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Rapid global characterization of immunoglobulin G1 following oxidative stress.

ABSTRACT: Although peroxide and leachable metal-induced chemical modifications are among the most important quality attributes in bioprocess development, there is no mainstream characterization method covering all common modifications theoretically possible on therapeutic proteins that also gives consistent results quickly. Here, we describe a method for rapid and consistent global characterization of leachable metals- or peroxide-stressed immunoglobulin (Ig) G1 monoclonal antibodies (mAbs). Using two independent protease digestions, data-independent acquisition and data-dependent acquisition liquid chromatography high-resolution mass spectrometry, we monitored 55 potential chemical modifications on trastuzumab, a humanized IgG1 mAb. Processing templates including all observed peptides were developed on Skyline to consistently monitor all modifications throughout the stress conditions for both enzymatic digestions. The Global Characterization Data Processing Site, a universal automated data processing application, was created to batch process data, plot modification trends for peptides, generate sortable and downloadable modification tables, and produce Jmol code for three-dimensional structural models of the analyzed protein. In total, 53 sites on the mAb were found to be modified. Oxidation rates generally increased with the peroxide concentration, while leachable metals alone resulted in lower rates of modifications but more oxidative degradants. Multiple chemical modifications were found on IgG1 surfaces known to interact with Fc?RIII, complement protein C1q, and FcRn, potentially affecting activity. The combination of Skyline templates and the Global Characterization Data Processing Site results in a universally applicable assay allowing users to batch process numerous modifications. Applying this new method to stability studies will promote a broader and deeper understanding of stress modifications on therapeutic proteins.

SUBMITTER: Chen Y

PROVIDER: S-EPMC6748588 | biostudies-literature | 2019 Aug/Sep

REPOSITORIES: biostudies-literature

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Rapid global characterization of immunoglobulin G1 following oxidative stress.

Chen Yao Y Doud Emma E Stone Todd T Xin Lun L Hong Wei W Li Yunsong Y

mAbs 20190704 6

Although peroxide and leachable metal-induced chemical modifications are among the most important quality attributes in bioprocess development, there is no mainstream characterization method covering all common modifications theoretically possible on therapeutic proteins that also gives consistent results quickly. Here, we describe a method for rapid and consistent global characterization of leachable metals- or peroxide-stressed immunoglobulin (Ig) G1 monoclonal antibodies (mAbs). Using two ind ...[more]

PMID: 31156028

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Project description:Background and objectivesInfant weight patterns predict subsequent weight outcomes. Rapid infant weight gain, defined as a >0.67 increase in weight-for-age z-score (WAZ) between two time points in infancy, increases obesity risk. Higher oxidative stress, an imbalance between antioxidants and reactive oxygen species, has been associated with low birthweight and paradoxically also with later obesity. We hypothesized that prenatal oxidative stress may also be associated with rapid infant weight gain, an early weight pattern associated with future obesity.MethodsWithin the NYU Children's Health and Environment Study prospective pregnancy cohort, we analyzed associations between prenatal lipid, protein, and DNA urinary oxidative stress biomarkers and infant weight data. Primary outcome was rapid infant weight gain (>0.67 increase in WAZ) between birth and later infancy at the 8 or 12 month visit. Secondary outcomes included: very rapid weight gain (>1.34 increase in WAZ), low (<2500 g) or high (≥4000 g) birthweight, and low (< -1 WAZ) or high (>1 WAZ) 12 month weight.ResultsPregnant participants consented to the postnatal study (n = 541); 425 participants had weight data both at birth and in later infancy. In an adjusted binary model, prenatal 8-iso-PGF2α, a lipid oxidative stress biomarker, was associated with rapid infant weight gain (aOR 1.44; 95% CI: 1.16, 1.78, p = 0.001). In a multinomial model using ≤0.67 change in WAZ as a reference group, 8-iso-PGF2α was associated with rapid infant weight gain (defined as >0.67 but ≤1.34 WAZ; aOR 1.57, 95% CI: 1.19, 2.05, p = 0.001) and very rapid infant weight gain (defined as >1.34 WAZ; aOR 1.33; 95% CI: 1.02, 1.72, p < 0.05) Secondary analyses detected associations between 8-iso-PGF2α and low birthweight outcomes.ConclusionsWe found an association between 8-iso-PGF2α, a lipid prenatal oxidative stress biomarker, and rapid infant weight gain, expanding our understanding of the developmental origins of obesity and cardiometabolic disease.

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Rapid global characterization of immunoglobulin G1 following oxidative stress.

Publications

Rapid global characterization of immunoglobulin G1 following oxidative stress.

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