Comprehensive Oncogenic Features of Coronavirus Receptors in Glioblastoma Multiforme

Abstract

The COVID-19 pandemic caused by SARS-CoV-2 infection has placed health systems under excessive pressure and especially elderly people with cancer. Glioblastoma multiforme (GBM) is a malignant brain tumor with an increasing incidence in elderly individuals, and thereby GBM patients are a vulnerable population during the COVID-19 outbreak. Accumulating studies have implied that SARS-CoV-2 might invade the brain directly via coronavirus receptors. However, little is known about SARS-CoV-2 infection in the clinical development of GBM. Here, we explored the oncogenic roles of six coronavirus receptors (ACE2, DPP4, ANPEP, AXL, TMPRSS2, and ENPEP) in GBM using bioinformatics and experimental approaches. We found that ANPEP and ENPEP were significantly increased at both the mRNA and protein levels in GBM compared with normal brain tissue. Kaplan-Meier survival curves and Cox regression analysis demonstrated that high expressions of ANPEP and ENPEP are associated with poor prognosis and survival. Moreover, all receptors are positively correlated with the immune infiltration levels of monocyte. Furthermore, we identified 245 genes between COVID-19 and coronavirus receptors-correlated genes in GBM and performed a thorough analysis of their protein-protein interaction network, functional signaling pathway and molecular process. Our work explores for the first time the association of coronavirus receptors with GBM and suggests ANPEP and ENPEP as potential therapeutic targets of GBM irrespective of COVID-19.

Keywords: ACE2; ANPEP; ENPEP; GBM; SARS-CoV-2.

Figure 1

Expression profile of coronavirus receptors in different regions of the brain. (A) Human brain datasets. Heatmap of the expression profiles of ACE2, DPP4, ANPEP, AXL, TMPRSS2, and ENPEP extracted from the consensus human brain datasets of the Human Protein Atlas (n = 441) (B) Representative image of immunohistochemistry images of coronavirus receptors in the cerebral cortex of human brain (source: The Human Protein Atlas; https://www.proteinatlas.org/humanproteome/brain) (n = 2).

Figure 2

Expression pattern of coronavirus receptors in glioblastoma multiforme (GBM). (A) CoV receptor mRNA in glioblastoma cell lines LN018, LN215, LN229, LN319, and BS149 (recurrent glioblastoma) from GDS4468. (B) Coronavirus receptor mRNA between normal (n = 5) and GBM (n = 156) tissues extracted from the TCGA database by the UALCAN program. (C) Representative images of immunohistochemistry images of receptors in normal (n = 3) and GBM (n = 27) tissue chips. (D) Histologic scores. Mean +SEM. Significance comparison is to normal people. Scale bar = 50 μm. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001, NS: no significance P > 0.05.

Figure 3

Prognosis and survival analysis of coronavirus receptors in GBM. (A) Kaplan–Meier survival curves in the TCGA database. Red indicates high expression, and blue indicates low expression (n = 168). (B) Forest plot for the univariate Cox proportional hazard regression model in the CGGA database (n = 216).

Figure 4

The functional heatmap table of the correlation between coronavirus receptor expressions and immune infiltration levels of different cell types in GBM by TIMER2.0. (A) Monocyte. (B) DCs. (C) NK cell. (D) Eosinophil. n = 153. GBM, glioblastoma.

Figure 5

Coronavirus receptor-related gene enrichment analysis in GBM. (A) An intersection analysis of the top 100 receptor-correlated genes with GBM among different receptor groups according to the E Venn diagram. (B) Bubble chart for KEGG enrichment pathway analysis based on total receptor-related genes. (C) The cnetplot of all genes in the yellow module that depicts the linkages of genes and the most important signaling pathways. (D) Functional and molecular processes related to coronavirus receptor-related genes in GBM.

Figure 6

Enrichment analysis of COVID-19-related genes and coronavirus receptor-related genes in GBM. (A) An intersection analysis between 7,230 COVID-19 genes downloaded from the Comparative Toxicogenomics Database (CTD) and the top 100 related genes of each receptor in GBM. A total of 245 common genes were identified between COVID-19 and coronavirus receptor-correlated genes in GBM. (B) Hierarchical clustering analyses of the direct interaction proteins across 245 common genes with coronavirus receptors. The color key represents the combined score calculated by the selected parameters of the homolog and experimentally determined interaction of the STRING program. Red, yellow and blue refer to high, medium and low combined scores, respectively. (C) Bubble chart for KEGG enrichment pathway analyses of common genes. (D) Bubble chart for GO functional and molecular processes of common genes.

Figure 7

Model for the regulation of ANPEP and ENPEP in GBM against coronavirus infections. Previous studies describe ACE2 and TMPRSS2 as the receptor or co-receptor for SARS-CoV and SARS-CoV-2; AXL for SARS-CoV-2; DPP4 for MERS-CoV; ANPEP for human coronavirus-229E; and DPP4, ENPEP and AENPEP as the candidate receptor for SARS-CoV-2. ANPEP and ENPEP are distributed in endothelial cells of the blood–brain barrier, through which coronaviruses enter the CNS. Protein-protein docking analysis of ANPEP or ENPEP to RBD of SARS-CoV-2 combined with the upregulations of ANPEP and ENPEP in GBM may cause the increase of susceptibility of GBM to SARS-CoV-2. The high levels of ANPEP and ENPEP in GBM is associated with poor survival and high immune infiltration. The overlap of increased risk of GBM to SARS-CoV-2, poor survival, and high immune infiltration may result in the severity of patients with GBM infected by SARS-CoV-2.