Cancer stem cells (CSC) have been considered responsible for the therapeutic-resistance/recurrence of cancers, and thus proposed as the therapeutic target. To clarify molecular mechanisms and develop clinical targets against CSC, we established CSC-clones from glioblastoma patient’s tissues as glioma stem cells(GSCs), having a potential of differentiation/promotion of glioblastoma, and subjected to a unique integrated Phospho-glycomics. Using original GSC-iPEACH database, we tried to identify specific phospho-glyco networks associated to the maintenance/differentiation of GSC.
We established 12 GSC clones from patient’s gliomas and glioma cell lines having the potential to differentiate into glioblastomas, extracted their Proteins and mRNAs, and subjected to iTRAQ/TMT, DNA array, Rectin/glyco-gene array, and quantitative phospho-proteome (HAMMOC) analyses. All of the data was integrated by iPEACH, and used for GO and knowledge-based network analyses. Biological validations were performed with immunocyto/histochemistry, western-blotting, mouse xenograft analyses.
The data integration and extraction of specific phospho- and glyco-proteins, and their related genes revealed that, during the GSC differentiation, cell surface glyco-proteins and RAS-MAPK/PI3K signalings were significantly up-regulated, meanwhile, SOX2, CD133, and specific proteoglycans/synthetic-enzymes/metabolic pathways were obviously down-regulated. Interestingly, GSC differentiation was significantly associated with the decrease of chondroitin Sulfate(CS)-modified proteins/related enzymes. By the CS-degradation enzyme treatment, GSC differentiation was dramatically induced and this was associated with the up-regulation of specific cellular phosphokinase dependent signalings. Importantly, these differentiation processes were also associated with the interaction of CS-proteins and adhesion molecules such as integrins, and suppressed by integrin-inhibitors/CS administrations significantly. Combination treatments of a cancer-drug Temozolomide and these GSC-differentiation inhibitors suppressed glioma progression, increased the chemosensitivities, and led the longer survival of mouse xenograft-models.
Functional integrated phospho-glycomics for the first time demonstrates that the GSC induces the specific glyco-phospho network/signalings to regulate GSC stemness/differentiation, and these information will provide the new therapeutic strategies/clinical targets against malignant gliomas.