High Content Phenotypic Profiling in Oesophageal Adenocarcinoma Identifies Selectively Active Pharmacological Classes of Drugs for Repurposing and Chemical Starting Points for Novel Drug Discovery

Oesophageal adenocarcinoma (OAC) is a highly heterogeneous disease, dominated by large-scale genomic rearrangements and copy number alterations. Such characteristics have hampered conventional target-directed drug discovery and personalized medicine strategies contributing to poor outcomes for patients. We describe the application of a high-content Cell Painting assay to profile the phenotypic response of 19,555 compounds across a panel of six OAC cell lines and two tissuematched control lines. We built an automated high-content image analysis pipeline to identify compounds that selectively modified the phenotype of OAC cell lines. We further trained a machine-learning model to predict the mechanism-of-action of OAC selective compounds using phenotypic fingerprints from a library of reference compounds.
We identified a number of phenotypic clusters enriched with similar pharmacological classes e.g. Methotrexate and three other antimetabolites which are highly selective for OAC cell lines. We further identify a small number of hits from our diverse chemical library which show potent and selective activity for OAC cell lines and which do not cluster with the reference library of compounds, indicating they may be selectively targeting novel oesophageal cancer biology. Overall our results demonstrate that our OAC phenotypic screening platform can identify existing pharmacological classes and novel compounds with selective activity for OAC cell phenotypes.