Extra Spindle Pole Bodies-Like 1 Serves as a Prognostic Biomarker and Promotes Lung Adenocarcinoma Metastasis

Abstract

Extra spindle pole bodies-like 1 (ESPL1), a cysteine endopeptidase, plays a vital role in chromosome inheritance. However, the association of ESPL1 with prognosis and immune infiltration in lung adenocarcinoma (LUAD) has not yet been explored. Here, we analyzed the expression level, prognostic values, diagnostic value, and immune infiltration level in LUAD using various databases. Immunohistochemistry (IHC) and quantitative real-time PCR (qRT-PCR) assays were used to detect the expression of ESPL1 in LUAD tissues and cell lines. In this study, we found that ESPL1 was upregulated in LUAD and a higher expression of ESPL1 was correlated with unfavorable prognosis in LUAD. Meanwhile, Cox hazard regression analysis results suggested that ESPL1 may be an independent prognostic factor for LUAD. Moreover, we demonstrated that ESPL1 expression was significantly correlated with immune infiltration of Th2 and dendritic cells in LUAD. We also confirmed that DNA copy number amplification and DNA hypo-methylation were positively correlated with ESPL1 expression in LUAD. Additionally, DNA copy number amplification was significantly associated with adverse clinical outcomes in LUAD. Kyoto Encyclopedia of Genes and Genomes (KEGG) and gene set enrichment analysis (GSEA) confirmed that ESPL1 was mainly involved in the DNA replication and glycolysis signaling pathway. Finally, we revealed that ESPL1 was highly expressed in LUAD tissues and cell lines. Knockdown of ESPL1 significantly inhibited cell migration and the invasion abilities of LUAD. Our study comprehensively confirmed that ESPL1 expression may serve as a novel prognostic biomarker for both the clinical outcome and immune cell infiltration in LUAD.

Keywords: DNA methylation; extra spindle pole bodies-like 1; immune infiltration; lung adenocarcinoma; prognosis biomarker.

Figure 1

The expression level and prognosis of ESPL1 in pan-cancer. (A) Pan-cancer expression of ESPL1 between tumor tissues from the TCGA database. (B, C) Kaplan–Meier overall survival of ESPL1 in pan-cancers. NS: p > 0.05, **p < 0.01; ***p < 0.001.

Figure 2

Correlation between ESPL1 mRNA expression and clinicopathological parameters. (A–D) ESPL1 was upregulated in LUAD and LUSC based on the TCGA database. (E) Protein level of ESPL1 in lung cancer tissue examined by the HPA database. (F, G) Correlation between ESPL1 expression and clinical stage, TNM stage, gender, age, residual tumor, smoker, OS event, and DSS event in patients with LUAD. *p < 0.05; **p < 0.01; ***p < 0.001.

Figure 3

ESPL1 knockdown inhibited cell migration and invasion of LUAD. (A, B) IHC assay used to determine the protein of ESPL1 in clinical lung cancer samples. (C) The relative expression of ESPL1 in LUAD cell lines including H1650, H358, A549, and H1299 examined by real-time RT-PCR and human bronchial epithelial (BEAS2B) cell line was used as control. (D) Establishment of ESPL1 knockdown in A549 and H1299 cells, verified by real-time RT-PCR. (E–H) Downregulation of ESPL1 inhibited A549 and H1299 cell migration and invasion by transwell and wound healing assays, Quantification data were also indicated for each assay. Scale bar = 50 μm. **p < 0.01; ***p < 0.001.

Figure 4

Prognostic and diagnostic values of ESPL1 in LUAD. (A–C) Correlation between ESPL1 expression and OS, DSS, and PFS in patients with LUAD examined by the TCGA datasets. (D) ROC curve analysis of the diagnostic values of ESPL1 in LUAD.

Figure 5

Validation of the overall survival of ESPL1 in LUAD. (A–C) Correlation between ESPL1 expression and OS in patients with lung cancer examined by GEO datasets.

Figure 6

Validation of the overall survival of ESPL1 in diverse clinical subtypes. (A–C) Validation of the overall survival of ESPL1 in diverse clinical subtypes, including stage I–II, T1/T2, T3/T4, N0/N1, M0, R0, age>65, female, and smoker.

Figure 7

CNV and DNA methylation analysis. (A, B) Correlation between ESPL1 expression and CNV of ESPL1 in LUAD. (C, D) Correlation between prognosis and CNV of ESPL1 in LUAD examined by the TCGA database. (E) DNA methylation level of ESPL1 in LUAD. (F) Correlation between pathological stage and DNA methylation of ESPL1 in LUAD examined by the TCGA database. (G) Correlation between ESPL1 expression and DNA methylation in LUAD examined by the TCGA database.

Figure 8

Gene–gene and protein–protein interaction network. (A, B) Gene–gene and protein–protein interaction network constructed by the genemania and STRING databases. (C) CancerSEA used to explore the potential role that ESPL1 might play in single LUAD cells.

Figure 9

KEGG enrichment analysis of ESPL1. (A–D) Identified genes with positive co-expression with ESPL1 using the TCGA database. (E–H) GO and KEGG enrichment analysis of ESPL1 in LUAD.

Figure 10

GSEA Identification of ESPL1-related signaling pathways. (A–D) GSEA identification of ESPL1-related signaling pathways in LUAD.

Figure 11

Correlation analysis of ESPL1 expression and infiltration levels of immune cells in LUAD. (A) ESPL1 expression in diverse immune subtypes. (B) Correlation analysis of ESPL1 CNV and infiltration levels of immune cells in LUAD. (C–E) Correlation analysis of ESPL1 expression and infiltration levels of immune cells in LUAD. *p < 0.05; **p < 0.01; ***p < 0.001.

Figure 12

Correlation between ESPL1 expression and immune modulator. (A–D) Examine the relationship between ESPL1 expression, immunostimulators, immunoinhibitors, chemokine, and MHC molecule by using the TISIDB database.