examine the contribution of CPR in the ROS damage associated with AD. He believes that a better understanding of the interactions between APP, HO2 and CPR will help elucidate the contribution of neuronal oxidant stress damage and its role in AD.

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Joseph El Khoury, M.D.
Massachusetts General Hospital
Charlestown, Massachusetts
Project: Mechanism of Microglial Recruitment in Alzheimer's Disease

The senile plaque, composed of aggregations of beta amyloid protein, is a pathological hallmark of Alzheimer's disease. Accumulation of beta amyloid in the brain leads to the activation of microglia cells. Microglia cells are inflammatory cells that can produce substances that are toxic to neurons and lead to neuronal degeneration. Studies have shown that there are key steps for neuronal damage from the accumulation of microglia in senile plaques. First, the binding of microglia to beta amyloid occurs. Second, microglia are induced to produce chemoattractants that recruit additional microglia to the beta amyloid deposit. Finally, the production of neurotoxins from the recruited microglia cause cellular damage. Dr. El Khoury is studying the mechanisms of the recruitment of microglia to senile plaques in AD using an in vitro cellular model for microglia interactions, post-mortem human brain tissue from AD patients and transgenic mice. He plans to identify chemoattractants produced by microglia and the receptors that are involved, the expression of the chemoattractants and their receptors in brain tissue and in mouse models of AD, and the effects of disrupting the expression of chemoattractants and the receptors in the recruitment of microglia. Understanding the mechanisms of recruitment of microglia to plaques and how they become activated to produce neurotoxins may help to identify new targets for the development of treatments for AD.

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Christian Haas, Ph.D.
Ludwig Maximilians University
Munich, Germany
Project: Generation, Metabolism and Biological Function of AICD

Presenilin (PS) supports the Intramembraneous cleavage of several substrates, including Notch I-IV and the ß-amyloid precursor protein (ßAPP). In the case of Notch, the PS-supported cleavage results in the production of the Notch intracellular domain (NICD), which is required for signal transduction and the regulation of target gene transcription. In recent work, Dr. Haas and his colleagues identified the corresponding fragment of ßAPP called the amyloid precursor protein intracellular domain (AICD) and found that it is generated by a molecular mechanism that is very similar to NCID. He is now investigating the molecular mechanisms of AICD generation and analyzing the putative similarities of AICD and NICD generation as well as the influence of familial Alzheimer's disease-related mutations on the production of these two cytoplasmic fragments. It is hoped that the expression of the recombinant fragment in tissue culture cells under conditions where its degradation is blocked will help identify genes that are involved. Based on the phenotype obtained from Caenorhabditis elegans (a worm) and zebrafish, conclusions regarding the target genes can be made and genetic modifiers can be isolated. If successful, this project will not only help to identify a biological function of ßAPP but is also important for the development of therapies to block the generation of amyloid ß-peptide using gamma-secretase inhibitors.

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Bruce Lamb, Ph.D.
Case Western Reserve University
Cleveland, Ohio
Project: Genetic Control of Amyloid Precursor Protein Processing

Clinical and neurological studies suggest that human Alzheimer's disease (AD) is highly variable in terms of the length, severity and age-related progression of the disease. Accumulating evidence suggests that its development and progression is subject to a variety of environmental and genetic influences. Studies have shown that early-onset familial AD may be caused by mutations in various AD genes. These genetic forms of AD all share a common pathology that involves alterations in the processing and metabolism of the amyloid beta peptide (Aß). The release of Aß from its precursor and the production, transport, turnover and deposition of Aß in the brain is still unclear. Dr. Lamb is focusing on a unique animal model for AD with entire copies of human AD genes placed in mice. These "genomic-based" transgenic animals will be used to study the processing and metabolism of Aß in mice with different genetic backgrounds. The long-term goal of this proposal is to complement genetic studies in humans to identify novel genetic pathways involved in the modulation of Aß processing and metabolism.

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Philippe Marabaud, Ph.D.
Mount Sinai School of Medicine
New York, New York
Project: Role of Presenilin-1-Mediated Cleavage of E-Cadherin

Mutations in presenilin-1 (PS1) are responsible for familial Alzheimer's disease (FAD), also known as early-onset Alzheimer's disease. PS1 is found at the plasma membrane and is involved in cell-cell contacts. It interacts with E-cadherin, a cell surface molecule that facilitates cell-cell adhesions and has a crucial role in the structure of the synapse, the region involved in neuron communication. E-cadherin forms complexes with cytosolic proteins, like catenins, to link the actin cytoskeleton. The remodeling of cell-cell interactions is a central determinant for many functions, including tissue repair, cell migration and cell death. The molecular mechanisms involved in the disassembly of cell-cell associations are not clearly understood. It has been shown E-cadherin processing is mediated by PS1/gamma secretase activity. Dr. Marambaude is investigating whether PS1-mediated cleavage of E-cadherin disconnects the cadherin with the cytoskeleton and plays a role in the disassembly of cell-cell adhesions. He also plans to study the signaling pathways that are activated by the PS1/gamma secretase-mediated processing of E-cadherin and the subsequent disassembly of cadherin/catenin complexes. Finally, he will examine whether mutated PS1 in FAD abnormally affects its activity to cleave E-cadherin and thereby participating in the neuronal loss observed in Alzheimer's disease. This project will help to determine the usefulness of treatments using gamma-secretase inhibitors in altering the E-cadherin-dependent adhesion and signaling systems.

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Hemant Paudel, Ph.D.
Lady Davis Institute for Medical Research
Montreal, Quebec, Canada
Project: Disequilibrium of Tau Protein Phosphorylation and Alzheimer's Disease

In Alzheimer's disease, neurofibrillary tangles (NTs) develop in nerve cells undergoing degeneration, and their distribution provides a reliable correlation to the degree of dementia. The molecular basis for the formation of NTs is not known. NTs contain paired helical filaments (PHFs) as the major fibrous component. The microtubule-associated protein tau is the major constituent of PHFs. It has been shown that tau abnormalities lead to neurodegeneration, microtubule disruption and dementia. This is thought to result from an imbalance in the phosphorylation of tau. The regulatory mechanisms that control tau phosphorylation and the determination of how this regulation fails in AD are not known. Dr. Paudel has recently identified a multiprotein complex in the brain that might be involved. He plans to characterize the proteins within this complex and determine their activities in tau phosphorylation with the goal of understanding the cascade of events that leads to abnormal tau phosphorylation, tau aggregation, microtubule disruption and neurodegeneration. Completion of this study will provide important information about the biochemical and cellular mechanisms of NT formation in the AD brain and help to design future treatment strategies to retard NT formation in the brains of AD patients.

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Salvador Soriano-Castell, Ph.D.
University of California at San Diego
San Diego, California
Project: Effects of Presenilin 1 Mutations on b-Catenin Signaling

The death of neurons in the brain is a symptom of Alzheimer's disease (AD) that is believed to be a major factor in the cognitive impairment affecting Alzheimer's patients. The death of neurons may result from the accumulation of plaques between neurons as well as from the formation of neurofibrillary tangles within neurons. However, why these factors are toxic is not known. One theory is that the high-stress environment found in AD can trigger neurons into starting cell division, but the cells lack the appropriate components to divide and, as a consequence, the process results in cell death. Dr. Soriano-Castell will focus on the early-onset form of familial AD that is linked to mutations in the presenilin proteins (PS). He proposes that PS1 deficiency and PS1 mutations upregulate b-catenin signaling in the adult brain. This leads to increased levels of the cyclin D1, which is a component used by neurons in their attempt to start dividing in response to environmental challenge. The high levels of cyclin D1 are thought to contribute to abnormal cell division and cell cycle signaling, resulting in the neuronal loss seen in AD. Dr. Soriano-Castell is studying transgenic animals to determine whether b-catenin signaling through cyclin D1 and abnormal cell divisions occur in the neurons of animals with AD. Understanding what causes neuronal cell death will help in designing treatments to prevent neuronal death or delay it sufficiently to improve the quality of life for Alzheimer's patients.

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David Sweatt, Ph.D.
Baylor College of Medicine
Houston, Texas
Project: Alpha7 Nicotinic Receptors and MAP Kinase in AD Models

Early stage Alzheimer's disease is generally observed as hippocampal dysfunction with the inability of the brain to consolidate short-term into long-term memories. These memory deficits are typically present before any generalized dementia or overall cognitive decline is observed. Dr. Sweatt hypothesizes that for the early stages of AD, which are characterized by subtle deficits in memory consolidation; there will be abnormalities in the normal biochemical machinery that underlies memory. He suggests that the MAP kinase signaling cascade plays a critical role in learning and memory. He is examining the transgenic mouse model of AD for MAP kinase signaling defects. He will also examine how the amyloid beta peptide is linked to the MAP kinase cascade. Preliminary results suggest that a receptor for amyloid beta peptide on the cell surface of neurons is the alpha7 nicotinic acetylcholine receptor. Dr. Sweatt will use genetic approaches to examine whether deletion of the alpha7 receptor gene leads to amelioration of memory deficits exhibited in transgenic mice with AD and decreases in biochemical markers. The identification of the alpha 7 receptor involvement in AD allows for the future study of agents that block the receptor as potential therapeutics for AD.

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Jianjun Wang, Ph.D.
Southern Illinois University
Carbondale, Illinois
Project: NMR Studies of Apolipoprotein E-beta 4 Interactions

The way that amyloid beta (Aß) peptide is deposited into insoluble plaques in Alzheimer's disease is still unknown. There is some evidence that apolipoprotein E (apoE) binds to Aß peptide and modulates its aggregation into plaques. ApoE has three isoforms, apoE2, apoE3 and apoE4, and reports have shown that each apoE form may affect Aß aggregation in a specific manner. It has been suggested that apoE4 increases amyloid deposition, and it is commonly accepted that the conversion of Aß protein from a soluble form to an insoluble aggregate is associated with a structural or conformational change in the protein. The Aß peptide changes from an alpha-helix random coil form to a beta sheet structure. Dr. Wang suggests that apoE may play an important role in this conversion. He is using nuclear magnetic resonance (NMR) techniques to study the binding of Aß and apoE and the subsequent conformational changes to Aß. The primary goal of this project is to determine if apoE serves to promote or inhibit Aß conversion to the aggregated insoluble form. This could lead to developing inhibitors for Aß peptide aggregation and, ultimately, to better treatments or even a way to prevent AD.

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