Biomolecular recognition of the glycan neoantigen CA19-9 by distinct antibodies

Abstract

Glycans decorate cell surface, secreted glycoproteins and glycolipids. Altered glycans are often found in cancers. Despite their high diagnostic and therapeutic potentials, glycans are polar and flexible molecules that are quite challenging for the development and design of high-affinity binding antibodies. To understand the mechanisms by which glycan neoantigens are specifically recognized by antibodies, we analyze the biomolecular recognition of a single tumor-associated carbohydrate antigen CA19-9 by two distinct antibodies using X-ray crystallography. Despite the plasticity of glycans and the very different antigen-binding surfaces presented by the antibodies, both structures reveal an essentially identical extended CA19-9 conformer, suggesting that the stability of the conformer selects the antibodies. Starting from the bound structure of one of the antibodies, we use the AbLIFT computational method to design a variant with seven core mutations that exhibited tenfold improved affinity for CA19-9. The results reveal strategies used by antibodies to specifically recognize glycan antigens and show how automated antibody-optimization methods may be used to enhance the clinical potential of existing antibodies.

Reliable energy-based antibody humanization and stabilization

“…Finally, we note that CUMAb is based on the fundamental insight that antibody stability and activity are determined by the entire Fv, including the framework positions on which the CDRs rest 16, 56 and the interfaces between the light and the heavy chains 14, 57 . This insight may help address other important challenges in antibody engineering, leading to general, reliable, and rapid antibody design strategies.…”

What Have We Learned from Design of Function in Large Proteins?

“…Enzyme active sites and protein-protein binding sites share in common a high density of amino acid interactions. We therefore also applied FuncLib to optimize proteinprotein interactions, finding that it can improve proteinbinding affinity [50] and antibody stability and affinity [83,84] by optimizing atomic interactions across the interacting surfaces. Furthermore, improving the interactions across the homooligomeric interfaces in a trimeric bacterial enzyme called PodA (Figure 4(c)) led to improved stability and an order of magnitude increase in its production yields [85].…”