Abstract 322: Nonclinical characterization of the secondary mechanisms of action of PF-08046054 (PDL1V), a novel vedotin ADC directed to PD-L1 that selectively depletes tumor cells without depleting healthy immune cells

PDL1V is a novel vedotin antibody drug-conjugate (ADC) currently in Phase I clinical development for advanced solid tumors as a monotherapy or in combination with pembrolizumab (NCT05208762). The primary mechanism of action (MOA) of PDL1V is direct cytotoxicity to PD-L1-expressing tumor cells via the intracellular delivery of MMAE. While PD-L1 is an established checkpoint protein, pharmacokinetic data suggest steric checkpoint inhibition by PDL1V may be limited by the dose and schedule being evaluated in the clinic. Here, we characterized additional antitumor MOAs of PDL1V, including bystander killing and immunogenic cell death (ICD), and evaluated the effects of PDL1V on immune cells. PDL1V is potently cytotoxic to a variety of PD-L1-positive cancer cell lines in vitro. Using a flow cytometry-based viability assay, we showed that PDL1V also drives dose-dependent bystander effect-based cytotoxicity of PD-L1-negative tumor cells when co-cultured with PD-L1-positive tumor cells. ICD, a regulated activation of immune cell recruitment to the TME, is caused by tumor cell release of damage-associated molecular patterns (DAMPs) such as cell surface chaperone calreticulin (CRT), extracellular adenosine triphosphate (eATP), and High Mobility Group Box 1 protein (HMGB1). PD-L1-positive cancer cells treated with PDL1V in vitro released all three DAMPs, suggesting that targeted delivery of the MMAE payload drives induction of ICD. As an immune checkpoint protein, PD-L1 is expressed on antigen presenting cells and T cells. Depletion of these cells could affect the tolerability of PDL1V as a monotherapy, or limit efficacy in combination with anti-PD-1 therapies. To investigate the potential for immune cell depletion, macrophages, CD4+ T cells, and CD8+ T cells were evaluated in an in vitro fluorescence-based cytotoxicity assay. PDL1V had minimal cytotoxic activity across all three cell-types, despite expression of PD-L1. These data support PDL1V as selectively cytotoxic to PD-L1-positive and negative cancer cells of the tumor microenvironment, and not cytotoxic to PD-L1-positive immune cells involved in natural antitumor immunity. Since T cell-driven antitumor immunity is a key driver for the efficacy of PD-(L)1 blockade, these data suggest PDL1V is a promising combination partner with anti-PD-1 immunotherapies such as pembrolizumab. Future studies will evaluate the effects of PDL1V on immunosuppressive cells, including regulatory T cells and myeloid derived suppressor cells, and the pharmacodynamic effects of PDL1V in syngeneic mouse xenograft models. Ethics Approval: All procedures performed on animals were in accordance with regulations and established guidelines and were reviewed and approved by an Institutional Animal Care and Use Committee or through an ethical review process. Citation Format: Megan Atkins Ramirez, Midori Clarke, Steven Jin, Ellie Moskovitz, Andres Forero-Torres, Garrett Cornelison. Nonclinical characterization of the secondary mechanisms of action of PF-08046054 (PDL1V), a novel vedotin ADC directed to PD-L1 that selectively depletes tumor cells without depleting healthy immune cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 322.