The antibody-based delivery of interleukin-12 to solid tumors boosts NK and CD8+ T cell activity and synergizes with immune check-point inhibitors

Abstract

We describe the cloning and characterization of a novel fusion protein (termed L19-mIL12), consisting of murine interleukin-12 in single-chain format, sequentially fused to the L19 anti-body in tandem diabody format. The fusion protein bound avidly to the cognate antigen (the alternatively-spliced EDB domain of fibronectin), retained the activity of the parental cyto-kine and was able to selectively localize to murine tumors in vivo, as shown by quantitative biodistribution analysis. L19-mIL12 exhibited a potent anti-tumor activity in immunocompetent mice bearing CT26 carcinomas and WEHI-164 sarcomas, which could be boosted by combination with check-point blockade, leading to durable cancer eradication. L19-mIL12 also inhibited tumor growth in mice with Lewis lung carcinoma (LLC), but in this case cancer cures could not be obtained, both in monotherapy and in combination. A microscopic analysis and a depletion experiment of tumor-infiltrating leukocytes illustrated the contribution of NK cells and CD8+ T cells for the anti-cancer activity observed in both tumor models. Upon L19-mIL12 treatment, the density of regulatory T cells (Tregs) was strongly increased in LLC, but not in CT26 tumors. A FACS analysis also revealed that the majority of CD8+ T cells in CT26 tumors were specific to the retroviral AH1 antigen.\n\nNOVELTY AND IMPACT STATEMENTIn this study, we describe the generation of a novel fusion protein consisting of murine inter-leukin-12 fused to the L19 antibody (specific to the EDB domain of fibronectin). L19-mIL12 revealed favourable tumor to organ ratios 24h after intravenous administration and it was able to cure 60% of CT26 tumor-bearing mice. From a clinical perspective, the rapid clearance from circulation should ease the administration to patients as infusions could be stopped upon onset of side-effects.