Human CD96 correlates to NK cell exhaustion and predicts the prognosis of human hepatocellular carcinoma.

Immune checkpoint blockade has become a promising therapeutic approach to reverse immune cell exhaustion. Co-inhibitory CD96 and TIGIT, together with co-stimulatory CD226, bind to common ligand CD155. The balancing between three receptors fine tunes the immune responses against tumors. In this study, we investigated the expression of CD96, TIGIT and CD226 in 55 fresh human hepatocellular carcinoma (HCC) samples, 236 paraffin-embedded HCC samples, and 20 normal human livers. The cumulative percentage, absolute count, and MFI of CD96+ NK cells are significantly increased in the intratumoral tissues of HCC, and break the balance between three receptors. Human CD96+ NK cells are functionally exhausted with impaired IFN-γ and TNF-α production, high gene expression of IL-10 and TGF-β1, and low gene expression of T-bet, IL-15, perforin and granzyme B. In addition, blocking CD96-CD155 interaction specifically increases lysis of HepG2 cells by NK cells. HCC patients with high level of CD96 or CD155 expression within tumor are strongly associated with deteriorating disease condition, shorter disease-free survival (DFS) and overall survival (OS) times. Patients with higher cumulative percentage of CD96+ NK cells within tumor also exhibit shorter DFS. High plasma level of TGF-β1 in HCC patients up-regulates CD96 expression and dynamically shifts the balance between CD96, TIGIT and CD226 in NK cells. Blocking TGF-β1 specifically restores normal CD96 expression and reverses the dysfunction of NK cells. Conclusion: these findings indicate that human intratumoral CD96+ NK cells are functionally exhausted and patients with higher intratumoral CD96 expression exhibit poorer clinical outcomes. Blocking CD96-CD155 interaction or TGF-β1 restores NK cell immunity against tumors by reversing NK cell exhaustion, suggesting a possible therapeutic role of CD96 in fighting liver cancer. This article is protected by copyright. All rights reserved.