# High CD73 Expression Is Associated with Poor Prognosis in Biliary Tract Cancer Through Reduced Stromal Tumor-Infiltrating Lymphocytes

## TL;DR

High CD73 expression in biliary tract cancer is linked to worse survival due to reduced immune cell infiltration in the tumor environment.

## Contribution

This study identifies CD73 as a novel prognostic marker in biliary tract cancer through AI-based analysis of immune cell infiltration patterns.

## Key findings

- High CD73 expression in tumor cells correlates with reduced stromal TIL infiltration and worse survival.
- Stromal TIL density is associated with improved patient outcomes in biliary tract cancer.
- CD73 expression selectively suppresses cytotoxic T-cell infiltration in the tumor microenvironment.

## Abstract

Biliary tract cancer (BTC) is a highly aggressive malignancy with a poor prognosis, and effective treatment options remain limited. In this study, we used artificial intelligence-based analysis of pathological images and found that patients with high expression of CD73 in tumor cells showed reduced infiltration of tumor-infiltrating lymphocytes (TILs), including cytotoxic T cells (CTL), in the stroma, which was associated with worse clinical outcomes. These findings suggest that an immunosuppressive tumor microenvironment (TME) mediated by CD73 may contribute to poor prognosis in BTCs. Targeting CD73 may therefore represent a promising therapeutic strategy and provide new treatment opportunities for patients with this disease.

Background: Biliary tract cancer (BTC) is an aggressive malignancy with limited therapeutic options and a poor prognosis. CD73 is upregulated under hypoxic conditions and promotes tumor progression. However, its clinical role in BTC and interaction with tumor-infiltrating lymphocytes (TILs) remain unclear. This study aimed to elucidate the association between CD73 expression and prognosis in BTC, as well as its impact on the tumor microenvironment (TME) and TILs. Methods: This retrospective study included 100 patients who underwent curative BTC surgery at Hokkaido University Hospital between 2018 and 2023. Formalin-fixed tumor specimens were analyzed using DeepPathFinder™ (biomy Inc., Tokyo, Japan), an AI-based digital pathology platform enabling objective quantification of CD73 expression and lymphocyte infiltration within tumoral (T) and stromal (S) compartments. Immunohistochemistry for CD3, CD8, Foxp3, and CD163 was used to identify T-cell subsets and macrophages. Associations between CD73, TIL subsets, and overall survival (OS) were assessed using the Kaplan–Meier, Cox regression, and Spearman correlation analyses. Results: High T-CD73 expression was associated with shorter OS (hazard ratio [HR] = 1.97, p = 0.041), whereas S-CD73 showed no prognostic relevance. Conversely, high S-TIL density was correlated with improved survival (HR = 0.49, p = 0.032). T-CD73 expression was negatively correlated with stromal CD3+ and CD8+ T-cell densities, indicating selective suppression of stromal cytotoxic T-lymphocyte (CTL) infiltration. No significant correlations were observed between Foxp3+ T cells and CD163+ M2 macrophages. Conclusions: CD73 upregulation in tumor cells impairs stromal CTL and TIL activity, leading to a poor prognosis. Spatial distribution, rather than total TIL number, better reflects effective anti-tumor immunity.