A platform for SpyCatcher conjugation to native antibodies

Area of Science:

• Bioconjugation Chemistry
• Protein Engineering
• Therapeutic Antibody Development

Background:

• Protein-antibody conjugates are crucial for targeted therapies.
• Existing 'off-the-shelf' antibody conjugation platforms are limited.
• The SpyTag/SpyCatcher system offers robust protein conjugation via isopeptide bonds.

Purpose of the Study:

• To develop a novel platform for rapid and stable 'off-the-shelf' antibody conjugation.
• To fuse SpyCatcher with native antibodies using specialized linkers.
• To demonstrate the platform's efficiency and broad applicability.

Main Methods:

• Fusion of SpyCatcher with native antibodies via cysteine-reactive tetra-divinylpyrimidine (TetraDVP)-SpyTag linkers.
• Conjugation of trastuzumab (HER2-targeting antibody) to SpyCatcher proteins.
• Testing conjugation efficiency and binding affinity on HER2-positive cells.
• Conjugation of additional IgG1 and IgG4 antibodies (durvalumab, brentuximab, cetuximab, gemtuzumab).
• Development of a scalable solid-phase synthesis for TetraDVP linkers.

Main Results:

• Robust protein-antibody conjugates were produced with high conversion rates.
• Conjugates retained specific binding to HER2-positive cells.
• The TetraDVP-SpyTag linker demonstrated broad applicability across multiple native IgG1 and IgG4 antibodies.
• Scalable solid-phase synthesis of TetraDVP linkers achieved high yields and purity.

Conclusions:

• The developed platform enables precise, single-step antibody bioconjugation.
• This method significantly advances protein-antibody conjugate synthesis for therapeutics and diagnostics.
• The platform offers a powerful tool for antibody-based drug development.