Abstract 5699: A new method for evaluating T cell response after stimulation on gastrointestinal cancer patients
Authors: Tanaka Tomoko , Yamashita Kimihiro , Sugita Yutaka , Fukuoka Eiji , Arimoto Akira , Kakeji Yoshihiro 
Published: August 2018

Abstract
Background: In anti-tumor immunity, T cells play a central role to eliminate cancer cells. Immune responses of T cells are initiated by communication with antigen presenting cells (APCs). It became clear that an immunological synapse (IS) is formed at the T cell-APC interface and is made up of a supramolecular activation cluster (SMAC) containing the T cell receptors (TCR) microclusters (MCs) and CD28 MCs. TCR MCs consist of TCR, kinases, and adaptor molecules, and are the minimal unit of T cell signal transduction. CD28 MCs also function as the signalosomes of CD28 activation. By using total internal reflection fluorescence microscopy to observe the interaction of T cell and planar bilayer, it was elucidated that TCR activation is regulated by the amount of MCs aggregation, which shows the extent of IS formation. Based on this finding, we hypothesized that ex vivo T cells could be examined IS formation and that exhausted T cells of advanced cancer patients should be low in response to a stimulus.
Method: To simplify IS formation of T cells, we used human monoclonal T cells by αCD3 and/or αCD28 antibody and replaced to evaluate the aggregation of MCs by detecting the CD3 or CD28 molecules on the surface of T cells. This phenomenon was defined as IS-like formation. In this method, IS-like formation was detected by confirming an image of each cell with imaging cytometry. After these conditions was established, it was verified whether it is possible to measure IS-like formation rate of T cells. Additionally, the correlation between IS-like formation and interferon-γ cytokine production after this activation was examined. In the same manner as above, IS - like formation rates of T cells derived from PBMCs of healthy donors or esophageal and gastric cancer patients were measured.
Result: We have succeeded in observing that CD3 or CD28 molecules on a T cell aggregated in one area after stimulation as IS-like formation by imaging cytometry. In dot plot study, positive area of CD3 or CD28 decreased after the stimulation, which showed the aggregation of these molecules. Accordingly, we could measure IS-like formation rate of a population of T cell clones. In addition, the rate of IS-like formation showed a positive correlation with interferon-γ production. Similar results were obtained T cell from PBMC in healthy donors. Moreover, IS-like formation rate of T cell from cancer patients decreased compared with healthy donors.
Conclusion: IS-like formation of T cell succeeded to observe by CD3 and/or CD28 antibody stimulation. We established the evaluation method of IS-like formation degree by imaging cytometry. Moreover, IS-like formation rate correlated with cytokine production of IFN-g. We established new method to evaluate the strength of T cell after activation.
Citation Format: Tomoko Tanaka, Kimihiro Yamashita, Yutaka Sugita, Eiji Fukuoka, Akira Arimoto, Yoshihiro Kakeji. A new method for evaluating T cell response after stimulation on gastrointestinal cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5699.