Glioblastoma (GBM) is a highly aggressive brain cancer with poor prognosis and low survival rate. Invasive cancer stem-like cells (CSCs) are responsible for tumor recurrence because they escape current treatments. Our main goal was to study the proteome of three GBM subpopulations to identify key molecules behind GBM cell phenotypes and potential cell markers for migrating cells. We used SuperQuant-an enhanced quantitative proteome approach-to increase proteome coverage. We found 148 proteins differentially regulated in migrating CSCs and 199 proteins differentially regulated in differentiated cells. We used Ingenuity Pathway Analysis (IPA) to predict upstream regulators, downstream effects and canonical pathways associated with regulated proteins. IPA analysis predicted activation of integrin-linked kinase (ILK) signaling, actin cytoskeleton signaling, and lysine demethylase 5B (KDM5B) in CSC migration. Moreover, our data suggested that microRNA-122 (miR-122) is a potential upstream regulator of GBM phenotypes as miR-122 activation was predicted for differentiated cells while its inhibition was predicted for migrating CSCs. Finally, we validated transferrin (TF) and procollagen-lysine 2-oxoglutarate 5-dioxygenase 2 (PLOD2) as potential markers for migrating cells.