Pathogenic generation of the 42-amino acid variant of the amyloid ␤-peptide (A␤) by ␤-and ␥-secretase cleavage of the ␤-amyloid precursor protein (APP) is believed to be causative for Alzheimer disease (AD). Lowering of A␤ 42 production by ␥-secretase modulators (GSMs) is a hopeful approach toward AD treatment. The mechanism of GSM action is not fully understood. Moreover, whether GSMs target the A␤ domain is controversial. To further our understanding of the mode of action of GSMs and the cleavage mechanism of ␥-secretase, we analyzed mutations located at different positions of the APP transmembrane domain around or within the A␤ domain regarding their response to GSMs. We found that A␤ 42 -increasing familial AD mutations of the ␥-secretase cleavage site domain responded robustly to A␤ 42 -lowering GSMs, especially to the potent compound GSM-1, irrespective of the amount of A␤ 42 produced. We thus expect that familial AD patients carrying mutations at the ␥-secretase cleavage sites of APP should respond to GSM-based therapeutic approaches. Systematic phenylalanine-scanning mutagenesis of this region revealed a high permissiveness to GSM-1 and demonstrated a complex mechanism of GSM action as other A␤ species (A␤ 41 , A␤ 39 ) could also be lowered besides A␤ 42 . Moreover, certain mutations simultaneously increased A␤ 42 and the shorter peptide A␤ 38 , arguing that the proposed precursor-product relationship of these A␤ species is not general. Finally, mutations of residues in the proposed GSM-binding site implicated in A␤ 42 generation (Gly-29, Gly-33) and potentially in GSM-binding (Lys-28) were also responsive to GSMs, a finding that may question APP substrate targeting of GSMs.Alzheimer disease (AD) 3 is the most common neurodegenerative disorder worldwide. The ␤-amyloid precursor protein (APP), a type I membrane protein, plays a central role in the pathogenesis of the disease (1). Sequential cleavage of APP by ␤-and ␥-secretase generates the amyloid-␤ (A␤) peptide, which deposits as plaques in the brain of affected patients and represents one of the principal pathological hallmarks of the disease (1). ␥-Secretase is an intramembrane-cleaving protease complex, which cleaves the APP transmembrane domain (TMD) in a progressive, stepwise manner via cleavages at the ⑀-, -, and ␥-sites until it is sufficiently shortened to allow the release of A␤ from the membrane (2-4). A␤ peptides generated by ␥-secretase cleavage differ in their C termini. The major product released is A␤ 40 , whereas A␤ 38 and A␤ 42 represent minor species (1). The highly aggregation-prone, neurotoxic A␤ 42 is believed to be causative for AD by initiating a cascade of pathogenic events, which ultimately causes neurodegeneration and dementia (1). Increased production of A␤ 42 underlies the vast majority of mutations associated with familial AD (FAD), which manifests with a very early disease onset. The majority of FAD mutations have been found in PS1, the catalytic subunit of ␥-secretase (5), whereas only a few mutations were found in its h...

Sequential processing of the ␤-amyloid precursor protein by ␤-and ␥-secretase generates the amyloid ␤-peptide (A␤), which is widely believed to play a causative role in Alzheimer disease. Selective lowering of the pathogenic 42-amino acid variant of A␤ by ␥-secretase modulators (GSMs) is a promising therapeutic strategy. Here we report that mutations in presenilin (PS), the catalytic subunit of ␥-secretase, display differential responses to non-steroidal anti-inflammatory drug (NSAID)-type GSMs and more potent second-generation compounds. Although many pathogenic PS mutations resisted lowering of A␤ 42 generation by the NSAID sulindac sulfide, the potent NSAID-like second-generation compound GSM-1 was capable of lowering A␤ 42 for many but not all mutants. We further found that mutations at homologous positions in PS1 and PS2 can elicit differential A␤ 42 responses to GSM-1, suggesting that a positive GSM-1 response depends on the spatial environment in ␥-secretase. The aggressive pathogenic PS1 L166P mutation was one of the few pathogenic mutations that resisted GSM-1, and Leu-166 was identified as a critical residue with respect to the A␤ 42 -lowering response of GSM-1. Finally, we found that GSM-1-responsive and -resistant PS mutants behave very similarly toward other potent second-generation compounds of different structural classes than GSM-1. Taken together, our data show that a positive A␤ 42 response for PS mutants depends both on the particular mutation and the GSM used and that attenuated A␤ 42 responses to low potency GSMs can be overcome for many PS mutants by second generation GSMs. The amyloid ␤-peptide (A␤)4 is a 37-43-amino acid secreted peptide and an invariant pathological hallmark of Alzheimer disease (AD). The 42-amino acid variant A␤ 42 has been suggested to be causative for the disease by triggering the amyloid cascade, a sequence of pathogenic events that ultimately leads to neurodegeneration and dementia in affected patients (1). The pathogenic peptide is generated by a sequential cleavage of the ␤-amyloid precursor protein (APP) by ␤-and ␥-secretase (2). After ␤-secretase cleavage, ␥-secretase cleaves the C-terminal fragment of APP that is left in the membrane by an intramembrane cleavage to release the various A␤ species (3-5). Although A␤ 42 is normally a minor species produced by this cleavage besides the major A␤ 40 species, its production is enhanced by familial AD (FAD) mutations in presenilin (PS) 1 and PS2, the catalytic component of ␥-secretase (6), as well as by a subset of FAD mutations in APP. Targeting ␤-and ␥-secretase by specific inhibitors is one of the current approaches toward an effective AD treatment (7). With respect to ␥-secretase, however, ␥-secretase inhibitors also block the cleavage of Notch1, a major physiological ␥-secretase substrate and, thus, the generation of the Notch1 intracellular domain (NICD), which is a crucial signaling molecule controlling cell differentiation (7). Interfering with the cleavage of this substrate accounts for adverse side effects in...

No abstract