Targeting Isoaspartate-Modified A Beta Rescues Behavioral Deficits In Transgenic Mice With Alzheimer'S Disease-Like Pathology

Background Amyloid beta (A beta)-directed immunotherapy has shown promising results in preclinical and early clinical Alzheimer's disease (AD) trials, but successful translation to late clinics has failed so far. Compelling evidence suggests that post-translationally modified A beta peptides might play a decisive role in onset and progression of AD and first clinical trials targeting such A beta variants have been initiated. Modified A beta represents a small fraction of deposited material in plaques compared to pan-A beta epitopes, opening up pathways for tailored approaches of immunotherapy. Here, we generated the first monoclonal antibodies that recognize l-isoaspartate-modified A beta (isoD7-A beta) and tested a lead antibody molecule in 5xFAD mice. Methods This work comprises a combination of chemical and biochemical techniques as well as behavioral analyses. A beta peptides, containing l-isoaspartate at position 7, were chemically synthesized and used for immunization of mice and antibody screening methods. Biochemical methods included anti-isoD7-A beta monoclonal antibody characterization by surface plasmon resonance, immunohistochemical staining of human and transgenic mouse brain, and the development and application of isoD7-A beta ELISA as well as different non-modified A beta ELISA. For antibody treatment studies, 12 mg/kg anti-isoD7-A beta antibody K11_IgG2a was applied intraperitoneally to 5xFAD mice for 38 weeks. Treatment controls implemented were IgG2a isotype as negative and 3D6_IgG2a, the parent molecule of bapineuzumab, as positive control antibodies. Behavioral studies included elevated plus maze, pole test, and Morris water maze. Results Our advanced antibody K11 showed a K-D in the low nM range and > 400fold selectivity for isoD7-A beta compared to other A beta variants. By using this antibody, we demonstrated that formation of isoD7-A beta may occur after formation of aggregates; hence, the presence of the isoD7-modification differentiates aged A beta from newly formed peptides. Importantly, we also show that the Tottori mutation responsible for early-onset AD in a Japanese pedigree is characterized by massively accelerated formation of isoD7-A beta in cell culture. The presence of isoD7-A beta was verified by K11 in post mortem human cortex and 5xFAD mouse brain tissue. Passive immunization of 5xFAD mice resulted in a significant reduction of isoD7-A beta and total A beta in brain. Amelioration of cognitive impairment was demonstrated by Morris water maze, elevated plus maze, pole, and contextual fear conditioning tests. Interestingly, despite the lower abundance of the isoD7-A beta epitope, the application of anti-isoD7-A beta antibodies showed comparable treatment efficacy in terms of reduction of brain amyloid and spatial learning but did not result in an increase of plasma A beta concentration as observed with 3D6 treatment. Conclusions The present study demonstrates, for the first time, that the antibody-mediated targeting of isoD7-modified A beta peptides leads to attenuation of AD-like amyloid pathology. In conjunction with previously published data on antibodies directed against pGlu-modified A beta, the results highlight the crucial role of modified A beta peptides in AD pathophysiology. Hence, the results also underscore the therapeutic potential of targeting modified amyloid species for defining tailored approaches in AD therapy.