NIH Research Festival
Rabbits are able to produce antibodies that recognize diverse epitopes including poorly immunogenic epitopes in tumor antigens. To design a humanization method for rabbit monoclonal antibodies (Rabmabs), we structurally aligned five Rabmabs in complex with their antigens in the PDB database and identified possible antigen-binding residues and their contacting residues. We found that the combined three complementarity determining region (CDR) systems (KABAT, IMGT and Paratome) can cover the most of the antigen-binding residues and their contacting residues. We then tested our humanization strategy by grafting the combined Kabat/IMGT/paratome CDRs into a human germline framework, using four anti-mesothelin RAbmabs as examples. The resulting humanized Fv had similar or improved binding affinity for antigen-expressing cells. The immunotoxin composed of humanized Fv fused to a toxin fragment of Pseudomonas exotoxin showed stronger cytotoxicity against target tumor cells than the immunotoxins derived from their original rabbit Fvs. Our data suggest that grafting the combined KABAT/IMGT/Paratome CDRs to a stable human germline framework is a logical and robust approach for humanization of rabbit antibodies for clinical applications.
Scientific Focus Area: Molecular Biology and Biochemistry
This page was last updated on Friday, March 26, 2021