Data Independent Acquisition Mass Spectrometry Enhanced Personalized Glycosylation Profiling of Haptoglobin in Hepatocellular Carcinoma.
Journal
Journal of proteome research
Journal Volume
23
Journal Issue
8
Pages
3571 - 3584
ISSN
1535-3907
Date Issued
2024-08-02
Author(s)
Abstract
Aberrant glycosylation has gained significant interest for biomarker discovery. However, low detectability, complex glycan structures, and heterogeneity present challenges in glycoprotein assay development. Using haptoglobin (Hp) as a model, we developed an integrated platform combining functionalized magnetic nanoparticles and zwitterionic hydrophilic interaction liquid chromatography (ZIC-HILIC) for highly specific glycopeptide enrichment, followed by a data-independent acquisition (DIA) strategy to establish a deep cancer-specific Hp-glycosylation profile in hepatitis B virus (HBV, = 5) and hepatocellular carcinoma (HCC, = 5) patients. The DIA strategy established one of the deepest Hp-glycosylation landscapes (1029 glycopeptides, 130 glycans) across serum samples, including 54 glycopeptides exclusively detected in HCC patients. Additionally, single-shot DIA searches against a DIA-based spectral library outperformed the DDA approach by 2-3-fold glycopeptide coverage across patients. Among the four N-glycan sites on Hp (N-184, N-207, N-211, N-241), the total glycan type distribution revealed significantly enhanced detection of combined fucosylated-sialylated glycans, which were the most dominant glycoforms identified in HCC patients. Quantitation analysis revealed 48 glycopeptides significantly enriched in HCC ( < 0.05), including a hybrid monosialylated triantennary glycopeptide on the N-184 site with nearly none-to-all elevation to differentiate HCC from the HBV group (HCC/HBV ratio: 2462 ± 766, < 0.05). In summary, DIA-MS presents an unbiased and comprehensive alternative for targeted glycoproteomics to guide discovery and validation of glyco-biomarkers.
Subjects
data-independent acquisition (DIA)
glycoprotein
haptoglobin
hepatocellular carcinoma (HCC)
SDGs
Type
journal article