Comparative study of trastuzumab modification analysis using mono/multi-epitope affinity technology with LC-QTOF-MS

Area of Science:

• Biochemistry
• Analytical Chemistry
• Materials Science

Background:

• Dynamic tracking of monoclonal antibody (mAb) biotransformation in vivo is essential for drug efficacy.
• Modifications at the antigen-binding site can lead to immune recognition issues, complicating analysis.
• Existing affinity materials face limitations in detecting key site modifications.

Purpose of the Study:

• To develop a novel multi-epitope affinity technology for enhanced in vivo mAb biotransformation analysis.
• To overcome limitations of traditional methods in detecting critical mAb modifications.
• To establish a sensitive bioanalytical platform for tracking trastuzumab modifications in biological fluids.

Main Methods:

• Fabrication of a composite material: metal-organic framework (MOF)@Au@peptide@aptamer.
• Simultaneous immobilization of complementarity determining region (CDR) mimotope peptide (HH24) and non-CDR mimotope aptamer (CH1S-6T).
• Integration with liquid chromatography-quadrupole time of flight-mass spectrometry (LC-QTOF-MS) for analysis.

Main Results:

• The MOF@Au@peptide@aptamer composite demonstrated superior enrichment of trastuzumab variants compared to mono-epitope methods.
• The platform successfully monitored deamidation and isomerization at specific sites (LC-Asn-30, HC-Asn-55).
• Faster modification trends were observed in serum compared to buffer, attributed to biological factors.

Conclusions:

• Multi-epitope affinity technology effectively addresses biases in traditional methods for mAb modification analysis.
• The novel bioanalytical platform enables accurate in vivo tracking of mAb modifications in diverse biological fluids.
• This approach holds significant potential for advancing mAb research and clinical applications.