Glioblastomas (GBM) are the most aggressive form of primary brain tumors in humans. A key feature of malignant gliomas is their cellular heterogeneity. In particular, the presence of an undifferentiated cell population defined Glioblastoma Stem cells (GSCs) has been described. Increased expression of anti-apoptotic and chemo-resistant genes in GCSs subpopulation, favours their high resistance to a broad spectrum of drugs.
CSN5 and CSN6, the critical subunits of the constitutive photomorphogenesis 9 (COP9) signalosome (CSN), has received attention as a regulator of the degradation of cancer-related proteins such as p53, c-myc and c-Jun, through the ubiquitin-proteasome system, suggesting its relevance in cancerogenesis. However, the biological functions and molecular mechanisms of CSN in glioblastoma (GBM) and in particular in GSCs remain poorly understood. Recently has been proposed that CSN6 positively regulated EGFR stability via reduced levels of EGFR ubiquitination. As reported in yeast, the deletion of the CSN5 gene causes the activation of the transcription of two autophagic genes (ATG16 and ATG33) and an increase of the vacuolar size, suggesting the autophagy activation in the growth phase in which autophagy is usually not required. In cancer cells, autophagy has been recognized as an adaptive response to starvation or stress conditions. However, prolonged autophagy leads to a high turnover rate of proteins and organelles above the survival threshold, resulting in cancer cells death. This adaptation can be utilized as a therapeutic tool for the cancer treatment because the use of molecules promoting autophagy could significantly increase the cell death in the tumors. In this project we propose to test the role played by CSN5 and CSN6 in GSCs derived from human biopsies. Moreover, the use of specific inhibitors of CSN5 activity will be tested in order to evaluate the effects on cell proliferation, EGFR activity and autophagic processes induction.
In recent years the interest on the neddylation pathway as an antitumor target is significantly increasing since the inhibitor of the activating enzyme Nedd8 (MLN4924) has reached phase 1 and 2 in some clinical trials. In addition, a new molecule, CSN5i-3 was recently described as a CSN5 inhibitor with anticancer activity. Indeed, since that an up regulation of the human CSN5 enzyme is present in many tumors, its inibitors may be considered a promising tool for cancer therapy in the future. Considering that glioblastoma multiforme is the most aggressive brain tumor in human and given the high inefficacy the surgical and pharmacological therapies in the treatment of this tumor, in this project we propose to test on glioblastoma cancer stem cells (GB7, GB8) obtained from human bipesies of two different patients, two inhibitors of Csn5 previously selected from a screening performed in yeast.
Taking advantage of the phenotypes observed in yeast using the Csn5 inhibitors, we will be able to address our experiments to the autophagic pathway and the mitochondrial function, a field still unexplored in a glioblastoma model. Moreover we also evaluated the ability of these inhibitors to counteract the EGFR pathway stability, an other pathway that may support the glioblastoma cancer stem cells survival and proliferation.