Enhanced B7-H4 expression in gliomas with low PD-L1 expression identifies super-cold tumors

BACKGROUND: Characterizing expression profiles of different immune checkpoint molecules are promising for personalized checkpoint inhibitory immunotherapy. Gliomas have been shown as potential targets for immune checkpoint inhibitors recently. Our study was performed to determine coexpression levels of two major B7 immune regulatory molecules programmed death ligand 1 (PD-L1) and B7-H4, both of which have been demonstrated to inhibit antitumor host immunity in gliomas. METHODS: We assessed tumor tissues from stage II–IV primary gliomas (n=505) by immunohistochemistry (IHC) for protein levels of both PD-L1 and B7-H4. Gene coexpression analysis assessing clusters based on extent of PD-L1/B7-H4 classifier genes expression were investigated in two transcriptome datasets (The Cancer Genome Atlas and Chinese Glioma Genome Atlas). In addition, levels of immune cell infiltrates were estimated with IHC and RNA-seq data for assessing the tumor immune microenvironment of PD-L1/B7-H4 subgroups. RESULTS: High expression of PD-L1 and B7-H4 in gliomas was 23% and 20%, respectively, whereas coexpression of two proteins at high levels was limited to 2% of the cases. Comparable results were seen in RNA-seq datasets where PD-L1 mRNA expression levels negatively correlated with that of B7-H4. Gene coexpression modules clustered within each grade of gliomas demonstrated lack of double-high modules (cluster with high expression of both PD-L1 and B7-H4 classifier genes). B7-H4 mRNA expression levels showed negative correlation with extent of immune cell infiltration and High-B7-H4 module gliomas (high B7-H4 but low PD-L1 classifier genes expression) had less tumor-infiltrating lymphocytes (TILs) and tumor-associated macrophages (TAMs). IHC assessment also showed few TILs and TAMs in High-B7-H4 subgroup gliomas. CONCLUSIONS: The majority of gliomas express PD-L1 or B7-H4, however, coexpression of both at high levels is minimal. The high-B7-H4 patients could be considered as ‘super-cold’ gliomas with significantly deficient in TILs, suggesting that B7-H4 might inhibit T-cell trafficking into the central nervous system. This study demonstrated that PD-L1 and B7-H4 may serve as mutually compensatory immune checkpoint molecules in gliomas for immune targeted or active-specific immunotherapy. The distinct B7-H4 pathways modulating T-cell function and immune evasion in glioma patients deserved to be further explored in the future during immunotherapy.