GENE-THERAPY WITH A SINGLE-CHAIN INTERLEUKIN-12 FUSION PROTEIN INDUCES T-CELL-DEPENDENT PROTECTIVE IMMUNITY IN A SYNGENEIC MODEL OF MURINE NEUROBLASTOMA

A major goal of tumor immunotherapy is the effective eradication of es tablished metastases associated with the induction of a T cell-mediate d protective immunity. We achieved this in a poorly immunogenic murine neuroblastoma model by gene therapy with a single chain interleukin 1 2 (scIL-12) fusion protein that assures equal expression of its p35 an d p40 subunits. Thus, NXS2 hybrid neuroblastoma cells (C1300 x dorsal root ganglion cells), which form experimental bone marrow and liver me tastases in syngeneic A/J mice, were transduced with a gene encoding m urine interleukin 12, monomerized by introduction of a protein linker between the p35 and p40 protein chains of this heterodimeric cytokine. We demonstrate for the first time that subcutaneous vaccination with these transduced cells induces a protective immunity, as indicated by the complete absence of liver and bone marrow metastasis after challen ge with NXS2 wild-type tumor cells. Furthermore, vaccination of animal s with established liver and bone marrow metastases completely eradica ted liver metastases and suppressed bone marrow metastases. The local and systemic immune response against scIL-12-transduced NXS2 cells is largely dependent on CD8(+) T cells. This was demonstrated in vivo by depletion of immunocompetent A/J mice with monoclonal anti-CD4 and ant i-CD8 antibodies and in vitro by specific major histocompatibility com plex, class I-restricted CD8(+) T cell-mediated killing of NXS2 and th eir parental C1300 neuroblastoma cells. In conclusion, we demonstrate successful anti-tumor immunotherapy with an scIL-12 fusion protein tha t could facilitate clinical application of interleukin 12 gene therapy .