BACKGROUND: Formation of deposits of the insoluble amyloid beta-peptide is believed to be causally related with neurodegeneration in Alzheimer disease (AD). The beta-peptide originates from a larger amyloid precursor protein (APP) by the action of proteolytic enzymes. The first proteolytic event leading to amyloid formation is the cleavage of APP by the membrane-bound aspartyl protease BACE-1, also known as memapsin-2. Inhibition of BACE-1 is thought to be a therapeutic approach to AD. Measuring BACE-1 activity in biological samples would be useful to elucidate the mechanism of AD and for development of AD drugs. METHODS: We developed a sensitive and specific activity assay for BACE-1. The assay is based on a genetically engineered proenzyme that is specifically activated by BACE-1. The resulting active enzyme is measured with a chromogenic substrate. The use of 2 coupled reactions produces a detection limit as low as 0.4 pmol/L. RESULTS: The assay detected BACE-1 activity in extracts of human brain tissue as well as, unexpectedly, in human cerebrospinal fluid (CSF). Gel electrophoresis and Western blotting identified the BACE-1 present in CSF as a truncated soluble form of the originally membrane-bound BACE-1. CONCLUSION: Detection of the soluble form of BACE-1 in CSF, a relatively easily accessible biological fluid, may be useful for monitoring the effects of drug candidates in vivo and may have diagnostic or prognostic applications.
BACKGROUND: Formation of deposits of the insoluble amyloid beta-peptide is believed to be causally related with neurodegeneration in Alzheimer disease (AD). The beta-peptide originates from a larger amyloid precursor protein (APP) by the action of proteolytic enzymes. The first proteolytic event leading to amyloid formation is the cleavage of APP by the membrane-bound aspartyl protease BACE-1, also known as memapsin-2. Inhibition of BACE-1 is thought to be a therapeutic approach to AD. Measuring BACE-1 activity in biological samples would be useful to elucidate the mechanism of AD and for development of AD drugs. METHODS: We developed a sensitive and specific activity assay for BACE-1. The assay is based on a genetically engineered proenzyme that is specifically activated by BACE-1. The resulting active enzyme is measured with a chromogenic substrate. The use of 2 coupled reactions produces a detection limit as low as 0.4 pmol/L. RESULTS: The assay detected BACE-1 activity in extracts of human brain tissue as well as, unexpectedly, in human cerebrospinal fluid (CSF). Gel electrophoresis and Western blotting identified the BACE-1 present in CSF as a truncated soluble form of the originally membrane-bound BACE-1. CONCLUSION: Detection of the soluble form of BACE-1 in CSF, a relatively easily accessible biological fluid, may be useful for monitoring the effects of drug candidates in vivo and may have diagnostic or prognostic applications.
Authors: Harald Hampel; Richard Frank; Karl Broich; Stefan J Teipel; Russell G Katz; John Hardy; Karl Herholz; Arun L W Bokde; Frank Jessen; Yvonne C Hoessler; Wendy R Sanhai; Henrik Zetterberg; Janet Woodcock; Kaj Blennow Journal: Nat Rev Drug Discov Date: 2010-07 Impact factor: 84.694
Authors: Harald Hampel; Yong Shen; Dominic M Walsh; Paul Aisen; Les M Shaw; Henrik Zetterberg; John Q Trojanowski; Kaj Blennow Journal: Exp Neurol Date: 2009-10-06 Impact factor: 5.330
Authors: Justyna A Dobrowolska; Maria S Michener; Guoxin Wu; Bruce W Patterson; Robert Chott; Vitaliy Ovod; Yuriy Pyatkivskyy; Kristin R Wildsmith; Tom Kasten; Parker Mathers; Mandy Dancho; Christina Lennox; Brad E Smith; David Gilberto; Debra McLoughlin; Daniel J Holder; Andrew W Stamford; Kevin E Yarasheski; Matthew E Kennedy; Mary J Savage; Randall J Bateman Journal: J Neurosci Date: 2014-06-11 Impact factor: 6.167
Authors: Inmaculada Lopez-Font; Claudia P Boix; Henrik Zetterberg; Kaj Blennow; Javier Sáez-Valero Journal: Mol Neurobiol Date: 2019-07-09 Impact factor: 5.590
Authors: David R Taylor; Edward T Parkin; Sarah L Cocklin; James R Ault; Alison E Ashcroft; Anthony J Turner; Nigel M Hooper Journal: J Biol Chem Date: 2009-06-29 Impact factor: 5.157