MicroRNAs expression analysis in high grade gliomas

Several biomarkers had been proposed as useful parameters to better define the prognosis or to delineate new target therapy strategies for glioblastoma (GBM) patients. MicroRNAs could represent interesting molecules, for their role in tumorigenesis and cancer progression and for their specific tissue expression. Although many studies have tried to identify a specific microRNAs signature for glioblastoma, by now an exhaustive GBM microRNAs profile is far to be well defined. In this work we set up a real-time qPCR, based on LNA primers, to investigate the expression of 19 microRNAs in brain tumors, focusing our attention on GBMs. MiRNAs expression in 30 GBM paired FFPE-Fresh/Frozen samples was firstly analyzed. The good correlation obtained comparing miRNAs results confirmed the feasibility of performing miRNAs analysis starting from FFPE tissues. This leads to many advantages, as a good disposal of archival tumor and normal brain specimens and the possibility to verify the percentage of tumor cells in the analyzed sample. In the second part we compared 3 non-neoplastic brain references to use as control in miRNAs analysis. Normal adjacent the tumor, epileptic specimens and a commercial total RNA were analyzed for miRNAs expression and results showed that different non-neoplastic controls could lead to important discrepancies in GBM miRNAs profiles. Analyzing 50 FFPE GBMs using all 3 non-neoplastic references, we defined a putative GBM miRNAs signature: mir-10b, miR-21 and miR-27a resulted upregulated, while miR-7, miR-9, miR-26a, miR-31, miR-101, miR-137, miR-222 and miR-330 were downregulated. Comparing miRNAs expression among GBM group and gliomas of grade I, II and III, we obtained 3 miRNAs (miR-10b, mir-34a and miR-101) showing a different regulation status between high grade and low grade gliomas. Intriguingly, miR-10b was upregulated in high grade and significantly downregulated in low grade gliomas, suggesting that could be a candidate for a GBM target therapy.