Abstract 3817: Improved T cell activation bioassays to facilitate the development of bispecific antibodies and engineered T cell immunotherapies

Abstract

T cells play a critical role in cell-mediated immunity and can mediate long-term, antigen-specific, effector and memory responses. A variety of immunotherapy strategies have been developed recently that are aimed at inducing, strengthening or engineering T cell responses. These have emerged as promising approaches for the treatment of diseases such as cancer and autoimmunity. Current methods used to measure TCR-mediated T cell proliferation and cytokine production rely on primary PBMCs as a source of T cells, which must be stimulated via co-culture with APCs or anti-TCR/CD3 antibodies. These assays are laborious and highly variable due to their reliance on donor primary cells, complex assay protocols and unqualified assay reagents. As a result, these assays are difficult to establish in quality-controlled drug development settings. To overcome this barrier, we developed two reporter-based bioluminescent T cell activation bioassays that can be used for the development of bispecific antibodies and engineered T cell immunotherapies. The assays consist of Jurkat T cells genetically engineered to express luciferase downstream of either NFAT or IL-2 response elements. The T cell activation bioassays reflect the mechanisms of action of biologics designed to induce TCR and/or CD28-mediated T cell activation, as demonstrated using anti-CD3 and/or anti-CD28 antibodies as well as blinatumomab, a bispecific antibody that simultaneously binds CD3 expressed on T cells and CD19 expressed on malignant B cells. The bioassays are pre-qualified according to ICH guidelines and demonstrate assay specificity, precision, accuracy and linearity required for routine use in potency and stability studies. Finally, our data illustrate the use of reporter-based T cell activation bioassays for characterizing and measuring the activity of engineered chimeric antigen receptor T cells. Citation Format: Pete Stecha, Denise Garvin, Jim Hartnett, Gopal B. Krishnan, Frank Fan, Mei Cong, Zhi-jie Jey Cheng. Improved T cell activation bioassays to facilitate the development of bispecific antibodies and engineered T cell immunotherapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3817.

Development of a reporter gene method to measure the bioactivity of anti-CD38 × CD3 bispecific antibody

“…In recent years, cell-based reporter gene assays were reported for measuring the bioactivity of monoclonal and bispecific antibodies, with advantages of robust performance and easy operation [ 32–34 ]. Promega Corporation developed a cell-based luciferase reporter gene assay to monitor the Jurkat T cell activation through binding to CD3 on the Jurkat T cells engineered with an nuclear factor of activated T cells response element (NFAT-RE) driving luciferase expression [ 35 , 36 ]. However, the CD38 expressed on the Jurkat T cells interferes the assay when the reporter gene bioassay was performed to measure the bioactivity of the Y150 bsAb.…”