Design and Synthesis of Clinical Candidate PF-06751979: A Potent, Brain Penetrant, β-Site Amyloid Precursor Protein Cleaving Enzyme 1 (BACE1) Inhibitor Lacking Hypopigmentation.

A major challenge in the development of β-site amyloid precursor protein cleaving enzyme 1 (BACE1) inhibitors for the treatment of Alzheimer's disease is the alignment of potency, drug-like properties, and selectivity over related aspartyl proteases such as Cathepsin D (CatD) and BACE2. The potential liabilities of inhibiting BACE2 chronically have only recently begun to emerge as BACE2 impacts the processing of the premelanosome protein (PMEL17) and disrupts melanosome morphology resulting in a depigmentation phenotype. Herein, we describe the identification of clinical candidate PF-06751979 (64), which displays excellent brain penetration, potent in vivo efficacy, and broad selectivity over related aspartyl proteases including BACE2.

Aminomethyl-Derived Beta Secretase (BACE1) Inhibitors: Engaging Gly230 without an Anilide Functionality.

A growing subset of β-secretase (BACE1) inhibitors for the treatment of Alzheimer's disease (AD) utilizes an anilide chemotype that engages a key residue (Gly230) in the BACE1 binding site. Although the anilide moiety affords excellent potency, it simultaneously introduces a third hydrogen bond donor that limits brain availability and provides a potential metabolic site leading to the formation of an aniline, a structural motif of prospective safety concern. We report herein an alternative aminomethyl linker that delivers similar potency and improved brain penetration relative to the amide moiety.

Utilizing structures of CYP2D6 and BACE1 complexes to reduce risk of drug-drug interactions with a novel series of centrally efficacious BACE1 inhibitors.

In recent years, the first generation of β-secretase (BACE1) inhibitors advanced into clinical development for the treatment of Alzheimer's disease (AD). However, the alignment of drug-like properties and selectivity remains a major challenge. Herein, we describe the discovery of a novel class of potent, low clearance, CNS penetrant BACE1 inhibitors represented by thioamidine 5.

Discovery of a series of efficient, centrally efficacious BACE1 inhibitors through structure-based drug design.

The identification of centrally efficacious β-secretase (BACE1) inhibitors for the treatment of Alzheimer's disease (AD) has historically been thwarted by an inability to maintain alignment of potency, brain availability, and desired absorption, distribution, metabolism, and excretion (ADME) properties. In this paper, we describe a series of truncated, fused thioamidines that are efficiently selective in garnering BACE1 activity without simultaneously inhibiting the closely related cathepsin D or negatively impacting brain penetration and ADME alignment, as exemplified by 36. Upon oral administration, these inhibitors exhibit robust brain availability and are efficacious in lowering central Amyloid β (Aβ) levels in mouse and dog.

Behavioral characterization of A53T mice reveals early and late stage deficits related to Parkinson's disease.

Parkinson's disease (PD) pathology is characterized by the formation of intra-neuronal inclusions called Lewy bodies, which are comprised of alpha-synuclein (α-syn). Duplication, triplication or genetic mutations in α-syn (A53T, A30P and E46K) are linked to autosomal dominant PD; thus implicating its role in the pathogenesis of PD. In both PD patients and mouse models, there is increasing evidence that neuronal dysfunction occurs before the accumulation of protein aggregates (i.

A dysregulated endocannabinoid-eicosanoid network supports pathogenesis in a mouse model of Alzheimer's disease.

Although inflammation in the brain is meant as a defense mechanism against neurotoxic stimuli, increasing evidence suggests that uncontrolled, chronic, and persistent inflammation contributes to neurodegeneration. Most neurodegenerative diseases have now been associated with chronic inflammation, including Alzheimer's disease (AD). Whether anti-inflammatory approaches can be used to treat AD, however, is a major unanswered question.

Ablation of the locus coeruleus increases oxidative stress in tg-2576 transgenic but not wild-type mice.

Mice transgenic for production of excessive or mutant forms of beta-amyloid differ from patients with Alzheimer's disease in the degree of inflammation, oxidative damage, and alteration of intermediary metabolism, as well as the paucity or absence of neuronal atrophy and cognitive impairment. Previous observers have suggested that differences in inflammatory response reflect a discrepancy in the state of the locus coeruleus (LC), loss of which is an early change in Alzheimer's disease but which is preserved in the transgenic mice. In this paper, we extend these observations by examining the effects of the LC on markers of oxidative stress and intermediary metabolism.

Neuroprotective profile of novel SRC kinase inhibitors in rodent models of cerebral ischemia.

Src kinase signaling has been implicated in multiple mechanisms of ischemic injury, including vascular endothelial growth factor (VEGF)-mediated vascular permeability that leads to vasogenic edema, a major clinical complication in stroke and brain trauma. Here we report the effects of two novel Src kinase inhibitors, 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[3-(4-methyl-1-piperazinyl)propoxy]-3-quinolinecarbonitrile (SKI-606) and 4-[(2,4-dichloro-5-methoxyphenyl)amino]-6-methoxy-7-[4-(4-methypiperazin-1-yl)but-1-ynyl]-3-quinolinecarbonitrile (SKS-927), on ischemia-induced brain infarction and short- and long-term neurological deficits. Two well established transient [transient middle cerebral artery occlusion (tMCAO)] and permanent [permanent middle cerebral artery occlusion (pMCAO)] focal ischemia models in the rat were used with drug treatments initiated up to 6 h after onset of stroke to mimic the clinical scenario.

Begacestat (GSI-953): a novel, selective thiophene sulfonamide inhibitor of amyloid precursor protein gamma-secretase for the treatment of Alzheimer's disease.

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.

Discovery of begacestat, a Notch-1-sparing gamma-secretase inhibitor for the treatment of Alzheimer's disease.

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.

Enhanced clearance of Abeta in brain by sustaining the plasmin proteolysis cascade.

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.

Induced expression of death domain genes NALP1 and NALP5 following neuronal injury.

Proteins in apoptotic pathways represent potential points of intervention in neurodegenerative disease. We identified several genes of interest that contain death domain such as CARD or Pyrin. We found that ASC and NALP10 were constitutively expressed in cerebellar neurons.

Pancortin-2 interacts with WAVE1 and Bcl-xL in a mitochondria-associated protein complex that mediates ischemic neuronal death.

The actin-modulating protein Wiskott-Aldrich syndrome protein verprolin homologous-1 (WAVE1) and a novel CNS-specific protein, pancortin, are highly enriched in adult cerebral cortex, but their functions are unknown. Here we show that WAVE1 and pancortin-2 interact in a novel cell death cascade in adult, but not embryonic, cerebral cortical neurons. Focal ischemic stroke induces the formation of a protein complex that includes pancortin-2, WAVE1, and the anti-apoptotic protein Bcl-xL.

Phospho-dependent functional modulation of GABA(B) receptors by the metabolic sensor AMP-dependent protein kinase.

GABA(B) receptors are heterodimeric G protein-coupled receptors composed of R1 and R2 subunits that mediate slow synaptic inhibition in the brain by activating inwardly rectifying K(+) channels (GIRKs) and inhibiting Ca(2+) channels. We demonstrate here that GABA(B) receptors are intimately associated with 5'AMP-dependent protein kinase (AMPK). AMPK acts as a metabolic sensor that is potently activated by increases in 5'AMP concentration that are caused by enhanced metabolic activity, anoxia, or ischemia.

Expression of NALP1 in cerebellar granule neurons stimulates apoptosis.

NAcht Leucine-rich-repeat Protein 1 (NALP1) contains a putative nucleotide binding site, a region of leucine-rich repeats, and death domain folds at both termini providing protein/protein association functions such as caspase recruitment. We report here that NALP1 gene expression was induced in primary cerebellar granule neurons (CGN) upon injury. Up-regulation of NALP1 was also observed in a model of transient focal ischemia induced by middle cerebral artery occlusion.

Temporal assessment of caspase activation in experimental models of focal and global ischemia.

Rodent models of focal and global ischemia were used to examine caspase activation. Several readouts were employed on identical tissue to provide correlative measurement of caspase induction, activation and enzymatic activity. In a rat focal ischemia model, caspase-3 enzymatic activity, as recorded by DEVD-AMC cleavage, peaked in penumbral cortex at 6-12 h following ischemia, correlating with increases in caspase 3-cleaved substrates of PARP and alpha-spectrin and subsequent disappearance of caspase-3 zymogen.

Characterization of the WAVE1 knock-out mouse: implications for CNS development.

Developing neurons must respond to a wide range of extracellular signals during the process of brain morphogenesis. One mechanism through which immature neurons respond to such signals is by altering cellular actin dynamics. A recently discovered link between extracellular signaling events and the actin cytoskeleton is the WASP/WAVE (Wiscott-Aldrich Syndrome protein/WASP-family verprolin-homologous protein) family of proteins.