A newly generated functional antibody identifies Tn antigen as a novel determinant in the cancer cell-lymphatic endothelium interaction

Abstract

Malignant transformation of epithelial cells is frequently associated with the alteration of glycosylation pathways. Tn is a common tumor-associated carbohydrate antigen present in 90% of human carcinomas and its expression correlates with metastatic potential and poor prognosis. Despite its relevance, the functional role of Tn in tumor biology has not been firmly established probably for the lack of appropriate experimental tools. Our aims were to produce highly reactive monoclonal antibodies against Tn making use of synthetically produced Tn and to test their usefulness for in vivo imaging as well as to define their potential functional activity in tumor cell spread. We immunized mice with Tn clustered on cationized BSA and screened the positive hybridomas with Tn-biotinylated alginate. Enzyme-linked immuno sorbent assay and immunofluorescence assays revealed that the most reactive anti-Tn IgM mAb (2154F12A4) selectively recognized Tn on the MCF7 breast cancer cell line since its binding to the cell membrane was completely abolished by preincubation with purified Tn. Importantly, QDot 800-conjugated mAb injected in MCF7-tumor bearing mice specifically bound to primary tumor lesions as well as to metastases in lymph nodes. In addition, this mAb was able to inhibit cancer cell adhesion to lymphatic endothelium suggesting a novel involvement of Tn in the lymphatic dissemination of cancer cells and hypothesizing future applications in inhibiting lymphatic metastases.

PD-1 Suppresses Development of Humoral Responses That Protect against Tn-Bearing Tumors

“…Mucin-elicited IgG may also contribute via antibody-dependent cellular cytotoxicity and antibodies may function by altering mucin signaling and/or adhesion properties important for limiting metastases. Indeed, the decreased dissemination observed in immune PD-1 −/− mice and in serum transfer experiments supports the latter (39). Finally, although mucin/Tn-specific Ab contributed to protection, it is possible B cells may regulate additional aspects of antitumor immunity, such as antigen presentation (40–42) and/or cytokine production.…”

A Comparative Study of Adhesion of Melanoma and Breast Cancer Cells to Blood and Lymphatic Endothelium

“…19 In addition, the adhesion of MCF7 cells to human microvascular lung lymphatic endothelial cells (HMVEC-LLy) was reduced following treatment with an anti-Tn antibody, a common tumor-associated carbohydrate antigen present in most carcinomas. 20 Studies conducted thus far have utilized either BEC or LEC to examine tumor-endothelial adhesion. To the best of our knowledge, no studies have conducted a direct side-to-side comparison of blood versus lymphatic adhesion.…”

A glycobiology review: Carbohydrates, lectins and implications in cancer therapeutics

“…Nearly all types of malignant cells and cells of many types of diseased tissues demonstrate alterations in their glycosylation patterns when compared to their normal counterparts (Hakomori, 1985, 1989, 1996; Dennis, 1992; Mody et al, 1995; Fukuda, 1996; Orntoft and Vestergarrd, 1999; Oppenheimer, 2006; Blomme et al, 2009; Danussi et al, 2009; Patsos et al, 2009; Pettersson-Kastberg et al, 2009; Powlesland et al, 2009; Shida et al, 2009). The alterations in glycosylation patterns are often a result of altered activities of glycosyltransferases and glycosidases as is evident in the specific and preferential display of certain glycoconjugates on cancer cells (Mody et al, 1995; Couldrey and Green, 2000; Gorelik et al, 2001; Oppenheimer, 2006; Schwartz-Albiez et al, 2008; Blomme et al, 2009; Goetz et al, 2009; Patsos et al, 2009; Powlesland et al, 2009; Rek et al, 2009; Shida et al, 2009).…”

“…The above example and the five mechanisms of protein modification demonstrate the ability of glycosylation to alter cellular activity at many levels. Therefore, alteration in glycosylation may eventually lead to chronic diseases such as cancer or may be a result, not a cause, of associated changes (Blomme et al, 2009; Danussi et al, 2009; Patsos et al, 2009; Pettersson-Kastberg et al, 2009; Powlesland et al, 2009; Shida et al, 2009). Covalent modification of proteins and enzymes through phosphorylation and dephosphorylation is a sophisticated control system that cells utilize to maintain homeostasis (Voet et al, 1999).…”

“…This altered phosphorylation pattern then changes enzyme cascade and signaling pathways which consequently influence expression levels of genes that encode, for example, glycosyltransferases and glycosidases. The altered activity levels of glycosyltransferases and glycosidases account for the specific and preferential display of certain glycoconjugates on cancer cells (Mody et al, 1995; Couldrey and Green, 2000; Gorelik et al, 2001; Oppenheimer, 2006; Schwartz-Albiez et al, 2008; Blomme et al, 2009; Danussi et al, 2009; Patsos et al, 2009; Pettersson-Kastberg et al, 2009; Powlesland et al, 2009; Shida et al, 2009). In hepatocellular carcinoma (HCC), for example, the activities of α-1,6-fucosyltransferase (α-1,6-FT), β-1,6-N-acetylglucosominyltransferase V (GlcNAc-TV) and β-1,4-N-acetylglucosaminyltransferase III (GlcNAc-TIII) are significantly increased compared to their normal counterparts (Blomme et al, 2009).…”