Shifting Gears Towards the Red: Novel Boron-Based Fluorophores for Bioimaging Applications

Abstract

Advancements in near-infrared (NIR) fluorescence imaging have enabled greater tissue penetration depths, high spatial resolution, reduced photon scattering, and minimal interference from tissue autofluorescence. Hence, NIR fluorophores exist as viable candidates for biological imaging applications, as well as providing unique insights into complex biological processes to better understand disease etiology. In this work, a series of novel boron-based coumarin and rhodamine-like fluorophores were developed and tested. Initially, modifications to the coumarin scaffold were investigated to develop more red-shifted dyes, whereby incorporation of a p-conjugated bridge was determined to be a critical component. Confocal microscopy studies with A549 lung cancer cells showed clear differences in the intra-cellular distributions of the fluorophores. The lipophilic carborane coumarin derivatives exhibited superior selectively within lipid droplets. In contrast, the polar boronic acid hydrazone-coumarins displayed intracellular localisation within the endoplasmic reticulum. A library of boron-containing rhodamine-like probes were also synthesised. All compounds exhibited near-infrared emission wavelengths with large Stokes shifts. Furthermore, modifications to the terminal boron moiety were not found to impact the overall red-shift of the molecules, although increasing the donor group strength favourably enhanced this shift. The low brightness of some of the probes, related to rhodamine spirocyclisation, meant that conclusive intracellular localisations could not be confirmed. The near-IR emitting nature of the rhodamine-like probes was recognised as a highly advantageous tool for bioimaging applications. Two fluorescent-labelled ligands with varying linker lengths, of the allosteric adamantyl benzamide P2X7R antagonist were prepared, based on the para-MIDA ester rhodamine-like fluorophore. Both probes were utilised in preliminary fluorescence studies, whereby the near-IR emitting nature of the fluorophore was retained despite conjugation to biomolecules. As well, a longer emission wavelength was observed with a shorter linker length. This research ultimately highlights the versatility of boron as a unique element in fluorescence and biological imaging applications.