Current Alzheimer Research - Volume 10, Issue 3, 2013

The drawbacks of amyloid immunotherapy, including the development of encephalitis, the lack of clinical improvement and of any effect on neurofibrillary tangles (NFTs), coupled with the central role of NFTs in dementia, may point that clearance of amyloid pathology is not sufficient for improving the dementia symptoms in Alzheimer's disease (AD) patients. This further supported the concept that immunotherapy targeting the NFT proteinous aggregates may be preferential. Yet, the encephalitogenicity of full-length tau protein under a proinflammatory CNS milieu, reported by us in immunized mice, demands to carefully and selectively target pathological tau, while not the normal functional tau, and assuring both efficacy (anti-NFT effect) as well as safety (free of encephalitis) of a potential vaccine. Accumulating evidence from animal studies shows that tau-immunotherapy, targeting selectively pathological tau, particularly the phosphorylated- tau isoforms, reduces the tau-pathology and improves the symptoms of dementia. These findings are based on studies from different research groups, including our laboratory, conducted in different animal models and using various immunization protocols. There is also evidence that the decrease in NFTs is antibody-mediated involving the endosomal/ lysosomal pathway. No adverse effects were reported by the research groups, including also our study in which mice were immunized with a single injection of phosphorylated-tau peptide under a CNS proinflammatory milieu. In this review, I discuss the studies reported in this field, focusing on different approaches, different immunization protocols and mechanistic aspects, with a focus on the promising efficacy of the tau-immunotherapy, while addressing the safety issues already in the preclinical stage, before progressing to clinical trials.

During the last years the development of approaches to multitarget drug design and discovery is gaining acceptance. The cannabinoids are potentially excellent multi-target drug candidates because of their interesting pharmacological profiles, among which stands out the dual capacity of cannabinoid ligands to act as cannabinoid agonist and cholinesterase inhibitors. In this article, inhibition, kinetics studies and docking simulations with a representative set of cannabinoids are presented. The results of these studies showed the inhibitory capacity of some agonist cannabinoids with selectivity at AChE or BuChE enzymes. The kinetic and modelling studies allowed us to postulate the potential mode of action and the binding site of the cannabinoids. In general, the studied cannabinoids showed a mixed type inhibition mode of action. The exception to this behaviour was found for the agonist CP-55,940 that showed a non-competitive inhibition, suggesting that this cannabinoid only binds to the peripheral site.

Background: The histamine H3 receptor plays a critical role in the negative neuromodulation of neurotransmitters involved in cognitive function. H3 receptor antagonists/inverse agonists have been shown to exert pro-cognitive effects in pre-clinical models. GSK239512 is a potent and selective H3 receptor antagonist developed for the treatment of cognitive dysfunction in neurodegenerative disorders. In this study we examined the safety, tolerability, pharmacokinetics and pro-cognitive effects of GSK239512 (oral) in patients with mild to moderate Alzheimer's disease using ascending dose titration regimens. Methods: The study was conducted in two parts. Part A was a single-blind, placebo run-in, flexible dose titration over 9 days in two cohorts, each consisting of two patients. Part B was a double-blind, randomised, placebo controlled, parallel group, which investigated 3 flexible dose titration regimens over 4 weeks in 3 cohorts, each consisting of eight patients. Results: Overall, the 5/10/20/40μg and 10/20/40/80μg regimens were well-tolerated. The regimen of 20/40/80/150μg showed the poorest tolerability likely due to the higher starting dose. There were no clinically significant abnormalities in haematology, clinical chemistry, urinalysis parameters and cardiovascular parameters. GSK239512 had positive effects on tasks of attention and memory with effect sizes between 0.56 and 1.37. Conclusions: GSK239512 displayed asatisfactory level of tolerability in patients with Alzheimer's disease with evidence for positive effects on attention and memory. The findings suggest that a titration regimen with a starting dose of 5-10μg and a maximum dose of 80μg is likely to be a well-tolerated and potentially efficacious regimen for future clinical trials in patients with Alzheimer's disease. These findings await replication in a larger study.

We have examined the anti-angiogenic compound, angiostatin as a modulator of inflammatory reactivity and vascular responses and for neuroprotection in an animal model of Alzheimer's disease (AD). Intra-hippocampal amyloidbeta (Aβ1-42) injection, relative to controls phosphate buffer saline (PBS) or reverse peptide Aβ42-1, increased gliosis in the molecular layer (ML) of rat hippocampus. Vascular remodeling was indicated from increased microvessel immunoreactivity (ir) in ML suggesting the possibility of an angiogenic response to peptide injection. Administration of Aβ1-42 also induced a loss of neurons in the granule cell region of hippocampus relative to controls. Treatment of peptide-injected rats with angiostatin was associated with a spectrum of modulatory effects including reduced microgliosis (by 34%), diminished microvessel ir (by 36%) and increased neuronal viability (by 31%) compared with peptide injection alone. Angiostatin treatment was ineffective in reducing astrogliosis induced by Aβ1-42 and applied alone the compound had no significant effect to alter gliosis, microvessel ir or neuronal viability compared with PBS control. In vitro, angiostatin significantly attenuated secretion of the pro-angiogenic agent, vascular endothelial growth factor (VEGF) in lipopolysaccharide (LPS)-stimulated THP-1 cells. Our findings provide novel evidence for a broad spectrum of angiostatin effects in an animal model of AD including actions to reduce inflammatory reactivity, stabilize vascular remodeling and confer neuroprotection. The overall effects of angiostatin are consistent with actions of the compound to inhibit microglial secretion of VEGF.

Alzheimer's disease (AD) is characterized by senile plaques (SP) of extracellular amyloid β peptides(Aβ), neurofibrillary tangles (NFT) of intracellular hyper-phosphorylated tau and widespread loss of neurons. Apoptosis is the main reason of neuronal loss. It is proved that Aβ triggers apoptotic cell death via the activation of caspase-dependent and - independent cell death effectors, respectively. Valproic acid (VPA) is a widely used mood stabilizer and antiepileptic drug. Our previous study showed that VPA treatment significantly reduced SP formation and improved memory deficits in transgenic AD model mice. The present study intended to explore the protective effect of VPA on neuronal loss in transgenic AD model mice and the possible mechanisms involved. Histological and ultra-structural analysis showed that VPA partially decreased the swollen mitochondria and neurophil and promoted neurite outgrowth in AD mice model. Meanwhile, VPA greatly rescued the neuronal loss in the brain of AD mice. TUNEL staining showed that VPA significantly reduced the number of apoptotic cells. Western blot analysis revealed that VPA notably down-regulated the expression of Caspase-3, Caspase-9 and Caspase-12, reduced the level of cytochrome C and Bax. The expression of the antiapoptotic protein Bcl-2 was increased after VPA treatment. Flow cytometry revealed that VPA significantly decreased intracellular level of Ca2+ and elevated mitochondrial membrane potential. Altogether, these results indicate that VPA protected AD mice via suppression of upstream factors of apoptosis, namely inhibition of both mitochondrial and endoplasmic reticulum pathway of apoptosis.

Our previous study has shown the preventive effects of quetiapine, an atypical antipsychotic drug, on memory impairment and brain pathological changes in a mouse model of Alzheimer's disease (AD). The aim of the present study was to evaluate the therapeutic effects of quetiapine on memory deficit and neuropathology in an amyloid precursor protein (APP)/presenilin-1 (PS1) double transgenic mouse model of AD. The APP/PS1 mice started to have detectable brain β-amyloid (Aβ) at 3 months of age. Non-transgenic and transgenic mice were treated with quetiapine (0, 2.5, or 5 mg/(kg day)) in drinking water from the age of 4 months. After 8 months of continuous quetiapine administration, memory deficit was reversed and brain Aβ plaque pathology was attenuated in the AD mice. Quetiapine also decreased the soluble Aβ peptide levels in brain and cerebrospinal fluid (CSF), and attenuated the decreased synaptic protein levels in the AD mice. Furthermore, quetiapine normalized the abnormal activity of glycogen synthase kinase-3β (GSK-3β), an AD-involved kinase, in the AD mice. These results suggest that quetiapine can treat and alleviate the neuropathology in an APP/PS1 transgenic mouse model of AD, and indicate that quetiapine may have therapeutic effects in the treatment of AD.

Spirolides are marine toxins that are not currently in the routine monitoring assays. Nicotinic receptors seem to be the target of these compounds making them a promising pharmacological tool for related diseases as dementias as previously shown in vitro. In the present work, the bioavailability of 13-desMethyl spirolide C (13-desMeC) in the brain and in vivo effects were tested. Bioavailability was studied by ultra-performance liquid chromatography-mass spectrometry and its effect over Alzheimer hallmarks was studied by Proton magnetic resonance spectroscopy (H-MRS) and western blot. Only 2 minutes after its intraperitoneal injection it is found in brain and remains detectable even 24 hours post administration. Based on previous works that showed beneficial effects in an in vitro model of Alzheimer's disease (AD), we studied the effect in the same mice, 3xTg-AD, in vivo. We found that 13-desMeC (11.9 ug/kg, i.p.) induced positive effects on AD markers with an increase in N-acetyl aspartate (NAA) levels. These results were supported by an increase in synaptophysin levels and also a decrease in the intracellular amyloid beta levels in the hippocampus of treated 3xTg- AD versus non treated mice remarking the positive effects of this molecule in a well known model of AD. These data indicate for the first time that 13-desMeC cross the blood brain barrier and shows in vivo beneficial effects against AD after administration of low intraperitoneal doses of this marine toxin. This toxin may inspire a novel medical treatment of age-related diseases.

Diets containing a high proportion of fat with respect to protein plus carbohydrates are capable of inducing ketone body production in the liver, which provides an energetic alternative to glucose. Some ketogenic diets have been tested as therapeutic strategies for treating metabolic disorders related to a deficiency in glucose-driven ATP generation. However, ketone bodies are not capable of providing extra tricarboxylic acid cycle intermediates, limiting the anabolic capacity of the cell. Therefore, it is reasonable to hypothesize that supplementing a ketogenic diet with anaplerotic compounds such as triheptanoin may improve ketogenic diet effectiveness. The present study tests this hypothesis in APP/PS1 (APPswe/PS1dE9) transgenic mice, used as a model of familial Alzheimer's disease because impaired energy supply to neurons has been linked to this neurodegenerative process. Triheptanoin supplementation to a ketogenic diet for three months and starting at the age of three months reduces the memory impairment of APP/PS1 mice at the age of 6 months. The Aβ production and deposition were not significantly altered by the ketogenic diet, supplemented or not by triheptanoin. However, mice fed with triheptanoin-rich ketogenic diet have shown decreased astroglial response in the vicinity of Aβ plaques and decreased expression of the pro-inflammatory cytokine interferon-γ in astrocytes. These findings correlate with transcriptional up-regulation of the ROS detoxifying mechanisms Sirt1 and Pparg, thus linking triheptanoin with improved mitochondrial status. Present findings support the concept that ketogenic diets supplemented with anaplerotic compounds can be considered potential therapeutic strategies at early stages of Alzheimer's disease.

Cerebral hypoglycemia/hypometabolism is associated with Alzheimer's disease (AD) and is routinely used to assist clinical diagnosis of AD by brain imaging. However, whether cerebral hypoglycemia/hypometabolism contributes to the development of AD or is a response of reduced neuronal activity remains unclear. To investigate the causal relationship, we cultured the differentiated N2a neuroblastoma cells in glucose/pyruvate-deficient media (GDM). Shortly after the N2a cells cultured in the GDM, the mitochondria membrane potential was reduced and the AMP-activated-proteinkinase (AMPK), an energy sensor, was activated. Treatment of GDM not only increased the levels of tau phosphorylation at Ser262 and Ser396, but also increased the levels of active forms of GSK3α and GSK3β, two known kinases for tau phosphorylation, of the N2a cells. The levels of activated Akt, a mediator downstream to AMPK and upstream to GSK3α/β, were reduced by the GDM treatment. The effect of hypoglycemia was further examined in vivo by intracerebroventricular (icv) injection of streptozotocin (STZ) to the Wistar rats. STZ selectively injuries glucose transporter type 2-bearing cells which are primarily astrocytes in the rat brain, hence, interrupts glucose transportation from blood vessel to neuron. STZicv injection induced energy crisis in the brain regions surrounding the ventricles, as indicated by higher pAMPK levels in the hippocampus, but not cortex far away from the ventricles. STZ-icv treatment increased the levels of phosphorylated tau and activated GSK3β, but decreased the levels of activated Akt in the hippocampus. The hippocampus-dependent spatial learning and memory was impaired by the STZ-icv treatment. In conclusion, our works suggest that hypoglycemia enhances the AMPK-Akt-GSK3 pathway and leads to tau hyperphosphorylation.

The APOE genotype is a known susceptibility factor for Alzheimer's disease (AD). It is apparent that the presence of the APOE ε40 allele increases the risk for developing AD, lowers the age of onset in AD, and may influence the pathological burden seen in AD. In this study, we asked whether BACE1 levels differ by APOE genotype in the AD and non-demented (ND) brain. We isolated mid-frontal cortex (MFC) and mid-temporal cortex (MTC) from post-mortem ND and AD subjects that were APOE ε3/3, ε3/4, ε4/4 carriers. All AD subjects met NINDS-ADRDA and NIA-Reagan criteria for a diagnosis of AD. The MFC and MTC were homogenized and the lysates underwent ELISA and Western blotting for BACE1. The ELISA revealed that total BACE1 levels were lower in the MFC of AD compared to ND subjects. Furthermore, in APOE ε4 carriers BACE1 levels were lower than ε3/3 carriers in the ND frontal cortex. No difference in BACE1 levels was observed in AD MFC and in ND and AD MTC tissues. The ELISA results were confirmed by Western blotting. Our data suggest that brain BACEl levels may be influenced by the apolipoprotein E genotype before the onset of AD, providing an alternative explanation for the lower amyloid beta 42 levels in CSF in ND and AD subjects.

The soluble Abeta oligomers in brain are highly correlated with memory related synaptic dysfunctions in Alzheimer's disease (AD). However, more recent studies implicate the involvement of Abeta dimers and trimers in memory related AD pathology. Apparently, Abeta oligomers can bind with cellular prion protein at the membrane receptors, forming annular amyloid pores and membrane ion channels to induce aberrant spine cytoskeletal changes. Hence synapse targeting of Abeta oligomers involves activation of many receptors such as N-Methyl-D-aspartate (NMDA), alpha-amino-3- hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), nicotinic acetylcholine (nAChRs), p75 neurotrophin (p75NTR) following aberrant clustering of metabotropic glutamate receptors (mGluR5) leading to neuronal loss and LTP failure. In particular, NMDA and AMPA receptor activation by soluble amyloid oligomers involves calcium mediated mitochondrial dysfunction, decreased Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) levels at the synapses accompanying dramatic loss of synaptic proteins such as postsynaptic density-95 (PSD-95), dynamin-1 and synaptophysin. This kind of receptor-Abeta oligomer interaction might eventually affect the neuronal membrane integrity by altering dielectric barrier, various synaptic proteins, spine morphology and density and P/Q calcium currents that might provoke a cascade of events leading to neuronal loss and memory failure. In this review, we try to explain in detail the various possible mechanisms that connect Abeta oligomers with synapse damage and memory failure.

Synaptic loss is the major neuropathological correlate of memory decline as a result of Alzheimer´s Disease (AD). Synaptic failure appears to depend on the toxic actions of small and soluble amyloid-β (Aβ) oligomers. However, few studies have addressed the mechanism by which aging makes synapses more vulnerable to Aβ toxicity. In the present study we analyzed mitochondrial function and morphology and markers of oxidative stress in isolated presynaptic nerve endings from the hippocampus that were exposed to Aβ peptide at different ages. We found an age-related decline in mitochondrial activity, reduced antioxidant contents and increased oxidative stress markers in resting and depolarized synaptic terminals. Ultrastructural changes including an increase in mitochondrial size and a significant reduction of synaptic vesicles contents were also observed. In addition, synaptosomes obtained from 24 month old rats were more sensitive to Aβ toxicity. These data provide evidence of morphological and biochemical synaptic changes associated with aging that may contribute to exacerbate the damaging effects of Aβ.

Alzheimer's disease (AD) and frontotemporal dementia (FTD) are the most common types of dementia in the presenile population. Episodic memory impairment, the clinical hallmark of AD, can also be encountered in patients with FTD, complicating accurate diagnosis. Several studies in FTD have correlated memory deficits with neuroimaging findings, but lacked to compare neuroimaging results in FTD patients with and without memory impairment, while this latter analysis may give us insight into the underlying mechanisms of memory impairment in FTD. The aim of the present study was to compare 99mTc-HMPAO SPECT hypoperfusion patterns between FTD with episodic memory impairment (n = 13), FTD patients without episodic memory impairment (n = 10) as well as early onset (< 70 yrs) AD patients (n = 13), and controls (n = 15). We performed our analyses by means of Statistical Parametric Mapping software (SPM5), and showed that FTD patients with episodic memory impairment had lower perfusion in the right temporal lobe compared with FTD patients without memory impairment. Lower perfusion in this region correlated with worse memory performance on the Clinical Dementia Rating scale in FTD patients. With equal performances on memory tests, patients with early onset AD showed posterior temporal and parietal lobe hypoperfusion in comparison with patients with FTD and memory impairment, while vice versa hypoperfusion in the anterior frontotemporal regions was found in FTD patients with memory impairment in comparison with AD.

Midlife habits may be important for the later development of Alzheimer's disease (AD). We estimated the contribution of midlife prayer to the development of cognitive decline. In a door-to-door survey, residents aged ≥65 years were systematically evaluated in Arabic including medical history, neurological, cognitive examination, and a midlife leisure-activities questionnaire. Praying was assessed by the number of monthly praying hours at midlife. Stepwise logistic regression models were used to evaluate the effect of prayer on the odds of mild cognitive impairment (MCI) and AD versus cognitively normal individuals. Of 935 individuals that were approached, 778 [normal controls (n=448), AD (n=92) and MCI (n=238)] were evaluated. A higher proportion of cognitively normal individuals engaged in prayer at midlife [(87%) versus MCI (71%) or AD (69%) (p<0.0001)]. Since 94% of males engaged in prayer, the effect on cognitive decline could not be assessed in men. Among women, stepwise logistic regression adjusted for age and education, showed that prayer was significantly associated with reduced risk of MCI (p=0.027, OR=0.55, 95% CI 0.33-0.94), but not AD. Among individuals endorsing prayer activity, the amount of prayer was not associated with MCI or AD in either gender. Praying at midlife is associated with lower risk of mild cognitive impairment in women.