Development of Non-Cell Based Potency Assays for Bispecific Antibodies

Purpose

In recent years, the development of Bispecific Antibodies (BsAb) has become an increasingly explored approach in cancer immunotherapy. Unlike traditional chemotherapeutics which indiscriminately kill all rapidly dividing cells, BsAb immunotherapy is a targeted approach to cancer treatment. Since 2014, nine BsAbs have been approved by the FDA, including 2 approvals in 2023 for the treatment of B-cell lymphoma. Potency assays are essential in the development of biologics such as BsAbs. During the development of BsAbs, potency assays measuring the ability of the drug to bind to its intended targets are a critical part of the manufacturing process. However, the development of such assays for BsAbs is complicated by the need to effectively assess interactions between the BsAb and both target antigens. In this study, two potency assays were developed for a Phase I BsAb, henceforth referred to as Antibody X. Antibody X was designed to bridge effector T-cells and target tumor cells to promote T-cell activation and the subsequent lysis of tumor cells. Antibody X contains antigen binding sites targeting a cell surface protein X (CSPX) on T-cells, and a cell surface protein Y (CSPY) on certain tumor cells (Figure 1). CSPX plays a vital role in Tcell activation while CSPY is a tumor-associated antigen expressed in various solid tumor types. To align with the mechanism of action (MOA) of Antibody X, two potency assays were developed to measure the binding of the BsAb to each target antigen independently. Following assay development, validation was completed for both the CSPX- and CSPY-based potency assays and facilitated continuation of product development.