TCR repertoire analysis revealed the highly conserved TCR repertoire in the bilateral tumor in mice

Temporal analysis of T-cell receptor (TCR) repertoire has been used to monitor treatment-induced changes in antigen-specific T cells in patients with cancer. However, lack of experimental model that allows the temporal analysis of TCR repertoire in same individual in homogeneous population limit the understanding of causal relationship between changes in TCR repertoire and antitumor responses. A bilateral tumor model, in which tumor cells were inoculated into the bilateral backs of mice, can be used for temporal analysis in TCR repertoire. In this study, we examined the prerequisite for this strategy: TCR repertoire are conserved between the bilateral tumor with same growth rate. The bilateral tumors with equivalent tumor size and draining lymph nodes (dLN) were collected 13 days after the tumor inoculation to analyze the TCR repertoire of CD4+ and CD8+ T cells. Most of the tumor-infiltrating T-cell clones were highly conserved between the bilateral tumors, and the extent of clonal expansion was equivalent. In addition, the similarity between bilateral tumors were equivalent to the heterogeneity in one side of the tumor. The similarity of TCR repertoire in the bilateral dLN was markedly lower than that of the tumor, suggesting that tumor-reactive T-cell clones induced independently in each dLN were integrated during recirculation and then infiltrated the tumor. These findings suggest that our bilateral tumor model is suitable for temporal monitoring of TCR repertoire to evaluate temporal and treatment-induced changes in tumor-reactive T-cell clones. Overall design: Lewis lung carcinoma (LLC) cells (5 × 105 cells /mouse) were inoculated subcutaneously (s.c.) into the bilateral backs of C57BL/6 mice (n=4). On day13, mice were sacrificed and the tumors and tumor-draining lymph nodes (DLN) were collected from the left and the right side. Tumor was devided into two pieces, and the TCR repertoire of CD4+ and CD8+ tumor infiltrating lymphocytes (TILs) in each piece were analyzed independently. Moreover, TCR repertoire of CD4+CD44hi and CD8+CD44hi T cells in the dLN were also analyzed.