ZL-1222, a PD-1-targeted potency-reduced IL-12 immunocytokine, overcomes PD-1 resistance and enhances antitumor immunity with an accepted safety profile

Immune checkpoint blockade (ICB) restores T-cell function in the tumor microenvironment (TME), yet many patients with solid tumors exhibit limited response due to resistance. Interleukin-12 (IL-12) is particularly effective at reprogramming the TME and promoting durable, antigen-spreading immunity; however, its clinical use is constrained by systemic toxicity. To address this, ZL-1222, a next-generation IL-12 immunocytokine, is being developed having two single-chain anti-PD-1 antibody fragments that are attached to the N terminus of a knobs-into-holes silenced human IgG 1 Fc domain. An IL-12 mutein engineered to decrease binding to the IL-12 receptor is fused to the C-terminus of one Fc arm. By targeting PD-1⁺ tumor-infiltrating lymphocytes (TILs), ZL-1222 delivers IL-12 directly to T/NK cells within the TME, enabling cis-activation that enhances antitumor immunity while minimizing systemic exposure and associated toxicities. In vitro, ZL-1222 demonstrates strong PD-1 antagonist activity. The IL-12 mutein shows a ~1000-fold reduction in IFN-gamma production in a human mixed lymphocyte reaction (MLR), a standard functional readout of IL-12 activity. In the presence of PD-1 + cells, ZL-1222 synergistically increases IFN-gamma production and tumor-cell killing, outperforming the combination of anti-PD-1 antibody plus IL-12 mutein. In vivo, the ZL-1222 surrogate (m45) shows strong antitumor activity in PD-1-resistant B16F10.OVA and EMT6 syngeneic models. In the CT26 model, depletion of CD8 + T cells or NK cells, but not CD4 + T cells, attenuates the antitumor effects of m45, indicating that tumor-growth-inhibition (TGI) depends on CD8 + T and NK cells. The surrogate also induces antitumor immune memory and inhibits tumor growth in a dose-dependent manner (0.03 to 3 mg/kg) with a strong pharmacokinetics (PK)/TGI correlation. Minimal mouse body-weight loss (<10%) was observed, consistent with the absence of severe treatment-related adverse effects. In a pilot toxicity study, cynomolgus monkeys tolerated single doses of ZL-1222 up to 10 mg/kg without evidence of systemic cytokine storm (IL-6 < 50 pg/mL; TNFalpha not detectable). In conclusion, ZL-1222, a PD-1-targeted, potency-reduced IL-12 immunocytokine, achieves potent antitumor activity in PD-1-resistant models via delivery to PD-1⁺ TILs, engaging CD8⁺ T and NK cells in the TME while minimizing systemic toxicity. It induces durable immune memory, demonstrates dose-dependent efficacy with robust PK/TGI correlation, and shows favorable tolerability in non-human primates, supporting its potential as a next-generation immunotherapy for solid tumors.