Development of SmartProbes for the optical imaging of pulmonary inflammation

The prospect of an optical biopsy to interrogate disease within the human lung offers the ability to improve the outcomes of clinical trials, provide early validation within drug discovery programmes, while also offering diagnostic capabilities. Optical molecular imaging using smart fluorescent probes offers a safe and affordable technique to allow the analysis of multiple lung pathologies and diseases such as fibrosis and chronic obstructive pulmonary disease (COPD). The aim of this project was to develop a number of families of SmartProbes and validate their biological utility. Macrophages are a valuable imaging target due to their key role in maintaining homeostasis and responding to inflammatory stimuli in the lung. Imaging these cells will allow us to dynamically observe macrophage biology and the multitude of host cell interactions within the lung. SmartProbes were targeted to macrophages via the cell specific markers - the macrophage mannose and folate receptors. The synthesis of SmartProbes targeting the folate receptor, including the application of new solid phase methodology to simplify the synthesis of these probes, is reported. In vitro studies were carried out in cancer cells and primary human monocyte-derived macrophages. An anti-MMR nanobody that targets the mannose receptor was fluorescently labelled to give a near-infrared SmartProbe. Its specificity and selectivity was assessed with primary immune cells and ex vivo clinical lung samples. The epithelial barrier is a layer of cells that line the lung and undergoes remodelling during diseases such as COPD and idiopathic pulmonary fibrosis (IPF). Hence, methods to visualise epithelial integrity were sought. SmartProbes that were pro fluorophores for beta-galactosidase were synthesised. Beta-galactosidase probes were applied in epithelial cell models and murine tissue, allowing the detection of senescence associated beta-galactosidase (SA-βGal) positive senescent cells in 3D tissue samples