Interconversion of Unexpected Thiol States Affects the Stability, Structure, and Dynamics of Antibody Engineered for Site-Specific Conjugation.
Orozco, Carolina T
Edgeworth, Matthew J
Devine, Paul WA
Hines, Alistair R
Phillips, Jonathan J
Bond, Nicholas J
American Chemical Society
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Orozco, C. T., Edgeworth, M. J., Devine, P. W., Hines, A. R., Cornwell, O., Thompson, C., Wang, X., et al. (2021). Interconversion of Unexpected Thiol States Affects the Stability, Structure, and Dynamics of Antibody Engineered for Site-Specific Conjugation.. Bioconjugate Chemistry, 32 (8), 1834-1844. https://doi.org/10.1021/acs.bioconjchem.1c00286
Antibody-drug conjugates have become one of the most actively developed classes of drugs in recent years. Their great potential comes from combining the strengths of large and small molecule therapeutics: the exquisite specificity of antibodies and the highly potent nature of cytotoxic compounds. More recently, the approach of engineering antibody-drug conjugate scaffolds to achieve highly controlled drug to antibody ratios has focused on substituting or inserting cysteines to facilitate site-specific conjugation. Herein, we characterize an antibody scaffold engineered with an inserted cysteine that formed an unexpected disulfide bridge during manufacture. A combination of mass spectrometry and biophysical techniques have been used to understand how the additional disulfide bridge forms, interconverts, and changes the stability and structural dynamics of the antibody intermediate. This quantitative and structurally resolved model of the local and global changes in structure and dynamics associated with the engineering and subsequent disulfide-bonded variant can assist future engineering strategies.
This work was supported by AstraZeneca and the Engineering and Physical Sciences Research Council Centre for Doctoral Training in Sensor Technologies and Applications under Grant EP/L015889/1.
Engineering and Physical Sciences Research Council (EP/L015889/1)
External DOI: https://doi.org/10.1021/acs.bioconjchem.1c00286
This record's URL: https://www.repository.cam.ac.uk/handle/1810/330557
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Licence URL: https://creativecommons.org/licenses/by-nc-nd/4.0/