Conformation-sensitive Antibodies against Alzheimer Amyloid-β by Immunization with a Thioredoxin-constrained B-cell Epitope Peptide

Abstract

Immunotherapy against the amyloid-␤ (A␤) peptide is a valuable potential treatment for Alzheimer disease (AD). An ideal antigen should be soluble and nontoxic, avoid the C-terminally located T-cell epitope of A␤, and yet be capable of eliciting antibodies that recognize A␤ fibrils and neurotoxic A␤ oligomers but not the physiological monomeric species of A␤. We have described here the construction and immunological characterization of a recombinant antigen with these features obtained by tandem multimerization of the immunodominant B-cell epitope peptide A␤1-15 (A␤15) within the active site loop of bacterial thioredoxin (Trx). Chimeric Trx(A␤15) n polypeptides bearing one, four, or eight copies of A␤15 were constructed and injected into mice in combination with alum, an adjuvant approved for human use. All three polypeptides were found to be immunogenic, yet eliciting antibodies with distinct recognition specificities. The anti-Trx(A␤15) 4 antibody, in particular, recognized A␤42 fibrils and oligomers but not monomers and exhibited the same kind of conformational selectivity against transthyretin, an amyloidogenic protein unrelated in sequence to A␤. We have also demonstrated that anti-Trx(A␤15) 4 , which binds to human AD plaques, markedly reduces A␤ pathology in transgenic AD mice. The data indicate that a conformational epitope shared by oligomers and fibrils can be mimicked by a thioredoxin-constrained A␤ fragment repeat and identify Trx(A␤15) 4 as a promising new tool for AD immunotherapy.Antipeptide antibodies are valuable tools for probing structure-function relationships in proteins as well as for therapeutic and diagnostic applications. When immunotherapy is the ultimate goal, the sequence span of the peptide, and thus the nature of the epitope(s) associated with it, as well as the conformational selectivity of the resulting antibodies may also be dictated by safety reasons. This is the case of the amyloid-␤ (A␤) 2 peptide, a major neuropathological hallmark (1-3) and a promising immunotherapeutic target of Alzheimer disease (AD) (4 -7). Following the encouraging results obtained with A␤42 vaccination in transgenic mouse models of AD (8 -10), a Phase II clinical trial utilizing preaggregated A␤42 as an antigen and the QS-21 saponin as an immunoadjuvant (AN-1792 vaccine) was halted due to the occurrence of meningoencephalitis in ϳ6% of treated patients (11-13). T-cell-mediated autoimmunity produced by the C-terminal portion of A␤ (14) along with a predominantly pro-inflammatory (T helper 1) immunoresponse and cross-reactivity with the presumably physiological monomeric forms of A␤ (A␤38 -43 peptides plus the amyloid-␤ precursor protein have been hypothesized as the main causes of these adverse effects (15-17). Of further concern is the fact that not only plaques but also soluble A␤ oligomers should be bound by a therapeutically effective antibody (18 -20). The latter represent an intermediate conformation prior to fibril formation and are presently considered as the most proximate causative agents of . S...

Influence of Oxidation and Multimerization on the Immunogenicity of a Thioredoxin-L2 Prophylactic Papillomavirus Vaccine

“…One of these epitopes, spanning the amino acid (aa) region 17 to 38 of HPV-16 L2 (L2 ), has gained special attention, as antibodies recognizing this region show neutralizing activity against a broad range of different papillomavirus (PV) types (15)(16)(17). This major cross-neutralizing epitope, which we mapped to the aa 20 to 38 region of L2 (L2 [20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35][36][37][38] ), contains two cysteine residues (positions 22 and 28) that are conserved in the L2 proteins of all known PVs. These cysteine residues are buried and disulfide bonded in mature HPV virions, and it has been suggested that disulfide-bond reduction, after viral entry, may be critical for endosomal escape and infectivity (18).…”

Protein conformational changes induced by adsorption onto material surfaces: an important issue for biomedical applications of material science

“…Remodeling protein conformation therefore exposes different structural determinants that can be recognized by different antibodies. Such antibodies have been used to discriminate between the inactive and active conformations of a protein [72], to neutralize viruses [73] and to differentiate between the aggregated or monomeric form of proteins [74][75][76]. When proteins are adsorbed on material surfaces, some epitopes are masked, which reduces the binding of the corresponding antibodies [77][78][79].…”

Molecules that Target beta‐Amyloid

“…In a similar approach, bacterial thioredoxin (Trx) was used as a scaffold to link four repeats of Aβ [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15] . 114 The anti-Trx (Aβ15) 4 antibody generated against this epitope bound to Aβ 1-42 fibrils and oligomers but not monomers and reduced Aβ pathology in transgenic AD mice.…”