posted on 2021-08-09, 12:06authored byCarolina
T. Orozco, Matthew J. Edgeworth, Paul W. A. Devine, Alistair R. Hines, Owen Cornwell, Christopher Thompson, Xiangyang Wang, Jonathan J. Phillips, Peter Ravn, Sophie E. Jackson, Nicholas J. Bond
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.