Understanding the DNA damage driven immune response in oesophageal adenocarcinoma

: opportunities for improved stratification and treatment

Abstract

Background: We previously identified a good-prognosis DNA repair deficient oesophageal adenocarcinoma (OAC) subgroup with improved response to neoadjuvant chemotherapy using a gene expression signature, the DNA Damage Immune Response (DDIR) assay. DDIR positive tumours were characterised by increased CD8+ T lymphocyte infiltration and PD-L1 expression. In this study, we examined subgroups within DDIR positive and DDIR negative OAC and investigated whether mutated genes associated with the DDIR positive subgroup could be potential therapeutic targets to improve response to treatment in the DDIR negative subgroup.

Methods: Bioinformatic analysis of three clinically annotated OAC gene expression datasets was undertaken to further characterise DDIR positive disease. Multi-omics data from The Cancer Genome Atlas was used to identify mutations associated with DDIR positive disease. To investigate the effect of knockdown of these mutated genes in DDIR negative disease, we performed an siRNA knockdown screen in ESO26 OAC cells +/-doxorubicin treatment, followed by RNA sequencing.

Results: Three OAC gene expression datasets were analysed: (1) Turkington (microarray); (2) TCGA-ESCA (RNA-seq); (3) OCCAMS (RNA-seq). Four distinct clusters were observed in each dataset: (1) DDIR-high (upregulation of immune genes and responsiveness to PD-L1 inhibitors); (2) DDIR mid/high; (3) DDIR mid/low; (4) DDIR low (treatment resistance). Mutations in 18 genes were significantly associated with DDIR positivity. In the siRNA knockdown screen in ESO26 cells, DDIR activation was observed with doxorubicin treatment and was potentiated by target gene knockdown.

Conclusions: We have identified four distinct OAC subgroups, consistent across three gene expression datasets. We have comprehensively examined activation of DDIR-related genes in response to DNA damaging chemotherapy and showed that this is cGAS-STING dependent in ESO26 cells. Activation of DDIR-related genes following doxorubicin treatment was potentiated by target gene knockdown indicating potential strategies to enhance the effectiveness of immunotherapy in OAC.

Thesis is embargoed until 31 July 2028.

Date of Award	Jul 2023
Original language	English
Awarding Institution	Queen's University Belfast
Sponsors	Wellcome Trust and the Health Research Board, Health Service Executive National Doctors Training annd Planning & Health and Scoial Care Research and Development Division, Northern Ireland
Supervisor	Kienan Savage (Supervisor), Richard Turkington (Supervisor) & Jaine Blayney (Supervisor)