Challenges with Development of a Pharmacokinetics Assay to Measure a Variably Glycosylated Fusion Protein

Abstract

Aim: Development of recombinant fusion proteins as drugs poses unique challenges for bioanalysis. This paper describes a case study of a glycosylated fusion protein, where variable glycosylation, matrix interference and high sensitivity needs posed unique challenges. Results: Six different assay configurations, across four different platforms were evaluated for measurement of drug concentrations. Two platforms that achieved the assay requirements were Simoa HD-1 and immune-capture LC–MS/MS-based assay. Conclusion: Both, Simoa HD-1 and the mass spectrometry-based methods were able to detect total drug by providing the adequate matrix tolerance, required sensitivity and detection of all the various glycosylated fusion proteins to support clinical sample analysis. The mass spectrometry-based method was selected due to robustness and ease of method transfer.

Keywords:

• assay
• PK
• sialic acid
• variants

Acknowledgments

The authors thank M Beardsley, P Day and T Baginski for their contributions to the characterization of the sialic acid variants.

Financial & competing interests disclosure

Funding for this work was provided by Genentech, Inc. Gizette Sperinde, Luna Liu, Keyang Xu, Tracy Bentley, Sid Sukumaran, Jeff Lutman, Catherine Huang, Kathi Williams, Martha Hokom and Sally K. Fischer are employees of Genentech, Inc., and stockholders of the Roche group. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.

Ethical conduct of research

The authors state that they have obtained appropriate institutional review board approval or have followed the principles outlined in the Declaration of Helsinki for all human or animal experimental investigations. In addition, for investigations involving human subjects, informed consent has been obtained from the participants involved.