A Genetic Approach to Display and Dissect the Cancer‐Associated O‐Glycoepitome

Abstract

A hallmark of cancer is aberrant protein O-glycosylation with expression of truncated immature O-glycans including the classical Tn, STn, and T pancarcinoma antigens. The truncation of O-glycans not only present epitopes recognized by the natural low affinity antibodies to the TTn glycan haptens, which are the basis for the polyagglutination phenomena, but also expose composite aberrant O-glycopeptide epitopes that are not normally expressed and can elicit high affinity IgG antibodies. Several examples of monoclonal antibodies with high selectivity for truncated O-glycopeptide epitopes, which in effect selectively target the cancer form of O-glycoproteins, have been produced in the past. Studies demonstrate that such epitopes serve as highly cancer-specific targets potentially amenable for potent T-cell engaging strategies, however, the current insight into the abundance and distribution of aberrant O-glycopeptide epitopes and the optimal glycoforms for these remain limited. To systematically explore antibody epitopes available in the cancer-associated O-glycoproteome, we have developed a cell-based platform for display and production of human O-glycoproteins with homogeneous truncated glycoforms. This platform includes CHO and HEK293 cells stably engineered with homogeneous glycosylation capacities for the Tn, STn, and T glycoforms, and a panel of reporter expression constructs containing segments of human mucins and mucin-like domains from O-glycoproteins for display of these with different O-glycans. An overview of these efforts will be presented.