Auto-antibodies to cancer antigens hold great promise as sensitive amplified biomarkers for early detection of cancer. Most high through-put strategies to discover such auto-antibodies largely fail to allow identification of antibodies specific for cancer-associated posttranslational modified variants of normal proteins. We hypothesized that aberrant processed proteins are likely auto-antibody targets. MUC1 is over-expressed and aberrantly glycosylated in many cancers and we sought to evaluate the potential of natural cancer-induced auto-antibodies to such aberrant O-glycoforms of MUC1 as sensitive diagnostic biomarkers of disease. We first demonstrated, using an antibody-based glycoprofiling ELISA, that circulating mucins in cancer patients exclude truncated aberrant cancer-associated glycoforms. We then developed an O-glycopeptide microarray and used this to demonstrate detection of IgG antibodies to MUC1 aberrant O-glycopeptide epitopes in patients vaccinated with 25Tn-MUC1-106-mer conjugated to KLH. Finally, screening of sera from breast, ovarian and prostate cancer patients led to identification of three distinct aberrant MUC1 O-glycopeptide epitopes that are targeted by cancer-associated IgG auto-antibodies. The results suggest that auto-antibodies to aberrant O-glycopeptide epitopes may represent a fruitful and previously unaddressed source of sensitive biomarkers for early detection of cancer. The methods developed for chemoenzymatic synthesis of O-glycopeptides in combination with microarrays allow for broader data-mining of the entire cancer O-glycopeptidome.

Purpose: Adhesion of disseminating tumor cells to the blood vascular endothelium is a pivotal step in metastasis. Previous investigations have shown that galectin-3 concentrations are increased in the bloodstream of patients with cancer and that galectin-3 promotes adhesion of disseminating tumor cells to vascular endothelium in vitro and experimental metastasis in vivo. This study determined the levels of galectin-1, -2, -3, -4, -8, and -9 in the sera of healthy people and patients with colon and breast cancer and assessed the influence of these galectins on cancer-endothelium adhesion.Experimental Design: Serum galectins and auto-anti-MUC1 antibodies were assessed using ELISA and mucin protein (MUC1) glycan microarrays, and cancer-endothelium adhesion was determined using monolayers of human microvascular lung endothelial cells.Results: The levels of serum galectin-2, -3, -4, and -8 were significantly increased up to 31-fold in patients with cancer and, in particular, those with metastases. As previously shown for galectin-3, the presence of these galectins enhances cancer-endothelium adhesion by interaction with the Thomsen-Friedenreich (TF; Galb1,3GalNAca-) disaccharide on cancer-associated MUC1. This causes MUC1 cell surface polarization, thus exposing underlying adhesion molecules that promote cancer-endothelium adhesion. Elevated circulating galectin-2 levels were associated with increased mortality in patients with colorectal cancer, but this association was suppressed when anti-MUC1 antibodies with specificity for the TF epitope of MUC1 were also present in the circulation.Conclusions: Increased circulation of several members of the galectin family is common in patients with cancer and these may, like circulating galectin-3, also be involved in metastasis promotion. Clin Cancer Res; 17(22); 7035-46. Ó2011 AACR.