AIM: To identify a small molecule L655,240 as a novel β-secretase (BACE1) inhibitor and to investigate its effects on β-amyloid (Aβ) generation in vitro. METHODS: Fluorescence resonance energy transfer (FRET) was used to characterize the inhibitory effect of L655,240 on BACE1. Surface plasmon resonance (SPR) technology-based assay was performed to study the binding affinity of L655,240 for BACE1. The selectivity of L655,240 toward BACE1 over other aspartic proteases was determined with enzymatic assay. The effects of L655,240 on Aβ40, Aβ42, and sAPPβ production were studied in HEK293 cells stably expressing APP695 Swedish mutant(K595N/M596L) (HEK293-APPswe cells). The activities of BACE1, γ-secretase and α-secretase were assayed, and both the mRNA and protein levels of APP and BACE1 were evaluated using real-time PCR (RT-PCR) and Western blot analysis. RESULTS: L655,240 was determined to be a competitive, selective BACE1 inhibitor (IC(50)=4.47±1.37 μmol/L), which bound to BACE1 directly (K(D)=17.9±0.72 μmol/L). L655,240 effectively reduced Aβ40, Aβ42, and sAPPβ production by inhibiting BACE1 without affecting the activities of γ-secretase and α-secretase in HEK293-APPswe cells. L655,240 has no effect on APP and BACE1 mRNA or protein levels in HEK293-APPswe cells. CONCLUSION: The small molecule L655,240 is a novel BACE1 inhibitor that can effectively decreases Aβ production in vitro, thereby highlighting its therapeutic potential for the treatment of Alzheimer's disease.
AIM: To identify a small molecule L655,240 as a novel β-secretase (BACE1) inhibitor and to investigate its effects on β-amyloid (Aβ) generation in vitro. METHODS: Fluorescence resonance energy transfer (FRET) was used to characterize the inhibitory effect of L655,240 on BACE1. Surface plasmon resonance (SPR) technology-based assay was performed to study the binding affinity of L655,240 for BACE1. The selectivity of L655,240 toward BACE1 over other aspartic proteases was determined with enzymatic assay. The effects of L655,240 on Aβ40, Aβ42, and sAPPβ production were studied in HEK293 cells stably expressing APP695 Swedish mutant(K595N/M596L) (HEK293-APPswe cells). The activities of BACE1, γ-secretase and α-secretase were assayed, and both the mRNA and protein levels of APP and BACE1 were evaluated using real-time PCR (RT-PCR) and Western blot analysis. RESULTS:L655,240 was determined to be a competitive, selective BACE1 inhibitor (IC(50)=4.47±1.37 μmol/L), which bound to BACE1 directly (K(D)=17.9±0.72 μmol/L). L655,240 effectively reduced Aβ40, Aβ42, and sAPPβ production by inhibiting BACE1 without affecting the activities of γ-secretase and α-secretase in HEK293-APPswe cells. L655,240 has no effect on APP and BACE1 mRNA or protein levels in HEK293-APPswe cells. CONCLUSION: The small molecule L655,240 is a novel BACE1 inhibitor that can effectively decreases Aβ production in vitro, thereby highlighting its therapeutic potential for the treatment of Alzheimer's disease.
Authors: Ellen W Baxter; Kelly A Conway; Ludo Kennis; François Bischoff; Marc H Mercken; Hans L De Winter; Charles H Reynolds; Brett A Tounge; Chi Luo; Malcolm K Scott; Yifang Huang; Mirielle Braeken; Serge M A Pieters; Didier J C Berthelot; Stefan Masure; Wouter D Bruinzeel; Alfonzo D Jordan; Michael H Parker; Robert E Boyd; Junya Qu; Richard S Alexander; Douglas E Brenneman; Allen B Reitz Journal: J Med Chem Date: 2007-08-08 Impact factor: 7.446