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Title: Assessing the activation of epidermal growth factor receptors by automated fluorescence lifetime imaging microscopy (FLIM) in relation to cancer treatments
Author: Kong, H. A.
Awarding Body: University of London
Current Institution: University College London (University of London)
Date of Award: 2008
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The decision to treat cancer patients with Iressa (Gefitinib, tyrosine kinase inhibitor of EGFR) and Herceptin (Trastuzumab, monoclonal antibody for HER2) is frequently based on EGFR or HER2 receptor over-expression. However, even in these selective groups of patients, the response rate is poor and unpredictable. The underlying mechanisms contributing to drug resistance, as well as predicting the success of these drugs in cancer patients are still poorly understood. One of the reasons for poor response rates based on HER (ErbB) levels is that over-expression 'per se' fails to consider receptor activation for example through autocrine expression of one of its several ligands. Over-expression as a criterion for treatment fails to account for patients having receptor activation without up-regulation. It was hypothesized that using Forster Resonance Energy Transfer (FRET) to measure the functional status of EGFR in tumour arrays should provide more quantitative prognostic information than using immunohistochemistry alone. After validating a high throughput fluorescence lifetime microscopy (FLIM) system in a series of cell lines, it was shown that EGFR phosphorylation, reflected in high FRET efficiency, is correlated with worsening Disease Free Survival (DFS) in a set of head and neck tumour arrays. The FRET methodology was applied further to assess the phosphorylation of HER2 and other HER family receptors in various breast cancer cell lines in relation to Iressa and Herceptin treatments. Monotherapy with Iressa while targeting EGFR and decreasing phosphorylation of HER3, induced proteolytic cleavage of HER4 and dimerisation between HER2 and HER4, leading to HER2 phosphorylation, as a result of ligand release. Therefore the activation of alternative pathways like HER2 and HER4 may mediate resistance to Iressa. It was also demonstrated that Herceptin while targeting HER2, paradoxically induced the phosphorylation of all HER receptors due to antagonist-induced ligand secretion. Therefore, it has been shown that Iressa and Herceptin treatment in breast cancer cells induces activation of alternative HER pathways, thus providing an insight into possible mechanisms of resistance for targeted therapies in breast cancer. The results suggest alternative treatments to overcome resistance to these targeted therapies in patients. FRET was also applied to assess HER2 phosphorylation in a set of HER2 positive breast tumour arrays using automated FLIM. It was shown that FRET maybe used to stratify HER2 positive breast cancer patients into different prognostic groups. It is proposed to utilise this assay for prospective stratification of patients in randomised trials of EGFR and HER2 inhibition. The methodology shows great promise and can also be applied to assess the activation of other signalling pathways (e.g. PKB and MAPK) in relation to various cancer treatments.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available