Intratumoral IL-12 delivery empowers CAR-T cell immunotherapy in a pre-clinical model of glioblastoma

Giulia Agliardi*, Anna Rita Liuzzi*, Alastair Hotblack, Donatella De Feo, Nicolás Núñez, Cassandra L. Stowe, Ekaterina Friebel, Francesco Nannini, Lukas Rindlisbacher, Thomas A. Roberts, Rajiv Ramasawmy, Iwan P. Williams, Bernard M. Siow, Mark F. Lythgoe, Tammy L. Kalber, Sergio A. Quezada, Martin A. Pule, Sonia Tugues, Karin Straathof§ and Burkhard Becher§. *These authors contributed equally § These authors contributed equally Corresponding authors: Burkhard Becher. E-Mail: [e-mail] Karin Straathof. E-Mail: [e-mail] Glioblastoma multiforme (GBM) is the most common and aggressive form of primary brain cancer, for which effective therapies are urgently needed. Chimeric antigen receptor (CAR)- based immunotherapy represents a promising therapeutic approach, but it is often impeded by highly immunosuppressive tumor microenvironments (TME). Here, in an immunocompetent, orthotopic GBM mouse model, we show that CAR-T cells targeting tumorspecific epidermal growth factor variant III (EGFRvIII) alone fail to control fully established tumors but, when combined with a single, locally delivered dose of IL-12, achieve durable antitumor responses. IL-12 not only boosts cytotoxicity of CAR-T cells, but also reshapes the TME, driving increased infiltration of proinflammatory CD4+ T cells, decreased numbers of regulatory T cells (Treg), and activation of the myeloid compartment. Importantly, the immunotherapy-enabling benefits of IL-12 are achieved with minimal systemic effects. Our findings thus show that local delivery of IL-12 may be an effective adjuvant for CAR-T cell therapy for GBM.