Design of Small Fluorescent Probes for Imaging Alzheimer’s Disease Pathology

Abstract Background Despite enormous efforts, early diagnosis of Alzheimer’s disease (AD) remains difficult. To date, there is no reliable diagnostic tool to identify biomarkers in the developing stages of AD [1]. In recent years, our group has focused part of the research on the development of small fluorescent molecules capable of effectively labelling AD biomarkers such as amyloid β plaques and tau protein [2, 3]. Methods We designed and synthesised a library of small fluorescent molecules that were tested in vitro for their binding affinity ( K d ) and change in optical properties upon binding to amyloid β fibrils. In addition, the most promising molecules were used to perform ex vivo labelling of post‐mortem AD brain tissue examined by fluorescence microscopy. Results Our synthesized fluorescent probes consist of three building blocks, i.e., electron‐donating (EDG) and electron‐withdrawing groups (EWG), linked to different unsaturated hydrocarbon (HC) linkers (Figure 1), which enable the so‐called “push‐pull” effect leading to fluorescence. First, we investigated how the individual building blocks affect the spectroscopic properties (absorption, emission, excitation, quantum yield) of the prepared molecules in different solvents, representing a simplified polarity model of the AD biomarker binding sites (Figure 2). Furthermore, the compounds evaluated in vitro bound to amyloid β 1‐42 fibrils with submicromolar K d values. Moreover, the selected compounds were shown to successfully stain amyloid β plaques as well as tau proteins on AD brain slices (Figure 2). Conclusion Based on the results we conducted structure‐spectroscopic‐affinity relationship study and determined how the building blocks of our fluorescent probes affect the optical as well as affinity properties when bound to amyloid β fibrils. Through in vitro and ex vivo studies, our probes are being systematically tested to identify the best candidate capable of achieving our final goal – the early detection of AD biomarkers in the patient’s blood, which would lead to a generally applicable, minimally invasive, cost‐effective, and long‐term traceable diagnostic assay. 1. Hansson, O., et al., Alzheimers. Dement., 2022 , 18 , 2669‐2686. 2. Šarlah, D., et al., Molecules, 2016 , 21 , 267. 3. Rejc, L., et al., J. Med. Chem., 2017 , 60 ,8741‐8757.