Figure legends:
Figure 1. An overview of the study and schematic of methodological workflow. A) Glioma tissue samples were collected from 11 high grade glioma patients and 5 low grade glioma patients. After the lysate preparation and protein digestion, label free quantification was performed. B) The raw files generated from the mass spectrometer was analysed using MaxQuant followed by statistical data analysis. C) Significant proteins were selected from the global proteomics data and multiple reaction monitoring assay was performed for validation. D)In-silico molecular docking of target proteins was performed to identify the potential drug candidates.
Figure 2. Quantitative proteomic profiling and pathway enrichment analysis of glioma samples. A) Heatmap of top 25 differentially expressed proteins in HGG and LGG. B) PCA plot showing two separate clusters for HGG and LGG with a score of 33% for PC1, 13.3% for PC2 and 10.1% for PC3. C) Graphical representation of the significance in the differential expression was quantitatively performed using volcano plot: log10 (p-value) vs. log2 (fold change). D) Gene set enrichment analysis revealed positively and negatively enriched pathways. E) Epithelial Mesenchymal Transition pathway is found to be highly enriched pathways in HGG with normalized enrichment score 1.77. F) Oxidative phosphorylation process is found to be highly enriched pathways in LGG with normalized enrichment score -1.87.
Figure 3. Protein-protein interaction analysis of significant proteins in high grade gliomas and their mapped pathways. Focal adhesion, ECM receptor interaction, N-Glycan biosynthesis and Complement coagulation cascade pathways are found to be highly upregulated in HGGs. Violin plots showing differentially expressed proteins in HGG and LGG (**1.00e-03 < p ≤ 1.00e-02; ***1.00e-04 < p ≤ 1.00e-03; ****p ≤ 1.00e-04).
Figure 4. Validation of significant proteins by multiple reaction monitoring assay. A) each sample was spiked with fixed amount of heavy labelled peptide and the Coefficient of variance is 10%. RPN1 and RPN2 are the subunits of oligosaccharyltransferase (OST). Box plots showing differentially expressed proteins in HGG and LGG (**1.00e-03 < p ≤ 1.00e-02; ***1.00e-04 < p ≤ 1.00e-03).
Figure 5. In-silico molecular docking of catalytic subunits of oligosaccharyltransferase complex with FDA approved drugs.A) The 3D representation of predicted binding pocket of Entrectinib. The binding affinity of Entrectinib with STT3A is -10.7 kcal/mol and with STT3B is -10.6 kcal/mol. B) The 3D representation of predicted binding pocket of Irinotecan. The binding affinity of Irinotecan with STT3A is -11 kcal/mol and with STT3B is -10.6 kcal/mol. C) The predicted 2D interaction map of Entrectinib with STT3A. D) The predicted 2D interaction map of Irinotecan with STT3A.
Figure 6. Molecular mechanism associated with OST mediated aggressive behaviour of high-grade gliomas. Malignant transformation of glioma cells involves extra-cellular matrix remodelling which is accompanied by OST mediated aberrant glycosylation. Glycosylation also results in activation of TGF- β signalling pathways. TGF- β is a strong inducer of EMT transcription factors, drives the EMT in both SMAD and non-SMAD signalling pathways.