Differentiating N-linked glycan structural isomers in metastatic and nonmetastatic tumor cells using sequential mass spectrometry

Abstract

In an effort to understand the role of molecular glycosylation in cancer a murine model has been used to characterize and fingerprint malignancies in established cell lines that manifest all the hallmarks of metastatic disease: spontaneous development, local invasion, intravasation, immune system survival, extravasation, and secondary tumor formation involving liver, kidney, spleen, lung, and brain. Using astrocyte cell controls, we compared N-linked glycosylation from a nonmetastatic brain tumor cell line and two different metastatic brain tumor cells. Selected ions in each profile were disassembled by ion trap mass spectrometry (MS(n)) which exhibited multiple structural differences between each tissue. These unique structures were identified within isomeric compositions as pendant nonreducing termini of di- and trisaccharide fragments, probably transparent to a tandem MS approach but distinctively not to sequential ion trap MS(n) detection.

The Diversity of O-Linked Glycans Expressed during Drosophila melanogaster Development Reflects Stage- and Tissue-specific Requirements for Cell Signaling

“…GC-MS analysis of the permethylated oligosaccharide alditol verified HexA-Hex at the nonreducing terminal as well as a singly substituted HexNAc-ol at the reducing terminal. The other, less prevalent form of the core 1 trisaccharide is branched by the addition of HexA to the 4-position of the reducing terminal GalNAc (62,63). Although the linear form of the trisaccharide has recently been described on proteins expressed by the Drosophila S2 cell line in culture, the branched core 1 trisaccharide is novel, and neither glycan has been detected previously in embryos (40).…”

Tandem Mass Spectrometry of Tagged and Permethylated Polysaccharides

“…Tandem MS of permethylated glycans carries the advantage that ions formed from cleavage of glycosidic bonds have a unique mass value that distinguishes an internal fragment from a single bond cleavage. Multistage dissociation of permethylated glycans has been used to determine detailed linkage information for saccharide units formed by gas phase disassembly of the glycans (Zhang et al, 2005;Prien et al, 2008Prien et al, , 2009Jiao et al, 2011). The key to multistage dissociation of glycans is the selection of a series of precursor ions that isolate structural branches for subsequent stages.…”

Oligosaccharide analysis by graphitized carbon liquid chromatography–mass spectrometry

“…Three isomeric O-glycan structures of composition Hex3-HexNAc1 of Caenorhabditis elegans were successfully characterized using this separation procedure. Notably, multistage tandem mass spectrometry (MS n ) of sodium adducts of permethylated glycans allows an even more detailed structure characterization The resulting branched fragmentation paths may comprise dozens of fragmentation spectra, and the integration of this information enables discrimination between structural isomers [65,66]. Analysis of complex oligosaccharide mixtures in this manner requires in-depth knowledge for interpretation of the spectra, but software tools to support the interpretation of the spectra for glycan structure elucidation are currently under development [67][68][69].…”