Beyond Monospecifics: Analytical Innovations for Bispecific Antibody Development

Mahmood Haj-Yahya, Analytical Method , Teva Pharmaceuticals, Netanya, Israel (mahmood.haj-yahya@teva.co.il)

Bispecific antibodies (BsAbs) are a promising class of biotherapeutics capable of engaging two distinct antigens, offering enhanced efficacy and reduced dosing compared to conventional monoclonal antibodies. Their structural complexity, however, presents unique analytical challenges during early development and clone selection.

In this study, we present a multi-tiered analytical framework developed and evaluated at TEVA for the high-throughput characterization of BsAbs at the harvest stage. The study evaluates three complementary platforms: Amplified Luminescent Proximity Homogeneous Assay Linked Immunosorbent Assay (AlphaLISA), Octet RH96 (Bio-Layer Interferometry), and Two-Dimensional High-Performance Liquid Chromatography (2D-HPLC).

AlphaLISA, although rapid and suitable for early screening, was primarily designed to detect heterodimeric BsAbs and not optimized for the detection of homodimeric impurities. As such, it offers direct and specific quantification of heterodimers but demonstrates limited sensitivity in impurity profiling.

In comparison, the Octet-based method demonstrates enhanced utility during the early screening stage, particularly for assessing homodimeric antibody impurities within heterodimeric BsAb samples. Consequently, the Octet-based method has been chosen as the preferred tool for high-throughput initial screening.

Our analytical strategy utilizes the Octet RH96 for initial functional screening followed by 2D-HPLC for high-sensitivity, orthogonal confirmation. The 2D-HPLC method effectively separates heterodimers from homodimeric impurities based on hydrophobicity, providing compositional clarity at the harvest stage.

Together, these methods enable functional screening beyond titer alone, allowing for the identification and selection of optimal minipools and clones with high bispecific integrity. Actual case studies supporting this strategy will be presented.

This integrated approach enhances high-throughput decision-making during clone selection, ensuring the quality and efficacy of BsAb candidates as they progress toward clinical development.

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Short Biography of Presenting Author

Mahmood Haj-Yahya is a Team Leader in the Analytical Development Department at Teva Pharmaceuticals, where he leads a multidisciplinary team focused on the analytical characterization of innovative biologics and biosimilars. His current work centers on the development and implementation of advanced analytical methods for biotherapeutic antibodies, supporting early CMC activities.

With over 15 years of experience in chemical biology and protein science, Mahmood brings deep expertise in chromatography, experimental design, data interpretation, and cross-functional collaboration.

Before joining Teva, Mahmood conducted postdoctoral research at EPFL in Switzerland, where he developed novel approaches for the semisynthesis of posttranslationally modified proteins related to neurodegenerative diseases. He holds a Ph.D. in Bioorganic Chemistry from Ben-Gurion University and has authored 15 peer-reviewed publications and two patents.