YBX1 as a prognostic biomarker and potential therapeutic target in hepatocellular carcinoma: A comprehensive investigation through bioinformatics analysis and in vitro study

Abstract

Backgrounds: Y-box binding protein 1 (YBX1) is a DNA/RNA binding protein known to contribute to the progression of various malignancies, however, a comprehensive pan-cancer analysis to investigate YBX1 across a broad spectrum of cancer types has not yet been conducted.

Methods: We utilized the TIMER database for a comprehensive pan-cancer analysis and assessed YBX-1 expression via the TCGA and GEO databases. The relationship between YBX-1 expression and tumor-infiltrating cells was examined using TIMER and the R programming language. To evaluate the prognostic value of YBX1, we performed Kaplan-Meier plots and Cox regression analyses. Through LinkedOmics, we identified genes significantly correlated with YBX-1. The WEB-based Gene SeT AnaLysis Toolkit was used for KEGG pathway enrichment analysis. Additionally, using shRNA-mediated knockdown, we explored the potential role of YBX1 in tumor cell biology.

Results: Our study identifies pronounced overexpression of YBX-1 across multiple cancer types, correlating with adverse outcomes, notably in liver hepatocellular carcinoma (LIHC). A distinct association between elevated YBX-1 expression and heightened immune cell infiltration suggests YBX-1's potential role in reshaping the tumor microenvironment. Intriguingly, our KEGG pathway analysis indicated a tight nexus between YBX-1 expression and lipid metabolism. Moreover, the suppression of YBX-1 via shRNA revealed diminished cellular proliferation and marked reductions in crucial molecules steering the fatty acid synthesis pathway, implicating YBX-1's potential regulatory role in lipid metabolism within LIHC.

Conclusions: YBX-1 serves as a favorable prognostic indicator in various cancers, particularly in liver hepatocellular carcinoma. Targeting YBX1 in HCC offers potential therapeutic strategies. This work paves the way for fresh insights into targeted therapeutic approaches for cancers, especially benefiting liver hepatocellular carcinoma patients.

Keywords: Fatty acid metabolism; Liver hepatocellular carcinoma; Pan cancer; YBX1.

Fig. 1

YBX1 expression profile in pan-cancer. (A) YBX1 gene expression in adjacent normal and tumor tissues across various tumor types in TCGA dataset as analyzed via TIMER. (B) The expression of YBX1 in pan-cancer and paired normal tissues in TCGA dataset. (C) Protein expression of YBX1 across cancers from Clinical Proteomic Tumor Analysis Consortium (CPTAC) database. **, P < 0.01; ***, P < 0.001; ns, no significance. TPM, transcripts per million.

Fig. 2

Exploring the potential relationship between YBX1 and different levels of immune cell infiltration in pan cancer. (A) Correlation analysis of immune cells between YBX1 and pan-cancer cells using TIMER method. (B) The correlation of YBX1 and the immune checkpoint-related genes SIGLEC15, TIGIT, CD274, HAVCR2, PDCD1, CTLA4, LAG3, PDCD1LG2 in pan-cancer plotted with the R language as a heat map. (C-D) Correlation analysis of YBX1 expression with MSI and TMB. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Fig. 3

Survival analysis of YBX1 in HNSC, STAD, COAD, LIHC, LUSC, BRCA, KICH, KIRC, THCA, CHOL cancers. (A) Overall survival (OS). (B) Progression-free survival (PFS). (C) Disease-free survival (DFS). (D) Disease-specific survival (DSS).

Fig. 4

YBX-1 expression in LIHC. (A-C) Expression level of YBX1 in tumor and normal tissues in TCGA-LIHC database and GTEx database. (D) YBX1 mRNA expression in LIHC tissues and paired adjacent normal tissues from GSE64041 dataset. (E) Diagnostic value of YBX1 in LIHC shown with a ROC curve. (F) Protein expression of YBX-1 in hepatocellular carcinoma from CPTAC database. (G) Representative immunohistochemical staining for YBX1 expression in HCC tissues and corresponding normal tissues by using tissue microarrays. (H) Immunohistochemical staining scores of YBX1 expression in HCC tissues (n = 48) and adjacent normal tissues (n = 48) from the tissue microarray. ***, P < 0.001.

Fig. 5

Kaplan-Meier curves of (A) OS, (B) PFS, (C) DFS, (D) DSS in HCC.

Fig. 6

Gene co-expression, GO-BP and KEGG pathway analysis of YBX-1. (A) KEGG pathway analysis of YBX-1 correlated genes in LIHC. (B) The top 50 genes positively and negatively correlated with YBX-1 in LIHC extracted from Linkdomics. (C) The top-level Gene Ontology biological processes obtained from Metascape database. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Fig. 7

Immune association analysis in HCC. (A) Association between YBX-1 expression and immune infiltration levels in LIHC using the TIMER database. (B) Differential levels of 24 immune cell subtypes in high and low YBX-1 expression groups in HCC. (C) Correlation analysis between immune cell infiltration levels and YBX-1 expression in HCC using Spearman's method. *, p < 0.05; **, p < 0.01; ***, p < 0.001; ns, no significance.

Fig. 8

YBX-1 regulate cell growth through mediating fatty acid metabolism. (A) Heat map to show the relationship between YBX-1 expression and fatty acid metabolic pathway. (B) Colony formation in LV-sh NC and LV-sh YBX-1 cells. (C-D) The cell growth curves of HepG2 LV-sh NC, HepG2 LV-sh YBX-1, Huh LV-NC and Huh7 LV-sh YBX-1. (E-F) Huh7 and HepG2 Cells were exposed to different concentrations of SU056 for 72 h, and the OD450 was measured after incubation with CCK-8. (G) Western Blot analysis of fatty acid metabolic related proteins. *, p < 0.05; **, p < 0.01; ***, p < 0.001; ns, no significance.