Unique Impacts of Methionine Oxidation, Tryptophan Oxidation, and Asparagine Deamidation on Antibody Stability and Aggregation

Alam, Magfur E.; Slaney, Thomas R.; Wu, Lina; Das, Tapan K.; Kar, S.; Barnett, Gregory V.; Leone, Anthony; Tessier, Peter M.

doi:10.1016/j.xphs.2019.10.051

Cited by 42 publications

(19 citation statements)

References 76 publications

(98 reference statements)

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“…The vast excess of untapped theoretical diversity means that additional motifs can be easily eliminated from future libraries, as they become identified. Noteworthy exceptions are methionine and tryptophan oxidation, 74 since removal of every CDR containing one of these amino acids would dramatically reduce final diversity.…”

Section: Discussionmentioning

confidence: 99%

Drug-like antibodies with high affinity, diversity and developability directly from next-generation antibody libraries

Teixeira

Erasmus

D’Angelo

et al. 2021

mAbs

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Therapeutic antibodies must have “drug-like” properties. These include high affinity and specificity for the intended target, biological activity, and additional characteristics now known as “developability properties”: long-term stability and resistance to aggregation when in solution, thermodynamic stability to prevent unfolding, high expression yields to facilitate manufacturing, low self-interaction, among others. Sequence-based liabilities may affect one or more of these characteristics. Improving the stability and developability of a lead antibody is typically achieved by modifying its sequence, a time-consuming process that often results in reduced affinity. Here we present a new antibody library format that yields high-affinity binders with drug-like developability properties directly from initial selections, reducing the need for further engineering or affinity maturation. The innovative semi-synthetic design involves grafting natural complementarity-determining regions (CDRs) from human antibodies into scaffolds based on well-behaved clinical antibodies. HCDR3s were amplified directly from B cells, while the remaining CDRs, from which all sequence liabilities had been purged, were replicated from a large next-generation sequencing dataset. By combining two in vitro display techniques, phage and yeast display, we were able to routinely recover a large number of unique, highly developable antibodies against clinically relevant targets with affinities in the subnanomolar to low nanomolar range. We anticipate that the designs and approaches presented here will accelerate the drug development process by reducing the failure rate of leads due to poor antibody affinities and developability. Abbreviations: AC-SINS: affinity-capture self-interaction nanoparticle spectroscopy; CDR: complementarity-determining region; CQA: critical quality attribute; ELISA: enzyme-linked immunoassay; FACS: fluorescence-activated cell sorting; Fv: fragment variable; GM-CSF: granulocyte-macrophage colony-stimulating factor; HCDR3: heavy chain CDR3; IFN2a: interferon α-2; IL6: interleukin-6; MACS: magnetic-activated cell sorting; NGS: next generation sequencing; PCR: polymerase chain reaction; SEC: size-exclusion chromatography; SPR: surface plasmon resonance; TGFβ-R2: transforming growth factor β-R2; VH: variable heavy; VK: variable kappa; VL: variable light; Vl: variable lambda;

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Section: Discussionmentioning

confidence: 99%

Drug-like antibodies with high affinity, diversity and developability directly from next-generation antibody libraries

Teixeira

Erasmus

D’Angelo

et al. 2021

mAbs

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“…Trp oxidation can cause color changes due to kynurenine formation, 72 reduce physical stability, 73 and oxidation of Trp residues in CDR loops can reduce binding affinity. 65 Structural flexibility, solvent exposure, secondary structure, and side-chain and backbone conformations affect Trp oxidation.…”

Section: Oxidationmentioning

confidence: 99%

In silico prediction of post-translational modifications in therapeutic antibodies

Vatsa¹

2022

mAbs

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Monoclonal antibodies are susceptible to chemical and enzymatic modifications during manufacturing, storage, and shipping. Deamidation, isomerization, and oxidation can compromise the potency, efficacy, and safety of therapeutic antibodies. Recently, in silico tools have been used to identify liable residues and engineer antibodies with better chemical stability. Computational approaches for predicting deamidation, isomerization, oxidation, glycation, carbonylation, sulfation, and hydroxylation are reviewed here. Although liable motifs have been used to improve the chemical stability of antibodies, the accuracy of in silico predictions can be improved using machine learning and molecular dynamic simulations. In addition, there are opportunities to improve predictions for specific stress conditions, develop in silico prediction of novel modifications in antibodies, and predict the impact of modifications on physical stability and antigen-binding.

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“…Especially, sulfonic acid has been reported to bind with calcium ion and CaOx crystals ( 60 , 61 ). Moreover, methionine oxidation of immunoglobulin, one of the high-abundance urinary proteins, could increase the protein hydrophilic properties and affinity to cationic surface ( 62 ), and tryptophan oxidation could promote immunoglobulin aggregation ( 63 ). Therefore, alterations of protein structure and ionic charge could also unveil the calcium- and/or oxalate-binding sites and enhance the crystal–protein complex formation that would enhance the stone nidus formation.…”

Section: Discussionmentioning

confidence: 99%

Oxidative Modifications Switch Modulatory Activities of Urinary Proteins From Inhibiting to Promoting Calcium Oxalate Crystallization, Growth, and Aggregation

Chaiyarit

Thongboonkerd

2021

Molecular & Cellular Proteomics

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This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

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Unique Impacts of Methionine Oxidation, Tryptophan Oxidation, and Asparagine Deamidation on Antibody Stability and Aggregation

Cited by 42 publications

References 76 publications

Drug-like antibodies with high affinity, diversity and developability directly from next-generation antibody libraries

Drug-like antibodies with high affinity, diversity and developability directly from next-generation antibody libraries

In silico prediction of post-translational modifications in therapeutic antibodies

Oxidative Modifications Switch Modulatory Activities of Urinary Proteins From Inhibiting to Promoting Calcium Oxalate Crystallization, Growth, and Aggregation

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