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Site-Specific Bioconjugation Using SMARTag® Technology: A Practical and Effective Chemoenzymatic Approach to Generate Antibody–Drug Conjugates

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Bioconjugation

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2033))

Abstract

As a critical feature of the next generation of antibody–drug conjugates (ADCs), site-specific bioconjugation approaches can help to optimize stability, pharmacokinetics, efficacy, and safety as well as improve manufacturing consistency. The SMARTag® technology platform offers a practical and efficient chemoenzymatic solution for site-specific protein modifications. A bioorthogonal aldehyde handle is introduced through the oxidation of a cysteine residue, embedded in a specific peptide sequence (CxPxR), to the aldehyde-bearing formylglycine (fGly). This enzymatic modification is carried out by the formylglycine-generating enzyme (FGE). The broad recognition of this short sequence by FGE within the context of heterologous proteins allows for the introduction of fGly residues at chosen sites in proteins expressed in prokaryotic and eukaryotic systems. The protocol presented here describes the methods for expressing fGly-containing antibodies in eukaryotic cells and subsequent site-specific conjugation with a payload-linker using aldehyde-specific Hydrazino-Iso-Pictet–Spengler (HIPS) chemistry.

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Authors and Affiliations

  1. Catalent Pharma Solutions, Richmond, CA, USA

    Junjie Liu, Robyn M. Barfield & David Rabuka

Authors
  1. Junjie Liu
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  2. Robyn M. Barfield
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  3. David Rabuka
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Corresponding author

Correspondence to David Rabuka .

Editor information

Editors and Affiliations

  1. Cellular and Molecular Immunology Laboratory, Vrije Universiteit Brussel, Brussels, Belgium

    Sam Massa

  2. In Vivo Cellular and Molecular Imaging Laboratory (ICMI), Vrije Universiteit Brussel, Brussels, Belgium

    Nick Devoogdt

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Liu, J., Barfield, R.M., Rabuka, D. (2019). Site-Specific Bioconjugation Using SMARTag® Technology: A Practical and Effective Chemoenzymatic Approach to Generate Antibody–Drug Conjugates. In: Massa, S., Devoogdt, N. (eds) Bioconjugation. Methods in Molecular Biology, vol 2033. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9654-4_10

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  • DOI: https://doi.org/10.1007/978-1-4939-9654-4_10

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9653-7

  • Online ISBN: 978-1-4939-9654-4

  • eBook Packages: Springer Protocols

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