Abstract LB204: Cis-delivery of a potency-reduced IL-12 via an anti-PD-1 single-chain antibody exhibits potent anti-tumor activity

Abstract Immune checkpoint blockade (ICB) works by alleviating immunosuppression on tumor-infiltrating lymphocytes (TILs) and has shown success in treating various cancer types. However, many patients with solid malignancies derive limited clinical benefit due to primary or acquired resistance. Interleukin-12 (IL-12), a proinflammatory cytokine produced by activated antigen-presenting cells, increases IFN-γ production and promotes Th1 cell differentiation, leading to enhanced T and NK cell proliferation and cytotoxicity. Numerous preclinical studies in mice have demonstrated that IL-12 can exert significant anti-tumor activity. However, its clinical application has been hindered by a narrow therapeutic index and severe adverse events associated with systemic IL-12 administration.ZL-1222 is a PD-1-targeted immunocytokine comprising an anti-PD-1 single-chain antibody (ScAb), a potency-reduced IL-12, and a non-effective Fc fragment with specific modifications to extend serum half-life. The potency-reduced IL-12 preferentially supports T cell function with reduced activity on NK cells due to differences in IL-12Rβ1 expression, thereby mitigating systemic toxicity in mice (Glassman et al. 2021). Since PD-1 is predominantly expressed on CD8+ and CD4+ TILs and is enriched in the tumor microenvironment (TME) compared to circulating T cell subsets, PD-1 targeting would further enhances IL-12-mediated T cell activity via a cis-effect within the TME while reducing systemic toxicity.In vitro, ZL-1222 effectively blocks PD-1/PD-L1 signaling in a co-culture/reporter cell assay and enhances mutant IL-12 activity in a PD-1-dependent manner in an IL-12 receptor reporter assay. In a SARS-CoV-2 Spike protein recall assay, ZL-1222 demonstrates significantly higher IFN-γ production in human PBMCs compared to the combination of potency-reduced IL-12 and the αPD-1 ScAb. Additionally, in a co-culture of human cancer cells and primary PBMCs, ZL-1222 demonstrates superior IFN-γ release and tumor killing compared to the combination treatment.In vivo, systemic administration of ZL-1222 mouse surrogates, m34 or m45, shows strong antitumor activity with reduced mouse body weight loss in the B16F10 syngeneic tumor model in human PD-1 knock-in C57Bl/6 mice and the B16F10.OVA syngeneic model in wild-type C57Bl/6 mice compared to treatment with wild type IL-12.These results position ZL-1222 as a promising next-generation IL-12 immunocytokine for cancer immunotherapy, potentially combining potent antitumor activity with improved systemic safety. Citation Format: Linda N Liu. Cis-delivery of a potency-reduced IL-12 via an anti-PD-1 single-chain antibody exhibits potent anti-tumor activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_2):Abstract nr LB204.