Imaging and quantifying analysis the binding behavior of PD-L1 at molecular resolution by atomic force microscopy

Area of Science:

• Immunology
• Biophysics
• Cancer Research

Background:

• Immunotherapy is a key cancer treatment.
• Programmed cell death ligand-1 (PD-L1) and programmed cell death protein-1 (PD-1) interactions are crucial in tumor immune evasion.
• Targeting the PD-L1/PD-1 pathway is a major focus in cancer immunotherapy.

Purpose of the Study:

• To quantitatively investigate the molecular interactions between PD-L1, its antibody, and PD-1 using single-molecule force spectroscopy.
• To map the spatial distribution of PD-L1 on cancer cells.
• To understand the physical factors influencing PD-L1 interactions for improved immunotherapy design.

Main Methods:

• Atomic force microscopy-based single-molecule force spectroscopy (SMFS).
• Quantitative measurement of unbinding forces for PD-L1/anti-PD-L1 and PD-L1/PD-1 interactions.
• Force-volume mapping to determine PD-L1 localization on T24 cells.
• Assessment of PD-L1 expression changes after interferon-gamma (INF-γ) treatment.

Main Results:

• The PD-L1/anti-PD-L1 antibody complex showed weaker binding (easier dissociation) compared to the PD-L1/PD-1 interaction.
• Unbinding forces measured on T24 cells were comparable to those on artificial substrates.
• PD-L1 was found to be randomly distributed on the surface of T24 cells.
• Interferon-gamma (INF-γ) treatment significantly increased PD-L1 recognition events, indicating upregulation of PD-L1 expression.

Conclusions:

• The study provides the first quantitative, single-molecule analysis of PD-L1 interactions with its antibody and PD-1.
• Findings reveal differences in binding affinities crucial for understanding immunotherapy efficacy.
• The results offer valuable insights into the physical parameters relevant for designing novel cancer immunotherapies targeting the PD-L1 pathway.