Establishing a Link Between Amino Acid Sequences and Self-Associating and Viscoelastic Behavior of Two Closely Related Monoclonal Antibodies

Yadav, Sandeep; Sreedhara, Alavattam; Kanai, Sonoko; Li, Jun; Lien, Samantha; Lowman, Henry B.; Kalonia, Devendra S.; Shire, Steven J.

doi:10.1007/s11095-011-0410-0

Cited by 112 publications

(173 citation statements)

References 52 publications

(77 reference statements)

Supporting

Mentioning

156

Contrasting

Order By: Relevance

“…3,4 Such intermolecular interactions increase the propensity of antibody molecules to undergo reversible self-association (RSA) where non-covalent multimers can form at high protein concentrations and then dissociate upon dilution. [5][6][7] The RSA of mAbs presents various pharmaceutical challenges, including the formation of protein aggregation precursors that can lead to the irreversible formation of oligomers. 8,9 In addition, the RSA of mAbs gives rise to a network of associated higher-order species that can affect the viscoelastic properties of the solution, 10 resulting in increased viscosity, [5][6][7]11 solution turbidity, 12 and, under certain conditions, even phase transitions.…”

Section: Introductionmentioning

confidence: 99%

Hydrogen exchange mass spectrometry reveals protein interfaces and distant dynamic coupling effects during the reversible self-association of an IgG1 monoclonal antibody

Arora

Hickey

Majumdar

et al. 2015

mAbs

View full text Add to dashboard Cite

Volkin (2015) Hydrogen exchange mass spectrometry reveals protein interfaces and distant dynamic coupling effects during the reversible self-association of an IgG1 monoclonal antibody, mAbs, 7:3, 525-539, DOI: 10.1080DOI: 10. /19420862.2015 To link to this article: https://doi.org/10. 1080/19420862.2015 There is a need for new analytical approaches to better characterize the nature of the concentration-dependent, reversible self-association (RSA) of monoclonal antibodies (mAbs) directly, and with high resolution, when these proteins are formulated as highly concentrated solutions. In the work reported here, hydrogen exchange mass spectrometry (HX-MS) was used to define the concentration-dependent RSA interface, and to characterize the effects of association on the backbone dynamics of an IgG1 mAb (mAb-C). Dynamic light scattering, chemical cross-linking, and solution viscosity measurements were used to determine conditions that caused the RSA of mAb-C. A novel HX-MS experimental approach was then applied to directly monitor differences in local flexibility of mAb-C due to RSA at different protein concentrations in deuterated buffers. First, a stable formulation containing lyoprotectants that permitted freeze-drying of mAb-C at both 5 and 60 mg/mL was identified. Upon reconstitution with RSA-promoting deuterated solutions, the low vs. high protein concentration samples displayed different levels of solution viscosity (i.e., approx. 1 to 75 mPa.s). The reconstituted mAb-C samples were then analyzed by HX-MS. Two specific sequences covering complementarity-determining regions CDR2H and CDR2L (in the variable heavy and light chains, respectively) showed significant protection against deuterium uptake (i.e., decreased hydrogen exchange). These results define the major protein-protein interfaces associated with the concentration-dependent RSA of mAb-C. Surprisingly, certain peptide segments in the V H domain, the constant domain (C H 2), and the hinge region (C H 1-C H 2 interface) concomitantly showed significant increases in local flexibility at high vs. low protein concentrations. These results indicate the presence of longer-range, distant dynamic coupling effects within mAb-C occurring upon RSA.

show abstract

Section: Introductionmentioning

confidence: 99%

Hydrogen exchange mass spectrometry reveals protein interfaces and distant dynamic coupling effects during the reversible self-association of an IgG1 monoclonal antibody

Arora

Hickey

Majumdar

et al. 2015

mAbs

View full text Add to dashboard Cite

show abstract

“…6g, h, f) despite replacing the charge residues outside the CDR, thereby still exhibiting a lower viscosity. The apparent linkage of viscosity with self-association is still apparent since M-7 and M-10 have lower degree of selfassociation compared to MAb 1, as shown by the corrected weight average molecular weight observed from preparative SE-AUC (Yadav et al 2011c) (Fig. 8b).…”

Section: Dilute Solution Static and Dynamic Light Scattering Measuremmentioning

confidence: 95%

“…The influence of surface charge distribution in governing the different high concentration solution behavior of MAbs 1 and 2 was assessed in recent work by studying the chargeswap mutants of MAb 1 and MAb 2 (Yadav et al 2011c(Yadav et al , 2012a. The details of these charge-swap mutants, including specific amino acid changes, are discussed in Yadav et al (2011c).…”

Section: Dilute Solution Static and Dynamic Light Scattering Measuremmentioning

confidence: 99%

“…The details of these charge-swap mutants, including specific amino acid changes, are discussed in Yadav et al (2011c). In these studies, MAb 1 showed a relatively more heterogeneous charge surface than for MAb 2 wherein the negative surface on the CDR can interact to attract the positively charged regions (Fig.…”

Section: Dilute Solution Static and Dynamic Light Scattering Measuremmentioning

confidence: 99%

“…The measurement was conducted at pH 6.0, histidine hydrochloride buffer in 15 and 150 mM ionic strength at 12,000 rpm. Reproduced from Yadav et al (2011c) self-association at high concentration, coarse-grained computational models of MAb 1 and MAb 2 were constructed. Two reduced coarse-grained (12-and 26-site) models were constructed for each antibody using either a compact Yshaped or an extended Y-shaped configuration, and coarsegrained molecular dynamics simulations as described by Chaudhri et al (2012) were carried out.…”

Section: Dilute Solution Static and Dynamic Light Scattering Measuremmentioning

confidence: 99%

See 2 more Smart Citations

Assessment and significance of protein–protein interactions during development of protein biopharmaceuticals

Yadav¹,

Li²,

Scherer³

et al. 2013

Biophys Rev

Self Cite

View full text Add to dashboard Cite

Early development of protein biotherapeutics using recombinant DNA technology involved progress in the areas of cloning, screening, expression and recovery/purification. As the biotechnology industry matured, resulting in marketed products, a greater emphasis was placed on development of formulations and delivery systems requiring a better understanding of the chemical and physical properties of newly developed protein drugs. Biophysical techniques such as analytical ultracentrifugation, dynamic and static light scattering, and circular dichroism were used to study protein-protein interactions during various stages of development of protein therapeutics. These studies included investigation of protein self-association in many of the early development projects including analysis of highly glycosylated proteins expressed in mammalian CHO cell cultures. Assessment of protein-protein interactions during development of an IgG1 monoclonal antibody that binds to IgE were important in understanding the pharmacokinetics and dosing for this important biotherapeutic used to treat severe allergic IgE-mediated asthma. These studies were extended to the investigation of monoclonal antibodyantigen interactions in human serum using the fluorescent detection system of the analytical ultracentrifuge. Analysis by sedimentation velocity analytical ultracentrifugation was also used to investigate competitive binding to monoclonal antibody targets. Recent development of high concentration protein formulations for subcutaneous administration of therapeutics posed challenges, which resulted in the use of dynamic and static light scattering, and preparative analytical ultracentrifugation to understand the self-association and rheological properties of concentrated monoclonal antibody solutions.

show abstract

Application of Biophysical and High‐Throughput Methods in the Preformulation of Therapeutic Proteins—facts and Fictions

Abdul‐Fattah

Mahler

2014

Biophysical Methods for Biotherapeutics

View full text Add to dashboard Cite

Establishing a Link Between Amino Acid Sequences and Self-Associating and Viscoelastic Behavior of Two Closely Related Monoclonal Antibodies

Abstract: Exposed charged residues in the CDR of MAb1 are critical in determining the self-associating and highly viscous behavior observed at high concentrations.

Cited by 112 publications

References 52 publications

Hydrogen exchange mass spectrometry reveals protein interfaces and distant dynamic coupling effects during the reversible self-association of an IgG1 monoclonal antibody

Hydrogen exchange mass spectrometry reveals protein interfaces and distant dynamic coupling effects during the reversible self-association of an IgG1 monoclonal antibody

Assessment and significance of protein–protein interactions during development of protein biopharmaceuticals

Application of Biophysical and High‐Throughput Methods in the Preformulation of Therapeutic Proteins—facts and Fictions

Contact Info

Product

Resources

About