Site-Specific Chemoenzymatic Conjugation of High-Affinity M6P Glycan Ligands to Antibodies for Targeted Protein Degradation

• DOI: 10.1021/acschembio.1c00751
• Publication Date: 2022-03-22T19:33:37Z
• Authors: Xiao Zhang; Huiying Liu; Jia He; Chong Ou; Thomas C. Donahue; Musleh M. Muthana; Lishan Su; Lai-Xi Wang
• Keywords: Mannose 6-phosphate; Endoglycosidase

Lysosome-targeting chimeras (LYTACs) offer an opportunity for the degradation of extracellular and membrane-associated proteins interest. Here, we report efficient chemoenzymatic method that enables a single-step site-specific conjugation high-affinity mannose-6-phosphate (M6P) glycan ligands to antibodies without need protein engineering conventional click reactions would introduce "unnatural" moieties, yielding homogeneous antibody-M6P conjugates targeted proteins. Using trastuzumab cetuximab as model antibodies, showed wild-type endoglycosidase S (Endo-S) could efficiently perform antibody deglycosylation simultaneous transfer M6P-glycan from synthetic oxazoline deglycosylated in one-pot manner, giving structurally well-defined conjugates. A two-step procedure, using Endo-S2 followed by transglycosylation with mutant (D184M), was also provide M6P glycan-antibody The approach highly specific Fc remodeling when both Fab domains were glycosylated, exemplified selective Fc-glycan cetuximab. SPR binding analysis indicated possessed nanomolar range affinities cation-independent receptor (CI-MPR). Preliminary cell-based assays M6P-trastuzumab M6P-cetuximab able selectively degrade HER2 EGFR, respectively. This modular glycan-remodeling strategy is expected find wide applications antibody-based lysosome-targeted membrane