Study of tumor-infiltrating T-cell reactivity to metastatic gastrointestinal cancers.

Abstract

e14179 Background: A high density of tumor-infiltrating lymphocytes (TIL) found in gastrointestinal cancers has been associated with improved prognosis. Evidence supporting specific autologous tumor recognition by TIL remains sparse and limits the development of innovative immunotherapy approaches. Methods: Metastases from 15 GI cancer patients (13 colorectal, 1 gastric, 1 cholangiocarcinoma) processed into fragments and fresh tumor cell suspensions (FrTu) were used to grow TIL and cancer cell lines. TIL phenotype was assessed by flowcytometry and reactivity by coculture with available cryopreserved FrTu and cancer cell lines. Limiting dilution techniques were used to isolate reactive CD8+ TIL clones. T-cell receptors (TCR) were sequenced using PCR amplification. Results: TIL were expanded successfully in all 15 cases. TIL phenotype studied from 7 freshly resected colorectal cancer metastases revealed a higher percentage of immunosuppressive T cells (CD4+CD25HIFoxP3+) compared to adjacent normal tissue and blood (respectively 5.4 ± 1.0%, 1.3 ± 0.5%, and 1.0 ± 0.4% of CD3+ cells). Interferon-gamma release by TIL after their first outgrowth upon coculture with cryopreserved FrTu was significant in all 6 cases studied, with positive results ranging from 7% (3/42) to 89% (8/9) of patient’s distinct cultures. Two newly established cancer cell lines allowed extensive investigation of TIL reactivity. In a case of gastric cancer, 65% of the CD3+CD8+ TIL expanded from liver metastases expressed the same TCR. TIL clones expressing this TCR could recognize the autologous cancer cell line with a high degree of specificity, suggesting that within this tumor a productive immune response occurred. In a case of metastatic cholangiocarcinoma, two TIL clones with distinct TCR represented less than 1% of the CD3+CD8+ cells grown from an omental metastasis and the malignant ascites, but could nonetheless lyse the autologous tumor with specificity. Conclusions: Despite the immunosuppressive microenvironment of GI cancer metastases, CD8+ TIL that recognize autologous tumor with specificity can be identified and expanded in vitro. The adoptive transfer of TIL for the therapy of metastatic GI cancer patients is currently being studied.