Abstract 3274: Novel cell-based bioassays for mono- and multi-specific antibodies targeting PD-1/PD-L1 for combination immunotherapy

Abstract Programmed cell death protein 1 (PD-1) is an immune checkpoint (IC) receptor that negatively regulates T cell function. Blocking the interaction of PD-1 with its ligands PD-L1 or PD-L2 unleashes the immune system and enhances anti-tumor responses. Immunotherapy using PD-1 or PD-L1 blocking antibodies has led to a paradigm shift in cancer drug discovery, due to its durable effect against a wide variety of cancers. However, despite its tremendous clinical success in cancer treatment, a significant fraction of patients and tumor types remain unresponsive to this therapy. This has led to the development of combination therapies linking PD-1 checkpoint inhibitors with inhibitors of several other IC receptors. A major challenge in immunotherapy drug development is the lack of quantitative and reproducible functional assays. Existing methods rely heavily on primary immune cells are highly variable and labor-intensive. Here, we report the development of a suite of cell line-based reporter bioassays for monoclonal antibodies targeting human PD-1/PD-L1, or bispecific antibodies targeting PD-1/PD-L1 and a co-stimulatory receptor (e.g., 4-1BB, OX40, ICOS) or an immune inhibitory receptor (e.g., CTLA-4, LAG-3, TIGIT). A mouse PD-1/PD-L1 assay and a human PD-1/PD-L2 assay are also developed and can be used in parallel with human PD-1/PD-L1 assay to enable preclinical studies and assess assay specificity. Each of these assays consist of two engineered cell lines: a T effector cell line that express a luciferase reporter driven by specific promoter/response elements responding to the intracellular signals mediated by the T cell receptor (TCR), with modulation from PD-1 and the second immune receptor, and an engineered artificial antigen presenting cell (aAPC). Importantly, these combination bioassays demonstrate the synergetic effect of PD-1 blockade with blockade of a second IC inhibitor receptor or activation of a costimulatory receptor, thus reflecting the mechanisms of action for the therapeutic antibodies of interest. The bioassays are sensitive and quantitative, are prequalified according to ICH guidelines, and demonstrate the performance characteristics (specificity, precision, accuracy, and linearity) to be used as potency assays for product release and stability studies during immunotherapy drug development. Citation Format: Jun Wang, Julia Gilden, Denise Garvin, Pete Stecha, Jim Hartnett, Frank Fan, Mei Cong, Jamison Grailer. Novel cell-based bioassays for mono- and multi-specific antibodies targeting PD-1/PD-L1 for combination immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3274.