During the last decade, adoptive transfer of autologous γδ T cells has been paid much attention to as an alternative immunotherapy for several types of cancer. It has been reported that human γδ T cells have potent cytotoxic activity against many types of tumor cell lines or primary cultured cells in vitro. Clinical studies in patients with metastatic renal cell carcinoma and non-small cell lung carcinoma showed that infusion of autologous γδ T cells is feasible and well tolerated. One of the problems in this therapy, however, is the difficulty for culturing peripheral γδ T cells in patients with cancer. Compared with healthy individuals, stimulation with phosphoantigens or bisphosphonates results in less expansion of γδ T cells in some patients. The tendency can be remarkably seen especially in those who have lower number of γδ T cells in peripheral blood. It is still unclear whether it is a unique phenomenon caused by something of malignancy or just due to the culture methodology. In this study, we investigated the characteristics of peripheral Vγ9Vδ2 T cells in patients with gastrointestinal cancer including their phenotype, the percentage in PBMC, and the proliferation activity in response to synthetic phosphoantigen, 2-methyl-3-butenyl-1-pyrophosphate (2M3B1PP).

PBMC were prepared from patients, and then the phenotype and the number of Vγ9Vδ2 T cells were measured by flow cytometry. To expand Vγ9Vδ2 T cells in vitro, PBMC were cultured in AIM-V medium supplemented with 100 IU/mL IL-2, 100μM 2M3B1PP, and 10% autologous plasma. The expansion of Vγ9Vδ2 T cells were also examined using flow cytometry.

We found that the percentage of the Vγ9Vδ2 T cells in peripheral T cells varied a lot depending on the patients. While their percentage in healthy individual was approx. 1.5-9.0%, it exceeded 10% in some patients, and even over 20% in several patients. Phenotypic analysis revealed that almost all the Vγ9Vδ2 T cells in these particular patients were effector-memory type cells with CD45RA-/CD27-/CCR7- and they massively expanded by the stimulation with 2M3B1PP. These results suggest that Vγ9Vδ2 T cells in these patients have been constantly exposed and stimulated with some specific antigens. On the other hand, the percentage of Vγ9Vδ2 T cells in T cells of some patients was less than 1.0%, and in the conventional culture methods the cells' response to 2M3B1PP was too low for them to proliferate themselves. However, their proliferation rate markedly increased when PBMC were inoculated at the high density (i.e. 1.3 x 104 Vγ9Vδ2 T cells per cm2 or more). The high density culture for 14 days has made it possible to expand the number of Vγ9Vδ2 T cells to 100-1000 folds. We conclude that Vγ9Vδ2 T cells in patients with gastrointestinal cancer have the equivalent level of proliferation ability to healthy individual even though the initial percentage is low, and thus these cells could be a possible effector cell type in a novel immunotherapy for gastrointestinal cancer.

Citation Format: Yasunobu Kobayashi, Mayuko Sakai, Izumi Fujita, Hiroshi Komine, Masanori Matsushita, Koichi Shimizu, Atsushi Aruga, Hirohito Kobayashi, Keishi Tanigawa. Phenotype and expansion profile of peripheral γδ T cells in patients with gastrointestinal cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2793. doi:10.1158/1538-7445.AM2014-2793