Figure S2 shows CEA691-specific cytotoxic activity of CTL lines isolated from PC patients. A. T cell line from CA11 were measured for their capability to recognise and kill T2 cells coated with 1mM (10-6 M) of CEA691, compared to control peptide, in 4-h CFSE cytotoxicity assay . B. The percentage specific cytotoxicity of CEA 691 loaded T2 cells (1mM peptide) at various E:T ratio, using CTL lines from 3 PC patients (CA07, 11, and 18). C. The cytotoxicity of T cells from CA11 against T2 cells coated with 10-5 to 10-12 M CEA691 peptide and control peptide, at E:T ratio 20:1 (left). The cytotoxicity curve was summarised on the bottom (D).
ARTICLE ABSTRACTPurpose: Carcinoembryonic antigen (CEA) is a candidate target for cellular immunotherapy of pancreatic cancer. In this study, we have characterized the antigen-specific function of autologous cytotoxic T lymphocytes (CTL) specific for the HLA-A2–restricted peptide, pCEA691-699, isolated from the peripheral T-cell repertoire of pancreatic cancer patients and sought to determine if ex vivo PD-L1 and TIM-3 blockade could enhance CTL function.Experimental Design: CD8+ T-cell lines were generated from peripheral blood mononuclear cells of 18 HLA-A2+ patients with pancreatic cancer and from 15 healthy controls. In vitro peptide-specific responses were evaluated by flow cytometry after staining for intracellular cytokine production and carboxy fluorescein succinimydyl ester cytotoxicity assays using pancreatic cancer cell lines as targets.Results: Cytokine-secreting functional CEA691-specific CTL lines were successfully generated from 10 of 18 pancreatic cancer patients, with two CTL lines able to recognize and kill both CEA691 peptide–loaded T2 cells and CEA+ HLA-A2+ pancreatic cancer cell lines. In the presence of ex vivo PD-L1 blockade, functional CEA691-specific CD8+ T-cell responses, including IFNγ secretion and proliferation, were enhanced, and this effect was more pronounced on Ag-specific T cells isolated from tumor draining lymph nodes.Conclusions: These data demonstrate that CEA691-specific CTL can be readily expanded from the self-restricted T-cell repertoire of pancreatic cancer patients and that their function can be enhanced by PD-L1 blockade. Clin Cancer Res; 23(20); 6178–89. ©2017 AACR.