Clinical importance of high-mannose, fucosylated, and complex N-glycans in breast cancer metastasis

BACKGROUND. Although aberrant glycosylation is recognized as a hallmark of cancer, glycosylation in clinical breast cancer (BC) metastasis has not yet been studied. While preclinical studies show that the glycocalyx coating of cancer cells is involved in adhesion, migration, and metastasis, glycosylation changes from primary tumor (PT) to various metastatic sites remain unknown in patients. METHODS. We investigated N-glycosylation profiles in 17 metastatic BC patients from our rapid autopsy program. Primary breast tumor, lymph node metastases, multiple systemic metastases, and various normal tissue cores from each patient were arranged on unique single-patient tissue microarrays (TMAs). We performed mass spectrometry imaging (MSI) combined with extensive pathology annotation of these TMAs, and this process enabled spatially differentiated cell-based analysis of N-glycosylation patterns in metastatic BC. RESULTS. N-glycan abundance increased during metastatic progression independently of BC subtype and treatment regimen, with high-mannose glycans most frequently elevated in BC metastases, followed by fucosylated and complex glycans. Bone metastasis, however, displayed increased core-fucosylation and decreased high-mannose glycans. Consistently, N-glycosylated proteins and N-glycan biosynthesis genes were differentially expressed during metastatic BC progression, with reduced expression of mannose-trimming enzymes and with elevated EpCAM, N-glycan branching, and sialyation enzymes in BC metastases versus PT. CONCLUSION. We show in patients that N-glycosylation of breast cancer cells undergoing metastasis occurs in a metastatic site–specific manner, supporting the clinical importance of high-mannose, fucosylated, and complex N-glycans as future diagnostic markers and therapeutic targets in metastatic BC. FUNDING. NIH grants R01CA213428, R01CA213492, R01CA264901, T32CA193145, Dutch Province Limburg “LINK”, European Union ERA-NET TRANSCAN2-643638.