Mass Spectrometry-Based Analysis of Serum N-glycan Biomarkers for Rapid Lung Cancer Detection

Abstract Blood serum is a readily accessible sample frequently used for screening assays and early disease detection. Protein glycosylation has been shown to change under various pathological conditions. In this study, the N -glycan profiles of the serum samples from patients primarily diagnosed with non-small and small cell lung cancer were scrutinized using two different preparation approaches to validate alterations detected by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. An increase in fucosylated tri- and tetra-antennary sialylated glycans with 2,3- and 2,6-linked NeuAc residues was observed in the serum samples of the majority of patients, including those with inflammation. Analysis of desialylated glycan pools revealed the presence of additional tetra-antennary glycans, as well as an overall increase in glycan antennarity and elongation involving multiple N-acetyllactosamine units. In patients, these glycans carry between two and four fucose units, predominantly linked only to the antennae. In contrast, control samples exhibited multiantennary glycans bearing either none or one fucose residue linked equally to the antenna and chitobiose core. A comparison of serum and lung tumor samples from the same patients revealed a negative correlation for several N -glycans, along with differences in branching and the isomeric variations of fucosylation in tri- and higher-antennary structures. This supports the hypothesis that alterations in serum glycans reflect more of an epiphenomenon rather than tumor-intrinsic processes. Nonetheless, both increased antenna fucosylation and presence of branched complex multifucosylated glycans in serum may serve as reliable markers for malignancy screening and warrant further clinical evaluation of these patients.