pH-Dependent Conformational Change of CDR of Cytosol-Penetrating Antibody Mediate the Endosomal Escape

摘要

Endosomal escape after endocytosis is a critical step for protein-based agents to exhibit their effects in the cytosol of cells. However, antibodies internalized into cells by endocytosis cannot reach the cytosol due to their inability to escape from endosomes. Here, we elucidate a unique endosomal escape mechanism of the full-length IgG format TMab4 antibody which can reach the cytosol of living cells after internalization. Internalized TMab4 via HSPG(as a cell surface receptor)-mediated endocytosis was dissociated from HSPG by activated heparanase in acidified early endosomes and then local structural changes of the endosomal escape motif of TMab4 in response to the acidified endosomal pH were critical for the formation of membrane pores through which TMab4 escaped into the cytosol. We generated a TMab4 variants for validation of structural determinants of endosomal escape and one of them show 3-fold improved endosomal escape efficiency. Our finding of the endosomal escape mechanism of the cy to sol-penetrating antibody and its improvement will establish a platform technology that enables a intact IgG format antibody to directly target cytosolic proteins.