title: Probing the O-glycoproteome of gastric cancer cell lines for biomarker discovery.
abstract: Circulating O-glycoproteins shed from cancer cells represent important serum biomarkers for diagnostic and prognostic purposes. We have recently shown that selective detection of cancer-associated aberrant glycoforms of circulating O-glycoprotein biomarkers can increase specificity of cancer biomarker assays. However, the current knowledge of secreted and circulating O-glycoproteins is limited. Here, we used the COSMC KO "SimpleCell" (SC) strategy to characterize the O-glycoproteome of two gastric cancer SimpleCell lines (AGS, MKN45) as well as a gastric cell line (KATO III) which naturally expresses at least partially truncated O-glycans. Overall, we identified 499 O-glycoproteins and 1236 O-glycosites in gastric cancer SimpleCells, and a total 47 O-glycoproteins and 73 O-glycosites in the KATO III cell line. We next modified the glycoproteomic strategy to apply it to pools of sera from gastric cancer and healthy individuals to identify circulating O-glycoproteins with the STn glycoform. We identified 37 O-glycoproteins in the pool of cancer sera, and only nine of these were also found in sera from healthy individuals. Two identified candidate O-glycoprotein biomarkers (CD44 and GalNAc-T5) circulating with the STn glycoform were further validated as being expressed in gastric cancer tissue. A proximity ligation assay was used to show that CD44 was expressed with the STn glycoform in gastric cancer tissues. The study provides a discovery strategy for aberrantly glycosylated O-glycoproteins and a set of O-glycoprotein candidates with biomarker potential in gastric cancer.
authors: From the ‡Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark; §IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal;
authors: Diana
authors: D
authors: Campos
authors: §IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal;
authors: Daniela
authors: D
authors: Freitas
authors: §IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal;
authors: Joana
authors: J
authors: Gomes
authors: §IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal;
authors: Ana
authors: A
authors: Magalhães
authors: From the ‡Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark;
authors: Catharina
authors: C
authors: Steentoft
authors: §IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal;
authors: Catarina
authors: C
authors: Gomes
authors: From the ‡Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark;
authors: Malene B
authors: MB
authors: Vester-Christensen
authors: ¶Experimental Pathology and Therapeutics Group, Portuguese Institute of Oncology, Rua Dr. António Bernardino de Almeida 4200-072 Porto, Portugal; ‖QOPNA, Department of Chemistry of the University of Aveiro, Campus Universitário de Santiago 3810-193 Aveiro, Portugal;
authors: José Alexandre
authors: JA
authors: Ferreira
authors: **Department of Pathology, Portuguese Institute of Oncology, Rua Dr. António Bernardino de Almeida 4200-072 Porto, Portugal;
authors: Luis P
authors: LP
authors: Afonso
authors: ¶Experimental Pathology and Therapeutics Group, Portuguese Institute of Oncology, Rua Dr. António Bernardino de Almeida 4200-072 Porto, Portugal;
authors: Lúcio L
authors: LL
authors: Santos
authors: ‡‡Faculty of Medicine of the University of Porto, Al. Prof. Hernâni Monteiro, 4200-319 Porto, Portugal;
authors: João
authors: J
authors: Pinto de Sousa
authors: From the ‡Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark;
authors: Ulla
authors: U
authors: Mandel
authors: From the ‡Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark;
authors: Henrik
authors: H
authors: Clausen
authors: From the ‡Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark; seva@sund.ku.dk celsor@ipatimup.pt.
authors: Sergey Y
authors: SY
authors: Vakhrushev
authors: §IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Rua Dr. Roberto Frias s/n, 4200-465 Porto, Portugal; ‡‡Faculty of Medicine of the University of Porto, Al. Prof. Hernâni Monteiro, 4200-319 Porto, Portugal; §§Institute of Biomedical Sciences Abel Salazar, ICBAS, Rua de Jorge Viterbo Ferreira n.228, 4050-313 Porto, Portugal seva@sund.ku.dk celsor@ipatimup.pt.
authors: Celso A
authors: CA
authors: Reis
mesh_terms: D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000097:blood / Q000378:metabolism*; D045744:Cell Line, Tumor; D005260:Female; D006023:Glycoproteins / Q000097:blood / Q000378:metabolism*; D006801:Humans; D008297:Male; D008875:Middle Aged; D017350:N-Acetylgalactosaminyltransferases / Q000097:blood / Q000378:metabolism*; D020543:Proteome; D013274:Stomach Neoplasms / Q000097:blood / Q000378:metabolism*; D000097763:Polypeptide N-acetylgalactosaminyltransferase
chemical_list: D014408:Biomarkers, Tumor; D006023:Glycoproteins; D017350:N-Acetylgalactosaminyltransferases; D020543:Proteome; D000097763:Polypeptide N-acetylgalactosaminyltransferase