Fluorescence\nDetection and Inhibition Mechanisms of\nDNTPH on Aβ42 Oligomers Characterized as Products in the Four\nStages of Aggregation

Aβ42 aggregation was implicated in the pathogenesis\nof Alzheimer’s\ndisease (AD) without effective treatment available currently. Future\nefforts in clinical trials should instead focus on applying those\nantiamyloid treatment strategies to the preclinical stage and “the\nearlier, the better”. How to identify and inhibit Aβ42\noligomers in the different stages of aggregation is therefore becoming\nthe key to controlling primary aggregation and consequent AD development.\nAggregation-induced emission probe DNTPH was demonstrated recently,\nenabling detection of amyloid at wavelengths up to 710 nm and exhibiting\nstrong inhibitory effects on Aβ fibrosis at low dose. However,\nthe detection and inhibition mechanisms of Aβ oligomers at various\nearly stages of aggregation remain unknown. To this end, we built\nfour different morphologies of Aβ42 pentamers characterized\nby products in monomeric aggregate (P<sub>M</sub>), primary nucleation\n(P<sub>P</sub>), secondary nucleation (P<sub>S</sub>), and fibril\nstages (P<sub>F</sub>) to explore the distinguishable ability and\ninhibition mechanisms of DNTPH with different concentrations upon\nbinding. The results showcased that DNTPH does detect the four different\nAβ42 oligomers with conspicuous fluorescence (λ<sub>P<sub>M</sub></sub> = 657 nm, λ<sub>P<sub>P</sub></sub> = 639 nm,\nλ<sub>P<sub>S</sub></sub> = 630 nm, and λ<sub>P<sub>F</sub></sub> = 648 nm) but fails to distinguish them, indicating that additional\nimprovements are required further for the probe to achieve it. The\ninhibition mechanisms of DNTPH on the four Aβ42 aggregation\nare however of amazing differences. For P<sub>M</sub> and P<sub>P</sub>, aggregation was inhibited by altering the secondary structural\ncomposition, i.e., by decreasing the β-sheet and toxic turn\n(residues 22–23) probabilities, respectively. For P<sub>S</sub>, inhibition was achieved by segregating and keeping the two disordered\nmonomeric species (P<sub>SM</sub>) away from the ordered secondary\nseed species (P<sub>SF</sub>) and consequently blocking further growth\nof the P<sub>SF</sub> seed. The inhibition mechanism for P<sub>S</sub> is first probed and proposed so far, as far as we know, and the\ncorresponding aggregation stage of P<sub>S</sub> is the most important\none among the four stages. The inhibition of P<sub>F</sub> was triggered\nby distorting the fibril chains, disrupting the ordered fibril surface\nfor the contact of monomers. In addition, the optimal inhibitory concentrations\nof DNTPH for P<sub>M</sub>, P<sub>P</sub>, and P<sub>F</sub> were\ndetermined to be 1:3, while for P<sub>S</sub>, it was 1:5. This outcome\noffers a novel perspective for designing drugs targeting Aβ42\noligomers at different aggregation stages.