SiaQuant Unveils Serum α2,3/α2,6 Sialylation Heterogeneities and Predicts Neoadjuvant Chemotherapy Response in Locally Advanced Cervical Cancer

Sialylation significantly influences tumor progression, invasion, and metastasis. Accurately characterizing sialylated <i>N</i>-glycopeptides (SGPs), particularly the linkage-specific analysis of α2,3,α2,6 sialic acids, remains a challenging yet crucial task in glycoproteomics. Notably, to date, there is a notable lack of detailed studies on α2,3/α2,6 sialylation in serum. Here, we present SiaQuant, an integrated strategy that employs liquid chromatography-ion mobility-tandem mass spectrometry (LC-IM-MS/MS), capitalizing on the distinctive separation of characteristic isomeric glycan fragments in ion mobility to accurately analyze serum α2,3/α2,6 sialylation patterns. It first provides proteome-wide insights into α2,3/α2,6 sialylation in serum, revealing three-dimensional heterogeneities across <i>N</i>-glycans, <i>N</i>-glycosites, and <i>N</i>-glycoproteins. Additionally, SiaQuant identifies potential candidate biomarkers for neoadjuvant chemotherapy (NACT) response in locally advanced cervical cancer (LACC), where the elevated level of α2,3/α2,6 sialylation was found to be associated with NACT resistance. In summary, SiaQuant offers the most comprehensive site- and linkage-specific <i>N</i>-glycosylation profiling of serum and shows great potential in clinical usage.