Page 167 - Molecular features of low-grade developmental brain tumours

MIR-519D & MIR-4758 CAN IDENTIFY GG FROM DNT AND ASTROCYTOMAS
classification includes histopathological criteria for LEATs 1,7. However, the different GNTs are difficult to distinguish by their histological features even by experienced pathologists 1,3. Improving the classification and the ability to distinguish LEATs from IDH1/2-wildtype low- grade gliomas is becoming more important to avoid unnecessary treatment with chemo- radiation therapy 1,44. Due to the current limitations in the classification of LEATs there is a need for developing accurate molecular techniques that can be used for diagnosis. In this study we evaluated the role of miRNAs in distinguishing GGs from DNTs and other gliomas. We found two miRNAs, miR-519d and miR-4758, upregulated in GGs compared to control tissue, DNTs and other gliomas. Therefore, these two miRNAs could be considered as additional markers in the classification of LEATs, especially in distinguishing GGs from DNTs and other low and high grade gliomas.
Recently, it was suggested that glioneuronal tumours can be separated into two groups: one group enriched for BRAF mutations with an astrocytic expression phenotype and one group enriched for FGFR1 mutations with an oligodendrocyte precursor expression phenotype 23. It would be interesting to see how miR-519d and miR-4758 are expressed in the two groups as suggested by Stone et al., 2018. Here, we evaluated the expression of both miRNAs in BRAF mutated GGs and GGs in which no mutation could be identified and we were unable to find any differences.
Using online prediction databases we identified the PI3K/AKT3/P21 pathway as a predicted target of both miR-519d and miR-4758. Next, we showed that the genes related to the PI3K/AKT3/P21 pathway were differentially expressed in GGs compared to control tissue, indicating that this pathway is deregulated in GGs. Overexpression experiments with miR- 519d and miR-4758 indicate that overexpression of miR-519d increased proliferation, which could be explained by the downregulation of CDKN1A, a key player of the PI3K/AKT3/P21 pathway. This is in agreement with previous studies showing that downregulation of CDKN1A, a direct target of miR-519d, results in increased proliferation 40,41,45. Overexpression of miR- 4758 rescued the expression of CDKN1A and the associated increase in proliferation due to miR-519d, suggesting a tumour suppressive role for miR-4758. However, transfection of miR- 4758 did not downregulate the predicted target gene CDKN1B, highlighting the limitations and the challenges posed by the interpretation of miRNA overexpression experiments to achieve insights into the function of specific miRNAs in the regulation of the cell cycle. Therefore, the exact mechanism behind the rescue effect of miR-4758 is yet to be fully elucidated and requires further investigation.
The upregulation of CDKN1A in GGs along with the slow growing phenotype of GGs could potentially be attributed to the opposing oncogenic properties of miR-519d and miR- 4758. It must be noted that based on our experiments we cannot exclude both miRNAs targeting other genes related to the PI3K/AKT3/P21 pathway affecting GG proliferation.
Overall, our results indicate that miR-519d and miR-4758 are potential regulators of the cell cycle and therefore could play a role in formation of GGs. Furthermore, we showed that these miRNAs could be used as markers to distinguish GGs from DNTs and other low and high grade gliomas, which has implications for determining targeted therapeutic strategies
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