Identification of lncRNAs Associated with Different Outcomes in Brain Tumors

Each year, over 15,000 patients will die from brain cancer. While there have been recent therapeutic advances for brain cancers, the 5-year survival rate hasn’t drastically improved over the past decade. To better understand the molecular determinants of brain cancer, we sought to identify and characterize all of the long non-coding RNAs (lncRNAs) in both grade II and III gliomas as well as grade IV glioblastomas (GBM). We determined the expression of all lncRNAs in over 750 brain cancer and normal RNA-seq datasets from the Cancer Genome Atlas (TCGA). Using a multivariate Cox regression survival model, we identified several hundred lncRNAs that are associated with patient survival. In order to determine whether these lncRNAs play a causative role in brain cancer oncogenic properties, we sought to determine whether the expression of these lncRNAs plays a role in the oncogenic phenotype of brain tumors. To verify that the lncRNAs are expressed sufficiently, their expression was measured via RT-PCR in both A172 and U87 glioblastoma cell lines. These tests left us with 9 lncRNAs to test for phenotypic significance. In both cell lines, the relative nuclear and cytoplasmic localizations have also been determined via cell fractionation followed by RT-PCR, revealing that most of the lncRNAs of interest were primarily located in the cytoplasm. We designed multiple siRNAs to each target lncRNA that was sufficiently expressed in our glioblastoma cell lines. Using U87 and A172 cell lines, several different assays for oncogenic phenotypes were completed after knockdown, including: cell proliferation assays, scratch wound-healing assays and transwell migration assays. A knockdown of approximately 90% has been achieved for each lncRNA, but future work is needed to determine the relative contribution of each lncRNA to brain tumor development and progression.