Characterizing Therapeutic Proteins for Drug Discovery and Development

Importance of the topic

Protein therapeutics such as monoclonal antibodies and antibody-drug conjugates offer high specificity and efficacy in treating a wide range of diseases. Advanced structural characterization is critical to ensure safety, efficacy and manufacturability throughout drug discovery and development.

Goals and overview of the study

LifeArc partnered with Thermo Fisher Scientific to integrate a high resolution accurate mass mass spectrometry platform in house. The objective was to accelerate and deepen the characterization of therapeutic proteins using a single flexible workflow covering intact mass analysis, sub-unit analysis, peptide mapping, native mass spectrometry and antibody-drug conjugate assessment.

Methodology and instrumentation

LifeArc implemented a complete biopharmaceutical characterization pipeline on a single platform. Used instrumentation included:

• Thermo Scientific Vanquish Flex Binary UHPLC system
• Thermo Scientific Q Exactive Plus hybrid Quadrupole-Orbitrap mass spectrometer with BioPharma option
• Thermo Scientific BioPharma Finder software

Method development employed weekly tuning and calibration, NISTmAb reference material for system suitability, and standard data acquisition modes: high mass range, protein mode and standard mode.

Main results and discussion

The platform delivered robust, stable performance requiring minimal maintenance. Denaturing intact mass and native mass spectra revealed clear charge envelopes and isoform distributions. Sub-unit analysis achieved isotopic resolution up to 140 000 FWHM at m/z 200, enabling detection of deamidation and other modifications. Peptide mapping workflows provided site-specific information on post-translational modifications and sequence variants. Native SEC-MS experiments characterized noncovalent protein complexes, and high resolution data enabled precise drug-to-antibody ratio (DAR) assessment in antibody-drug conjugates.

Benefits and practical applications

The integrated platform allowed LifeArc to perform comprehensive analyses with microgram sample amounts, reducing turnaround time and outsourcing costs. Walk-up usability enabled rapid training of users across biology, chemistry and biotherapeutics. Detailed structural insights guided process optimization, improved candidate selection and supported regulatory filings.

Future trends and applications

Further integration of separation techniques, native mass spectrometry of higher order assemblies, and advanced data analysis software will extend capabilities for complex biologics and multispecific antibodies. Combining hydrogen deuterium exchange and ion mobility with HRAM MS promises deeper structural and dynamics insights for next generation therapeutics.

Conclusion

By adopting a high resolution accurate mass proteomics platform, LifeArc achieved a seamless, cost efficient workflow for characterizing therapeutic proteins. The solution streamlined intact mass, sub-unit, peptide and native analyses, empowering rapid decision making and advancing the development of biopharmaceutical candidates.

References

No formal literature references were cited in the original document.