β-site amyloid precursor protein-cleaving enzyme 1(BACE1) inhibitor treatment induces Aβ5-X peptides through alternative amyloid precursor protein cleavage

Erik Portelius, Robert A Dean, Ulf Andreasson, Niklas Mattsson, Anni Westerlund, Maria Olsson, Ronald Bradley Demattos, Margaret M Racke, Henrik Zetterberg, Patrick C May, Kaj Blennow, Erik Portelius, Robert A Dean, Ulf Andreasson, Niklas Mattsson, Anni Westerlund, Maria Olsson, Ronald Bradley Demattos, Margaret M Racke, Henrik Zetterberg, Patrick C May, Kaj Blennow

Abstract

Introduction: The β-secretase enzyme, β-site amyloid precursor protein-cleaving enzyme 1 (BACE1), cleaves amyloid precursor protein (APP) in the first step in β-amyloid (Aβ) peptide production. Thus, BACE1 is a key target for candidate disease-modifying treatment of Alzheimer's disease. In a previous exploratory Aβ biomarker study, we found that BACE1 inhibitor treatment resulted in decreased levels of Aβ1-34 together with increased Aβ5-40, suggesting that these Aβ species may be novel pharmacodynamic biomarkers in clinical trials. We have now examined whether the same holds true in humans.

Methods: In an investigator-blind, placebo-controlled and randomized study, healthy subjects (n =18) were randomly assigned to receive a single dose of 30 mg of LY2811376 (n =6), 90 mg of LY2811376 (n =6), or placebo (n =6). We used hybrid immunoaffinity-mass spectrometry (HI-MS) and enzyme-linked immunosorbent assays to monitor a variety of Aβ peptides.

Results: Here, we demonstrate dose-dependent changes in cerebrospinal fluid (CSF) Aβ1-34, Aβ5-40 and Aβ5-X after treatment with the BACE1-inhibitor LY2811376. Aβ5-40 and Aβ5-X increased dose-dependently, as reflected by two independent methods, while Aβ1-34 dose-dependently decreased.

Conclusion: Using HI-MS for the first time in a study where subjects have been treated with a BACE inhibitor, we confirm that CSF Aβ1-34 may be useful in clinical trials on BACE1 inhibitors to monitor target engagement. Since it is less hydrophobic than longer Aβ species, it is less susceptible to preanalytical confounding factors and may thus be a more stable marker. By independent measurement techniques, we also show that BACE1 inhibition in humans is associated with APP-processing into N-terminally truncated Aβ peptides via a BACE1-independent pathway.

Trial registration: ClinicalTrials.gov NCT00838084. Registered: First received: January 23, 2009, Last updated: July 14, 2009, Last verified: July 2009.

Figures

Figure 1
Figure 1
Mass spectra displaying the effect of treatment on multiple Aβ species in cell media. (A) DMSO (vehicle),(B) 1.25 μM of the BACE1-inhibitor LY2811376, (C) 2.5 μM of the BACE1-inhibitor LY2811376, (D) 1.25 μM of the BACE-inhibitor BACE IV and (E) 2.5 μM of the BACE-inhibitor BACE IV in media from SH-SY5Y cells. *Represent unidentified peaks. Aβ, β-amyloid; BACE, β-site APP-cleaving enzyme; DMSO, dimethyl sulfoide.
Figure 2
Figure 2
Mass spectra displaying multiple Aβ species recovered from human CSF specimens by immunoprecipitation with the anti-Aβ antibodies 6E10 and 4G8. (A) Pre-treatment, (B) 12, (C) 24, and (D) 36 hours post treatment with 90 mg of LY2811376. The right-hand panels are magnified spectra displaying the increase in Aβ5-40 and decrease in Aβ1-34 in response to treatment. Aβ, β-amyloid; CSF, cerebrospinal fluid.
Figure 3
Figure 3
The relative mass spectrometric change from baseline and ELISA-derived concentrations in response to a single dose of 30 mg or 90 mg of the BACE inhibitor LY2811376. (A) Mass spectrometric change in the CSF Aβ1-34 time course after LY2811376 treatment and (B) mass spectrometric change in the CSF Aβ5-40 time course after LY2811376 treatment. (C) ELISA-derived concentrations of the CSF Aβ5-40 time course after LY2811376 treatment and (D) ELISA-derived concentrations of the CSF Aβ5-X time course after LY2811376 treatment. Open circles represent placebo, grey squares represent treatment with 30 mg LY2811376 and closed triangles represent treatment with 90 mg LY2811376. Data are presented as mean ± SD and n =6 for both graphs. Aβ, β-amyloid; BACE, β-site APP-cleaving enzyme; CSF, cerebrospinal fluid; n, number; SD, standard deviation.
Figure 4
Figure 4
Correlation between Aβ5-X and (A) sAPPα and (B) sAPPβ in the 90-mg treatment group. Aβ, β-amyloid; APP, amyloid precursor protein.

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