Characterising tissue-resident memory CD8+ T cells in Malignant Pleural Effusion

Thesis

Malignant pleural effusion (MPE) is the accumulation of fluid in the pleural space due to cancer. MPE is rich in immune infiltrate, including T cells and NK cells. T cells in MPE are topographically sequestered away from the chronic antigen stimulation of the tumour core. Hence, MPE-derived tumour-reactive lymphocytes may have better preserved effector function. In this study, the immunophenotype of CD56^brightCD3-CD16^- NK cells, CD4⁺ and CD8⁺ T cells were comprehensively characterised using flow cytometry from ex vivo MPE (n = 62). MPE-derived CD8+ T cells were found to be enriched for CD103⁺ T_RM-like cells. TRM-like CD103⁺CD8⁺ T cells derived from MPE displayed an intermediate expression of single and combined immune checkpoint receptors PD1, TIM3, TIGIT and NKG2a between CD103⁺CD8⁺ T cells derived from PBMC and pleural-biopsy. Tissue resident CD56^brightCD3-CD16- NK cells were also enriched in MPE and were found to abundantly express NKG2a. Conversely, MPE-derived CD4⁺ T cells were found to have phenotypes indicative of being derived from peripheral blood. The total CD8⁺ T cell and CD103⁺CD8⁺ T cell compartments in MPE were also comprehensively characterised at the transcriptomic level. In-depth single-cell RNAseq analysis of CD8⁺ T cells revealed the T_RM phenotype is defined by ZNF683 (Hobit). Single-cell RNAseq of T_RM CD103⁺CD8⁺ T cells revealed cells are diverse, clonally expanded and constituted by new cell states that are “Unique to the pleura”, which may have favourable characteristics for future cell therapy. Furthermore, gene signatures identified in “Early pleura” cells and “Pleural” cells stratified TCGA datasets of mesothelioma and lung adenocarcinoma patients for better overall survival probability. Tumour-reactive CD8⁺ T cells were isolated from ex vivo MPE using a novel isolation method and were shown to secrete cytokines (IFNγ and TNFα) and have robust in vitro killing capacity against autologous primary cancer cell lines. This work provides, for the first time, an in-depth characterisation of cytotoxic tissue-resident lymphocytes in MPE and develops the current understanding of MPE immunity by revealing MPE is a valuable source of tumour-reactive T cells for cell therapy.

Authors: Dominey-Foy, DCC

• DOI: 10.5287/ora-zvjyn5j50
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: Tumour-reactive CD8+ T cells; Anti-tumour immunity; Cell therapy
• Subjects: T cell biology; Malignant pleural effusion; Immunology