Qualification of a LC-HRMS platform method for biosimilar development using NISTmab as a model

Area of Science:

• Analytical Chemistry
• Biopharmaceutical Analysis
• Quality Control

Background:

• Biosimilars offer cost-effective alternatives to biopharmaceuticals.
• Rigorous analytical methods are crucial for ensuring biosimilar consistency and regulatory compliance.
• Liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) is a powerful technique for characterizing complex biologics.

Purpose of the Study:

• To validate an LC-HRMS based peptide mapping method for biosimilar characterization.
• To assess the method's performance according to ICH Q2R1 guidelines using NISTmAb as a reference.
• To ensure the method's suitability for identifying and characterizing monoclonal antibodies in biosimilar development.

Main Methods:

• Peptide mapping using Liquid Chromatography-High Resolution Mass Spectrometry (LC-HRMS).
• Method validation following International Council for Harmonisation (ICH) Q2R1 guidelines.
• Assessment of system suitability, specificity, accuracy, precision, robustness, and carryover using NISTmAb.

Main Results:

• The validated LC-HRMS method demonstrated high accuracy (mass error < 2.42 ppm) and precision (%RSD < 0.000219%) for NISTmAb peptides.
• The method successfully differentiated NISTmAb from similar molecules and spiked conditions, confirming specificity.
• Robustness was confirmed across varying storage conditions, column temperatures, and mobile phase compositions, with negligible carryover.

Conclusions:

• The validated LC-HRMS peptide mapping method is a versatile and reliable tool for biosimilar characterization.
• This method effectively assesses critical quality attributes, including sequence coverage and post-translational modifications (PTMs).
• The established method supports the development and quality assurance of biosimilar monoclonal antibodies.