Imaging asparaginyl endopeptidase (AEP) in the live brain as a biomarker for Alzheimer’s disease

Abstract

Background Discovery of early-stage biomarkers is a long-sought goal of Alzheimer’s disease (AD) diagnosis. Age is the greatest risk factor for most AD and accumulating evidence suggests that age-dependent elevation of asparaginyl endopeptidase (AEP) in the brain may represent a new biological marker for predicting AD. However, this speculation remains to be explored with an appropriate assay method because mammalian AEP exists in many organs and the level of AEP in body fluid isn’t proportional to its concentration in brain parenchyma. To this end, we here modified gold nanoparticle (AuNPs) into an AEP-responsive imaging probe and choose transgenic APPswe/PS1dE9 (APP/PS1) mice as an animal model of AD. Our aim is to determine whether imaging of brain AEP can be used to predict AD pathology. Results This AEP-responsive imaging probe AuNPs-Cy5.5-A&C consisted of two particles, AuNPs-Cy5.5-AK and AuNPs-Cy5.5-CABT, which were respectively modified with Ala–Ala–Asn–Cys–Lys (AK) and 2-cyano-6-aminobenzothiazole (CABT). We showed that AuNPs-Cy5.5-A&C could be selectively activated by AEP to aggregate and emit strong fluorescence. Moreover, AuNPs-Cy5.5-A&C displayed a general applicability in various cell lines and its florescence intensity correlated well with AEP activity in these cells. In the brain of APP/PS1 transgenic mice , AEP activity was increased at an early disease stage of AD that precedes formation of senile plaques and cognitive impairment. Pharmacological inhibition of AEP with δ-secretase inhibitor 11 (10 mg kg−1, p.o.) reduced production of β-amyloid (Aβ) and ameliorated memory loss. Therefore, elevation of AEP is an early sign of AD onset. Finally, we showed that live animal imaging with this AEP-responsive probe could monitor the up-regulated AEP in the brain of APP/PS1 mice. Conclusions The current work provided a proof of concept that assessment of brain AEP activity by in vivo imaging assay is a potential biomarker for early diagnosis of AD. Graphical abstract

Pharmacological Inhibition of the Asparaginyl Endopeptidase (AEP) in an Alzheimer’s Disease Model Improves the Survival and Efficacy of Transplanted Neural Stem Cells

“…Elevation of AEP in AD and aged brains has been documented by many studies. AEP was reported as an upstream secretase that facilitates amyloidogenic APP processing and enhances Aβ secretion, i.e., an important player for disease onset of AD [36][37][38][39][40][41][42][43][44]. It is unsurprising that inhibition of AEP can decrease Aβ generation.…”

The Asparaginyl Endopeptidase Legumain: An Emerging Therapeutic Target and Potential Biomarker for Alzheimer’s Disease

“…Similarly, we treated SAMP8 and APP/PS1 mice with δ-secretase inhibitor 11 (10 mg kg −1 , p.o.) for 3 months; the brain AEP activity of animal models was suppressed along with a significantly lesser deposit of Aβ and ameliorated memory loss [ 15 , 49 ]. Considering that the oral plasma elimination half-life (t 1/2 ) of δ-secretase inhibitor 11 in mice is too short, about 2.31 h, although this AEP inhibitor is effective in treating animals, its practical significance in the application to human AD is very limited.…”

“…To perform the in vivo imaging analysis of brain AEP activity, we recently utilized gold nanoparticles (AuNPs) as carriers synthesized an aggregation reaction-based fluorescent AEP imaging probe, for which Cy5.5 conjugated AuNPs could be selectively activated by cellular AEPs and emit strong fluorescence [ 49 ]. This AEP-targeted probe had a general applicability in various cell lines, including glioma cells and degenerative neurons, and its florescence imaging intensity correlated well with the enzymatic activity of AEPs expressed in these cells.…”

“…Our AEP probes were based on a fluorescence imaging mechanism and tested on animals [ 49 ]. The avenue of developing new brain AEP imaging probes and methods is open.…”

The Mammalian Cysteine Protease Legumain in Health and Disease

“…Both ADAM10 and 17 are legumain substrates (Table 1) and are cleaved and degraded by legumain in the lysosomes [28], which subsequently could trigger AD onset or progression. In addition, legumain has been suggested as an AD prognostic biomarker and elevated legumain monitored by live animal imaging was shown to be an early sign of AD onset in an AD pre-clinical murine model [127].…”