Site-specific
labeling and conjugation of antibodies are highly
desirable for fundamental research and for developing more efficient
diagnostic and therapeutic methods. We report here a general and robust
chemoenzymatic method that permits a one-pot site-specific functionalization
of antibodies. A series of selectively modified disaccharide oxazoline
derivatives were designed, synthesized, and evaluated as donor substrates
of different endoglycosidases for antibody Fc glycan remodeling. We
found that among several endoglycosidases tested, wild-type endoglycosidase
from Streptococcus pyogenes of serotype
M49 (Endo-S2) exhibited remarkable activity in transferring the functionalized
disaccharides carrying site-selectively modified azide, biotin, or
fluorescent tags to antibodies without hydrolyzing the resulting transglycosylation
products. This discovery, together with the excellent Fc deglycosylation
activity of Endo-S2 on recombinant antibodies, allowed direct labeling
and functionalization of antibodies in a one-pot manner without the
need of intermediate and enzyme separation. The site-specific introduction
of varied numbers of azide groups enabled a highly efficient synthesis
of homogeneous antibody–drug conjugates (ADCs) with a precise
control of the drug-to-antibody ratio (DAR) ranging from 2 to 12 via
a copper-free strain-promoted click reaction. Cell viability assays
showed that ADCs with higher DARs were more potent in killing antigen-overexpressed
cells than the ADCs with lower DARs. This new method is expected to
find applications not only for antibody–drug conjugation but
also for cell labeling, imaging, and diagnosis.
Functionalization of therapeutic lysosomal enzymes with mannose-6-phosphate (M6P) glycan ligands represents a major strategy for enhancing the cation-independent M6P receptor (CI-MPR)-mediated cellular uptake, thus improving the overall therapeutic efficacy of...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.