Macrocyclic BACE-1 inhibitors acutely reduce Abeta in brain after po application.

A series of macrocyclic peptidic BACE-1 inhibitors was designed. While potency on BACE-1 was rather high, the first set of compounds showed poor brain permeation and high efflux in the MDRI-MDCK assay. The replacement of the secondary benzylamino group with a phenylcyclopropylamino group maintained potency on BACE-1, while P-glycoprotein-mediated efflux was significantly reduced and brain permeation improved. Several compounds from this series demonstrated acute reduction of Abeta in human APP-wildtype transgenic (APP51/16) mice after oral administration.

Macrocyclic peptidomimetic beta-secretase (BACE-1) inhibitors with activity in vivo.

The macrocyclic peptidic BACE-1 inhibitors 2a-c show moderate enzymatic and cellular activity. By exchange of the hydroxyethylene- to ethanolamine-transition state mimetic the peptidic character was reduced, providing the highly potent and selective inhibitor 3. Variation of the P' moiety resulted in the macrocyclic inhibitor 14. Both macrocycles show inhibition of BACE-1 in the brain of APP51/16 transgenic mice, 3 (NB-544) after intravenous and 14 (NB-533) after oral application.

Dynamics of Abeta turnover and deposition in different beta-amyloid precursor protein transgenic mouse models following gamma-secretase inhibition.

Human beta-amyloid precursor protein (APP) transgenic mice are commonly used to test potential therapeutics for Alzheimer's disease. We have characterized the dynamics of beta-amyloid (Abeta) generation and deposition following gamma-secretase inhibition with compound LY-411575 [N(2)-[(2S)-2-(3,5-difluorophenyl)-2-hydroxyethanoyl]-N(1)-[(7S)-5-methyl-6-oxo-6,7-dihydro-5H-dibenzo[b,d]azepin-7-yl]-L-alaninamide]. Kinetic studies in preplaque mice distinguished a detergent-soluble Abeta pool in brain with rapid turnover (half-lives for Abeta40 and Abeta42 were 0.7 and 1.7 h) and a much more stable, less soluble pool. Abeta in cerebrospinal fluid (CSF) reflected the changes in the soluble brain Abeta pool, whereas plasma Abeta turned over more rapidly. In brain, APP C-terminal fragments (CTF) accumulated differentially. The half-lives for gamma-secretase degradation were estimated as 0.4 and 0.1 h for C99 and C83, respectively. Three different APP transgenic lines responded very similarly to gamma-secretase inhibition regardless of the familial Alzheimer's disease mutations in APP. Amyloid deposition started with Abeta42, whereas Abeta38 and Abeta40 continued to turn over. Chronic gamma-secretase inhibition lowered amyloid plaque formation to a different degree in different brain regions of the same mice. The extent was inversely related to the initial amyloid load in the region analyzed. No evidence for plaque removal below baseline was obtained. gamma-Secretase inhibition led to a redistribution of intracellular Abeta and an elevation of CTFs in neuronal fibers. In CSF, Abeta showed a similar turnover as in preplaque animals demonstrating its suitability as marker of newly generated, soluble Abeta in plaque-bearing brain. This study supports the use of APP transgenic mice as translational models to characterize Abeta-lowering therapeutics.

In vivo detection of amyloid-beta deposits by near-infrared imaging using an oxazine-derivative probe.

As Alzheimer's disease pathogenesis is associated with the formation of insoluble aggregates of amyloid beta-peptide, approaches allowing the direct, noninvasive visualization of plaque growth in vivo would be beneficial for biomedical research. Here we describe the synthesis and characterization of the near-infrared fluorescence oxazine dye AOI987, which readily penetrates the intact blood-brain barrier and binds to amyloid plaques. Using near-infrared fluorescence imaging, we demonstrated specific interaction of AOI987 with amyloid plaques in APP23 transgenic mice in vivo, as confirmed by postmortem analysis of brain slices. Quantitative analysis revealed increasing fluorescence signal intensity with increasing plaque load of the animals, and significant binding of AOI987 was observed for APP23 transgenic mice aged 9 months and older. Thus, AOI987 is an attractive probe to noninvasively monitor disease progression in animal models of Alzheimer disease and to evaluate effects of potential Alzheimer disease drugs on the plaque load.

Redistribution of GABAB(1) protein and atypical GABAB responses in GABAB(2)-deficient mice.

GABAB receptors mediate slow synaptic inhibition in the nervous system. In transfected cells, functional GABAB receptors are usually only observed after coexpression of GABAB(1) and GABAB(2) subunits, which established the concept of heteromerization for G-protein-coupled receptors. In the heteromeric receptor, GABAB(1) is responsible for binding of GABA, whereas GABAB(2) is necessary for surface trafficking and G-protein coupling. Consistent with these in vitro observations, the GABAB(1) subunit is also essential for all GABAB signaling in vivo. Mice lacking the GABAB(1) subunit do not exhibit detectable electrophysiological, biochemical, or behavioral responses to GABAB agonists. However, GABAB(1) exhibits a broader cellular expression pattern than GABAB(2), suggesting that GABAB(1) could be functional in the absence of GABAB(2). We now generated GABAB(2)-deficient mice to analyze whether GABAB(1) has the potential to signal without GABAB(2) in neurons. We show that GABAB(2)-/- mice suffer from spontaneous seizures, hyperalgesia, hyperlocomotor activity, and severe memory impairment, analogous to GABAB(1)-/- mice. This clearly demonstrates that the lack of heteromeric GABAB(1,2) receptors underlies these phenotypes. To our surprise and in contrast to GABAB(1)-/- mice, we still detect atypical electrophysiological GABAB responses in hippocampal slices of GABAB(2)-/- mice. Furthermore, in the absence of GABAB(2), the GABAB(1) protein relocates from distal neuronal sites to the soma and proximal dendrites. Our data suggest that association of GABAB(2) with GABAB(1) is essential for receptor localization in distal processes but is not absolutely necessary for signaling. It is therefore possible that functional GABAB receptors exist in neurons that naturally lack GABAB(2) subunits.