Impact of tumor-derived CCL2 on T cell effector function
Introduction
In 1983 Botazzi et al. [1] reported that the amount of tumor derived chemotactic factor (TDCF) produced by tumors correlated with the level of macrophage infiltration. Subsequently, monocyte chemoattractant protein-1 (MCP-1, CCL2) was found to be responsible for most of the biological effects of TDCF [2]. CCL2 is a CC chemokine that is capable of acting upon macrophages as well as T cells and basophils [3]. Following these initial studies multiple reports documented that additional human and murine tumors constitutively express CCL2 [4], [5], [6]. Yet the role of tumor-derived CCL2 in tumorigenicity and anti-tumor immunity has not been resolved. In fact, the field has been complicated by contrasting results.
Several reports have indicated that tumor-derived CCL2 can enhance progression of breast cancer in humans and increase malignancy of murine mammary tumors [5], [6]. The ability of CCL2 to recruit macrophages may be one mechanism by which CCL2 can mediate some of these effects [7]. In contrast, several studies have reported that introduction of the CCL2 gene into tumor cells correlated with decreased tumorigenicity and facilitated immune mediated tumor rejection [8], [9], [10]. Thus, the precise role of CCL2 in anti-tumor immunity remains unsettled. While many investigators have examined the influence of tumor-derived CCL2 on macrophages, the impact of CCL2 on the T cell response to cancer has for the most part been overlooked. One study that did focus on T cells reported that MCA205-derived CCL2 impaired T cell effector function [11].
In this study we were interested in determining how tumor-derived CCL2 influenced the T cell response to the 4T1 murine mammary carcinoma model. 4T1 is a relatively weakly immunogenic tumor cell line that constitutively expresses CCL2 [12]. Previously we reported that mice bearing the 4T1 tumor possess splenic T cells with altered chemokine receptor function [12]. Here we wanted to know whether inhibiting the tumor cells from producing CCL2 would influence the T cell response and consequently the immunogenicity of the tumor. For this reason we generated cells that lacked detectable levels messenger RNA encoding CCL2 and CCL2 protein. Next, we compared the T cell response elicited by the CCL2 null cells to the response elicited by the cells that produced CCL2. Analysis revealed that blockade of tumor-derived CCL2 could increase T cell effector function, but not the immunogenicity of the tumor.
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Mice and tumor cell lines
For these studies 6–8 week old female BALB/c mice were used. The BALB/c mice were purchased from The Jackson Laboratory (Bar Harbor, ME). Food and water were provided ad libitum. All of the tumor cells used in this study (4T1, 4T1-9, 4T1-A4, 4T1-G7, and SM1) were maintained in complete RPMI (cRPMI) (RPMI 1640, BioWhittaker, Walkersvile, MD) supplemented with 10% heat-inactivated fetal bovine serum (Gibco BRL, Gaithersburg, MD), glutamine (2 mM, BioWhittaker), penicillin (100 U/mL, BioWhittaker),
Characterization of CCL2- and CCL2+ tumor cells
In order to investigate the role of tumor-derived CCL2 in the T cell response to cancer we compared CCL2- and CCL2+ tumors. For this purpose we used cells derived from a clone of 4T1. The A4 cells expressed similar levels of mRNA for CCL2 compared to the parental tumor, whereas no message was detected from the CCL2- tumor; G7 (Fig. 1A). Analysis of supernatants, collected from the tumor cells, for CCL2 protein revealed a similar pattern. A4 expressed high levels of CCL2 whereas G7 produced no
Discussion
Due to contrasting results in different animal models the role of CCL2 in anti-tumor immunity has been difficult to unravel. Some investigators have reported that tumor-derived CCL2 decreased tumorigenicity. For instance, Chinese Hamster Ovary (CHO) cells transfected with either the human or murine CCL2 gene lose their ability to form tumors in nude mice [8]. Similarly, a delay in tumor growth occurred when a B16 derived cell line, B78HI, was transfected with the CCL2 gene [16]. However, in
Acknowledgements
This project was funded by the Department of Defense Breast Cancer Research Program DAMD17-01-1-0288.
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