Regulation of the amyloid precursor protein processing by the 5-HT4 receptor and small G proteins

Abstracts: Pharmacological Treatments / 1 (Suppl 1) (2005) and apoE4). ApoE4 results in an increased and apoE2 a decreased risk for AD relative to apoE3. ApoE is a 299 amino acid protein that is the most abundant apolipoprotein in the central nervous system (CNS). In the CNS, it is primarily synthesized by glial cells and secreted in high density lipoprotein (HDL) like particles that contain cholesterol and phospholipids. Objective(s): While the mechanism(s) underlying apoE’s involvement in AD and CAA pathogenesis remains unknown, abundant evidence suggests that a major mechanism is via apoE’s ability to act as a chaperone for the amyloid-␤ (A␤) peptide to modulate its clearance as well as the likelihood that it will aggregate in the brain. Our objective has been to understand at an anatomic, cellular, and molecular level how interactions between apoE and A␤ predispose to AD and CAA as this may lead to novel diagnostic and treatment strategies. Methods: Using animal models that develop AD-like pathology as well as CAA, we have found that apoE appears to be involved in clearance of soluble forms of A␤ in the brain. In addition, apoE isoforms influence the development of A␤-containing plaques in brain parenchyma and CAA with the order of the effect being E4E3E2. Evidence suggests that increased expression of apoE2 and apoE3 decrease the amount of A␤ deposition. Understanding molecules and pathways that control the levels and expression of apoE may be novel targets for therapy. We have found that the protein ABCA1 is important for the normal production and levels of plasma HDL and is also critical for maintaining normal levels of apoE and apoE-associated cholesterol in the brain. Conclusions: Determining methods to alter expression or function of ABCA1 and/or other proteins that regulate apoE levels and its state of lipidation may be important future therapeutic targets to prevent, delay, and treat AD and CAA. PL2-02

SOLUBLE ABETA OLIGOMERS (ADDLs) IN ALZHEIMER’S DISEASE MEMORY LOSS, THERAPEUTICS, AND DIAGNOSTICS

William Klein; Northwestern University, Evanston, IL, USA Early diagnosis and treatment of AD ultimately will derive from identification of pathogenic mechanism. Such a mechanism must explain the singular hallmark of early AD - a profound inability to form new memories. For many years, the most promising hypothesis maintained that memory failure derived from neuron death, induced by insoluble deposits of amyloid fibrils. However, as recognized by neuropathologists as early as 1975, cognitive deficits in AD might derive as well from neuritic failure, not only neuron death. Significant support for the neuritic failure alternative came in 2002 with the discovery that cognitive loss in hAPP tg-mice is reversible and thus independent of cell death. Recovery of memory, accomplished with antibodies against Abeta, occurred without reduction in amyloid plaque burden. Memory recovery appears to stem from elimination of small soluble Abeta oligomers, also known as ADDLs. ADDLs are known to inhibit synaptic plasticity, are implicated in synapse degeneration, and are strikingly elevated in tg-mice models of AD, in AD brain, and in AD CSF; these various findings led to the hypothesis that memory loss in early AD is an ADDL-mediated synapse failure. This hypothesis has been strengthened by recent cell culture data showing synapses constitute ADDLs’ specific point of attack. Rather than exhibiting non-selective association with cells, ADDLs are ligands that bind with specificity to particular synapses. It is likely that this pattern found in culture recapitulates the perineuronal dendritic binding of ADDLs seen in brain sections, suggesting that synaptic association of ADDLs gives rise to diffuse amyloid. ADDL targeting of synapses in culture generates ectopic induction of Arc, an immediate early gene whose regulated expression is required for long-term memory formation. Ectopic over-expression of Arc has been linked to dysfunctional learning, potentially due to anomalies induced in receptor trafficking and spine geometry. Synaptic targeting by ADDLs and its impact on memory-linked synaptic events provides an appealing explanation for early AD memory loss and suggests that ADDLs provide a valid target for therapeutics and suitable biomarker for diagnostics.

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MONDAY, JUNE 20, 2005 SYMPOSIA S2-01 STATINS AND OTHER ALPHA-SECRETASE ACTIVATORS S2-01-01

ALPHA-SECRETASE AS A THERAPEUTIC TARGET

Falk Fahrenholz, Rolf Postina, Kristina Endres, Claudia Prinzen, Elzbieta Kojro; Johannes Gutenberg University, Mainz, Germany Background: An increase of the alpha-secretase activity in the brain provides an attractive strategy for the treatment of Alzheimer disease (AD), since proteolysis of the amyloid precursor protein (APP) within the sequence of A-beta peptides precludes their formation. In addition, alphasecretase cleavage releases the N-terminal extracellular domain APPsalpha with neurotrophic and neuroprotective properties. Objective(s): The aim of the investigations is to upregulate the non-amyloidogenic alphasecretase pathway in the brains of AD patients. Methods: Studies with peripheral and neuronal cell lines and with transgenic mice were performed to elucidate signal transduction pathways and to identify compounds which increase the alpha-secretase activity or the expression level. Conclusions: Members of the ADAM family (a disintegrin and metallo-proteinase) can act as alpha-secretases. Even moderate neuronal overexpression of ADAM10 in an AD mouse model increased the production of APPs-alpha, reduced the formation of soluble A-beta peptides, and prevented their deposition in plaques. Functionally, impaired long-term potentiation and cognitive deficits were alleviated. A functional promoter of the human ADAM10 gene was cloned which has its highest activity in neural cell lines. Elements essential for strong promoter activity were characterized, and compounds identified as inducers of human ADAM10 promoter activity. On the level of G-protein-coupled receptors localized in brain areas affected by AD, we identified neuropeptides and the MAP kinase pathway as strong activators of the alpha-secretase. Furthermore, we found upregulation of the alpha-secretase activity in cellular systems after reduction of cholesterol by statins. In summary, several compounds including nutritional factors and endogenous hormones were identified which stimulate the non-amyloidogenic pathway and therefore might have beneficial effects for the treatment of AD. S2-01-02

REGULATION OF THE AMYLOID PRECURSOR PROTEIN PROCESSING BY THE 5-HT4 RECEPTOR AND SMALL G PROTEINS

Frank Lezoualc’h, Marjorie Maillet, Sylvain J. Robert; Inserm U-446, University Paris XI, Faculty of Pharmacy, Chaˆtenay-Malabry, France Background: The serotonin 5-HT4 receptor mediates many physiological effects in the central nervous system. The recent molecular identification of 5-HT4 receptor isoforms with distinct C-terminal tails and the development of selective 5-HT4 receptor ligands have led to many important new insights into the signalling pathways and the physiological roles of these Gs protein-coupled-receptors in neurones. With respect to neurodegenerative disorders, it is suggested that 5-HT4 receptor agonists may represent a new avenue for the treatment of Alzheimer’s disease. Objective: Because strong evidence supports beneficial effects of 5-HT4 receptor agonists in memory and learning, we investigated the potential involvement of the 5-HT4 receptor in the amyloid precursor protein processing (APP). Methods: Experiments were performed in different cell lines and primary culture of embryonic mouse cortical neurons. Conclusions: Activation of the human 5-HT4(e) receptor isoform stimulates the secretion of sAPPalpha. The increase was inhibited by selective 5-HT4 receptor antagonists, GR113808 and SB204070. Secretion of sAPPalpha induced by the h5HT4(e) receptor seems not to be due to a general boost of the constitutive secretory pathway but rather to its specific effect on alpha secretases. In addition, we found that structural differences in the C-terminal tails of 5-HT4 receptor isoforms influence and contribute to the specificity of their functional response on APP processing. An interesting observation is that

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Abstracts: Lifestyle and Other Risk Factors / 1 (Suppl 1) (2005)

the effect of the 5-HT4 receptor on sAPPalpha release is independent of the protein kinase A (PKA). Indeed, the signalling pathway which couples the 5-HT4 receptor to sAPPalpha secretion involves small GTPases of the Ras and Rho families. Activation of the cAMP-regulated guanine nucleotide exchange factor (GEF), Epac stimulates alpha secretase activity through the small GTPases Rap1 and Rac in a cAMP-dependent but PKA-independent manner. These findings may open new avenues for the identification of new targets for the regulation of APP processing. S2-01-03

BRAIN AND BEHAVIOR IN THE FRAMINGHAM HEART STUDY

Charles DeCarli; University of California at Davis, Sacramento, CA, USA Previous studies show strong associations between elevations in middle life blood pressure and the prevalence of later life cognitive impairment and dementia. The mechanisms by which cerebrovascular risk factors lead to cognitive impairment remain unclear, but data suggest that cerebrovascular risk factors are associated with brain injury through accelerated brain atrophy, abnormalities of cerebral white matter and silent stroke that could impact on cognitive performance. Prior studies of the relationship between vascular risk factors and subclinical changes in brain structure and function have several limitations including restrictions to highly select populations such as ‘successful agers,’ or male, World War II veteran twins. Other population studies have used semi-quantitative rather than quantitative MRI techniques and have limited follow-up or prospective data available to associate risk with outcome. The Framingham Heart Study is a communitybased cohort that has been longitudinally evaluated for cardiovascular risk and the development of clinical stroke and dementia since 1949. Beginning in 1999, over 3,000 individuals have had quantitative MRI in conjunction with detailed neuropsychological testing. In this presentation, I will review data showing strong associations between a variety of cerebrovascular risk factors, alterations in brain morphology and cognitive outcome. From these data I will show that cerebrovascular disease remains a common, but preventable disorder where treatment can significantly reduce the risk of future cognitive impairment and dementia thereby yielding large public health benefit. S2-01-04

THE ROLE OF STATINS IN DELAYING ALZHEIMER’S DISEASE. CIRCULATING CHOLESTEROL LEVELS, APOE GENOTYPE AND INITIAL DEMENTIA SEVERITY INFLUENCE THE LEVEL OF BENEFIT PRODUCED BY ATORVASTATIN IN MILD-TO-MODERATE AD: RESULTS OF THE ADCLT

D. Larry Sparks1, Donald J. Connor1, Marwan N. Sabbagh1, Jean Lopez1, Patrick Browne2, Bob Petersen3, Dawn Wasser4, Sherry Johnson-Traver1; 1Sun Health Research Institute, Sun City, AZ, USA; 2Boswell Hospital, Sun City, AZ, USA; 3Case Western Reserve University, Cleveland, OH, USA; 4Del Webb Hospital, Sun City, West, AZ, USA Background: Previous epidemiological studies have suggested that prior statin use may reduce the risk of developing AD later in life. More recent incidence studies question the link between statin use and reduced risk of AD. We have shown that atorvastatin treatment produced significantly (p ⬍ 0.004) improved performance on cognition and memory after 6 months of treatment (ADAS-cog) among patients with mild-to-moderate AD. This superior effect persisted at one-year (p ⫽ 0.055). Evidence also suggests that atorvastatin may slow the progression of mild-to-moderate AD, thereby prolonging the quality of an afflicted individual’s life. Objective: To determine if the clinical benefit of atorvastatin treatment on ADAS-cog performance compared to placebo in mild-to-moderate AD is influenced by severity of cognitive impairment, circulating cholesterol levels, or Apolipoprotein E genotype. Methods: This was a double-blind, placebo-controlled, randomized (1:1) trial with a 1-year exposure to once

daily atorvastatin calcium (80 mg; two 40mg tablets) or placebo among individuals with mild-to-moderate AD (MMSE score of 12-28). Stable dose use of cholinesterase inhibitors, estrogen and vitamin E was allowed, as was the use of most other medications in the treatment of co-morbidities. Participants using cholesterol-lowering medications or being treated for major depression or a psychiatric condition were excluded. Of the 98 participants providing Informed Consent, 67 were randomized and 63 were evaluable. A primary outcome measure was change in the Alzheimer Disease Assessment Scale-Cognitive (ADAS-cog) sub-scale score. Secondary outcome measures included scores on the MMSE, circulating cholesterol levels and Apolipoprotein E genotype. Conclusions: Atorvastatin produced a significant positive effect on the ADAS-cog performance after 6-months of treatment. This positive effect was more prominent among individuals entering the trial with, 1) higher MMSE scores, 2) cholesterol levels above 200 mg/dL or 3) if they harbored an ApoE-4 allele. Individuals in the placebo group tended to perform worse if their cholesterol levels were above 200 mg/dL or they harbored an ApoE-4 allele. Accordingly, atorvastatin therapy may be of benefit in the treatment of mild-to-moderately affected AD patients, but the level of benefit produced may be predicated on earlier treatment, ApoE genotype or whether the patient exhibits elevated cholesterol levels.

MONDAY, JUNE 20, 2005 SYMPOSIA S2-02 LIFESTYLE AND OTHER RISK FACTORS S2-02-01

CHRONIC DISTRESS IN OLD AGE, AGERELATED NEUROPATHOLOGY, AND DENDRITIC ATROPHY IN THE CA3 SUBFIELD OF THE HIPPOCAMPUS

Robert S. Wilson; Rush University Medical Center, Chicago, IL, USA Background: Chronic psychological distress is associated with dementia in old age, but the basis of the association is uncertain. Objective(s): To investigate the neurobiological basis of chronic psychological distress in older persons. Methods: Older Catholic clergy members (n⫽178) from the Rush Religious Orders Study underwent annual clinical evaluations, including detailed cognitive function testing, and brain autopsy at death. A composite measure of chronic psychological distress was constructed from standard measures of two traits, neuroticism and anxiety proneness, completed at baseline, and depressive symptoms, assessed annually. On postmortem examination, amyloid-beta, tau-positive neurofibrillary tangles, and alpha-synuclein-positive Lewy bodies were quantified in six brain regions, and the number, volume, and location of cerebral infarctions were noted. In a subset of participants with low (n⫽16 below the 25th percentile) or high (n⫽16 above the 75th percentile) chronic distress, an index of dendritic area in the CA3 subfield of the hippocampus was calculated as the percent area with microtubule-associated protein 2 labeled processes divided by density of cresyl violet stained neurons per mm2. Results: Chronic distress was inversely related to level of global cognitive function proximate to death but was unrelated to amyloid load, density of tau tangles, Lewy bodies, or cerebral infarction. In the subset with either low or high chronic distress, dendritic area in the CA3 region was inversely related to the composite measure of chronic distress (r⫽-0.48, p⫽0.006) and to each of its components (r⫽⫺0.35, p⫽0.048 for neuroticism; r⫽⫺0.047, p⫽0.007 for trait anxiety; r⫽⫺0.56, p⬍ 0.001 for depressive symptomalogy). In separate logistic regression models, the association between chronic distress and dendritic area remained after controlling for age, sex, or education. Conclusions: Chronic distress does not appear to be related to the most common causes of cognitive impairment in old age but was associated with dendritic atrophy in the CA3 subfield of the hippocampus, a defining neurobiological consequence of chronic stress in animal models. Further research on these models and their application to loss of cognition in humans is needed.