Converging lines of evidence implicate the beta-amyloid peptide (Ab) as causative in Alzheimer's disease. We describe a novel class of compounds that reduce Ab production by functionally inhibiting g-secretase, the activity responsible for the carboxy-terminal cleavage required for Ab production. These molecules are active in both 293 HEK cells and neuronal cultures, and exert their effect upon Ab production without affecting protein secretion, most notably in the secreted forms of the amyloid precursor protein (APP). Oral administration of one of these compounds, N-[N-(3,5-di¯uoro-phenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester, to mice transgenic for human APP V717F reduces brain levels of Ab in a dose-dependent manner within 3 h. These studies represent the ®rst demonstration of a reduction of brain Ab in vivo. Development of such novel functional g-secretase inhibitors will enable a clinical examination of the Ab hypothesis that Ab peptide drives the neuropathology observed in Alzheimer's disease.

Passive immunization with an antibody directed against the N terminus of amyloid ␤ (A␤) has recently been reported to exacerbate cerebral amyloid angiopathy (CAA)-related microhemorrhage in a transgenic animal model. Although the mechanism responsible for the deleterious interaction is unclear, a direct binding event may be required. We characterized the binding properties of several monoclonal anti-A␤ antibodies to deposited A␤ in brain parenchyma and CAA. Biochemical analyses demonstrated that the 3D6 and 10D5, two N-terminally directed antibodies, bound with high affinity to deposited forms of A␤, whereas 266, a central domain antibody, lacked affinity for deposited A␤. To determine whether 266 or 3D6 would exacerbate CAA-associated microhemorrhage, we treated aged PDAPP mice with either antibody for 6 weeks. We observed an increase in both the incidence and severity of CAA-associated microhemorrhage when PDAPP transgenic mice were treated with the N-terminally directed 3D6 antibody, whereas mice treated with 266 were unaffected. These results may have important implications for future immune-based therapeutic strategies for Alzheimer's disease.

Neurodegenerative processes in Alzheimer disease (AD) are thought to be driven in part by the deposition of amyloid f3 (Af8), a 39-to 43-amino acid peptide product resulting from an alternative cleavage of amyloid precursor protein. Recent descriptions ofin vitro neurotoxic effects ofA18 support this hypothesis and suggest toxicity might be mediated by A,B-induced neuronal calcium disregulation. In addition, it has been reported that "aging" A18 results in increased toxic potency due to peptide aggregation and formation of a (8-sheet secondary structure. In addition, Aj8 might also promote neuropathology indirectly by activating immune/inflammatory pathways in affected areas of the brain (e.g., cortex and hippocampus). Here we report that A13 can modulate cytokine secretion ] from human astrocytoma cells (U-373 MG). Freshly prepared and aged Af8 modestly stimulated IL-6 and IL-8 secretion from U-373 MG cells. However, in the presence of interleukin-1g3 (IL-1p8), aged, but not fresh, Ai3 markedly potentiated (3-to 8-fold) cytokine release. In contrast, aged A(3 did not potentiate substance P (NK-1)-or histamine (H1)-stimulated cytokine production. Further studies showed that IL-1p8-induced cytokine release was potentiated by A13-(25-35), while A(8-(1-16) was inactive. Calcium disregulation may be responsible for the effects of A(3 on cytokine production, since the calcium ionophore A23187 similarly potentiated IL-1p8-induced cytokine secretion and EGTA treatment blocked either A(8 or A23187 activity. Thus, chronic neurodegeneration in AD-affected brain regions may be mediated in part by the ability of A(3 to exacerbate inflammatory pathways in a conformation-dependent manner.Alzheimer disease (AD) is a progressive dementia affecting 1-6% of the elderly (>65 years old) and is the fourth leading cause of death in developed countries (1). As a neurodegenerative disorder, AD is pathologically characterized by cortical atrophy, neuronal loss, and the presence of neurofibrillary tangles and senile amyloid plaques. Amyloid plaques are anatomically defined as (i) "diffuse" or "preamyloid" plaques that consist of amorphous, noncongophillic amyloid 13 (AP3) deposits containing few, if any, reactive astrocytes and microglia (diffuse plaques are not associated with neuronal pathology); and (ii) "senile" or "neuritic" plaques consisting of fibrillar, congophillic AP3 deposits containing reactive astrocytes and microglia and surrounded by neuritic degeneration. These characteristic lesions are localized to regions of the brain important for cognition, learning, and memory-e.g., cerebral cortex and hippocampus (1). The neuropathology of AD, driving the progressive loss of cognitive function and memory, may result from the deposition of A,B, a 39-to 43-amino acid peptide product of an alternative cleavage of amyloid precursor protein (APP) (2,3

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