Phenotype, Cytokine Production and Cytolytic Capacity of Fresh (Uncultured) Tumour-Infiltrating T Lymphocytes in Human Renal Cell Carcinoma

We investigated the phenotype and functional capacities of tumour-infiltrating lymphocytes (TIL), freshly isolated from primary renal cell carcinoma (RCC) specimens (n=20). Three-colour flow cytometry immunophenotyping revealed that RCC TIL consist mainly of CD3(+) T cells, with a clear predominance of CD4(-)CD8(+) over CD4(+) CD8(-) T cells, and a marked population of CD4(+) CD8(+) T cells. Natural killer (NK) cells were also strongly represented (> 25% in 15 of 20 tumour samples), while B cells constituted a minor TIL subset (<5% in 18 of 20 tumour samples). More importantly, the T and NK cells within the tumour displayed a significantly higher expression of the early activation marker CD69 than their counterparts in adjacent normal renal tissue and in peripheral blood. Expression of CD54 and of HLA-DR was also elevated on CD3(+) TIL, and HLA-DR expression was further vigorously up-regulated following ex vivo stimulation with anti-CD3, all suggesting enhanced immune activity within the tumour microenvironment. CD3(+) CD4(+) TIL displayed a normal capacity to up-regulate CD25 expression and to secrete both Th1-type (IL-2, tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma)) and Th2-type (IL-4, IL-5 and IL-10) cytokines upon triggering with anti-CD3. Furthermore, cytokine production was susceptible to modulation by CD28 costimulation. CD3(+) CD8(+) TIL, on the other hand, consistently demonstrated a poor up-regulation of CD25 upon triggering with anti-CD3, and displayed poor ex vivo cytolytic activity in an anti-CD3-redirected 4-h cytotoxicity assay against murine P815 cells. Collectively, our findings indicate that the CD3(+) CD4(+) TIL in RCC have normal functional capacities, whereas the proportionally major CD3(+) CD8(+) TIL are functionally impaired. The relevance of these findings to the in vivo local immune response in RCC is discussed.