Thiophene bridged aldehydes (TBAs) image ALDH activity in cells via modulation of intramolecular charge transfer

Aldehyde dehydrogenases (ALDHs) catalyze the oxidation of an aldehyde to a carboxylic acid and are implicated in the etiology of numerous diseases. However, despite their importance, imaging ALDH activity in cells is challenging due to a lack of fluorescent imaging probes. In this report, we present a new family of fluorescent probes composed of an oligothiophene flanked by an aldehyde and an electron donor, termed thiophene-bridged aldehydes (TBAs), which can image ALDH activity in cells. The TBAs image ALDH activity via a fluorescence sensing mechanism based on the modulation of intramolecular charge transfer (ICT) and this enables the TBAs and their ALDH-mediated oxidized products, thiophene-bridged carboxylates (TBCs), to have distinguishable fluorescence spectra. Herein, we show that the TBAs can image ALDH activity in cells via fluorescence microscopy, flow cytometry, and in a plate reader. Using TBA we were able to develop a cell-based high throughput assay for ALDH inhibitors, for the first time, and screened a large, 1460-entry electrophile library against A549 cells. We identified α,β-substituted acrylamides as potent electrophile fragments that can inhibit ALDH activity in cells. These inhibitors sensitized drug-resistant glioblastoma cells to the FDA approved anti-cancer drug, temozolomide. The TBAs have the potential to make the analysis of ALDH activity in cells routinely possible given their ability to spectrally distinguish between an aldehyde and a carboxylic acid.