T lymphocytes with chimeric receptor induce carcinoembryonic antigen-positive specific gastric carcinoma cells apoptosis

Abstract

To develop a method to generate T lymphocytes that identify tumor specific carcinoembryonic antigen (CEA), and induce anti-tumor response such as apoptosis. Peripheral blood samples were collected from 10 healthy persons and peripheral blood mononuclear cells (PBMCs) were isolated. 1, then CD8(+) T cells were isolated from the PBMCs by magnetic activated cell sorting (MACS) using magnetic beads-conjugated anti-CD8 monoclonal antibodies. Then the recombinant vector anti-CEA-scFv-CD3zeta-pcDNA3.0 was transfected into the CD8(+) T cells by lipofectamine 2000 and T lymphocytes with chimeric receptor were generated and cultivated. The T lymphocytes' activation was assessed by detecting the expression of CD69, an early signal of T cell activation. Human gastric carcinoma cells of the lines HGC-27 (CEA(+)) and SGC-7901 (CEA(-)) were cocultivated with the T lymphocytes with chimeric receptor. Flow cytometry (FCM) with annexin V-FITC/PI double staining was used to detect the expressions of annexin V, a signal of cell apoptosis, so as to observe the apoptosis of the gastric carcinoma cells. T cells activated by phytohemagglutinin (PHA) were used as positive controls, and T cells transfected with blank vector pcDNA3.0 were used as negative controls. T lymphocytes with chimeric receptor directed towards CEA(+) gastric carcinoma cells were generated. 12 and 24 hours after the co-culture of these T cells and HCG-27 cells, the CD69 expression rates were 40.5% +/- 3.4% and 48.3% +/- 2.8% respectively. However, the CD69 expression rates of the HGC-27 and SGC-7901 cells transfected with the T lymphocytes transfected with blank vector were both 0%. FCM showed that the apoptotic rates of the CEA(+) gastric tumor HGC-27 cells was 47.8% +/- 4.2%, significantly higher than that of the CEA(-) gastric tumor SGC-7901 cells (18.7% +/- 2.8%, P < 0.05). T lymphocytes with chimeric receptor targeting CEA specifically identify CEA positive gastric carcinoma cells and promote the apoptosis thereof, thus providing a promising method of cellular immunotherapy for gastric carcinomas.