IMMU-51. Treatment efficacy in murine and human glioma models with brain Compartment-Locked IL-12 does not per se rely on influx of peripheral immune cells and induces distinct transcriptome and secretome changes

Abstract Glioblastoma (GBM) is widely considered unresponsive to immunotherapy due to its immunosuppressive microenvironment and limited immune cell infiltration. However, recent single-cell transcriptomic studies reveal the presence of exhausted, potentially antigen-specific T cells in human GBM. We have developed a locally administered, brain-confined cytokine therapy with minimal systemic exposure using an engineered, neonatal Fc receptor (FcRn)-silenced, Compartment-Locked (CL) IL-12Fc fusion protein. This approach thus relies on presence and reactivation of tumor-resident, IL-12-responsive cells. In syngeneic, preclinical GBM models, intratumorally administered CL IL-12Fc induces robust antitumor efficacy, also under conditions of peripheral lymphopenia (induced via systemic temozolomide) or when T cell influx was pharmacologically inhibited. In 3D-perfused, patient-derived GBM explant cultures—with a largely preserved native tumor microenvironment and without addition of autologous peripheral immune cells—CL IL-12Fc triggered pro-inflammatory reprogramming. Proteomics-based secretome analysis revealed significant upregulation of interferon-gamma (IFNγ) pathway signatures. Single-cell RNA sequencing of GBM tissues prior to culture setup confirmed the presence of resident memory T cells in most patients. Notably, single cell transcriptome analysis of post-treatment explants demonstrated a robust IFNγ response in both tumor-associated macrophages, microglia and tumor cells. Our findings demonstrate that CL IL-12Fc can activate local immune responses within human GBM samples, independent of peripheral immune cell recruitment. This localized immune activation is an important prerequisite for the clinical translation of locally confined cytokine therapy. We aim to validate these findings using spatially resolved multiplex immunofluorescence histology. In summary we provide evidence that CL IL-12Fc can facilitate local immune activation without peripheral immune cell requirement.