Assessing in vivo dynamics of multiple quality attributes from a therapeutic IgG4 monoclonal antibody circulating in cynomolgus monkey

Abstract

Characterization of biopharmaceutical proteins and assessment and understanding of the critical quality attributes (CQAs) is a significant part of biopharmaceutical product development and is routinely performed in vitro. In contrast, systematic analysis of the quality attributes in vivo is not as widespread, although metabolism and clearance of multiple variants of therapeutic proteins administered to non-human primates and human subjects may have a different impact on safety, efficacy and exposure. The major hurdles of such studies are usually sample availability and technical capability. In this study, we used affinity purification coupled with liquid chromatography and mass spectrometric analysis of the digested protein for consistent and simultaneous detection of the full amino acid sequence of a therapeutic IgG4 monoclonal antibody, MAB1. This methodology allowed us to assess in vivo changes of all sequence-related modifications and quality attributes of MAB1 over the duration of a preclinical pharmacokinetic study in cynomolgus monkeys.

Quantitation and pharmacokinetic modeling of therapeutic antibody quality attributes in human studies

“…Previously, we used parallel reaction monitoring (PRM) to quantify peptides using fragment ions from a single parent as a way to further reduce the interference after the affinity purification. 22 In this study, due to the high specificity of affinity purification provided by the anti-id ( Fig. 1B ), MS quantitation based on extracted ion chromatograms (XIC) from full scan MS analysis was sufficient to avoid interferences and provide reliable quantitation over the concentration range of the PK samples ( Fig.…”

“…The results shown in this study are qualitatively similar to our previous cynomolgus monkey study of an IgG4 for the attributes common between the 2 molcules. 22 This suggests that non-human primate data may be predictive for clinical studies in human subjects for common quality attributes of therapeutic IgGs. These findings also propose that different IgG subclasses (IgG4 and IgG1) behave similarly in terms of some quality attributes dynamics in vivo.…”

“… 7-20 More recently, we reported a study in which simultaneous monitoring of multiple attributes of a therapeutic IgG4 monoclonal antibody in cynomolgus monkeys was performed. 22 There studies contribute to a quantitative assessment of impacts of individual therapeutic protein variants in vivo.…”

“…Building upon previous experience, 22 we extended our multiple-attribute profiling to human PK studies. To reduce the endogenous protein background, we utilized an affinity purification procedure using a capturing antibody that can specifically recognize the unique complementarity-determining region (CDR) of a therapeutic human IgG1 MAB2, along with magnetic beads with low non-specific binding properties.…”

A Quadrupole Dalton-based multi-attribute method for product characterization, process development, and quality control of therapeutic proteins

“…Such an approach is typically referred to as multi-attribute method (MAM). 7-10 , 23 , 24 Most of these methods combine high accuracy / high resolution MS with sophisticated data analysis software to identify and quantify various PTMs. These methods have shown comparable results with the traditional methods such as HILIC oligosaccharide analysis, CEX, and capillary electrophoresis sodium dodecyl sulfate (CE-SDS).…”