Correlation between CD4⁺CD25⁺T_reg Cells and CCR4 in Nasopharyngeal Carcinoma

Discussion

Patients with NPC have benefited from radiotherapy and chemotherapy, but more recent studies have examined the role of host immune responses in head and neck carcinoma progression, and suggested that T lymphocytes may play a role in controlling tumor growth^[8]. T lymphocytes are composed of two subgroups: suppressor T cells (Ts) CD8⁺ and helper T cells (Th) CD4⁺, which interact with each other balancing the immune function. CD4⁺ T cells regulate the immunity and help B lymphocytes to secrete antibodies, while CD8⁺ T cells mediate killing of target cells. The percentage of CD4⁺/CD8⁺ T cells in PB is the most widely used index of immunity in clinical practice. However, some studies have suggested that there is no good correlation between the percentage of CD4⁺/CD8⁺ T cells and the stage, tumor burden and survival of tumor patients. However, CD4⁺ T cell and CD8⁺ T-cell populations are composed of subpopulations with different specific functions. Moreover, since the distribution of different T-cell populations in tumor sites differs from that in PB, it is expected that immunosuppression may occur in tumor sites but not in other parts of the body. Therefore, the CD4⁺/CD8⁺ ratio is not a good index of the status of the immune system.

At tumor sites, lymphocytes infiltrate into primary or secondary tumors and play an important role in killing tumor cells and in reducing the potential for metastasis. Therefore, the quantity and functioning of TI lymphocytes represents the intensity and level of host-antitumor immunity. However, it has been reported that the quantity of CD4⁺CD25⁺ T_reg cells increases in tumor sites and in the PB of patients with ovarian carcinoma, lung cancer, breast cancer, pancreatic cancer and colon carcinoma^[9–13]. CD4⁺CD25⁺ T_reg cells act as an immune system regulator by direct cell contact and cytokine secretion^[14,15].

In our study, the data showed that the percentage of CD4⁺CD25⁺ T_reg cells in NPC specimens was much higher than that in PB or in the normal tissue of controls. We also found that there was a negative correlation between CD4⁺CD25⁺ T_reg cells and CD4⁺CD25^– T cells in the NPC specimens. Yang et al.^[12] reported that CD4⁺CD25⁺ T_reg cells could suppress not only the proliferation of CD4⁺CD25^– T cells but also interferon γ and interleukin-4 secretion by CD4⁺CD25^– T cells. Experimental depletion of CD4⁺CD25⁺ T_reg cells in mice with tumors improves immune-mediated tumor clearance and enhances the response to immune-based therapy. The quantity of CD4⁺CD25⁺ T_reg cells is greater than that of CD4⁺CD25^– T cells in gastric adenocarcinoma; the close connection indicates that CD4⁺CD25⁺ T_reg cells suppress CD4⁺CD25^–T cells by direct cellular contact^[16]. Therefore, there is a possibility that CD4⁺CD25⁺ T_reg cells promote tumor growth by suppressing CD4⁺CD25^– T cells. This needs further investigation with a larger number of samples.

The percentage of CD4⁺CD25⁺ T_reg cells in the patients with NPC at stage III+IV was higher than that at stage I + II, which is consistent with the results of the study on gastric and esophageal carcinomas^[17], but in contrast to a recent report by Lau et al.^[18] The main reason for this discrepancy is that the percentage of CD4⁺CD25⁺ T_reg cells in the study of Lau et al. was that in PB, whereas we analysed both tumor sites and PB. Thus our study suggests that the percentage of CD4⁺CD25⁺ T_reg cells in tumor sites is more appropriate for evaluating host immunity.

Recruitment of T cells to a particular site is a multistage process involving secretion of chemokine by tumor and rolling T cells^[19]. A limited number of CD4⁺CD25⁺ T_reg cells are found in normal tissues, but there are no data on their role in normal tissue^[20]. In this experiment, we found that the percentage of CCR4⁺ cells in TI and PB lymphocytes was significantly higher in patients with NPC than in controls, and the percentage of CCR4⁺ cells was positively related to that of CD4⁺CD25⁺ T_reg cells in both specimens and PB of the patients with NPC. The hypothesis that malignant tumors can attract CD4⁺CD25⁺ T_reg cells through CCR4 is logical. It has been reported that the levels of the CCR4 ligand, CCL22, are elevated in gastric adenocarcinoma^[16], ovarian carcinoma^[21] and Hodgkin’s disease^[22], and the CCL22 level is related to the prognosis of patients with cancer, and only in lymph nodes infiltrated by tumor cells would CD4⁺CD25⁺ T_reg cells suppress immunity^[23]. On the other hand, little CCR4 is expressed by T cells in lymph nodes free of tumor cell infiltration such as in Hodgkin’s disease^[24]. Another experiment proved that CCR4 was highly expressed on CD4⁺CD25⁺ T_reg cells^[25]. Therefore, it is logical to suggest that effective T cells can recognize tumor antigen and play an important role in anti-tumor activity by means of antigen-dependent-cell-mediated cytotoxicity at an early stage. On the other hand, in order to survive, the tumor would produce copious amounts of CCR4 ligands, CCL17 and CCL22, to attract CD4⁺CD25⁺ T_reg cells to suppress immunity in tumor sites.

Investigators also reported that mature dendritic cells (DCs) could secrete CCL22 and CCL17 to mediate the migration of T lymphocytes to tumor sites^[26]. There is competition among T lymphocytes, and CCR4 expressed on CD4⁺CD25⁺ T_reg cells would give them an advantage, enabling the CD4⁺CD25⁺ T_reg cells to suppress the proliferation and function of effective T cells^[27,28]. We suggest that DCs can present tumor antigens to effective T cells for their anti-tumor effects; meanwhile DCs also produce chemokines to attract CD4⁺CD25⁺ T_reg cells to tumor sites. Therefore, CCR4 plays an important role in recruitment and suppression of CD4⁺CD25⁺ T_reg cells at tumor sites.

CD4⁺CD25⁺ T_reg cells facilitate the growth of malignant tumors, so suppression and deletion of CD4⁺CD25⁺ T_reg cells will strengthen the host anti-tumor response. Shimizu et al.^[29] transferred a splenic cell suspension depleted of CD4⁺CD25⁺ T_reg cells to BALB/c athymic nude mice, before implantation with BALB/c-derived radiation leukemia cells. The tumors of the majority (> 90%) of the mice were significantly suppressed and the mice survived for a longer term (> 60 days) than control mice treated with normal rat IgG (< 40 days). Sutmuller et al.^[30] eliminated CD25⁺ T cells by anti-murine CD25 antibody in C57BL/6 female mice inoculated with a B16BL6 melanoma cell line. They found that depletion of CD25⁺ T cells enhanced tumor rejection, but this rejection was dependent on CD4⁺ T cells and CD8⁺ T cells. However, the anti-CD25 antibody could also reduce the anti-tumor effect of CD8⁺ T cells and natural killer cells. Cytokine-induced killer cells depleted of CD4⁺CD25⁺ T_reg cells include more CD8⁺ and CD3⁺CD56⁺cells than those with CD4⁺CD25⁺ T_reg cells, and they can kill tumor cells more effectively^[31]. Ishida et al.^[24] showed that a chimeric anti-CCR4 monoclonal antibody could deplete CCR4⁺ cells and inhibit the migration of CD4⁺CD25⁺ T_reg cells in vitro, and, therefore, they proposed that a therapeutic strategy with an anti-CCR4 monoclonal antibody combined with tumor-specific cytotoxicity T lymphocyte was worth examining. CD4⁺CD25⁺ T_reg cells have been extensively studied and an understanding of their role has advanced, therefore, a safer and more efficient way of treating NPC will be discovered.