In-depth evaluation of XPR1 as a new prognostic indicator for endometrial cancer

Abstract

Background: XPR1 is crucial in the development of some tumors, yet its association with endometrial cancer (EC) remains uncertain. We propose that XPR1 exhibits elevated expression in EC and is significantly linked to unfavorable patient outcomes, positioning it as a prospective prognostic biomarker.

Methods: This study investigated XPR1 expression in 554 Uterine Corpus Endometrial Carcinoma(UCEC) cases and 35 normal tissue samples using the The Cancer Genome Atlas(TCGA) database. Western blotting confirmed XPR1 protein presence in EC cell lines (ECC-1) and normal endometrial cells (EEC). The impact of XPR1 on EC cell proliferation and invasion was assessed using EdU proliferation and Transwell invasion assays. A Dot blot assay evaluated m6A methylation of XPR1 in ECC-1 and EEC. The relationship between XPR1 and EC patient prognosis was analyzed using Kaplan-Meier survival and Cox regression analyses. A nomogram was developed to predict 1-, 3-, and 5-year survival probabilities for EC patients, with its accuracy assessed via a calibration curve. Functional enrichment analyses using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were conducted to elucidate biological roles and pathways related to XPR1-associated genes. The 'corrplot' R package analyzed the correlation between XPR1 expression and m6A methylation in EC, as well as its link to immune infiltration. Statistical analyses were performed using R software, with a bilateral p-value < 0.05 considered statistically significant.

Results: This study revealed that XPR1 expression was significantly elevated in EC tissues compared to normal tissues (p < 0.001). Its expression correlated with clinical characteristics including patient age, BMI, clinical stage, histological grade, and tumor invasiveness, suggesting its potential as a prognostic marker.Kaplan-Meier analysis demonstrated that high XPR1 expression correlated with reduced overall survival (HR = 1.60, 95% CI: 1.06-2.42, p = 0.025). However, multivariable Cox regression analysis did not identify XPR1 as an independent prognostic factor.Functional experiments indicated that elevated XPR1 expression promoted proliferative and invasive capacities in endometrial carcinoma cells. XPR1 expression also associated with infiltration levels of immune cells (B cells, CD8 + T cells, CD4 + T cells, macrophages, neutrophils, and dendritic cells), suggesting potential involvement in tumor immune microenvironment regulation.Co-expressed genes with XPR1 were enriched in key biological processes including RNA processing, DNA metabolic regulation, and KEGG signaling pathways. Furthermore, XPR1 showed positive correlations with multiple m6A-related genes, and high XPR1 expression in EC coincided with elevated m6A methylation levels. Although these findings indicate that there is a correlation between XPR1 and m6A modification, it is uncertain that XPR1 can directly regulate m6A modification. The specific mechanism of XPR1 in m6A mediated post transcriptional regulation needs further study.

Conclusion: This study identified a significant association between elevated XPR1 expression and adverse clinical outcomes in endometrial carcinoma patients, with additional connections observed to m6A modification and tumor immune microenvironment dynamics. As a novel prognostic indicator, XPR1 demonstrated correlations with tumor aggressiveness features and biological processes-including cellular proliferation, invasion, and m6A/immune-related pathways-suggesting its potential biological relevance in endometrial carcinogenesis. However, its independent prognostic value remains undefined, and direct mechanistic roles in m6A-mediated post-transcriptional modulation warrant subsequent experimental validation.

Keywords: Endometrial cancer; Immune infiltration; M6A; XPR1.

Fig. 1

Expression of XPR1 in EC. A The expression of XPR1 in Pan cancer was analyzed based on TCGA database, (B) and (C) 35 normal samples and 554 UCEC tumor samples were obtained from TCGA database. After TPM data cleaning, the mRNA expression differences between normal samples and tumor samples were compared by paired and unpaired methods, (D) the protein expression of XPR1 in paracancerous tissues and tumor tissues was detected by Western blotting, (E) the immunohistochemical results of EC paracancerous tissues and cancer tissues were obtained from he human protein atlas database to analyze the expression differences between paracancerous tissues and cancer tissues, (F) the effect of XPR1 in EC was analyzed based on TCGA database, (G) the cbioportal was used.The database was used to analyze the mutation of XPR1 in EC.**indicate p < 0.01

Fig.2

The correlation between XPR1 expression levels and clinicopathological characteristics in EC patients. (A) age, (B) BMI, (C) Clinical stage, (D) Histologic grade, (E) Menopause status, (F) Primary therapy outcome, (G)Residual tumor, (H) Tumor invasion(%),(I)Diabetes,(J)Hormones therapy,(K)OS event,(L)PFI event*, ** and *** indicate p < 0.05, p < 0.01 and p < 0.001, respectively

Fig. 3

Prognostic analysis of XPR1 expression levels and clinicopathological features. (A-L)Prognostic subgroup analyses of EC patients with different clinicopathological status

Fig. 4

High XPR1 expression is not an independent risk factor for overall survival. A Nomogram for predicting OS at 1, 3 and 5 years, (B) Calibration curves for 1, 3 and 5 years

Fig.5

Functional enrichment of XPR1 related genes. A. Volcano map of differential genes, (B) Heat map showing the top 30 genes positively associated with XPR1 in EC, (C) Enrichment analyses of BP of XPR1co-expressed genes, (D) Enrichment analyses of CC of XPR1 co-expressed genes, (E) Enrichment analyses of MF of XPR1 co-expressed genes, (F) Enrichment analyses of XPR1co-expressed gene terms in KEGG. *** indicates p < 0.001

Fig. 6

The correlation between XPR expression level and m6A modification in EC. (A)TCGA UCEC cohort analyzed the association between XPR1 and the expression of 20 m6A-related genes,(B)Dot bolts. (C-M)Scatter plots were plotted to show the association between XPR1 and the expression of m6A-related genes including YTHDF1, YTHDF2, YTHDF3, YTHDC1, YTHDC2, WTAP, RBM15, RBM15B, ZC3H13, HNRNPC, METTL14

Fig. 7

The correlation between XPR1 expression and immune infiltration. (A)The timer database shows the correlation between XPR1 and related immune cells in EC,(B) The correlation between XPR1 and immune infiltrating cells in EC,(C) heatmap shows the relationships among tumor-infiltrating immune cells (TIICs),(D) Differential distribution of immune cells in patients with high XPR1 expression and low XPR1 expression.*, ** and *** indicate p < 0.05, p < 0.01 and p < 0.001, respectively

Fig. 8

XPR1 enhances the proliferation and metastasis of EC cells. A and B Validation of XPR1 protein expression knockdown and overexpression in ECC-1cells. C and D after silencing and overexpressing XPR1 in ECC-1 cells, the proliferation of ECC-1 cells were detected by EdU, (E) and (F) after silencing and overexpressing XPR1 in ECC-1 cells, the invasion of ECC-1 cells cells were detected by Transwell.**, ***and**** indicate p < 0.01, p < 0.001,andp < 0.0001 respectively