The SPPL3-Defined Glycosphingolipid Repertoire Orchestrates HLA Class I-Mediated Immune Responses

Highlights

• Iterative KO screens reveal a new pathway controlling HLA-I antigen presentation

• SPPL3 suppresses B3GNT5 activity affecting the cell surface GSL repertoire

• B3GNT5-generated GSLs limit the capacity of HLA-I to interact with natural ligands

• Inhibition of GSL synthesis in glioma enhances anti-tumor immune activation

Summary

HLA class I (HLA-I) glycoproteins drive immune responses by presenting antigens to cognate CD8⁺ T cells. This process is often hijacked by tumors and pathogens for immune evasion. Because options for restoring HLA-I antigen presentation are limited, we aimed to identify druggable HLA-I pathway targets. Using iterative genome-wide screens, we uncovered that the cell surface glycosphingolipid (GSL) repertoire determines effective HLA-I antigen presentation. We show that absence of the protease SPPL3 augmented B3GNT5 enzyme activity, resulting in upregulation of surface neolacto-series GSLs. These GSLs sterically impeded antibody and receptor interactions with HLA-I and diminished CD8⁺ T cell activation. Furthermore, a disturbed SPPL3-B3GNT5 pathway in glioma correlated with decreased patient survival. We show that the immunomodulatory effect could be reversed through GSL synthesis inhibition using clinically approved drugs. Overall, our study identifies a GSL signature that inhibits immune recognition and represents a potential therapeutic target in cancer, infection, and autoimmunity.