Analysis of NK cells and chemokine receptors in tumor infiltrating CD4 T lymphocytes in human renal carcinomas

Abstract

Recent data suggest that chemokines and chemokine receptors mediate leukocyte recruitment of all components of the antitumor response. This study aimed to phenotypically characterize the immune lymphocyte infiltrate in human renal cell carcinomas RCCs and at the invasive margin (tumor-host interface) and to define the association of these findings with established prognostic indicators. Tumor infiltrating lymphocytes TILs were obtained from 24 patients with RCC undergoing radical nephrectomy. Peripheral blood cells from 37 patients were also obtained before surgery. Our findings are consistent with the preferential recruitment of CD4+ Th1-polarized effector memory cells that express CXCR3/CCR5. These cells were the main component of TILs and expressed as CXCR3, CCR5, CD45RO, and CD95. Natural killer (NK) cells were found in significantly higher proportions in TILs of RCCs than in peripheral blood lymphocytes (PBLs) or in other tumors studied (colorectal and breast cancers), where these cells were found in small proportions. No differences in nuclear grade or other studied parameters were observed between the TILs and the lymphocytes present at the invasive margin, which showed a similar composition. However, differences were found according to the tumor stage. First, significantly fewer NK cells were observed in PBLs from metastatic patients. Second, a significantly lower proportion of CCR5/CXCR3/CD4+ cells and a higher proportion of CCR4/CD4+ cells were observed in metastatic patients, suggesting that preferential Th1-polarization may gradually change during the progression of renal cancer cells. Finally, the frequency of CD25/CD4+ cells was higher in metastatic patients. Although the sample of patients with metastasis was small, the overall results suggest a change in composition of the TILs that may potentially confer a selective advantage for tumor growth and may account for the suppression of an effective cytotoxic response.

Fig. 1

a Dot plot histograms of the flow cytometric analysis of CD16/CD56 (NK), CD62L/CD4, CD45RO/CD4, and CD25/CD4 in PB cells and TILs. These data demonstrate a high percentage of CD45RO+, CD62L^low T CD4 cells (memory cells), and NK cells in the TILs. b Dot plot histograms of the flow cytometric analysis of CXCR3/CD4, CCR5/CD4, CCR4/CD4, and CD95/CD4 cells in PB cells and TILs. The proportion of CCR5/CXCR3/CD4 cells was higher in TILs than in PBLs

Fig. 1

a Dot plot histograms of the flow cytometric analysis of CD16/CD56 (NK), CD62L/CD4, CD45RO/CD4, and CD25/CD4 in PB cells and TILs. These data demonstrate a high percentage of CD45RO+, CD62L^low T CD4 cells (memory cells), and NK cells in the TILs. b Dot plot histograms of the flow cytometric analysis of CXCR3/CD4, CCR5/CD4, CCR4/CD4, and CD95/CD4 cells in PB cells and TILs. The proportion of CCR5/CXCR3/CD4 cells was higher in TILs than in PBLs

Fig. 2

Association between the concentration of NK cells in PB (cells/μl) and percentage of these cells in the TILs in 24 renal carcinomas. All but four patients demonstrated statistically significant correlation between these two parameters. Four exceptional cases showed a high NK cell number in TILs accompanied by a very low blood level of NK cells. Correlation coefficient=0.77 (P<0.001), calculated excluding the four cases indicated in the box

Fig. 3

Analysis of the intratumoral lymphocyte subsets in renal cell carcinoma and colon carcinoma. Results are expressed as ratio of the mean percentage of cells +/- standard deviation of 23 RCCs (grey bars) and five colon carcinomas (black bars). Statistical analyses were performed using the Mann–Whitney test