Functional Characteristics and Molecular Mechanism of a New scFv Antibody Against Aβ42 Oligomers and Immature Protofibrils

Abstract

Amyloid β peptide (Aβ42) is a major determinant of Alzheimer's disease (AD). In this study, we studied a novel single-chain variable fragment (scFv), AS, generated from an antibody library of AD patients, which recognized and bound specifically to medium-size amyloid β peptide (Aβ42) oligomers and immature protofibrils (25-55 kDa) and, more importantly, reduced their level by blocking their formation or inducing their disassembly. Consequently, scFv AS ameliorated or prevented their cytotoxicity and protected SH-SY5Y cells and primary cultured neurons in vitro from their damage in a concentration-dependent manner. Comparison of its cytotoxicity-inhibiting and cytotoxicity-neutralizing activities indicated that scFv AS displayed its protective effect on target cells mainly due to its cytotoxicity-inhibitory activity though it could also neutralize the cytotoxicity. We also found that scFv AS could efficiently cross the in vitro BBB model with a delivery efficiency of over 70% after a 60-min post-administration. The scFv AS was a monovalent antibody with an affinity constant (KD) of 5.5 × 10(-6) M and a binding threshold of 6.25 × 10(-4) μM for Aβ42 oligomers. The molecular docking simulations of Aβ42 to scFv AS revealed that scFv AS tends to approached Aβ42 oligomers and immature protofibrils mainly by their hydrophobic interaction and then drew Aβ42 molecule into the gap between VL and VH domains of scFv AS by hydrophilic interaction between scFv AS and the N-terminal region (residues 1-15) of Aβ42 and the hydrophobic interactions between scFv AS and the middle region (residues 20-33) of Aβ42. The combination of scFv AS with Aβ42 was realized likely through an induced-fit process.

Application of Antibody Fragments Against Aβ With Emphasis on Combined Application With Nanoparticles in Alzheimer’s Disease

“…In this way, Aβ cytotoxicity was inhibited. ScFv AS could also transport across the BBB model in vitro (Zhang et al, 2015b). What is more, scFv HT6 was able to combine with medium-sized Aβ42 aggregation and lead to disaggregation of large Aβ42 aggregation into small Aβ42 oligomers .…”

“…And scFv MO6 could cross the BBB model in vitro (Zhang et al, 2015a). Moreover, scFv AS was identified and could combine with immature protofibrils as well as medium-size Aβ oligomers (Zhang et al, 2015b). ScFv AS could decrease levels of Aβ oligomers through preventing their formation or leading to their disaggregation (Zhang et al, 2015b).…”

Molecular interactions between monoclonal oligomer-specific antibody 5E3 and its amyloid beta cognates

“…The inspection of the top hundred docked complexes of Fv5E3 with the dodecamer by Gallion, and the cross-β sub-unit by Lührs et al suggests that the backbone of the GSNKG turn by itself can be an epitope for Fv5E3, as the K28 residues are not solvent-exposed in the chain interacting with Fv5E3. The unexpected hydrophobic interactions of Fv5E3 and the AβOs are reminiscent of the KW1 antibody fragment [72] or ScFv AS [73] mechanism for the detection of AβOs (S5 and S7 Tables of SI).…”

A human scFv antibody that targets and neutralizes high molecular weight pathogenic amyloid‐β oligomers

“…2008, Meli et al . 2009), AβOs and protofibrils (Zhang et al . 2015), and AβO subpopulations (Kasturirangan et al .…”

“…Since then, many groups have developed anti-Ab scFvs (Liu et al 2004;Solorzano-Vargas et al 2008;Medecigo et al 2010;Wang et al 2016). Conformationspecific anti-Ab scFvs in the literature include those specific for AbOs (Zameer et al 2008;Meli et al 2009;Wang et al 2009), AbOs and protofibrils (Zhang et al 2015), and AbO subpopulations (Kasturirangan et al 2012(Kasturirangan et al , 2013.…”