HGG-17. Malignant role of regulatory lncRNA LINC01271-mediated metastatic progression in Glioblastoma

Introduction Glioblastoma (GBM) is an aggressive and malignant brain tumor that accounts for only a 6.9% five-year survival rate. Programmed Death Ligand 1 (PD-L1) is an immunosuppressive molecule expressed on tumor cells and facilitates the growth and migration of GBM. High PD-L1 expression in GBM tumors is associated with poor survival. Recently, long non-coding RNAs (lncRNAs) have been found to correlate with PD-L1 in the regulation of GBM malignancy. Therefore, targeting lncRNAs-PD-L1 pathways can control the course of GBM as therapeutic potential. Method Human glioblastoma (U-87) cells were cultured in a 6-well plate and transfected with PD-L1-CRISPR-Cas9 plasmid for knock-in and knock-out. The PD-L1 overexpression and deletion of U-87 cells were observed under a microscope. Metastatic growth of U87 cells under PD-L1 overexpression conditions was assessed using MTT, microscopy, and scratch assay. Extracted RNA was subjected to a real-time PCR assay using human-specific PD-L1 and lncRNAs (INHEG, LINC01271, and UNC5B). Further, siRNA transfection was performed to silence LINC01271, which has been used to correlate with PD-L1 expression. Results A significant increase and decrease of PD-L1 expression was obtained in knock-in and knock-out U87 cells. Overexpression of PD-L1 promotes U-87 proliferation. The scratch assay showed the migration of U-87 was largely enhanced in PD-L1 overexpressed cells compared to the control, suggesting metastatic progression. The PD-L1-overexpressed U-87 cells showed a statistically significant 7-fold increase (p = 0.02*) in LINC01271 gene expression compared to the control and knock-out U-87 cells (p = 0.01*). In contrast, UNC5B and INHEG showed a 4.8-fold (p = 0.001**) and 2.9-fold (p = 0.03*) increase in PD-L1-overexpressed U-87 cells, respectively. Importantly, a higher PD-L1 expressed (p > 0.0001***) in U-87 cells correlates to the up-regulation of LINC01271. LINC01271-specific siRNA silencing significantly reduced the expression of PD-L1 (p = 0.0007***) compared to the control U-87. Conclusion The positive correlation in the expression of PD-L1 and LINC01271 indicates the oncogenic roles in GBM. PD-L1-dependent LINC01271 expression in GBM modulates the progression of GBM and its severe outcome.