CircZNF609 and circNFIX as possible regulators of glioblastoma pathogenesis via miR-145-5p/EGFR axis

Abstract

Glioblastoma is a rare and deadly malignancy with a low survival rate. Emerging evidence has shown that aberrantly expressed circular RNAs (circRNAs) play a critical role in the initiation and progression of GBM tumorigenesis. The oncogenic function of circZNF609 and circNFIX is involved in several types of cancer, but the role and underlying mechanism of these circRNAs in glioblastoma remain unclear. In this study, we hypothesized that circZNF609 and circNFIX may regulate EGFR through sponging miR-145-5p. Herein, we assessed the expression levels of circZNF609, circNFIX, miR-145-5p, and EGFR using quantitative polymerase chain reaction in glioblastoma patients and normal brain samples. The results showed that circZNF609, circNFIX, and EGFR expression levels were upregulated and miR145-5p was downregulated (p = 0.001, 0.06, 0.002, and 0.0065, respectively), while there was no significant association between clinicopathological features of the patients and the level of these genes expression. We also found a significant inverse correlation between miR145-5p and the expression of cZNF609, cNFIX and EGFR (p = 0.0003, 0.0006, and 0.009, respectively). These findings may open a new window for researchers to better understand the potential pathways involved in GBM pathogenesis. In conclusion, it may provide a new potential pathway for the development of effective drugs for the treatment of GBM patients.

Keywords: CircNFIX; CircZNF609; EGFR; miR-145-5p; Glioblastoma.

Figure 1

Volcano diagram of differentially expressed miRNAs in glioblastoma. |log2FC|> 1 and adj p-value < 0.05 were considered as cut-off values for the identification of DE-miRNAs. Blue and red dots denote the downregulated and upregulated miRNAs in glioblastoma samples; grey dots denote miRNAs without differential expression between GBM and normal samples. The arrow represents the miR-145-5p.

Figure 2

Predicted target genes of miR-145-5p. miRNet was used for establishing a network of miR-145-5p and their target genes. Blue rectangle represents has-mir-145-5p; purple and orange dots represent target genes and circRNAs; the dots shown with two colors represent circRNA-gene.

Figure 3

Circ-ZNF609 and circ-NFIX may act as ceRNAs to bind to miR-145-5p with EGFR. Interaction sites between (A) circ0000615, (B) circ0049658 and miR-145-5p according to IntaRNA. (C) Three interaction sites between miR-145-5p and EGFR based on bioinformatics website (miRanda and miRTarBase).

Figure 4

Schematic representation of (A) circ0000615 and (B) circ0049658 biogenesis. The back splice junction site of these circRNAs was confirmed by Sanger sequencing.

Figure 5

Relative expression of (A) circZNF609, (B) circNFIX, (C) miR-145-5p and (d) EGFR genes in glioblastoma samples, compared with normal brain tissues. ^** indicates a significant difference between two groups (p < 0.01) and ns indicates that not statistically significantly different among the groups (p > 0.05).

Figure 6

The correlation between expression levels of (A) circZNF609 and miR-145-5p, (B) circNFIX and miR-145-5p (C) miR-145-5p and EGFR, (D) circZNF609 and EGFR, and (E) circNFIX and EGFR genes in GBM tissues compared with normal brain samples.

Figure 7

Grouped graph of expression correlation of cZNF609, cNFIX, miR-145-5p and EGFR in GBM and normal tissues as calculated by Pearson correlation.

Figure 8

ROC curve analysis of (A) circNFIX and (B) circZNF609 expression for discrimination of GBM tumors from normal tissues.