Exploring the Effect of Aliphatic Substituents on Aryl Cyano Amides on Enhancement of Fluorescence upon Binding to Amyloid-β Aggregates

Abstract

The self-assembly of amyloid-β (Aβ) peptides into amyloid aggregates is a pathological hallmark of Alzheimer’s Disease. We previously reported a fluorescent Aryl Cyano Amide (ARCAM) probe that exhibits an increase in fluorescence emission upon binding to Aβ aggregates in solution and in neuronal tissue. Here, we investigate the effect of introducing small aliphatic substituents on the spectroscopic properties of ARCAM both free in solution and when bound to aggregated Aβ. We found that introducing substituents designed to hinder the rotation of bonds between the electron donor and acceptor on these fluorophores can affect the overall brightness of fluorescence emission of the probes in amyloid-free solutions, but the relative fluorescence enhancement of these probes in amyloid-containing solutions is dependent on the location of the substituents on the ARCAM scaffold. We also observed the capability to tune the excitation or emission wavelength of these probes by introducing electron-donating or -withdrawing substituents that putatively affect either the energy required for photoexcitation or the stability of the photoexcited state. These studies reveal new design principles for developing ARCAM-based fluorescent Aβ-binding probes with an enhanced fluorescence signal compared to background and tunable spectroscopic properties, which may lead to improved chemical tools for aiding in the diagnosis of amyloid-associated neurodegenerative diseases.

Excited State Rotational Freedom Impacts Viscosity Sensitivity in Arylcyanoamide Fluorescent Molecular Rotor Dyes

“…Quantum yields for probes 1 – 5 were reported previously . The following method was used for determining the quantum yields for probes 6 and 7 .…”

“…We also developed a series of ARCAM analogues with small, aliphatic substituents near the two rotatable single bonds that we thought most affected the planarity of the molecule (Figure , probes 2 – 5 ) and investigated how these substituents affected their fluorescence enhancement and binding affinity to aggregated forms of amyloid-β (Aβ) peptides, the major component of senile plaques in Alzheimer’s Disease . We observed that introducing substituents in the vinylic position of 1 (as in 2 and 3 ) lowered the overall fluorescence intensity of the free probes in solution but increased their fluorescence enhancement upon binding to Aβ compared to probe 1 .…”