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Title: Improving outcomes for resistant rectal cancer
Author: O'Cathail, Sean Micheal
ISNI:       0000 0004 8506 9516
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2019
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The management of locally advanced rectal cancer involves the use of neoadjuvant chemoradiotherapy but up to 20% of patients have no discernible response. Understanding the factors that mediate resistance to treatment is therefore an important clinical research question. The work in this thesis aimed to further understand potential biological contributors to therapeutic resistance by exploring the contribution of the KEAP1-NRF2 pathway in-vitro and evaluating it as a potential biomarker in-vivo. Using RNAi and clonogenic assays, NRF2 knockdown produced a radiosensitive phenotype in intrinsically resistant cell line models but had no effect in the most radiosensitive cell line. An attempt at CRIPSR isogenic validation of the RNAi was also made. Using a CRISPR isogenic activating mutation, radioresistance was induced in this previously sensitive cell line. Overexpression of NRF2 also induced chemoresistance to the commonly used radiosensitizers in clinical practice. RNA sequencing was carried out on both NRF2 downregulated (radiosensitive) and NRF2 upregulated (radioresistant) cell line models to better understand the mechanism of radioresistance. NRF2 downregulation produces a marked reduction in amino acid metabolism, particularly glutamine/asparagine, relative to control conditions. Overexpression of NRF2 produced the opposite effect. The data suggest an NRF2 regulated 'switch' of metabolism in favour of an anabolic, proliferative phenotype which produces radioresistance. To understand the clinical relevance of NRF2 signalling a bioinformatic approach was used. A metagene signature of 36 NRF2 regulated genes to aggregate pathway expression was derived and trained using publically available datasets. The resulting transcriptomic biomarker demonstrated that high NRF2 expression was associated with worse survival outcomes in 1,360 validation patients from public resources and the S:CORT collaboration. Interestingly NRF2 expression was significantly enriched in Consensus Molecular Subtype 4, suggesting a further refinement of the molecular transcriptomic classification of colorectal cancer. Lastly higher NRF2 expression was shown to be associated with a worse response to neoadjuvant chemotherapy, as measured by the neoadjuvant rectal score, in a curated cohort of 127 rectal cancers who received standard of care treatment. This validates the in-vitro cell line findings and supports an important role for NRF2 signalling in prognosis and therapeutic resistance in colorectal cancer.
Supervisor: Maughan, Tim ; Hawkins, Maria ; Lewis, Annie Sponsor: Cancer Research UK
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Oncology