Domain contributions to antibody retention in multimodal chromatography systems

Robinson, Julie; Roush, David J.; Cramer, Steve

doi:10.1016/j.chroma.2018.05.058

Cited by 34 publications

(47 citation statements)

References 41 publications

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“…In contrast, the F C domain exhibited significantly stronger retention on the two MM CEX systems with 0.63 and 0.9 M NaCl required for the elution from the Nuvia cPrime and Capto MMC resins, respectively. The higher retention on Capto MMC as compared to Nuvia cPrime could be due to the more solvent exposed aromatic moiety on the Capto ligand (Robinson, Roush, et al, 2018;. Further, the elevated salt elution concentration seen in the MM resins as compared to the single mode CEX system is likely due to more complex interactions occurring with the F C domain.…”

Section: Resultsmentioning

confidence: 99%

“…One such protein is the F C domain, which is highly conserved across a given class of mAbs and which plays an important role in a number of biotherapeutics such as bispecific antibodies and fusion proteins (Shukla, Hubbard, Tressel, Guhan, & Low, 2007). Robinson et al has examined the domain contributions of mAb binding using a strategic set of chromatographic experiments and shown a shift in domain dominance with pH and resin type (Robinson, Roush, & Cramer, 2018. Gagnon et al demonstrated that the binding of the F C domain was driven by calcium affinity interactions in hydroxyapatite (HA) chromatography (Gagnon, Cheung, & Yazaki, 2009).…”

Section: Introductionmentioning

confidence: 99%

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Identification of Preferred Multimodal Ligand Binding Regions on IgG1 FC using Nuclear Magnetic Resonance and Molecular Dynamics Simulations

Gudhka

Bilodeau

McCallum

et al. 2020

Preprint

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In this study, the binding of multimodal chromatographic ligands to the IgG1 FC domain were studied using nuclear magnetic resonance and molecular dynamics simulations. Nuclear magnetic resonance experiments carried out with chromatographic ligands and a perdeuterated 15N-labeled FC domain indicated that while single mode ion exchange ligands interacted very weakly throughout the FC surface, multimodal ligands interacted with specific clusters of residues with relatively high affinity, forming distinct binding regions on the Fc. The multimodal ligand binding sites on the FC were concentrated in the hinge region and near the interface of the CH2 and CH3 domains. Further, the multimodal binding sites were primarily composed of positively charged, polar and aliphatic residues in these regions, with histidine residues exhibiting some of the strongest binding affinities with the multimodal ligand. Interestingly, comparison of protein surface property data with ligand interaction sites indicated that the patch analysis on FC corroborated molecular level binding information obtained from the nuclear magnetic resonance experiments. Finally, molecular dynamics simulation results were shown to be qualitatively consistent with the nuclear magnetic resonance results and to provide further insights into the binding mechanisms. An important contribution to multimodal ligand-FC binding in these preferred regions was shown to be electrostatic interactions and pi-pi stacking of surface exposed histidines with the ligands. This combined biophysical and simulation approach has provided a deeper molecular level understanding of multimodal ligand-FC interactions and sets the stage for future analyses of even more complex biotherapeutics.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Identification of Preferred Multimodal Ligand Binding Regions on IgG1 FC using Nuclear Magnetic Resonance and Molecular Dynamics Simulations

Gudhka

Bilodeau

McCallum

et al. 2020

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…Porous polymer materials are traditionally synthesized through empirical optimization of a phase inversion process [1][2][3]. Pore formation occurs when a polymer solution undergoes a precipitation process involving temporal and local fluctuations in conditions that lead to a distribution of morphologies and the resultant PSD, in combination with surface chemistry controls the efficiency, selectivity, and capacity for membrane filtration and chromatography [4,5]. In the current state-of-the-art, a desired PSD is targeted through qualitative correlations of parameters in a ternary phase diagram with the process parameters of a membrane production line [6,7].…”

Section: Introductionmentioning

confidence: 99%

“Linking microstructure of membranes and performance”

Sorci

Woodcock

Andersen

et al. 2020

Journal of Membrane Science

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“…Robinson et al. recently reported that retention of mAbs and their fragments on different mixed‐mode adsorbents can be related to distinct surface properties elucidated by these surface property maps. In this study, SAP and EP maps were used in an attempt to interpret the chromatographic results.…”

Section: Introductionmentioning

confidence: 99%

“…SAP and EP maps have previously been shown to be powerful tools for explaining selectivity of different mixed-mode adsorbents for antigen binding fragments differing in hydrophobicity and surface charge [25,26]. Robinson et al [27] recently reported that retention of mAbs and their fragments on different mixed-mode adsorbents can be related to distinct surface properties elucidated by these surface property maps. In this study, SAP and EP maps were used in an attempt to interpret the chromatographic results.…”

Section: Introductionmentioning

confidence: 99%

Optimizing selectivity of anion hydrophobic multimodal chromatography for purification of a single‐chain variable fragment

Sakhnini

Pedersen

León

et al. 2019

Engineering in Life Sciences

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Single‐chain variable fragments (scFv) are widely used in several fields. However, they can be challenging to purify unless using expensive Protein L‐based affinity adsorbents or affinity tags. In this work, a purification process for a scFv using mixed‐mode (MM) chromatography was developed by design of experiments (DoE) and proteomics for host cell protein (HCP) quantification. Capture of scFv from human embryonic kidney 293 (HEK293) cell feedstocks was performed by hydrophobic charge induction chromatography (MEP HyperCel™), whereafter polishing was performed by anion hydrophobic MM chromatography (Capto Adhere™). The DoE designs of the polishing step included both binding and flow‐through modes, the latter being the standard mode for HCP removal. Chromatography with Capto Adhere™ in binding‐mode with elution by linear salt gradient at pH 7.5 resulted in optimal yield, purity and HCP reduction factor of 98.9 > 98.5%, and 14, respectively. Totally, 258 different HCPs were removed, corresponding to 84% of identified HCPs. The optimized conditions enabled binding of the scFv to Capto Adhere™ below its theoretical pI, while the majority of HCPs were in the flow‐through. Surface property maps indicated the presence of hydrophobic patches in close proximity to negatively charged patches that could potentially play a role in this unique selectivity.

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Domain contributions to antibody retention in multimodal chromatography systems

Cited by 34 publications

References 41 publications

Identification of Preferred Multimodal Ligand Binding Regions on IgG1 FC using Nuclear Magnetic Resonance and Molecular Dynamics Simulations

Identification of Preferred Multimodal Ligand Binding Regions on IgG1 FC using Nuclear Magnetic Resonance and Molecular Dynamics Simulations

“Linking microstructure of membranes and performance”

Optimizing selectivity of anion hydrophobic multimodal chromatography for purification of a single‐chain variable fragment

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