Integrated analysis identifies CD276 in fibroblasts as a malignancy predictor and regulator of neutrophil infiltration in hepatoblastoma

Background: Hepatoblastoma is the most prevalent liver cancer in children. Immunotherapy targeting immune checkpoint molecules has become pivotal in various cancer treatments. However, the clinical significance of immune checkpoint ligands in hepatoblastoma remains largely unclear due to various challenges. This study sought to first characterize the expression profile of the immune checkpoint ligand CD276 in hepatoblastoma and assess its potential as a predictor of malignant characteristics and regulator of neutrophil infiltration. Methods: Univariable and multivariable logistic regression analyses were performed to evaluate the clinical significance of immune checkpoint ligands in the bulk RNA-seq dataset and develop a novel predictive model for malignancy. Furthermore, single-cell RNA sequencing (scRNA-seq), immunohistochemistry (IHC), deconvolution analysis, and correlation analysis were employed to characterize the expression pattern of CD276 and explore its influence on the tumor immune microenvironment. Results: The bulk RNA-seq analysis revealed CD276 transcript levels were significantly elevated in hepatoblastoma tissues, especially in patients with more aggressive malignant phenotypes. Furthermore, we developed a predictive model based on a risk score and constructed a user-friendly nomogram to predict patient metastasis by integrating CD276 levels with clinical features. Both scRNA-seq and multiplex immunohistochemistry (mIHC) analyses confirmed that CD276 is highly expressed, predominantly in cancer-associated fibroblasts. Our results also demonstrated that CD276 levels correlate with immune infiltration in hepatoblastoma, and that CD276 regulates CXCL2 to modulate neutrophil infiltration, suggesting a potential mechanism underlying the role of CD276 in hepatoblastoma malignancy. In addition, we validated that CD276 in cancer-associated fibroblasts promotes hepatoblastoma growth in mice. Conclusions: Our findings highlight the critical role of CD276 in hepatoblastoma malignancy, potentially providing novel insights and therapeutic targets for the development of combinatorial immunotherapy strategies for this disease.