Abstract 4344: Interferon-α and PD-1 blockade mutually mask each other in a tumor-activated manner

Abstract (Purpose) Immune checkpoint blockade (ICB) therapies have revolutionized cancer treatment but are often constrained by tumor immune evasion through the loss of MHC I antigen presentation. Type I interferons exert potent antitumor effects that may convert non-responders into responders; however, their systemic administration is limited by severe immune-related adverse events (irAEs). To improve efficacy without compromising safety, we developed a tumor-activated IFNα-PD-1 that uses dual masking to keep both elements inactive systemically. (Methods) The blocking technology attenuates molecular bioactivity by integrating IFNα into the frameworks of anti-PD-1 antibodies. Through this dual-masking design, both cytokine and antibody functions are sterically constrained, preventing premature activation in circulation. Reactivation occurs upon proteolytic cleavage of a tumor-selective linker, a process termed spatial reclasp, which restores molecular flexibility and releases mutual steric hindrance between IFNα and the antibody. This conformational unmasking enables simultaneous recovery of cytokine signaling and checkpoint blockade exclusively within the tumor microenvironment. (Results) In ELISA assays, the inactivated lockers exhibited more than 20-fold reductions in antigen-binding activity relative to the parental antibodies, whereas protease activation restored binding to levels comparable with the parental forms. The activated fusion induced marked upregulation of MHC I expression in melanoma cells, confirming spatially restricted IFNα signaling. In vivo, the IFNα-PD1 fusion demonstrated superior tumor suppression and enhanced intratumoral T cell infiltration with minimal systemic toxicity. (Conclusions) This work establishes IFNα as a tumor-activated checkpoint locker that restores antitumor immunity while mitigating systemic toxicity. The dual-masking architecture provides a generalizable framework for engineering conditionally active immuno-oncology therapeutics, supporting future plug-and-play applications across ICB modalities and enabling safer, more potent next-generation cancer immunotherapies. Citation Format: Shi-Wei Chao, Chih-Hung Chuang. Interferon-α and PD-1 blockade mutually mask each other in a tumor-activated manner [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 4344.