All-Photonic Control of Biorelevant Processes using Molecular Photoswitches and Caged Compounds

Doctoral thesis, 2015

The development of light-controlled molecular tools for in situ regulation of biological activity holds great promise e.g. for resolving dynamic aspects of cellular signal transduction and to overcome inherent pharmaceutic limitations such as poor drug selectivity. In this thesis, photoswitchable molecules from the spiropyran, dithienylethene, and azobenzene families as well as an o-nitrobenzyl-type caged compound were used in a variety of constellations intended for biological use. The presented research ranges from spectroscopic studies on isolated molecular interactions in solution to photoactivated kinase inhibition in live zebrafish. The first part of this thesis focuses on novel approaches for photocontrolled drug release, based on a supramolecular competitive binding methodology. Two host-guest systems were investigated for this purpose; a dithienylehtene – porphyrin dimer, and a spiropyran – cucurbit[7]uril (CB7) system. In the first investigated system, it was found that drug release could be reversibly controlled by light. The spectral properties of the release scaffold also provided a fluorescence-based reporting function. In the second system, an unexpected loss of spiropyran bistability was observed upon complexation to CB7. Also, the undesired hydrolysis of the spiropyran photoswitch was completely halted when bound to the macrocycle. The spiropyran hydrolysis reaction was further examined in a separate study, also included in this thesis. The second and thirds parts focus on photocontrolled small-molecules introduced to living systems. This includes a study on the cytotoxic properties of a DNA-binding spiropyran in live human cancer cells. It was demonstrated how cell-death could be selectively triggered by photoisomerizing the spiropyran to the merocyanine form inside the cells. Part three focuses on photocontrolled inhibition of the, for cancer highly relevant, REarranged during Transfection (RET) kinase. An azobenzenederived photoswitch and an o-benzyl-type caged inhibitor based on the pyrazolopyrimidine scaffold were developed and evaluated. Both compounds exhibited light-controlled RET inhibition in an isolated enzyme- and live cell assay. The caged compound was also used to photonically activate inhibition of RET in live zebrafish, confirming uptake and decaging of the compound inside the live organism.