EEPD1 is identified as a predictor of prognosis and immune microenvironment through pan-cancer analysis and related to progression of colorectal cancer

Abstract

Background: EEPD1 is vital in homologous recombination, while its role in cancer remains unclear.

Methods: We performed multiple pan-cancer analyses of EEPD1 with bioinformatics methods, such as gene expression, gene alterations, Prognosis and enrichment analysis, tumor microenvironment, immune cell infiltration, TMB, MSI, immunotherapy, co-expression of genes, and drug resistance. Finally, RT-qPCR, EdU, and transwell assays helped investigate the impact of EEPD1 on CRC cells.

Results: EEPD1 was dysregulated and correlated with bad prognosis in several cancers. GSVA and GSEA revealed that EEPD1 was primarily associated with the "WNT_BETA_CATENIN_SIGNALING," "ribonucleoprotein complex biogenesis," "Ribosome," and "rRNA processing." The infiltration of CD8⁺ T cells, MAIT cells, iTreg cells, NK cells, Tc cells, Tex cells, Tfh cells, and Th1 cells were negatively correlated with EEPD1 expression. Additionally, EEPD1 is significantly associated with TMB and MSI in COAD, while enhanced CRC cell proliferation and migration.

Conclusions: EEPD1 was dysregulated in human cancers and correlated with various cancer patient prognoses. The dysregulated EEPD1 expression can affect tumor-infiltrating immune cells and immunotherapy response. Therefore, EEPD1 could act as an oncogene associated with immune cell infiltration in CRC.

Keywords: CRC; Cell migration; Cell proliferation; EEPD1; Immune infiltration.

Fig. 1

EEPD1 Pan-Cancer expression and genetic alteration. (A) EEPD1 Pan-cancer differential expression according to TCGA and GTEx database. (B) Mean expression of EEPD1 in multiple cancers. (C) EEPD1 expression in COAD tumor tissues and normal tissues. (D) EPD1 expression in paired COAD tumor tissues and normal tissue. (E) EEPD1 pan-cancer genetic alterations in cBioportal database.

Fig. 2

Association between EEPD1 and prognosis and clinic phenotypes. (A) Forest plot based on the univariate COX regression analysis of EEPD1 and OS in 33 cancers (B) Kaplan-Meier analysis of association between EEPD1 and prognosis in cancers. (C) Association between EEPD1 and tumor stage in cancers.

Fig. 3

Enrichment analysis of EEPD1 in colorectal cancer. (A) correlation analysis of EEPD1 in colorectal cancer. (B) GEVA of EEPD1 in colorectal cancer. (C–E) GSEA of EEPD1 based on GO, KEGG pathways and Reactome pathways.

Fig. 4

TME analysis of EEPD1. (A) ESTIMATE analysis of EEPD1 in caners. (B) ESTIMATE analysis of EEPD1 in CRC. (C) The correlation between the expression level of EEPD1 and TME in COAD.

Fig. 5

The correlation between the expression of EEPD1 and pan-cancer immune cell infiltration based on TIMER2 (Red boxes represent positive associations, green negative associations, and boxes with a cross indicate p > 0.05). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 6

(A)The correlation between EEPD1 expression and immune cell infiltration in colorectal cancer. (B)Circle plot of the correlation between EEPD1 expression and immune cell infiltration in colorectal cancer. cells with pearson correlation coefficient >0.15 with EEPD1 were selected for correlation circle mapping. Red represents positive correlation, green represents negative correlation, and the darker the color, the stronger the correlation. (C)The correlation between EEPD1 expression and TMB. (D) The correlation between EEPD1 expression and MSI. (E) The correlation between EEPD1 expression and tumor response to PD-L1 inhibitor. (F) Kaplan-Meier analysis of association between EEPD1 and prognosis in PD-L1 treated patients. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 7

The gene co-expression analysis of EEPD1 and (A) immune checkpoints, (B) chemokines, (C) chemokines receptors, (D) immune activation genes, (E) immunosuppressive genes, (F) MHC genes.

Fig. 8

The gene co-expression analysis of EEPD1 and (A) EMT down, (B) EMT up, (C) pyroptosis, (D) autophagy, (E) TGF-β1 signaling, (F) Wnt/β-catenin signaling, (G) ferroptosis, (H) MMR genes.

Fig. 9

The correlation between EEPD1 expression and IC50 of multiple drugs. (A–S) The EEPD1 expression is positive with IC50 of drugs. (T–Z) The EEPD1 expression is negative with IC50 of drugs.

Fig. 10

(A)The nomogram to predict OS of CRC patients. (B) Calibration plot for the nomogram. (C) ROC curve for EEPD1 and other clinical parameters. (D) ROC curve for nomogram.

Fig. 11

EEPD1 promote cell proliferation and metastasis in CRC cells lines. (A) EEPD1 was significantly up-regulated in CRC tumor tissues According to immunohistochemistry (B) EEPD1 was significantly up-regulated in CRC tumor tissues According to RT-qPCR. (C) The expression of EEPD1 in CRC cell lines was higher than that in normal colorectal cell lines. (D)EEPD1 was knock down in HCT8 and RKO cell lines. (E, F) knockdown of EEPD1 significantly reduced the EdU-positive rate of EdU assay in RKO and HCT8 cells. (G, H) knockdown of EEPD1 significantly reduced the number of cells that cross the TRANSWELL chamber. (I, J) knockdown of EEPD1 significantly promoted the expression of P21 and P16 and showed an upward trend in the expression of Bad.

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