Identification of Novel Tumor-Associated Cell Surface Sialoglycoproteins in Human Glioblastoma Tumors Using Quantitative Proteomics

Abstract

Glioblastoma multiform (GBM) remains clinical indication with significant “unmet medical need”. Innovative new therapy to eliminate residual tumor cells and prevent tumor recurrences is critically needed for this deadly disease. A major challenge of GBM research has been the identification of novel molecular therapeutic targets and accurate diagnostic/prognostic biomarkers. Many of the current clinical therapeutic targets of immunotoxins and ligand-directed toxins for high-grade glioma (HGG) cells are surface sialylated glycoproteins. Therefore, methods that systematically and quantitatively analyze cell surface sialoglycoproteins in human clinical tumor samples would be useful for the identification of potential diagnostic markers and therapeutic targets for malignant gliomas. In this study, we used the bioorthogonal chemical reporter strategy (BOCR) in combination with label-free quantitative mass spectrometry (LFQ-MS) to characterize and accurately quantify the individual cell surface sialoproteome in human GBM tissues, in fetal, adult human astrocytes, and in human neural progenitor cells (NPCs). We identified and quantified a total of 843 proteins, including 801 glycoproteins. Among the 843 proteins, 606 (72%) are known cell surface or secreted glycoproteins, including 156 CD-antigens, all major classes of cell surface receptor proteins, transporters, and adhesion proteins. Our findings identified several known as well as new cell surface antigens whose expression is predominantly restricted to human GBM tumors as confirmed by microarray transcription profiling, quantitative RT-PCR and immunohistochemical staining. This report presents the comprehensive identification of new biomarkers and therapeutic targets for the treatment of malignant gliomas using quantitative sialoglycoproteomics with clinically relevant, patient derived primary glioma cells.

Bioorthogonal Labeling, Bioimaging, and Photocytotoxicity Studies of Phosphorescent Ruthenium(II) Polypyridine Dibenzocyclooctyne Complexes

“…This is consistent with previous findings that more N ‐azidoglycans can be detected on the cell surface of wild‐type CHO cell lines than many other human and animal cell lines, such as HeLa (human cervix carcinoma cells), COS‐7 (monkey kidney fibroblasts), and S49 (mouse lymphoma cells),19 indicating that the capability of metabolizing Ac 4 ManNAz is highly cell line‐dependent 19. 20…”

Molecular Identity of Human Outer Radial Glia during Cortical Development

“…We confirmed by immunostaining that TNC and PTPRZ1 are expressed in a subset of ITGB5 positive oRG cells (Figure 4E). Many of the cell surface proteins enriched in oRG cells are also highly overexpressed in glioblastomas compared with normal human astrocytes including TNC, PTPRZ1, and LGALS3BP (Autelitano et al, 2014; Nie et al, 2015), and PTN stimulation of PTPRZ1 is sufficient to stimulate the coordinated processes of epithelial-to-mesenchymal transition in a glioblastoma cell line (Perez-Pinera et al, 2006). Therefore, we investigated whether human glioblastoma samples contain cell populations that co-express oRG markers.…”

Identification of Plasma Membrane Glycoproteins Specific to Human Glioblastoma Multiforme Cells Using Lectin Arrays and LC‐MS/MS

“…Autelitano et al. identified hundreds of cell surface N ‐linked sialylated glycoproteins that are differentially expressed in primary cultures derived from GBM patient tissues in comparison with fetal and adult astrocytes and neural progenitor cells isolated from human brain by labeling sialylated glycans followed by label‐free quantitative MS . While these chemical labeling strategies allowed for extensive identification of cell surface glycoproteins expressed in malignant gliomas, lectins that recognize and bind to specific carbohydrate structural epitopes have also been used to characterize the carbohydrate expression patterns of glioblastoma‐derived stem cells.…”