B7-H6 orchestrates oncogenic signaling and represents a therapeutic vulnerability in metastatic clear cell renal cell carcinoma

B7-H6 (NCR3LG1) is an immune checkpoint protein belonging to the B7 family of immunomodulatory proteins and serves as a ligand for the natural cytotoxicity triggering receptor NKp30 (NCR3) expressed on natural killer (NK) cells. In addition to its extracellular immunomodulatory domain, B7-H6 transmembrane protein contains a putative immunoreceptor tyrosine-based inhibitory motif (ITIM), as well as SH2- and SH3-binding motifs within its cytoplasmic domain. Here, we investigated the expression and functional impact of B7-H6 in metastatic clear cell renal cell carcinoma (ccRCC). B7-H6 expression was detected on tumor cells as well as on tumor-infiltrated leucocytes (TILs). Expression of B7-H6 on both tumor cells and TILs in the primary tumor was associated with synchronous metastasis (p = 0.003 and p = 0.005, respectively) and lymph node metastasis (p = 0.015 and p = 0.001, respectively). Kaplan-Meier survival analysis revealed that B7-H6 expression on tumor cells was associated with reduced overall survival (p = 0.016) in metastatic ccRCC. In vitro RNA interference-mediated silencing of B7-H6 in renal cancer cell lines decreased cell viability in both two-dimensional and three-dimensional cultures. Conversely, ectopic overexpression of full-length B7-H6 protein enhanced transcriptional activity of AP-1 (Activating Protein-1), G-CSF (Granulocyte-Colony Stimulating Factor) and NF-κB (Nuclear Factor Kappa-light-chain-enhancer of activated B cells), whereas truncating B7-H6 intracellular domain diminished these effects. Together, these results provide novel insights into the role of B7-H6 in cancer progression, and suggest that targeting B7-H6 may represent a promising therapeutic strategy to complement current anticancer treatments. B7-H6 is expressed on tumor cells and tumor-infiltrating leukocytes (TILs) in metastatic clear cell renal cell carcinoma. B7-H6 expression in tumor cells correlates with synchronous metastasis (p = 0.003) and dissemination to lymph nodes (p = 0.015) and reduced overall survival (p = 0.016). B7-H6 expression in TILs correlates with synchronous metastasis (p = 0.005) and dissemination to lymph nodes (p = 0.001). Silencing of B7-H6 reduces renal cancer cell viability in both 2- and 3-dimensional (2D and 3D) culture models. The cytoplasmic domain of B7-H6 modulates AP-1 (Activating Protein-1), G-CSF (Granulocyte-Colony Stimulating Factor) and NF-κB (Nuclear Factor Kappa-light-chain-enhancer of activated B cells) transcriptional activity, supporting a role in intracellular signaling.