In Situ Tracking of Membrane HER2- Antibody–Drug Conjugate Interaction with Surface PlasmonResonance Imaging

Abstract

Abstract Antibody–drug conjugates (ADCs) rely on specific recognition of tumor-associated membrane receptors to achieve targeted intracellular drug delivery, yet in situ characterization of their interaction dynamics remains limited. Here, we develop a single-cell plasmonic imaging platform to quantitatively resolve the interaction between the membrane human epidermal growth factor receptor 2 (HER2) and HER2-targeted therapeutics. This label-free approach enables continuous monitoring of the molecular interaction process, allowing real-time extraction of detailed binding kinetics. Using this platform, we systematically compare the binding behaviors of the HER2-targeting antibody trastuzumab (Herceptin) and clinically relevant ADCs (T-DM1 and T-DXd), revealing distinct kinetic signatures associated with the drug conjugation. Analysis across cell lines with different HER2 expressions reveals that increased receptor density does not necessarily enhance binding stability, suggesting a potential trade-off between receptor availability and effective interaction dynamics. To further evaluate the potential capability of tracking membrane-associated dynamics, polystyrene nanoparticle probes were employed to validate real-time imaging of endocytosis dynamics, distinguishing uptake behavior in live versus fixed cells.This work establishes cell-based plasmonic imaging as a quantitative and mechanistic approach for evaluating ADC-receptor interactions in situ, offering valuable insights for rational ADC design and precision therapeutic optimization.