Sequential Proteomic and N‑Glycoproteomic Analyses of Bronchoalveolar Lavage Fluids for Potential Biomarker Discovery of Lung Adenocarcinoma

Lung
adenocarcinoma (LUAD) is the most common histological
subtype
of nonsmall-cell lung cancer. Herein, a multiomics method, which combined
proteomic and N-glycoproteomic analyses, was developed to analyze
the normal and cancerous bronchoalveolar lavage fluids (BALFs) from
six LUAD patients to identify potential biomarkers of LUAD. The data-independent
acquisition proteomic analysis was first used to analyze BALFs, which
identified 59 differentially expressed proteins (DEPs). The bioinformatic
analyses of 59 DEPs have shown that a potential marker protein, beta-1,4-galactosyltransferase
1 (B4GALT1), was consistently downregulated in all cancerous lung
lobes (CLLs). As the downregulation of B4GALT1 may indicate changes
in protein N-glycosylation, site-specific N-glycoproteome analysis
of BALFs from the normal lung lobes (NLLs) and CLLs was further performed
by using a fully automated glycopeptide enrichment and separation
system. Comparing the glycan structures containing free GlcNAc in
BALFs between NLLs and CLLs qualitatively, the percentage of unique
glycan structure for free GlcNAc existing only in NLLs was 52.8%,
which was significantly higher than the 46.3% existing only in CLLs.
Furthermore, the sequential proteomic and N-glycoproteomic analyses
allowed us to identify a panel of functionally related potential biomarkers
consisting of one protein (B4GALT1) and four glycoproteins (NFKB1,
F2, LTF, and DLD).