In general, Intact form of IgG antibodies cannot reach the cytosol by escaping from the endosome after receptor-mediated endocytosis because they have no endosomal escape activity.
Our group recently reported the cytosol-penetrating antibody TMab4 cytotransmab. Also, we generated TMab4-3 which is an engineered version of TMab4 to improve the endosomal escape efficiency and its expression yield.
However, TMab4-3 is not yet suitable for targeted therapy because TMab4-3 has a binding activity against HSPG expressed on the surface of normal cells. In addition, it is necessary to improve the endosomal escape efficiency to increase the therapeutic efficacy of cytotransmab.
Here, I generated a tumor-specific cytotransmab with improved endosomal escape efficiency. First, I have created a new cytotransmab called as CT-41 with the elimination of HSPG binding activity. Second, a cyclic peptide for targeting EpCAM which is well known for over-expressed on surface of tumor cell was fused with N-terminus of CT-41 to confer tumor tissue specificity. Third, an additional endosomal escape motif for cytotransmab have been introduced into the EpCAM specific cytotransmab called as CT-ep41. Lastly, charged residue mutations are inserted near the hydrophobic endosomal escape motif to increase its developability and solubility.
As a result, CT12-ep61 as a final clone is showed a specificity to EpCAM-positive tumor cells and increased endosome escape efficiency about 34% compared to CT-ep41 and its expression level is higher about 8-fold compared to CT-ep41.
These results suggest that tumor-tissue specific cytotransmab can be applied as a tool for targeted cancer therapy.
