Investigating the tumour immune microenvironment as a determinant of antigen presentation and immune response in oesophageal cancer

Abstract

Oesophageal cancer (OC), the 12^th most common cancer (UK), presents with high clinical need and poor survival. OC is histologically divided into adenocarcinoma (OAC) and squamous cell carcinoma. Antigen presentation machinery (APM) is crucial for eliciting anti-tumoral immune responses by cancer-antigen recognition. However, the genomic landscape of APM genes and dysregulation of expression in OAC is unknown. A bioinformatic approach has been employed to elucidate the landscape of genomic defects in APM genes using large multi-omics datasets (TCGA and OCCAMS). Digital cytometry methods were applied to determine the impact of APM expression on immune composition within the tumour immune microenvironment (TiME). Additionally, immunohistochemistry (IHC) of a large tissue microarray series for candidate APM genes and immune cell markers (HLA-A/B/C/E/Class 2, CD3, CD4, CD8, Foxp3) was performed with digital histopathology analysis to validate findings. Lastly, cell modelling and single-cell RNA sequencing were implemented to determine the role of CSDE1 in regulating the expression of MHC class I genes in OAC cell lines. Genomic landscape analysis found APM mutation incidence to be infrequent and copynumber segments complex over the HLA locus of chromosome 6; these results did not inform survival analysis. However, the expression of APM genes showed a significant impact on overall survival (OS), with the expression of 12 out of 18 MHC class I, 12/20 MHC class II and 4/7 APM regulators were associated to altered OS. Following this analysis, the impact of APM gene expression on the TiME identified 10/20 MHC class I, 7/20 MHC class II and 3/7 APM gene expression regulators associated with altered immune composition. Using these approaches together, identified 17/45 assessed APM genes possessed both an association with survival outcomes and altered immune composition, including CSDE1, HLA-E, ERAP2, CD74, HLA-DRB1 and HLA-DRB5, among others. IHC demonstrated that the low expression of HLA-E and HLA class II and the high expression of TAP1 and CSDE1 corresponded to shorter OS. T cell density alone did not associate with OS. Yet the high expression of HLA-A/B/C combined with the low abundance of CD8+ T cells resulted in shorter OS compared to high HLA-A/B/C protein expression and greater CD8+ T cell density. CD3/8+ T-cell density correlated with greater HLA-ABC and HLA class 2 protein expression and a correspondingly increased T-cell infiltrate. Knockdown of CSDE1 expression produced greater JAK/STAT1 signalling and increased expression of HLA-A/B/C transcripts, suggesting a significant negative regulator role in the expression of MHC class I HLAs. CSDE1 was upregulated in single cancer cells compared to healthy comparative cells, with evidence suggesting overexpression may be acquired in premalignancy over increasing dysplasia (i.e., Barrett’s oesophagus). This work has translational value for OAC patients. Firstly, APM expression presents a potential marker of altered survival and immune composition, which has biomarker utility for immunotherapy implementation. Secondly, CSDE1 may form an actionable pathway to improve immune responses as a potential future target for vaccine therapies.

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Identifiers

ORCID for William Pratt: orcid.org/0000-0002-3214-6552

ORCID for Matthew Rose-Zerilli: orcid.org/0000-0002-1064-5350

ORCID for Tim Underwood: orcid.org/0000-0001-9455-2188

ORCID for Tim Elliott: orcid.org/0000-0003-1097-0222

ORCID for Paul Skipp: orcid.org/0000-0002-2995-2959

ORCID for Jacek Brodzki: orcid.org/0000-0002-4524-1081

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