Current Alzheimer Research - Volume 7, Issue 3, 2010

Alzheimer's disease (AD) drug discovery researchers face immense challenges in developing drugs that get into the brain, are safe and ultimately produce clinically meaningful results. Perhaps even more challenging is determining at what point in the disease cascade is best to intervene. We know that many different pathways initiate or exacerbate disease progression [1]. Amyloid-β (Aβ), the main component of amyloid plaques, has long been considered the leading target for AD intervention programs even though the amyloid cascade hypothesis remains controversial to this day [2, 3]. Innovative approaches to target Aβ are being developed and may be the best way to treat the disease at its earliest stages. Yet, as we learn more about the disease, it is becoming increasingly clear that alternative approaches may also be necessary to diversify the AD drug discovery portfolio. Increasing “shots on goal” will bring new ideas and novel targets into the pipeline, accelerating treatment development for AD. In the spirit of these objectives, the Alzheimer's Drug Discovery Foundation (ADDF) hosted the 10th International Conference on Alzheimer's Drug Discovery on September 14-15, 2009 in Jersey City, NJ. At the conference, attendees were treated not only to the breathtaking skyline view of New York City, but also to two days of diverse research aimed at AD drug discovery. The conference drew attendants from academia, industry and government. Sessions focused on Neuroprotection Strategies (chaired by Frank Longo, Stanford University), Anti-Amyloid and Protein Misfolding (chaired by Michael Wolf, Harvard Medical School), Anti- Tangles and Frontotemporal Dementia (chaired by Jeff Kuret, Ohio State University), and Alternative Strategies: New Targets for AD Therapy (chaired by Diana Shineman, ADDF). A comprehensive summary of the conference can be found on the conference web site (http:// www.worldeventsforum.com/addf/10th/html/summary.html) . The proceedings in this special issue highlight a few of exciting programs presented at this conference. This year's conference built upon previous efforts and focused more heavily on alternative targets for therapeutic development [4-6]. The ultimate goal for AD drug discovery, regardless of the molecular target, is to protect neurons and their processes so that memory and function can be maintained. There are many ways to target neuroprotection and strengthen neuronal defenses against the toxic insults of AD. For example, Karin Yurko-Mauro from Martek Biosciences presents an overview of clinical data on the health benefits of Docosahexaenoic acid (DHA), the main component of omega-3 fatty acids (Yurko-Mauro, page 190-196). Her paper notes that DHA supplementation resulted in a cardiovascular benefit (decreased heart rate, blood pressure and triglycerides) as well as some cognitive benefit in older individuals with memory complaints, specifically in the paired associated learning test. Still, further studies are needed to determine if DHA substantially impacts AD progression. It is easy to lose sight of the big picture when the field is often focused narrowly on specific molecular targets. In this issue, Michela Gallagher (Johns Hopkins University) reminds us that AD is a disease of neuronal systems, underlined by synaptic failure that begins very early in the disease process (Gallagher et al., page 197-199). This paper shows evidence that hyperactivity in the CA3 region of the hippocampus, the brain region critical for proper memory function, precedes memory deficits in animal models as well as in humans. Therapeutic development efforts are focused on inhibiting this hyperactivity to hopefully prevent downstream synaptic degeneration, memory loss and pathology seen in later stages of AD. Echoing the sentiments of Dr. Gallagher, James Malter (University of Wisconsin) also looks at AD as a systemsbased disease and presents work on the Fragile X Mental Retardation Protein (FMRP), which has recently been shown to regulate the dendritic translation of APP downstream of mGluR5 activation (Malter et al., page 200-206). mGluR5 antagonists can decrease Amyloid Precursor Protein (APP) translation and reduce Aβ40 in brain lysate. Work is ongoing to expand on these findings. While Dr. Malter's program targets upstream of Aβ generation at the level of APP, Michael Wolfe (Harvard Medical School), Philip Williams (University of Hawaii) and Michael Sierks (Arizona State University) are all focused on intervening directly at the level Aβ generation using very different approaches. In these proceedings, Dr. Wolfe presents a summary of the discovery and development of inhibitors of γ-secretase that block Aβ generation while sparing Notch to prevent the side effects seen with pan γ- secretase inhibition (Augelli-Szafran et al., page 207-209)...

Memory loss is a prominent health concern, second only to heart disease for older individuals. Docosahexaenoic acid (DHA), the principle omega-3 fatty acid in brain and heart, plays an important role in neural and cardiac function. Decreases in plasma DHA are associated with cognitive decline in healthy elderly and Alzheimer's patients. Higher DHA intake and plasma levels are inversely correlated with increased relative risk of Alzheimer's disease (AD) and fatal coronary heart disease. DHA provides well known cardiovascular benefits (e.g. lower triglycerides, increased HDL cholesterol, decreased resting heart rate) in older adults. Preclinically, DHA supplementation restores brain DHA levels, enhances learning and memory tasks in aged animals, and significantly reduces beta amyloid, plaques, and tau in transgenic AD models. To date, clinical studies with DHA+EPA supplementation have shown some positive effects in mild cognitive impairment but not in AD, suggesting that early intervention may be a key factor to providing effective therapies. A recent clinical study examined individual effects of 900mg/d algal DHA as a nutritional supplement for age-related cognitive decline (ARCD). This randomized, double-blind, placebo-controlled study (n=485) found significantly fewer CANTAB® Paired Associate Learning errors with algal DHA at six months versus placebo (diff. score -1.63±0.76, p=0.03). Positive effects on Verbal Recognition Memory (p<0.02) and significant decreases in resting heart rate with DHA (p<0.03) were observed, indicating improved learning and episodic memory functions and cardiovascular benefits for ARCD. Collectively, data reveal a potentially beneficial role for DHA in preventing or ameliorating cognitive decline and cardiovascular disease in the aged.

Risk for Alzheimer's disease escalates dramatically with increasing age in the later decades of life. It is widely recognized that a preclinical condition in which memory loss is greater than would be expected for a person's age, referred to as amnestic mild cognitive impairment, may offer the best opportunity for intervention to treat symptoms and modify disease progression. Here we discuss a basis for age-related memory impairment, first discovered in animal models and recently isolated in the medial temporal lobe system of man, that offers a novel entry point for restoring memory function with the possible benefit in slowing progression to Alzheimer's disease.

As the mechanisms underlying neuronal development and degeneration become clarified, a number of common effectors and signaling pathways are becoming apparent. Here we describe the identification of Aβ, long considered a pathologic mediator of Alzheimers Disease and Down Syndrome, as similarly over-expressed in the neurodevelopmental disease, Fragile X Syndrome. We also show that mGluR5 inhibitors, currently employed for the treatment of Fragile X, reduce Aβ production in rodent models of Fragile X and AD as well as reduce disease phenotypes including seizures. Thus seemingly disparate neurologic diseases may share a common pathologic instigator and be treatable with a common, currently available class of therapeutics.

Overwhelming evidence supports a central role for the amyloid β-peptide (Aβ) in the pathogenesis of Alzheimer's disease (AD), and the proteases that produce Aβ from its precursor protein APP are top targets for therapeutic intervention. Considerable effort has focused on targeting γ-secretase, which generates the C-terminus of Aβ; however, γ- secretase inhibitors cause serious toxicities due to interference with the Notch signaling pathway. We have been working toward compounds that directly alter γ-secretase activity to reduce Aβ production without affecting the proteolysis of Notch. Using purified enzyme and substrate, we have shown that γ-secretase can be selectively inhibited in this way by naphthyl-substituted γ-aminoketones and γ-aminoalcohols. These early hits, however, suffered from chemical instability and/or poor potency. Iterative design, synthesis and evaluation have led to the discovery of Notch-sparing γ-secretase inhibitors with substantially increased potencies in biochemical and cellular assays. These compounds are of low molecular weight and are under evaluation for drug-like properties. The discovery and development of these compounds will be discussed.

Despite the long history of drug discovery from natural sources, the marine environment, which covers 70% of the Earth's surface, is still relatively unexplored. Intense competition for limited resources drives the evolution of specific and potent chemical defenses distinct from their terrestrial counterparts. Based on this rationale, we recently began screening extracts derived from marine invertebrate and cyanobacterial samples for BACE-1 inhibitors in a chemiluminescent enzyme-fragment complementation (EFC) assay. The results of this broad screening are presented here, along with our progress towards the development of a secondary LC-MS homogeneous affinity assay. Incubation of the extracts active in the EFC assay with BACE1, subsequent isolation of the enzyme-inhibitor complex and then analysis of the small molecule inhibitor by LC-MS rapidly links a chemical structure to biological activity. This approach enables the rapid targetorientated discovery of BACE-1 inhibitors from marine sources.

Accumulation and deposition of beta amyloid (Aβ) play a critical role in the pathogenesis of Alzheimer's Disease (AD), and numerous approaches to control Aβ aggregation are being actively pursued. Brain Aβ levels are controlled by the action of several proteolytic enzymes such as neprilysin (NEP), insulin degrading enzyme (IDE) and plasmin. While up-regulation of these enzymes increased clearance of Aβ in transgenic mouse models of AD, these enzymes have other natural substrates and multiple cleavage sites in Aβ complicating their use for treating AD. Alternatively, immunotherapeutic approaches to clear Aβ are gaining interest. Active and passive immunization studies with Aβ can reduce plaque burden and memory loss, but clinical trials were stopped due to meningioencephalitis in some patients. Naturally occurring proteolytic antibodies have been shown to cleave Aβ, and their serum titers are increased in patients with AD reflecting a protective autoimmune response. These antibodies however cannot cross the blood brain barrier and depend entirely on peripheral clearance to clear Aβ. A potentially non-inflammatory approach to facilitate Aβ clearance and reduce toxicity is to promote hydrolysis of Aβ at its α-secretase site using affinity matured single chain antibody fragments (scFvs). Bispecific antibodies consisting of a proteolytic scFv and a targeting scFv can be engineered to selectively supplement and target extracellular α-secretase activity and to target toxic Aβ forms facilitating their degradation and clearance without generating an immune response. This strategy represents a suitable paradigm for treating other neurological diseases such as Parkinson's Disease, Lou Gehrig's Disease, and spongiform encephalopathies.

The accumulation of the amyloid-beta peptide (Aβ) continues to emerge as a central factor in Alzheimer's disease (AD). In recent years attention has been drawn to clearance mechanisms of Aβ as evidence suggests reduced clearance may be linked to late-onset AD. Direct degradation of Aβ by endopeptidases has emerged as one critical pathway of clearance. Of particular interest are endopeptidases that are sensitive to the neprilysin inhibitors thiorphan and phosphoramidon (i.e. “NEP-like”) as these inhibitors induce a dramatic increase in Aβ levels resulting in rapid plaque formation in wild-type rodents. This review focuses on neprilysin (NEP) and on another NEP-like endopeptidase termed neprilysin- 2 (NEP2). The involvement of these endopeptidases in AD and the state of their therapeutic development are discussed.

Alzheimer's disease is diagnosed by postmortem detection of pathological lesions that accumulate in specific brain regions. Although the presence of both β-amyloid plaques and tau-bearing neurofibrillary lesions defines Alzheimer's disease, the distribution of neurofibrillary lesions alone correlates strongly with neurodegeneration and cognitive decline. A whole-brain imaging test capable of detecting these lesions in premortem cases could have great potential for staging and differentially diagnosing Alzheimer's disease. Here we discuss the challenges in developing a whole-brain imaging approach for detection of this intracellular target.

Tau aggregation is an appealing target for therapeutic intervention. However, conformational change or aggregation needs to be targeted without inhibiting the normal biology of tau and its role in microtubule stabilization. The number of compound classes being tested at this time are very limited and include Congo red derivatives [1], anthraquinones (Pickhardt et al. 2005 [2], disputed in Crowe et al. 2007 [3]), 2,3-di(furan-2-yl)-quinoxalines , phenylthiazolyl-hydrazide (PTH) [4], polyphenols and porphyrins [5] and cyanine dyes [6-8]. Herein we have utilized a member of the cyanine dye family (C11) in an organotypic slice culture model of tangle formation. Our results demonstrate that C11 is capable of affecting tau polymerization in a biphasic, dose dependent manner. At submicromolar concentrations (0.001 μM) C11 reduced levels of aggregated tau. However, higher doses resulted in an increase in tau polymerization. These effects can also be seen at the level of individual filaments with changes in filament length and number mirroring the pattern seen via immunoblotting. In addition, this effect is achieved without altering levels of phosphorylation at disease and microtubule binding relevant epitopes.

Dephosphorylation (activation) of cofilin, an actin binding protein, is stimulated by initiators of neuronal dysfunction and degeneration including oxidative stress, excitotoxic glutamate, ischemia, and soluble forms of β-amyloid peptide (Aβ). Hyperactive cofilin forms rod-shaped cofilin-saturated actin filament bundles (rods). Other proteins are recruited to rods but are not necessary for rod formation. Neuronal cytoplasmic rods accumulate within neurites where they disrupt synaptic function and are a likely cause of synaptic loss without neuronal loss, as occurs early in dementias. Different rod-inducing stimuli target distinct neuronal populations within the hippocampus. Rods form rapidly, often in tandem arrays, in response to stress. They accumulate phosphorylated tau that immunostains for epitopes present in “striated neuropil threads,” characteristic of tau pathology in Alzheimer disease (AD) brain. Thus, rods might aid in further tau modifications or assembly into paired helical filaments, the major component of neurofibrillary tangles (NFTs). Rods can occlude neurites and block vesicle transport. Some rod-inducing treatments cause an increase in secreted Aβ. Thus rods may mediate the loss of synapses, production of excess Aβ, and formation of NFTs, all of the pathological hallmarks of AD. Cofilin-actin rods also form within the nucleus of heat-shocked neurons and are cleared from cells expressing wild type huntingtin protein but not in cells expressing mutant or silenced huntingtin, suggesting a role for nuclear rods in Huntington disease (HD). As an early event in the neurodegenerative cascade, rod formation is an ideal target for therapeutic intervention that might be useful in treatment of many different neurological diseases.

Positron emission tomography (PET) imaging of Alzheimer's Disease (AD) offers the potential to provide early onset diagnosis and subsequent intervention, including guided treatment regimens. One of the restricting factors in clinical application of PET technology is the limited availability of radioligands with affinity to specific targets of interest. Given the short half-life of the most popular positron emitter currently used (18F; ∼120 min.) extremely rapid and efficient radiochemistry methods are needed to ensure required compounds are prepared and purified for administration within the 2-3 half life practical limit. Recent efforts to combine microwave mediated synthesis with advanced catalysis in the synthesis of specific categories of AD imaging agents will be presented.

Cannabinoids have been shown to increase neurogenesis in adult brain, as well as protect neurons from excitotoxicity, calcium influx, inflammation, and ischemia. Recent studies have shown that synthetic cannabinoids can alleviate water maze impairments in rats treated with intracranial amyloid β protein (Aβ); however it is unknown whether this effect is due to the cannabinoids' anti-inflammatory properties or whether it affects Aβ processing. Here we investigate whether cannabinoids have any effect on Alzheimer's disease in vivo. We found that HU210, a potent synthetic cannabinoid, did not improve water maze performance or a contextual fear conditioning task in an APP23/PS45 double transgenic mouse model of AD. HU210 had no effect on APP processing and Aβ generation, as well as neuritic plaque formation in the brains of AD transgenic mice. Our study showed that synthetic cannabinoid HU210 had no beneficial effects on AD neuropathology and behavioral deficits of AD model mice, which advises caution of such drug's application in AD therapies.

High levels of cholesterol have been proposed as a risk factor for Alzheimer's disease (AD). Polymorphism of genes encoding proteins that regulate cholesterol metabolism have also been associated with the frequency of Alzheimer's development. Some studies have shown that cholesterol-lowering drugs reduce the frequency of AD development. The proposed role of cholesterol in AD has been challenged by several studies. In this review, we provide a brief account of the major pieces of evidence in support of and against the possible role of cholesterol in the development of AD, and the methodologies used. We highlight the interactions between cholesterol and amyloid beta (Aβ) and, with the peptide's precursor protein. Drawing from our teams' recent findings, we speculate on how Aβ peptides may influence the fluidity, stability of the membrane, as well as membrane morphological changes.

The MCI construct aims to investigate the grey area existing between normal aging and dementia, in order to identify in the preclinical stage patients at risk of developing dementia. The construct of the MCI has been proposed by taking the neuropathological staging of the Alzheimer's disease (AD) as reference and providing an explicit set of identifying criteria, but it has raised two main problems: (a) the variability of estimates concerning the actual rate of conversion from MCI to dementia, and (b) the number of subjects who return to normality. These problems stem in part from the operational difficulties met by the MCI identifying criteria concerned with: the memory tests used, the cut-off adopted to identify patients with an ‘objective’ memory disorder, the assessment of subjective memory disorders, and the integrity of daily living activities. After a short discussion of these operational difficulties, I will pass to shortly survey the laboratory data providing additional predictive value for the conversion of MCI to dementia (ApoE4, CSF biomarkers, Neuroimaging data, and Vascular risk factors) and some recent attempts to identify pre-MCI patients, with a purely subjective cognitive impairment. I will conclude my review by asking if we have a single MCI or a family of MCI constructs, each of whom could play a preferential role in specific clinical contexts.