Coagulation Factor IX Analysis in Bioreactor Cell Culture Supernatant Predicts Quality of the Purified Product

Overview

Journal Commun Biol

Specialty Biology

Date 2021 Mar 24

PMID 33758337

Citations 2

Authors

Lucia F Zacchi

Dinora Roche-Recinos

Cassandra L Pegg

Toan K Phung

Mark Napoli

Campbell Aitken

Vanessa Sandford

Stephen M Mahler

Yih Yean Lee

Benjamin L Schulz

Christopher B Howard

Affiliations

Soon will be listed here.

Abstract

Coagulation factor IX (FIX) is a complex post-translationally modified human serum glycoprotein and high-value biopharmaceutical. The quality of recombinant FIX (rFIX), especially complete γ-carboxylation, is critical for rFIX clinical efficacy. Bioreactor operating conditions can impact rFIX production and post-translational modifications (PTMs). With the goal of optimizing rFIX production, we developed a suite of Data Independent Acquisition Mass Spectrometry (DIA-MS) proteomics methods and used these to investigate rFIX yield, γ-carboxylation, other PTMs, and host cell proteins during bioreactor culture and after purification. We detail the dynamics of site-specific PTM occupancy and structure on rFIX during production, which correlated with the efficiency of purification and the quality of the purified product. We identified new PTMs in rFIX near the GLA domain which could impact rFIX GLA-dependent purification and function. Our workflows are applicable to other biologics and expression systems, and should aid in the optimization and quality control of upstream and downstream bioprocesses.

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