Selective modulation of amyloid-β peptide degradation by flurbiprofen, fenofibrate, and related compounds regulates Aβ levels

Gamma-secretase modulators (GSMs) include selected non-steroidal anti-inflammatory drugs such as flurbiprofen that selectively lowers the neurotoxic amyloid-β peptide Aβ_1-42. GSMs are attractive targets for Alzheimer's disease, in contrast to 'inverse GSMs,' such as fenofibrate, which selectively increase the level of Aβ_1-42. A methodology for screening of Aβ modulating drugs was developed utilizing an Aβ-producing neuroblastoma cell line stably transfected with mutant human amyloid precursor protein, immunoprecipitation of Aβ peptides, and mass spectroscopic quantitation of Aβ_1-37/Aβ_1-38/ Aβ_1-40/Aβ_1-42 using an Aβ internal standard. The unexpected conclusion of this work was that in this system, drug effects are independent of γ-secretase. The methodology recapitulated reported results for modulation of Aβ by GSMs. However, control experiments in which exogenous Aβ_1-40/Aβ_1-42 was added (i) to drug-treated wild-type cells or (ii) to conditioned media from these wild-type cells, gave comparable patterns of Aβ modulation. These results, suggesting that drugs modulate the ability of cell-derived factors to degrade Aβ, was interrogated by adding protease inhibitors and performing molecular weight cut-off fractionation. The results confirmed that modulation of Aβ_1-40/Aβ_1-42 was mediated by selective proteolysis. Treatment of N2a cells with flurbiprofen or fenofibric acid selectively enhanced Aβ_1-42 clearance by extracellular proteolysis; treatment with HCT-1026 or fenofibrate (esters of flurbiprofen and fenobric acid) inhibited clearance of Aβ_1-40 and Aβ_1-42.