ß-amyloid monomer scavenging by an anticalin protein prevents neuronal hyperactivity in mouse models of Alzheimer's Disease.

Hyperactivity mediated by synaptotoxic ß-amyloid (Aß) oligomers is one of the earliest forms of neuronal dysfunction in Alzheimer's disease. In the search for a preventive treatment strategy, we tested the effect of scavenging Aß peptides before Aß plaque formation. Using in vivo two-photon calcium imaging and SF-iGluSnFR-based glutamate imaging in hippocampal slices, we demonstrate that an Aß binding anticalin protein (Aß-anticalin) can suppress early neuronal hyperactivity and synaptic glutamate accumulation in the APP23xPS45 mouse model of ß-amyloidosis. Our results suggest that the sole targeting of Aß monomers is sufficient for the hyperactivity-suppressing effect of the Aß-anticalin at early disease stages. Biochemical and neurophysiological analyses indicate that the Aß-anticalin-dependent depletion of naturally secreted Aß monomers interrupts their aggregation to neurotoxic oligomers and, thereby, reverses early neuronal and synaptic dysfunctions. Thus, our results suggest that Aß monomer scavenging plays a key role in the repair of neuronal function at early stages of AD.

Structural basis of Alzheimer ß-amyloid peptide recognition by engineered lipocalin proteins with aggregation-blocking activity.

We describe the structural analysis of two Anticalin® proteins that tightly bind Aß40, a peptide involved in the pathophysiology of Alzheimer's disease. These anticalins, US7 and H1GA, were engineered on the basis of the human lipocalin 2, thus yielding compact single-domain binding proteins as an alternative to antibodies. Albeit selected under different conditions and mutually deviating in 13 amino acid positions within the binding pocket (of 17 mutated residues in total), both crystallised anticalins recognize the same epitope in the middle of the ß-amyloid peptide. In the two complexes with the Aß40 peptide, its central part comprising residues LysP16 to LysP28 shows well defined electron density whereas the flanking regions appear structurally disordered. The compact zigzag-bend conformation which is seen in both structures may indicate a role during conversion of the soluble monomeric form into pathogenic Aß state(s) and, thus, explain the aggregation-inhibiting effect of the anticalins. In contrast to solanezumab, which targets the same Aß region in a different conformation, the anticalin H1GA does not show cross-reactivity with sequence-related human plasma proteins. Consequently, anticalins offer promising reagents to prevent oligomerization of Aß peptides to neurotoxic species in vivo and their small size may enable new routes for brain delivery.