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Glioblastoma (GBM) is the most aggressive primary brain tumor with poor patient survival that is at least partly caused by malignant and therapy-resistant glioma stem-like cells (GSLCs) that are protected in GSLC niches. Previously, we have shown that the chemo-attractant stromal-derived factor-1α (SDF-1α), its C-X-C receptor type 4 (CXCR4) and the cysteine protease cathepsin K (CatK) are localized in GSLC niches in glioblastoma. Here, we investigated whether SDF-1α is a niche factor that through its interactions with CXCR4 and/or its second receptor CXCR7 on GSLCs facilitates their homing to niches. Furthermore, we aimed to prove that SDF-1α cleavage by CatK inactivates SDF-1α and inhibits the invasion of GSLCs. We performed mass spectrometric analysis of cleavage products of SDF-1α after proteolysis by CatK. We demonstrated that CatK cleaves SDF-1α at 3 sites in the N-terminus, which is the region of SDF-1α that binds to its receptors. Confocal imaging of human GBM tissue sections confirmed co-localization of SDF-1α and CatK in GSLC niches. In accordance, 2D and 3D invasion experiments using CXCR4/CXCR7-expressing GSLCs and GBM cells showed that SDF-1α had chemotactic activity whereas CatK cleavage products of SDF-1α did not. Besides, CXCR4 inhibitor plerixafor inhibited invasion of CXCR4/CXCR7-expressing GSLCs. In conclusion, CatK can cleave and inactivate SDF-1α. This implies that CatK activity facilitates migration of GSLCs out of niches. We propose that activation of CatK may be a promising strategy to prevent homing of GSLCs in niches and thus render these cells sensitive to chemotherapy and radiation. [ABSTRACT FROM AUTHOR]

Background: Glioblastoma multiforme (GBM) is among the most aggressive cancers with a poor prognosis in spite of a plethora of established diagnostic and prognostic biomarkers and treatment modalities. Therefore, the current goal is the detection of novel biomarkers, possibly detectable in the blood of GBM patients that may enable an early diagnosis and are potential therapeutic targets, leading to more efficient interventions. Experimental Procedures: MicroRNA profiling of 734 human and human-associated viral miRNAs was performed on blood plasma samples from 16 healthy individuals and 16 patients with GBM, using the nCounter miRNA Expression Assay Kits. Results: We identified 19 miRNAs with significantly different plasma levels in GBM patients, compared to the healthy individuals group with the difference limited by a factor of 2. Additionally, 11 viral miRNAs were found differentially expressed in plasma of GBM patients and 24 miRNA levels significantly correlated with the patients’ survival. Moreover, the overlap between the group of candidate miRNAs for diagnostic biomarkers and the group of miRNAs associated with survival, consisted of ten miRNAs, showing both diagnostic and prognostic potential. Among them, hsa miR 592 and hsa miR 514a 3p have not been previously described in GBM and represent novel candidates for selective biomarkers. The possible signalling, induced by the revealed miRNAs is discussed, including those of viral origin, and in particular those related to the impaired immune response in the progression of GBM. Conclusion: The GBM burden is reflected in the alteration of the plasma miRNAs pattern, including viral miRNAs, representing the potential for future clinical application. Therefore proposed biomarker candidate miRNAs should be validated in a larger study of an independent cohort of patients. [ABSTRACT FROM AUTHOR]

Background: Cancer genome and transcriptome analyses advanced our understanding of cancer biology. We performed transcriptome analysis of all known genes of peptidases also called proteases and their endogenous inhibitors in glioblastoma multiforme (GBM), which is one of the most aggressive and deadly types of brain cancers, where unbalanced proteolysis is associated with tumor progression. Methods: Comparisons were performed between the transcriptomics of primary GBM tumors and unmatched non-malignant brain tissue, and between GBM cell lines (U87-MG and U373) and a control human astrocyte cell line (NHA). Publicly-available data sets and our own datasets were integrated and normalized using bioinformatics tools to reveal protease and protease inhibitor genes with deregulated expression in both malignant versus non-malignant tissues and cells. Results: Of the 311 protease genes identified to be differentially expressed in both GBM tissues and cells, 5 genes were highly overexpressed, 2 genes coding for non-peptidase homologues transferrin receptor (TFRC) and G protein-coupled receptor 56 (GPR56), as well as 3 genes coding for the proteases endoplasmic reticulum aminopeptidase 2 (ERAP2), glutamine-fructose-6-phosphate transaminase 2 (GFPT2) and cathepsin K (CTSK), whereas one gene, that of the serine protease carboxypeptidase E (CPE) was strongly reduced in expression. Seventy five protease inhibitor genes were differentially expressed, of which 3 genes were highly overexpressed, the genes coding for stefin B (CSTB), peptidase inhibitor 3 (PI3 also named elafin) and CD74. Seven out of 8 genes (except CSTB) were validated using RT-qPCR in GBM cell lines. CTSK overexpression was validated using RT-qPCR in GBM tissues as well. Cathepsin K immunohistochemical staining and western blotting showed that only proteolytically inactive proforms of cathepsin K were overexpressed in GBM tissues and cells. Conclusions: The presence of high levels of inactive proforms of cathepsin K in GBM tissues and cells indicate that in GBM the proteolytic/collagenolytic role is not its primary function but it plays rather a different yet unknown role. [ABSTRACT FROM AUTHOR]