| Budget Amount *help |
¥7,700,000 (Direct Cost: ¥7,700,000)
Fiscal Year 1997: ¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 1996: ¥4,100,000 (Direct Cost: ¥4,100,000)
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| Research Abstract |
To determine the T-cell receptor (TCR) Valpha/Vbeta gene usage of the human autologous gastric tumor-specific cytotoxic T-lymphocytes (CTLs), we first established two pairs of tumor cell lines, HST2 and SSTW,from the malignant peritoneal effusions of signet ring cell carcinomas and their peripheral blood lymphocyte-derived tumor-specific CD8-positive CTL lines, TcHST2 and TcSSTW.TCR Valpha/Vbeta gene usage from these CTL was examined using the reversely transcribed-polymerase chain reaction method, demonstrating that the Valpha7, Valpha12, and Vbeta20 transcripts were commonly detected. The fact that repeated antigenic stimulation by mixed lymphocyte-autologous tumor cell cultures brought about the specific cytolysis and the restricted TCR usage of TCR Valpha7, Valpha12, and Vbeta20 strongly suggests that these TCR V region products participated in T-cell-cancer interaction. This restricted TCR V gene usage in the gastric signet ringcell carcinomas led us to examine further the frequen
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cy of TCR Valpha/Vbeta usage in 11 cases of in vivo tumor-infiltrating lymphocytes with this particular type of tumor. The date showed that Valpha7, Valpha12, Vbeta6, and Vbeta20 were also predominantly expressed among these tumor-infiltrating lymphocytes in vivo. However, it seemed that T-cells with these TCR V region products are not specific for the gastric signet ring cell carcinomas, since they also frequently infiltrate into noncancerous lesions, such as peptic ulcers. These data may suggest that T-cellswith certain TCR Valpha/Vbeta products could preferentially infiltrate into the stomach tissue, while some of these T-cells may be cytotoxic to the neoplastic autologous tumor cells.
To study the clonal dominance and the TCR structural stability of HST2-specific CTL from patient's peripheral blood lymphocytes (PBL), the PBL were newly stimulated with a mixed lymphocyte-autologous tumor cell (HST2) culture (MLTC) and cytotoxic T cell lines, such as HPBL3x, were obtained. RT-PCR and the nucleotide sequence data indicated that HPBL3x also showed TCR Valpha7 and Vbeta20 transcripts, and that HPBL3x TCR was composed of the exact same CDR3 gene structures as those of the TcHST2 clone. T cells with the same TCR structures were also detected in patient's non-treated peripheral blood, although they were infrequent. These data indicated that functional cytotoxic T cells with these distinct CDR3 equivalent structures were the dominant effector cells against HST2 autologous tumor cells. Moerover, the highly dominant and reproducible clonal expansion of T cells bearing heterodimeric TCR with identical variable, N diversity and constant region structures suggest that the molecular nature of governing antigenic peptide to TcHST2 may be stable and perhaps immunologically dominant in the interaction between CTL and HST2 autologous tumor cells. Less
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