Altered Cell Surface <i>N</i> -Glycosylation Implicates Hypersialylation in Breast Cancer Brain Metastasis

Breast cancer is the second most common cause of brain metastasis, often in advanced-stage disease. The mechanisms underlying breast cancer brain metastasis (BCBM), particularly how tumor cells cross the blood-brain barrier and adapt to the brain environment, remain unclear. Cell surface glycosylation plays diverse roles, and its dysregulation in cancer can disrupt signaling and promote metastasis. We investigated how changes in cell surface <i>N</i>-glycans contribute to BCBM by analyzing <i>N</i>-glycans released from human breast cancer cell lines (MDA-MB-231, MDA-MB-361, HTB-131, HTB-22), a brain-seeking variant (MDA-MB-231BR), and glioblastoma cells (CRL-1620) using nano liquid chromatography-tandem mass spectrometry (LC-MS/MS). Surface <i>N</i>-glycans were enzymatically released from live cells with PNGase F without compromising the membrane integrity. Results showed the 231BR cell line expressed higher levels of sialylated <i>N</i>-glycans than other cells, with <i>N</i>-glycan 4502 being the most abundant. Four sialylated structures (4501, 4502, 3501, and 5602) were significantly elevated in 231BR, suggesting a role in brain metastasis. This study expands our understanding of altered <i>N</i>-glycan profiles in BCBM and highlights potential molecular features linked to brain colonization. Further research on these <i>N</i>-glycans could clarify their function in mediating metastasis and identifying therapeutic targets.