Numerous studies have established a pivotal role for A␤42 in Alzheimer's disease (AD) pathogenesis. In contrast, although A␤40 is the predominant form of amyloid ␤ (A␤) produced and accumulates to a variable degree in the human AD brain, its role in AD pathogenesis has not been established. It has generally been assumed that an increase in A␤40 would accelerate amyloid plaque formation in vivo. We have crossed BRI-A␤40 mice that selectively express high levels of A␤40 with both Tg2576 (APPswe, K670NϩM671L) mice and BRI-A␤42A mice expressing A␤42 selectively and analyzed parenchymal and cerebrovascular A␤ deposition in the bitransgenic mice compared with their singly transgenic littermates. In the bitransgenic mice, the increased steady-state levels of A␤40 decreased A␤ deposition by 60 -90%. These results demonstrate that A␤42 and A␤40 have opposing effects on amyloid deposition: A␤42 promotes amyloid deposition but A␤40 inhibits it. In addition, increasing A␤40 levels protected BRI-A␤40/Tg2576 mice from the premature-death phenotype observed in Tg2576 mice. The protective properties of A␤40 with respect to amyloid deposition suggest that strategies that preferentially target A␤40 may actually worsen the disease course and that selective increases in A␤40 levels may actually reduce the risk for development of AD.

Selective lowering of Abeta42 levels (the 42-residue isoform of the amyloid-beta peptide) with small-molecule gamma-secretase modulators (GSMs), such as some non-steroidal anti-inflammatory drugs, is a promising therapeutic approach for Alzheimer's disease. To identify the target of these agents we developed biotinylated photoactivatable GSMs. GSM photoprobes did not label the core proteins of the gamma-secretase complex, but instead labelled the beta-amyloid precursor protein (APP), APP carboxy-terminal fragments and amyloid-beta peptide in human neuroglioma H4 cells. Substrate labelling was competed by other GSMs, and labelling of an APP gamma-secretase substrate was more efficient than a Notch substrate. GSM interaction was localized to residues 28-36 of amyloid-beta, a region critical for aggregation. We also demonstrate that compounds known to interact with this region of amyloid-beta act as GSMs, and some GSMs alter the production of cell-derived amyloid-beta oligomers. Furthermore, mutation of the GSM binding site in the APP alters the sensitivity of the substrate to GSMs. These findings indicate that substrate targeting by GSMs mechanistically links two therapeutic actions: alteration in Abeta42 production and inhibition of amyloid-beta aggregation, which may synergistically reduce amyloid-beta deposition in Alzheimer's disease. These data also demonstrate the existence and feasibility of 'substrate targeting' by small-molecule effectors of proteolytic enzymes, which if generally applicable may significantly broaden the current notion of 'druggable' targets.