Passive immunization against pyroglutamate-3 amyloid-β reduces plaque burden in Alzheimer-like transgenic mice: a pilot study

Abstract

Background: N-terminally truncated and modified pyroglutamate-3 amyloid-β protein (pE3-Aβ) is present in most, if not all, cerebral plaque and vascular amyloid deposits in human Alzheimer's disease (AD). pE3-Aβ deposition is also found in AD-like transgenic (tg) mouse brain, albeit in lesser quantities than general Aβ. pE3-Aβ resists degradation, is neurotoxic, and may act as a seed for Aβ aggregation.

Objective: We sought to determine if pE3-Aβ removal by passive immunization with a highly specific monoclonal antibody (mAb) impacts pathogenesis in a mouse model of Alzheimer's amyloidosis.

Methods: APPswe/PS1ΔE9 tg mice were given weekly intraperitoneal injections of a new anti-pE3-Aβ mAb (mAb07/1) or PBS from 5.8 to 13.8 months of age (prevention) or from 23 to 24.7 months of age (therapeutic). Multiple forms of cerebral Aβ were quantified pathologically and biochemically. Gliosis and microhemorrhage were examined.

Results: Chronic passive immunization with an anti-pE3-Aβ mAb significantly reduced total plaque deposition and appeared to lower gliosis in the hippocampus and cerebellum in both the prevention and therapeutic studies. Insoluble Aβ levels in hemibrain homogenates were not significantly different between immunized and control mice. Microhemorrhage was not observed with anti-pE3-Aβ immunotherapy.

Conclusions: Selective removal of pE3-Aβ lowered general Aβ plaque deposition suggesting a pro-aggregation or seeding role for pE3-Aβ.

Fig. 1

Characterization of pE3-Aβ mAb07/1. a Western blot of Aβ(1–40), Aβ(3–40) and pE3-Aβ(3–40), detected with mAb07/1 or mAb6E10. Peptides (20 ng each) were separated in gels containing 8 M urea. The anti-pE3-Aβ mAb does not detect the truncated precursor or full-length Aβ. b Analysis of antibody binding using surface plasmon resonance. The peptides were covalently linked and the antibody applied in a buffered solution. Significant binding of mAb07/1 was only observed with pE3-Aβ (3–40) immobilized on the surface. Other peptides analyzed are: MCP-1 and 2; gastrin; GnRH; neurotensin; orexin; TRH; the N-terminus of collagen, and fibronectin.

Fig. 2

In a prevention study initiated during the early stage of plaque deposition, weekly passive immunization of APPswe/PS1ΔlE9 mice with anti-pE3-Aβ mAb07/1 from 5.8 to 13.8 months of age significantly reduced pE3-Aβ as well as general Aβ (R1282 IR) and fibrillar amyloid (Thioflavin S) deposition in the hippocampus (a, b) and cerebellum (c, d) compared to that in PBS control mice. Immunohistochemical results (a, c) and Thioflavin S labeling were quantified by image analysis (b, d). Absolute values are provided in table 1. Scale bars, 200 µm. p values: * p > 0.05; ** p > 0.01; *** p > 0.001; n.s. = nonsignificant (p = 0.089).

Fig. 3

In a therapeutic study initiated well after the onset of cerebral Aβ deposition and gliosis, weekly passive immunization with anti-pE3-Aβ mAb07/1 in 23-monthold APPswe/PS1ΔE9 mice for 7 weeks resulted in the attenuation of pE3-Aβ and general Aβ (R1282 IR) deposition as well as fibrillar amyloid (Thioflavin S) in the hippocampus (a, b) and cerebellum (c, d) compared to PBS control mice. Immunohistochemical results (a, c) and Thioflavin S labeling were quantified by image analysis (b, d). Absolute values are provided in table 1. Scale bars, 200 µm. p values: * p > 0.05; *** p > 0.001; n.s. = nonsignificant (p = 0.11); p = 0.06 (strong trend).