Immunodominant epitope and properties of pyroglutamate-modified Aβ-specific antibodies produced in rabbits

Abstract

N-truncated and N-modified forms of amyloid beta (Aß) peptide are found in diffused and dense core plaques in Alzheimer's disease (AD) and Down's syndrome patients as well as transgenic mouse models of AD. Although the pathological significance of these shortened forms Aβ is not completely understood, previous studies have demonstrated that these peptides are significantly more resistant to degradation, aggregate more rapidly in vitro and exhibit similar or, in some cases, increased toxicity in hippocampal neuronal cultures compared to the full-length peptides. In the present study we further investigated the mechanisms of toxicity of one of the most abundant Ntruncated/modified Aβ peptide bearing amino-terminal pyroglutamate at position 3 (AβN3(pE)). We demonstrated that AβN3(pE) oligomers induce phosphatidyl serine externalization and membrane damage in SH-SY5Y cells. Also, we produced AβN3(pE)-specific polyclonal antibodies in rabbit and identified an immunodominant epitope recognized by anti-AβN3(pE) antibodies. Our results are important for developing new immunotherapeutic compounds specifically targeting AβN3(pE) aggregates since the most commonly used immunogens in the majority of vaccines for AD have been shown to induce antibodies that recognize the N-terminal immunodominant epitope (EFRH) of the full length Aβ, which is absent in N-amino truncated peptides.

Inhibiting β-Secretase Activity in Alzheimer's Disease Cell Models with Single-Chain Antibodies Specifically Targeting APP

“…This region also includes the strongly immunogenic N-terminal sequence of soluble Aβ. 32 Since we wanted an inhibitor that selectively binds APP but not Aβ, we first performed a negative selection using FACS to remove clones that bound Aβ. While some strong Aβ binders did remain, as evidenced by the recognition of Aβ by the strongest binders in the P1 pool (Fig.…”

New progress in active immunotherapy targeting to amyloid beta

“…Gevorkian and coworkers [37,38] reported the use of pyroglutamate-modified A as a new antigen in A immunotherapy. Zvirbliene and coworkers [39] showed the size of A oligomers may influence the immune response.…”

Pyroglutamate-Modified Amyloid Beta Peptides: Emerging Targets for Alzheimer´s Disease Immunotherapy

“…In addition, AβN3(pE) was shown to induce a redox-sensitive neuronal apoptosis involving caspase activation and an arachidonic acid-dependent pro-inflammatory pathway in primary neuronal cultures [41]. Recently, our group demonstrated that AβN3(pE) oligomers induce phosphatidylserine externalization and membrane damage in differentiated SH-SY5Y cells [56]. Moreover, AβN3(pE) was shown to inhibit long term potentiation in mouse hippocampal slices [54].…”

“…In recent years, a couple of laboratories started to include pyroglutamate-modified Aβ in their research agenda. Our group performed first studies on immunogenicity of AβN3(pE) and AβN11(pE) in rabbits and demonstrated that while AβN3(pE) peptide induces antibodies specifically binding to AβN3(pE), anti-AβN11(pE) antibodies recognize two pyroglutamate species, AβN3(pE) and AβN11(pE), and full-length Aβ as well [56, 88]. The latter results suggest that AβN3(pE), AβN11(pE) and full-length Aβ may share a common B cell epitope, and are important for designing immunogens capable of inducing antibodies targeting three main pathological species of the Aβ peptide present in human brain.…”

“…This should significantly enhance the efficacy of immunotherapy in the CNS of AD patients, because only approximately 0.1% of the antibody in the blood gains entry into the brain. We also performed epitope mapping of anti-AβN3(pE) and anti-AβN11(pE) antibodies and demonstrated the presence of one immunodominant epitope at the N-terminal part of AβN3(pE) and two major B cell epitopes in AβN11(pE) (one at the N-terminal part (aa 11-15) and another at the central part (aa 20-24) of the peptide) [56, 88]. Studies on immunogenic properties of identified epitopes/mimotopes are underway.…”