P-Glycoprotein Efflux And Other Factors Limit Brain Amyloid Beta Reduction By Beta-Site Amyloid Precursor Protein-Cleaving Enzyme 1 Inhibitors In Mice

Alzheimer's disease (AD) is a progressive neurodegenerative disease. Amyloid beta (A beta) peptides are hypothesized to cause the initiation and progression of AD based on pathologic data from AD patients, genetic analysis of mutations that cause early onset forms of AD, and preclinical studies. Based on this hypothesis, beta-site amyloid precursor protein (APP)-cleaving enzyme 1 (BACE1) inhibitors are an attractive therapeutic approach for AD because cleavage of the APP by BACE1 is required to form A beta. In this study, three potent BACE1 inhibitors are characterized. All three inhibitors decrease A beta formation in cultured cells with IC(50) values less than 10 nM. Analysis of APP C-terminal fragments by immunoblotting and A beta peptides by mass spectrometry showed that these inhibitors decreased A beta by inhibiting BACE1. An assay for A beta 1-40 in mice was developed and used to show that these BACE1 inhibitors decreased plasma A beta 1-40, but not brain A beta 1-40, in wild-type mice. Because these BACE1 inhibitors were substrates for P-glycoprotein (P-gp), a member of the ATP-binding cassette superfamily of efflux transporters, these inhibitors were administered to P-gp knockout (KO) mice. These studies showed that all three BACE1 inhibitors decreased brain A beta 1-40 in P-gp KO mice, demonstrating that P-gp is a major limitation for development of BACE1 inhibitors to test the amyloid hypothesis. A comparison of plasma A beta 1-40 and brain A beta 1-40 dose responses for these three compounds revealed differences in relative ED(50) values, indicating that factors other than P-gp can also contribute to poor brain activity by BACE1 inhibitors.