The Transmembrane Receptor TIRC7 Identifies a Distinct Subset of Immune Cells with Prognostic Implications in Cholangiocarcinoma

Abstract

Cholangiocarcinoma (CCA) is a heterogeneous malignancy with a dismal prognosis. Therapeutic options are largely limited to surgery and conventional chemotherapy offers limited benefit. As immunotherapy has proven highly effective in various cancer types, we have undertaken a quantitative immunohistopathological assessment of immune cells expressing the immunoinhibitory T cell immune response cDNA 7 receptor (TIRC7), an emerging immunoinhibitory receptor, in a cohort of 135 CCA patients. TIRC7⁺ immune cells were present in both the tumor epithelia and stroma in the majority of CCA cases with the highest levels found in intrahepatic CCA. While intraepithelial density of TIRC7⁺ immune cells was decreased compared to matched non-neoplastic bile ducts, stromal quantity was higher in the tumor samples. Tumors exhibiting signet ring cell or adenosquamous morphology were exclusively associated with an intraepithelial TIRC7⁺ phenotype. Survival analysis showed intraepithelial TIRC7⁺ immune cell density to be a highly significant favorable prognosticator in intrahepatic but not proximal or distal CCA. Furthermore, intraepithelial TIRC7⁺ immune cell density correlated with the number of intraepithelial CD8⁺ immune cells and with the total number of CD4⁺ immune cells. Our results suggest the presence and prognostic relevance of TIRC7⁺ immune cells in CCA and warrant further functional studies on its pharmacological modulation.

Keywords: PD-L1; TIRC7; biomarkers; cholangiocarcinoma; immune checkpoint inhibitors; immunotherapy; tumor microenvironment.

Figure 1

TIRC7⁺ immune cells in cholangiocarcinoma. Photographs of both a low (A) and particularly high (B) intraepithelial TIRC7⁺ immune cell density and low (C) and particularly high (D) numbers of TIRC7⁺ cells in the stromal compartment. Displayed are tumor samples. Note that the photographs were used for visualization and as such do not represent the expression average. Original magnification: 200×.

Figure 2

Quantitative analysis of TIRC7⁺ immune cells in tumor and nontumor tissue. Intraepithelial density of TIRC7⁺ immune cells was significantly reduced in tumor tissue as compared to matched non-neoplastic bile duct (BD) epithelia (available for a subset) across all CCAs (A) as well as in the subgroup analysis for iCCA (B), pCCA (C) and dCCA (D). Total number of TIRC7⁺ cells in the stromal compartment was significantly higher in tumor as compared to nontumor tissue when all CCAs were included (E), which can be attributed to significantly higher levels in iCCA cases (F). No significant differences were found regarding pCCA (G) and dCCA (H) tissue with respect to the stromal compartment. For all analyses the Mann–Whitney U test was used. Data are depicted as mean ± standard error of the mean.

Figure 3

Intergroup analysis of TIRC7⁺ immune cell quantity in tumor tissue. Intraepithelial density of TIRC7⁺ immune cells was significantly different among the three CCA subgroups with significantly higher levels in iCCA compared to pCCA cases (A). With respect to the stromal compartment, the total number of TIRC7⁺ cells differed between the groups, being significantly higher for pCCA as compared to iCCA (B). Overall group differences were assessed using the Kruskal–Wallis followed by Dunn’s test for post hoc pairwise comparisons. Data are depicted as mean ± standard error of the mean.

Figure 4

Survival analysis in relation to TIRC7 quantity. Cases with absent TIRC7⁺ intraepithelial cells showed a remarkably shortened survival as compared to TIRC7-positive cases across all CCA types (A). An analogous trend was also seen when using the median (B) and the mean (C) value of intraepithelial TIRC7⁺ immune cell density as stratification cut-offs. Subgroup analysis revealed a strong survival benefit of TIRC7-positive cases in iCCA (D), while no differences were found with respect to pCCA (E) and dCCA (F). p-values were computed using log-rank testing. Hazard ratios (HR) and its 95% confidence intervals were calculated using the Mantel Haenszel approach.

Figure 5

Association of intraepithelial TIRC7⁺ immune cell density with immune cell phenotype. Analysis of quantity of intraepithelial CD8⁺ (A) or total CD4⁺ (B) T cells in CCA cases with absent (0%), low (1–3%) or high (≥4%) intraepithelial TIRC7⁺ immune cell density shown by scatter dot plots with mean and SEM (CD8 Kruskal–Wallis p = 0.007; CD4 Kruskal–Wallis p = 0.005). Correlation of intraepithelial CD8⁺ (C) or total CD4⁺ (D) T cells with intraepithelial TIRC7⁺ immune cell density, as indicated (CD8 Spearman r = 0.300, p = 0.003; CD4 Spearman r = 0.367, p < 0.001). Overall group comparison was performed using the Kruskal–Wallis followed by Dunn’s test for post hoc pairwise comparisons (A,B). Correlation metrics were computed out using Spearman’s correlation analysis (C,D).