Oral Presentation HUPO 2019 - 18th Human Proteome Organization World Congress

Stage-specific N-glycosylation of abundant plasma proteins across four stages of colorectal cancer (64272)

Rebeca Kawahara 1 , Sayantani Chatterjee 1 , Nicolle Packer 1 , Giuseppe Palmisano 2 , Seong Beom Ahn 3 , Morten Thaysen-Andersen 1
  1. Department of Molecular Sciences, Macquarie University, Sydney, NSW, Australia
  2. Department of Parasitology, University of Sao Paulo, Sao Paulo, SP, Brazil
  3. Department of Biomedical Sciences, Macquarie University, Sydney, NSW, Australia

Glycobiology plays central roles in colorectal cancer (CRC) as exemplified by the aberrant glycosylation decorating circulating plasma proteins in CRC patient blood. Previous efforts have explored the aberrant CRC plasma N-glycome, but the site-specific information of the plasma proteins carrying altered glycosylation is crucially missing precluding their use as markers for diagnosis, prognosis and risk assessment of CRC. To this end, we have here used integrated mass spectrometry-based quantitative glycomics and glycoproteomics to characterise the N-glycosylation of plasma proteins directly in non-depleted blood obtained from CRC patients spanning four disease stages (Dukes I-IV, n=7/stage) and age/gender-matched healthy donors (n=8). PGC–LC-MS/MS glycomics profiled a total of 74 biosynthetically-related N-glycans across the sample cohort. Interestingly, 26 N-glycans including the abundant biantennary mono- and bi-α2,6-sialylated glycans with and without α1,6-fucosylation and the related but less abundant tri- and tetra-antennary core-fucosylated sialoglycans displayed stage-specific expression profiles that also differed quantitatively from the N-glycan profile of plasma proteins from healthy blood. TMT labelling, high-pH fractionation and HILIC glycopeptide enrichment were employed prior to high resolution LC-MS/MS to facilitate the identification and quantitation of ~2,000 intact N-glycopeptides covering 76 N-glycoproteins from the same sample cohort. Integrated glycomics and glycoproteomics data analysis and adjustment for stage-specific protein level variations revealed that the CRC stage-specific N-glycome changes were recapitulated at the individual glycosylation site level of the ten most abundant plasma glycoproteins including haptoglobin, Ig gamma-1 chain C region and fibrinogen. This study demonstrates the immense yet often under-utilised potential of employing integrated mass spectrometry-based glycomics and glycoproteomics to discover the aberrant glycosylation signatures that are associated with CRC progression. The study also points to non-depleted plasma as a useful biospecimen for accurate and effective stage-stratification of CRC patients.