Targeting cystatin F activation enhances NK cell cytotoxicity in glioblastoma modelsSenjor, Emanuela (Avtor) Habič, Anamarija (Avtor) Švajger, Urban (Avtor) Mitrović, Ana (Avtor) Proj, Matic (Avtor) Porčnik, Andrej (Avtor) Prestor, Borut (Avtor) Jerala, Miha (Avtor) Bošnjak, Matic (Avtor) Gobec, Stanislav (Avtor) Breznik, Barbara (Avtor) Kos, Janko (Avtor) Perišić, Milica (Avtor) glioblastomacystatin F3D modelsIntroduction: Glioblastoma (GBM) is a highly invasive brain tumor with limited treatment options and poor prognosis. Natural killer (NK) cells are key effectors of antitumor immunity, capable of eliminating cancer stem-like cells. However, GBM creates an immunosuppressive microenvironment that limits NK cell function. Here, we identify cystatin F as an immunosuppressive factor involved in regulating NK cell granule-mediated cytotoxicity. Methods: We analyzed cystatin F expression in GBM and its correlation with immune exhaustion markers. NK cell activity was compared between GBM patients and healthy donors. In vitro co-cultures of cystatin F-expressing microglial cells and glioblastoma stem-like cells were used to assess NK cell function. To block cystatin F activation from dimeric to active monomeric form, a small-molecule inhibitor of cathepsin V, the activating protease, was applied. Results: Cystatin F expression correlated with immune exhaustion and suppression markers in GBM. NK cells from patients showed reduced cytotoxicity compared to healthy donors. Co-cultures confirmed that cystatin F-expressing microglia impaired NK cell cytotoxicity, while inhibition of cathepsin V restored NK cell function in standard cytotoxicity assays, 3D spheroids, and microfluidic perfused models. Discussion: These results indicate that cystatin F mediates NK cell suppression in GBM. Targeting its activation enhances NK cell cytotoxicity, offering a potential strategy to improve NK-based immunotherapy for glioblastoma.20252025-11-26 09:56:19Neznano24363UDK: 577ISSN pri članku: 1664-3224DOI: 10.3389/fimmu.2025.1708281COBISS_ID: 258230275sl