We have developed a novel organotypic (OTC) model of bronchial dysplasia/early lung cancer. In this system, immortalised normal human bronchial epithelial cells are grown at the air-liquid interface on a collagen matrix with embedded fibroblasts. In OTC there is a clear phenotype in cells in which the activation of putative oncogenes at the air-liquid interface recapitulates a histological phenotype consistent with human bronchial dysplasia / early lung cancer.
In vivo, multiple potentially important genotypes may co-exist and influence the cellular phenotypes. We have introduced complexity into our model by combining two genetic lesions, which leads to a more marked phenotype. For this studentship we propose two main scientific objectives:
(I) To perform a medium throughput functional/phenotypic screen of an extended library of small molecule inhibitors in an in vitro model of bronchial dysplasia.
The student will first recapitulate the current OTC experiments and optimise them for a 96- transwell format (rather than current 12 well format) more suitable for screening therapeutic compounds. A 384-compound commercially available library that targets multiple key signal transduction pathways and comprises all classes of known anticancer compounds will be used. The impact of these compounds on the dysplastic phenotype will be interrogated.
(II) To extend the existing in vitro OTC model to cover more complex genotypes and assess the impact of these changes on hits from Aim I. Using existing lentiviral protocols and the recently introduced CRISPR technology further/other genetic lesions will be introduced into the same immortalised cells with the aim of producing the more complex genotypes. Further, the microenvironment will be manipulated. The impact of these refinements on the development of dysplasia and the effect of anticancer compounds will be interrogated.
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