Isomer-Specific Monitoring of Sialylated N-Glycans Reveals Association of α2,3-Linked Sialic Acid Epitope With Behcet's Disease

Nari Seo; Hyunjun Lee; Myung Jin Oh; Ga Hyeon Kim; Sang Gil Lee; Joong Kyong Ahn; Hoon-Suk Cha; Kyoung Heon Kim; Jaehan Kim; Hyun Joo An

doi:10.3389/fmolb.2021.778851

Isomer-Specific Monitoring of Sialylated N-Glycans Reveals Association of α2,3-Linked Sialic Acid Epitope With Behcet's Disease

Front Mol Biosci. 2021 Nov 23:8:778851. doi: 10.3389/fmolb.2021.778851. eCollection 2021.

Authors

Nari Seo^{1

2}, Hyunjun Lee³, Myung Jin Oh^{1

2}, Ga Hyeon Kim^{1

2}, Sang Gil Lee^{1

2}, Joong Kyong Ahn⁴, Hoon-Suk Cha⁵, Kyoung Heon Kim⁶, Jaehan Kim³, Hyun Joo An^{1

2}

Affiliations

¹ Graduate School of Analytical Science and Technology, Chungnam National University, Daejeon, South Korea.
² Asia Glycomics Reference Site, Daejeon, South Korea.
³ Department of Food and Nutrition, Chungnam National University, Daejeon, South Korea.
⁴ Division of Rheumatology, Department of International Medicine, Kangbuk Samsung Hospital, Sungkyunkwan University School of Medicine, Seoul, South Korea.
⁵ Division of Rheumatology, Department of Medicine, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea.
⁶ Department of Biotechnology, Graduate School, Korea University, Seoul, South Korea.

Abstract

Behcet's disease (BD) is an immune disease characterized by chronic and relapsing systemic vasculitis of unknown etiology, which can lead to blindness and even death. Despite continuous efforts to discover biomarkers for accurate and rapid diagnosis and optimal treatment of BD, there is still no signature marker with high sensitivity and high specificity. As the link between glycosylation and the immune system has been revealed, research on the immunological function of glycans is being actively conducted. In particular, sialic acids at the terminus of glycoconjugates are directly implicated in immune responses, cell-cell/pathogen interactions, and tumor progression. Therefore, changes in sialic acid epitope in the human body are spotlighted as a new indicator to monitor the onset and progression of immune diseases. Here, we performed global profiling of N-glycan compositions derived from the sera of 47 healthy donors and 47 BD patients using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) to preferentially determine BD target glycans. Then, three sialylated biantennary N-glycans were further subjected to the separation of linkage isomers and quantification using porous graphitized carbon-liquid chromatography (PGC-LC)/multiple reaction monitoring (MRM)-MS. We were able to successfully identify 11 isomers with sialic acid epitopes from the three glycan compositions consisting of Hex₅HexNAc₄NeuAc₁, Hex₅HexNAc₄Fuc₁NeuAc₁, and Hex₅HexNAc₄NeuAc₂. Among them, three isomers almost completely distinguished BD from control with high sensitivity and specificity with an area under the curve (AUC) of 0.945, suggesting the potential as novel BD biomarkers. In particular, it was confirmed that α2,3-sialic acid at the terminus of biantennary N-glycan was the epitope associated with BD. In this study, we present a novel approach to elucidating the association between BD and glycosylation by tracing isomeric structures containing sialic acid epitopes. Isomer-specific glycan profiling is suitable for analysis of large clinical cohorts and may facilitate the introduction of diagnostic assays for other immune diseases.

Keywords: Behcet’s disease; biomarker; glycan isomer; quantitation; sialic acid epitope.