Increased CD112 Expression in Methylcholanthrene-Induced Tumors in CD155-Deficient Mice

<div><p>Tumor recognition by immune effector cells is mediated by antigen receptors and a variety of adhesion and costimulatory molecules. The evidence accumulated since the identification of CD155 and CD112 as ligands for DNAM-1 in humans and mice has suggested that the interactions between DNAM-1 and its ligands play an important role in T cell– and natural killer (NK) cell–mediated recognition and lysis of tumor cells. We have previously demonstrated that methylcholanthrane (MCA) accelerates tumor development in DNAM-1–deficient mice, and the <i>Cd155</i> level on MCA-induced tumors is significantly higher in DNAM-1–deficient mice than in wild-type (WT) mice. By contrast, <i>Cd112</i> expression on the tumors is similar in WT and DNAM-1-deficient mice, suggesting that CD155 plays a major role as a DNAM-1 ligand in activation of T cells and NK cells for tumor immune surveillance. To address this hypothesis, we examined MCA-induced tumor development in CD155-deficient mice. Unexpectedly, we observed no significant difference in tumor development between WT and CD155-deficient mice. Instead, we found that <i>Cd112</i> expression was significantly higher in the MCA-induced tumors of CD155-deficient mice than in those of WT mice. We also observed higher expression of DNAM-1 and lower expression of an inhibitory receptor, TIGIT, on CD8⁺ T cells in CD155-deficient mice. These results suggest that modulation of the expression of receptors and CD112 compensates for CD155 deficiency in immune surveillance against MCA-induced tumors.</p></div

Relative cytokine mRNA levels in MCA-induced tumors.

<p>(A, B) Fibrosarcomas induced by 5 µg MCA in WT or CD155-deficient (KO) C57BL/6N (A) or BALB/c (B) mice were resected from each mouse and subjected to quantitative qRT-PCR for transcripts of the indicated cytokines. Horizontal bars represent means and error bars represent means ± SEM. <i>P</i> Values for Student’s <i>t</i> test are shown.</p

Expression of CD155 ligands on resting and activated T cells from WT and CD155-deficient mice.

<p>(A) Peripheral blood lymphocytes from WT (<i>n</i> = 3) or CD155-deficient (KO; <i>n</i> = 3) C57BL/6N mice were stained with anti-DNAM-1, anti-TIGIT, or anti-CD96 antibodies (WT; blue lines, KO; red lines) or control antibodies (WT; light blue lines, KO; pink lines), and analyzed by flow cytometry. The numbers (WT; in blue, KO; in red) indicate the mean fluorescence intensity (MFI) of DNAM-1, TIGIT, and CD96 staining. Representative data are shown. (B) MFI was used to analyze DNAM-1 expression on CD4⁺ T, CD8⁺ T, and NK cells as in (A). Error bars represent means ± SD. (C) CD8⁺ T cells purified from spleen were activated with anti-CD3 antibody and IL-2 for the indicated number of days. Cells were stained and analyzed by flow cytometry, as described in (A). (D) The expression of DNAM-1, TIGIT, and CD96 on CD8⁺ T cells is shown as MFI as in (C). *, <i>P</i><0.05; **, <i>P</i><0.01; ***, <i>P</i><0.001.</p

MCA-induced tumor development in WT and CD155-deficient mice.

<p>(A, B) WT (<i>n</i> = 18 and 16, respectively) or CD155-deficient (KO; <i>n</i> = 19 and 20, respectively) C57BL/6N mice were injected s.c. with 5 µg (A) or 100 µg (B) methylcholanthrane (MCA) on day 0. (C) WT (<i>n</i> = 15) or KO (<i>n</i> = 10) BALB/c mice were injected s.c. with 5 µg MCA on day 0. Tumor size in each mouse was measured once a week. Tumor size (top) and survival data (bottom) are shown.</p

A hypothetical model for interactions between activating/inhibitory receptors DNAM-1, TIGIT and CD96 on T or NK cells and CD155/CD112 ligands expressed on MCA-induced fibrosarcoma in WT or CD155-deficient mice.

<p>(A) DNAM-1 and TIGIT bind to both of CD155 and CD112, while CD96 interacts with CD155 only. Each receptor-ligand interaction transduces either activating or inhibitory signal, as shown by the red or blue arrow, respectively. The modulation of the receptors and ligand expression on CD155-deficient (KO) fibrosarcoma are indicated. (B) The sums of the activating and inhibitory signals are similar between WT and KO.</p

Relative <i>Cd112</i> mRNA levels in organs and MCA-induced tumors.

<p>(A) Tissues from WT (<i>n</i> = 3) or CD155-deficient (KO; <i>n</i> = 3) mice were subjected to quantitative qRT-PCR for <i>Cd112</i>. Error bars represent means ± SD. (B, C) Fibrosarcomas induced in WT or CD155-deficient C57BL/6N (B) or BALB/c (C) mice by 5 µg MCA were resected from each mouse and subjected to qRT-PCR for <i>Cd112</i>. Horizontal bars represent means and error bars represent means ± SEM. <i>P</i> Values for Student’s <i>t</i> test are shown.</p