Azobenzenes for the development of therapeutics

Abstract

Azobenzenes are photochromic molecules, commonly utilised for their ability to undergo light-mediated isomerisation. This unique property enables them to modulate biological systems upon their incorporation. Chapter 1 explores the utilisation of azobenzenes for peptide stapling, that has gained notable interest for therapeutic applications. However, many reported methodologies rely on the use of tissue-damaging UV light. Recent approaches that minimise the dependence on UV light have been successful at adjusting peptide conformation but fell short in achieving a significant modulation of the biological activity between the peptide isomer forms. Our work describes the development of novel azobenzene systems primed for peptide stapling via cysteine-SNAr. Specifically, a tetra-ortho-chloro azobenzene motif that can undergo bi-directional visible- light mediated isomerisation. As a proof-of-concept, we employed this newly developed peptide stapling methodology to target the disruption of p53/MDM2 protein-protein interaction. Through competitive fluorescence polarisation, a substantial discrepancy in binding affinity was observed between each isomer for the p53-interacting domain of human MDM2. Chapter 2 explores the applicability of azobenzenes for the development of antibody-drug conjugates (ADCs), a rapidly growing class of targeted cancer therapeutics. Previous reports have shown that ortho-methoxy azobenzenes undergo reductive cleavage in reducing environments, making them ideal candidates for the development of novel GSH-cleavable ADCs, responsive to elevated GSH levels present in the tumour microenvironment. Currently, such ADCs exclusively rely on reducible disulfides, that often suffer from instability in circulation. Our initial work describes the development of a model azobenzene-fluorophore system that readily undergoes selective release of the probe at elevated GSH levels but remains stable at low GSH levels and absence of GSH. Subsequent incorporation of the azobenzene motif, bearing a cytotoxic payload, into an anti-HER2 trastuzumab antibody afforded a novel GSH-cleavable ADC that demonstrated selectivity towards HER2-positive cell lines, with a wide therapeutic window.