Characterization of the Oligosaccharides Associated with the Human Ovarian Tumor Marker CA125

Abstract

CA125 is a mucin commonly employed as a diagnostic marker for epithelial ovarian cancer. Induction of humoral responses to CA125 leads to increased survival times in patients with this form of cancer, suggesting a potential role for this mucin in tumor progression. In this study, oligosaccharides linked to CA125 derived from the human ovarian tumor cell line OVCAR-3 were subjected to rigorous biophysical analysis. Sequencing of the Oglycans indicates the presence of both core type 1 and type 2 glycans. An unusual feature is the expression of branched core 1 antennae in the core type 2 glycans. CA125 is also N-glycosylated, expressing primarily high mannose and complex bisecting type N-linked glycans. High mannose type glycans include Man 5 -Man 9 GlcNAc 2 . The predominant N-glycans are the biantennary, triantennary, and tetraantennary bisecting type oligosaccharides. Remarkably, the N-glycosylation profiles of CA125 and the envelope glycoprotein gp120 (derived from H9 lymphoblastoid cells chronically infected with HIV-1) are very similar. The CA125-associated N-glycans have also recently been implicated in crucial recognition events involved in both the innate and adaptive arms of the cell-mediated immune response. CA125 may therefore induce specific immunomodulatory effects by employing its carbohydrate sequences as functional groups, thereby promoting tumor progression. Immunotherapy directed against CA125 may attenuate these immunosuppressive effects, leading to the prolonged survival of patients with this extremely serious form of cancer.

Mass spectrometry-based analysis of glycoproteins and its clinical applications in cancer biomarker discovery

“…However, due to the challenges mentioned above, purification, enrichment and fractionation approaches are of the essence prior to MS analysis. Several comprehensive reviews about sample pretreatment of glycoproteins have been published in recent years (Fanayan et al, 2012;Ongay et al, 2012;Lazar et al, 2013), so we will not discuss them here. …”

“…Moreover, methylated carbohydrate deem to ionize more efficiently than their native counterparts. In conjunction with ESI and collision-induced dissociation (CID), permethylated carbohydrate also yield a most detailed structural information (Patel, 1993;Colangelo and Orlando, 1999;Huang et al, 2001;Wheeler and Harvey, 2001;Huang et al, 2002;Zaia, 2004;Taylor et al, 2006).…”

“…This methylation procedure has now been used for derivatization of carbohydrates (Ciucanu and Costello, 2003;Kui Wong et al, 2003;Kang et al, 2005), other polyols, and fatty acids, including their hydroxyl derivatives (Morelle et al, 2004;). However, it has gained a particular popularity with complex oligosaccharides and glycolipids.…”

“…N-linked glycans are attached to the amide group of asparagine residues in a consensus Asn-X-Ser/Thr sequence (X can be any amino acid except proline) (Bause, 1983). Other known, but less well studied forms of glycosylation include glycosylphosphatidylinositol anchors attached to protein carboxyl terminus, C-glycosylation that occurs on tryptophan residues (Weim, 2009), and S-linked glycosylation through a sulfur atom on cysteine or methionine (Lote and Weiss, 1971;Vijayakrishnan et al, 2009) ( Aebersold and Mann, 2003). N-glycan moieties also play a vital role in the folding, processing, and secretion of proteins from the endoplasmic reticulum (ER) and the Golgi apparatus (Aebersold and Mann, 2003).…”

“…The structural aspects of glycans have been studied by mass spectrometry (MS) for many years, during the last decade the development of matrix-assisted laser desorption/ionization (MALDI) and electrospray ionization (ESI) MS for this class of compounds has accelerated substantially the acceptance of MS-based methodologies. In the structural analysis of complex glycans originating from various isolated glycoproteins (Carlson, 1968;Dell, 1987;Hellerqvist, 1990;Varki, 1993;Tarentino and Plummer, 1994;Tretter et al, 1991;Faid et al, 2006;Ohtsubo and Marth, 2006;Faid et al, 2007;Morelle et al, 2005a;Morelle et al, 2005b;Morelle et al, 2006b;Morelle et al, 2006c), MALDI-MS in conjunction with exoglycosidase digestion and a tandem (MS/MS) operation have become particularly popular (Küster, Wheeler, Hunter, Dwek and Harvey, 1997).…”