Analytical Pipeline for Discovery and Verification of Glycoproteins from Plasma-Derived Extracellular Vesicles as Breast Cancer Biomarkers

Glycoproteins
comprise more than half of current FDA-approved protein
cancer markers, but the development of new glycoproteins as disease
biomarkers has been stagnant. Here we present a pipeline to develop
glycoproteins from extracellular vesicles (EVs) through integrating
quantitative glycoproteomics with a novel reverse phase glycoprotein
array and then apply it to identify novel biomarkers for breast cancer.
EV glycoproteomics show promise in circumventing the problems plaguing
current serum/plasma glycoproteomics and allowed us to identify hundreds
of glycoproteins that have not been identified in blood. We identified
1,453 unique glycopeptides representing 556 glycoproteins in EVs,
among which 20 were verified significantly higher in individual breast
cancer patients. We further applied a novel glyco-specific reverse
phase protein array to quantify a subset of the candidates. Together,
this study demonstrates the great potential of this integrated pipeline
for biomarker discovery.