CDR Repair: A Novel Approach to Antibody Humanization

Dennis, Mark S.

doi:10.1007/978-0-387-76643-0_2

Cited by 11 publications

(5 citation statements)

References 40 publications

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“…Humanization of mAb7 to mAb7.v1 was performed as described previously ( 17 ). The variable regions of mAb7.v1 were cloned into a previously described Fab phage display vector ( 18 ).…”

Section: Methodsmentioning

confidence: 99%

Inhibitory Mechanism of an Allosteric Antibody Targeting the Glucagon Receptor

Mukund¹,

Shang²,

Clarke³

et al. 2013

Journal of Biological Chemistry

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Background: Allosteric regulators of GPCRs provide unique pharmacological properties.Results: The mechanism of allosteric inhibition of the glucagon receptor by an antibody, which is uniquely sensitive to a naturally occurring G40S mutation, is detailed.Conclusion: Allosteric sites on the glucagon receptor extracellular domain regulate receptor activity.Significance: Mechanisms of allosteric regulation of GPCRs aid discovery of drugs with improved selectivity.

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“…Humanization of mAb7 to mAb7.v1 was performed as described previously ( 17 ). The variable regions of mAb7.v1 were cloned into a previously described Fab phage display vector ( 18 ).…”

Section: Methodsmentioning

confidence: 99%

Inhibitory Mechanism of an Allosteric Antibody Targeting the Glucagon Receptor

Mukund¹,

Shang²,

Clarke³

et al. 2013

Journal of Biological Chemistry

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“…15 Similarly, the method of CDR repair utilizes soft-randomization of CDR position with care taken to focus the library on germline amino acids. 24 Finally, a method of generating a binary substitution library between mouse and germline CDR allows for germlining any non-essential CDR residue. 25 Beyond the potential risk of immunogenicity associated with non-human sequence content, the additional constraint of manufacturability must be considered when selecting germlines.…”

Section: Introductionmentioning

confidence: 99%

Beyond CDR-grafting: Structure-guided humanization of framework and CDR regions of an anti-myostatin antibody

Apgar

Mader

Agostinelli

et al. 2016

mAbs

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Antibodies are an important class of biotherapeutics that offer specificity to their antigen, long half-life, effector function interaction and good manufacturability. The immunogenicity of non-human-derived antibodies, which can be a major limitation to development, has been partially overcome by humanization through complementarity-determining region (CDR) grafting onto human acceptor frameworks. The retention of foreign content in the CDR regions, however, is still a potential immunogenic liability. Here, we describe the humanization of an anti-myostatin antibody utilizing a 2-step process of traditional CDR-grafting onto a human acceptor framework, followed by a structure-guided approach to further reduce the murine content of CDR-grafted antibodies. To accomplish this, we solved the co-crystal structures of myostatin with the chimeric (Protein Databank (PDB) id 5F3B) and CDR-grafted antimyostatin antibody (PDB id 5F3H), allowing us to computationally predict the structurally important CDR residues as well as those making significant contacts with the antigen. Structure-based rational design enabled further germlining of the CDR-grafted antibody, reducing the murine content of the antibody without affecting antigen binding. The overall "humanness" was increased for both the light and heavy chain variable regions.

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“…2 One set of techniques seeks to "humanize" an exogenous antibody, e.g., by complementarity determining region (CDR) grafting, 3 resurfacing, 4 specificity-determining residue (SDR) grafting, 5 superhumanization, 6 human string content optimization 7 and various library-based methods. [8][9][10][11] Such techniques have been shown to generate antibodies with significantly reduced immunogenicity. 12 Another set of techniques seeks to generate "fully human" antibodies, e.g., via display of human antibody libraries with phage 13 or yeast, 14 immunization of humanized mice, 15,16 or B cell cloning.…”

Section: Introductionmentioning

confidence: 99%

Antibody humanization by structure-based computational protein design

Choi

Hua

Sentman

et al. 2015

mAbs

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(2015) Antibody humanization by structure-based computational protein design, mAbs, 7:6, 1045-1057, DOI: 10.1080DOI: 10. /19420862.2015 To link to this article: https://doi.org/10. 1080/19420862.2015 Keywords: antibody, computational protein design, human string content, humanization, pareto optimization, protein structure analysisAbbreviations: MHC, major histocompatibility complex; HSC, Human String Content; RMSD, root mean square deviation; BLI, bio-layer interferometry; IDT, Integrated DNA Technologies; PBS, phosphate-buffered saline; PBS-T, PBS C 0.1% Tween-20; PBS-F, PBS C 0.1% Bovine serum albuminAntibodies derived from non-human sources must be modified for therapeutic use so as to mitigate undesirable immune responses. While complementarity-determining region (CDR) grafting-based humanization techniques have been successfully applied in many cases, it remains challenging to maintain the desired stability and antigen binding affinity upon grafting. We developed an alternative humanization approach called CoDAH ("Computationally-Driven Antibody Humanization") in which computational protein design methods directly select sets of amino acids to incorporate from human germline sequences to increase humanness while maintaining structural stability. Retrospective studies show that CoDAH is able to identify variants deemed beneficial according to both humanness and structural stability criteria, even for targets lacking crystal structures. Prospective application to TZ47, a murine antihuman B7H6 antibody, demonstrates the approach. Four diverse humanized variants were designed, and all possible unique VH/VL combinations were produced as full-length IgG1 antibodies. Soluble and cell surface expressed antigen binding assays showed that 75% (6 of 8) of the computationally designed VH/VL variants were successfully expressed and competed with the murine TZ47 for binding to B7H6 antigen. Furthermore, 4 of the 6 bound with an estimated K D within an order of magnitude of the original TZ47 antibody. In contrast, a traditional CDR-grafted variant could not be expressed. These results suggest that the computational protein design approach described here can be used to efficiently generate functional humanized antibodies and provide humanized templates for further affinity maturation.

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CDR Repair: A Novel Approach to Antibody Humanization

Cited by 11 publications

References 40 publications

Inhibitory Mechanism of an Allosteric Antibody Targeting the Glucagon Receptor

Inhibitory Mechanism of an Allosteric Antibody Targeting the Glucagon Receptor

Beyond CDR-grafting: Structure-guided humanization of framework and CDR regions of an anti-myostatin antibody

Antibody humanization by structure-based computational protein design

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