Alzheimer’s disease (AD) is characterized by cognitive decline and 5- to 10-fold increased seizure incidence. How seizures contribute to cognitive decline in AD or other disorders is unclear. We show that spontaneous seizures increase expression of ΔFosB, a highly stable Fos-family transcription factor, in the hippocampus of an AD mouse model. ΔFosB suppressed expression of the immediate early gene c-Fos, which is critical for plasticity and cognition, by binding its promoter and triggering histone deacetylation. Acute histone deacetylase (HDAC) inhibition or inhibition of ΔFosB activity restored c-Fos induction and improved cognition in AD mice. Administration of seizure-inducing agents to nontransgenic mice also resulted in ΔFosB-mediated suppression of c-Fos, suggesting that this mechanism is not confined to AD mice. These results explain observations that c-Fos expression increases after acute neuronal activity but decreases with chronic activity. Moreover, these results indicate a general mechanism by which seizures contribute to persistent cognitive deficits, even during seizure-free periods.

Alzheimer's disease is associated with cognitive decline and seizures. Growing evidence indicates that seizures contribute to cognitive deficits early in disease, but how they develop and impact cognition are unclear. To investigate potential mechanisms, we studied a mouse model that overexpresses mutant human amyloid precursor protein with high levels of amyloid beta (Aβ). These mice develop generalized epileptiform activity, including nonconvulsive seizures, consistent with alterations in corticothalamic network activity. Amyloid precursor protein mice exhibited reduced activity marker expression in the reticular thalamic nucleus, a key inhibitory regulatory nucleus, and increased activity marker expression in downstream thalamic relay targets that project to cortex and limbic structures. Slice recordings revealed impaired cortical inputs to the reticular thalamic nucleus that may contribute to corticothalamic dysfunction. These results are consistent with our findings of impaired sleep maintenance in amyloid precursor protein mice. Finally, the severity of sleep impairments predicted the severity of deficits in Morris water maze, suggesting corticothalamic dysfunction may relate to hippocampal dysfunction, and may be a pathophysiological mechanism underlying multiple behavioral and cognitive alterations in Alzheimer's disease.

On the basis of the previously reported benzimidazole 1,3'-bipyrrolidine benzamides (1), a new class of 2-(pyrrolidin-1-yl)ethyl-3,4-dihydroisoquinolin-1(2H)-one derivatives (3-50) were synthesized and evaluated as potent H(3) receptor antagonists. In particular, compound 39 exhibited potent in vitro binding and functional activities at the H(3) receptor, good selectivities against other neurotransmitter receptors and ion channels, acceptable pharmacokinetic properties, and a favorable in vivo profile.

Novel 5-cyclic amine-3-arylsulfonylindazoles were prepared, and several analogues within this class have been identified as high-affinity 5-HT6 receptor ligands with improved pharmacokinetic and pharmacological properties. One selected example, 18b, showed good brain penetrability and a generally favorable pharmacokinetic profile with procognitive efficacy in the rat novel object recognition assay. The synthesis and structure activity relationship of this potent class are discussed.

Alpha-7 nicotinic acetylcholine receptor (alpha 7 nAChR) agonists are promising therapeutic candidates for the treatment of cognitive impairment. We report a series of novel, potent small molecule agonists (4-18) of the alpha 7 nACh R deriving from our continuing efforts in the areas of Alzheimer's disease and schizophrenia. One of the compounds of the series containing a urea moiety (16) was further shown to be a selective agonist of the alpha 7 nAChR with excellent in vitro and in vivo profiles, brain penetration, and oral bioavailability and demonstrated in vivo efficacy in multiple behavioral cognition models. Structural modifications leading to the improved selectivity profile and the biological evaluation of this series of compounds are discussed.

The 5-HT6 receptor (5-HT6R) is a G-protein coupled receptor positively coupled to adenylate cyclase primarily localized within the central nervous system, particularly in brain regions associated with cognition. Antagonism of the 5-HT6R has been reported to modulate the release of multiple neurotransmitters including elevating extracellular levels of glutamate and acetylcholine in regions associated with cognitive function such as the cerebral cortex and hippocampal formation. Accumulating preclinical and clinical data suggest that 5-HT6R antagonists may function to improve cognitive performance and therefore serve as effective treatment options for Alzheimer's disease. SAM-531 (WAY-262531) is a new chemical entity that is a selective full antagonist at the 5-HT6R. Here we describe the pre-clinical profile of SAM-531 in support of its clinical development as a treatment for Alzheimer's disease. SAM-531 was characterized in vitro in multiple assays evaluating binding affinity, functional activity and receptor selectivity. In vivo characterization of SAM-531 consisted of an evaluation of the effect of treatment on brain neurochemistry using in vivo microdialysis as well as tests of rodent cognitive function such as contextual fear conditioning and novel object recognition. Cognitive testing was performed in assays utilizing both pharmacological and non-pharmacological deficit states. SAM-531 is a potent and selective full antagonist at the human 5-HT6R (Ki = 1.0 nM; IC50 = 10 nM). In rats, oral administration of SAM-531 caused significant increases in extracellular levels of glutamate and acetylcholine in the dorsal hippocampus. Treatment with SAM-531 blocked a scopolamine-induced deficit in a hippocampal-dependent learning and memory task, contextual fear conditioning. In addition, SAM-531 treatment blocked both glutmatergic- (MK-801) and cholinergic-mediated (scopolamine) deficits in recognition memory. Furthermore, treatment with SAM-531 enhanced retention of recognition memory when measured 48 hours after training, a time at which memory no longer can be measured in vehicle-treated animals. SAM-531 is a potent, selective and orally bioavailable 5-HT6R antagonist which modulates neurotransmitters associated with learning and memory and is active in multiple assays of cognitive dysfunction in the rodent. These studies provide pre-clinical support for the further development of the 5-HT6R antagonist SAM-531 as a potential treatment for Alzheimer's disease.

Using a focused screen of biogenic amine compounds we identified a novel series of H(3)R antagonists. A preliminary SAR study led to reduction of MW while increasing binding affinity and potency. Optimization of the physical properties of the series led to (S)-6n, with improved brain to plasma exposure and efficacy in both water intake and novel object recognition models.

The presenilin containing gamma-secretase complex is responsible for the regulated intramembraneous proteolysis of the amyloid precursor protein (APP), the Notch receptor, and a multitude of other substrates. gamma-Secretase catalyzes the final step in the generation of Abeta(40) and Abeta(42) peptides from APP. Amyloid beta-peptides (Abeta peptides) aggregate to form neurotoxic oligomers, senile plaques, and congophilic angiopathy, some of the cardinal pathologies associated with Alzheimer's disease. Although inhibition of this protease acting on APP may result in potentially therapeutic reductions of neurotoxic Abeta peptides, nonselective inhibition of the enzyme may cause severe adverse events as a result of impaired Notch receptor processing. Here, we report the preclinical pharmacological profile of GSI-953 (begacestat), a novel thiophene sulfonamide gamma-secretase inhibitor (GSI) that selectively inhibits cleavage of APP over Notch. This GSI inhibits Abeta production with low nanomolar potency in cellular and cell-free assays of gamma-secretase function, and displaces a tritiated analog of GSI-953 from enriched gamma-secretase enzyme complexes with similar potency. Cellular assays of Notch cleavage reveal that this compound is approximately 16-fold selective for the inhibition of APP cleavage. In the human APP-overexpressing Tg2576 transgenic mouse, treatment with this orally active compound results in a robust reduction in brain, plasma, and cerebral spinal fluid Abeta levels, and a reversal of contextual fear-conditioning deficits that are correlated with Abeta load. In healthy human volunteers, oral administration of a single dose of GSI-953 produces dose-dependent changes in plasma Abeta levels, confirming pharmacodynamic activity of GSI-953 in humans.

gamma-Secretase inhibitors have been shown to reduce the production of beta-amyloid, a component of the plaques that are found in brains of patients with Alzheimer's disease. A novel series of heterocyclic sulfonamide gamma-secretase inhibitors that reduce beta-amyloid levels in cells is reported. Several examples of compounds within this series demonstrate a higher propensity to inhibit the processing of amyloid precursor protein compared to Notch, an alternative gamma-secretase substrate.

Recent evidence has suggested a potential role for APP in the regulation of brain Apolipoprotein E (ApoE) and cholesterol metabolism. In support of this function we have previously presented data from a transcriptional profiling study which demonstrates dysregulation of cholesterol-related genes in the Tg2576 APP transgenic mouse model of AD. We have now expanded these findings to investigate the effects of APP transgene expression on ApoE metabolism. ApoE protein levels in the cortex and hippocampus of wildtype and APP transgenic mice have been examined using western blotting and an ApoE-specific immunoassay. In both brain regions ApoE protein levels were significantly increased in the transgenic animals compared to wildtype controls. This was not a result of increased ApoE transcription, as mRNA levels in the Tg2576 mice were unchanged. ApoE is one of a number of ligands for the low density lipoprotein-related receptor, LRP1. To investigate whether changes in ApoE protein levels arise as a result of decreased functionality of LRP1, we first investigated levels of other LRP1 ligands in APP transgenic mice. Immunohistochemical analysis demonstrated increased protein levels of two additional LRP1 ligands, tissue plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1). Together these findings suggest that LRP1 function may be altered in the APP transgenic mice leading to a reduction in the catabolism of LRP1 ligands. Previously, LRP1 expression has been shown to be regulated at the transcriptional level by the APP intracellular domain (AICD). To investigate whether this mechanism underlies the differences in ApoE, tPA and PAI-1 protein levels observed in the Tg2576 mice we examined LRP1 mRNA levels. Surprisingly, LRP1 mRNA was not altered in the Tg2576 mice compared to wildtype controls. An examination of LRP1 protein levels and/or LRP1 processing and their effects of LRP1 function is currently underway. Taken together these data provide evidence in support of the growing association between AD and cholesterol metabolism and further establish the biological link between APP and ApoE, the two major determinants of AD.

The amyloid hypothesis states that a variety of neurotoxic beta-amyloid (Abeta) species contribute to the pathogenesis of Alzheimer's disease. Accordingly, a key determinant of disease onset and progression is the appropriate balance between Abeta production and clearance. Enzymes responsible for the degradation of Abeta are not well understood, and, thus far, it has not been possible to enhance Abeta catabolism by pharmacological manipulation. We provide evidence that Abeta catabolism is increased after inhibition of plasminogen activator inhibitor-1 (PAI-1) and may constitute a viable therapeutic approach for lowering brain Abeta levels. PAI-1 inhibits the activity of tissue plasminogen activator (tPA), an enzyme that cleaves plasminogen to generate plasmin, a protease that degrades Abeta oligomers and monomers. Because tPA, plasminogen and PAI-1 are expressed in the brain, we tested the hypothesis that inhibitors of PAI-1 will enhance the proteolytic clearance of brain Abeta. Our data demonstrate that PAI-1 inhibitors augment the activity of tPA and plasmin in hippocampus, significantly lower plasma and brain Abeta levels, restore long-term potentiation deficits in hippocampal slices from transgenic Abeta-producing mice, and reverse cognitive deficits in these mice.

SAR on HTS hits 1 and 2 led to the potent, Notch-1-sparing GSI 9, which lowered brain Abeta in Tg2576 mice at 100 mg/kg po. Converting the metabolically labile methyl groups in 9 to trifluoromethyl groups afforded the more stable analogue 10, which had improved in vivo potency. Further side chain modification afforded the potent Notch-1-sparing GSI begacestat (5), which was selected for development for the treatment of Alzheimer's disease.

5-Hydroxytryptamine (5-HT)1A receptors play an important role in multiple cognitive processes, and compelling evidence suggests that 5-HT1A antagonists can reverse cognitive impairment. We have examined the therapeutic potential of a potent (Ki = 1.1 nM), selective (>100-fold), orally bioavailable, silent 5-HT1A receptor antagonist (KB = 1.3 nM) (R)-N-(2-methyl-(4-indolyl-1-piperazinyl)-ethyl)-N-(2-pyridinyl)-cyclohexane carboxamide (WAY-101405). Oral administration of WAY-101405 was shown to be effective in multiple rodent models of learning and memory. In a novel object recognition paradigm, 1 mg/kg enhanced retention (memory) for previously learned information, and it was able to reverse the memory deficits induced by scopolamine. WAY-101405 (1 mg/kg) was also able to reverse scopolamine-induced deficits in a rat contextual fear conditioning model. In the Morris water maze, WAY-101405 (3 mg/kg) significantly improved learning in a paradigm of increasing task difficulty. In vivo microdialysis studies in the dorsal hippocampus of freely moving adult rats demonstrated that acute administration of WAY-101405 (10 mg/kg) increased extracellular acetylcholine levels. The selective radioligand [3H]WAY-100635, administered i.v., was used for in vivo receptor occupancy studies, where WAY-101405 occupied 5-HT1A receptors in the rat cortex, with an ED50 value of 0.1 mg/kg p.o. Taken together, these studies demonstrate that WAY-101405 is a potent and selective, brain penetrant, orally bioavailable 5-HT1A receptor “silent” antagonist that is effective in preclinical memory paradigms at doses where approximately 90% of the postsynaptic 5-HT1A receptors are occupied. These results further support the rationale for use of this compound class in the treatment of cognitive dysfunction associated with psychiatric and neurological conditions.