Does inappropriate DNA replication provide a mechanism for acquisition of drug resistance?

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This project will use the acquisition of resistance to the AZ drug Osimertinib, the best-in-class EGFR inhibitor, in PC9 cells as a model system. i) We will first apply scRNAseq during Osimertinib treatment to characterise gene expression during treatment. A single cell approach will reveal the status of individual replicating cells, which is likely to be heterogeneous as cells do not react uniformly under extended treatment.ii) We have developed a state-of-the-art method for detecting replication stress and DNA damage, TrAEL-seq, which profiles DNA replication patterns with high sensitivity. TrAEL-seq will be used to replicating cells to understand replication stress caused by drug treatment.iii) Replication stress can give rise to extrachromosomal circular DNA (ecDNA), which frequently harbours gene amplifications that bestow drug resistance. It would be particularly exciting to connect drug-assocatied replication stress in DTPs directly to formation of ecDNA, which we will test using circular DNA sequencing.