Prognostic and Immunological Potential of Ribonucleotide Reductase Subunits in Liver Cancer

Abstract

Background: Ribonucleotide reductase (RR) consists of two subunits, the large subunit RRM1 and the small subunit (RRM2 or RRM2B), which is essential for DNA replication. Dysregulations of RR were implicated in multiple types of cancer. However, the abnormal expressions and biologic functions of RR subunits in liver cancer remain to be elucidated.

Methods: TCGA, HCCDB, CCLE, HPA, cBioPortal, and GeneMANIA were utilized to perform bioinformatics analysis of RR subunits in the liver cancer. GO, KEGG, and GSEA were used for enrichment analysis.

Results: The expressions of RRM1, RRM2, and RRM2B were remarkably upregulated among liver cancer tissue both in mRNA and protein levels. High expression of RRM1 and RRM2 was notably associated with high tumor grade, high stage, short overall survival, and disease-specific survival. Enrichment analyses indicated that RRM1 and RRM2 were related to DNA replication, cell cycle, regulation of nuclear division, DNA repair, and DNA recombination. Correlation analysis indicated that RRM1 and RRM2 were significantly associated with several subsets of immune cell, including Th2 cells, cytotoxic cells, and neutrophils. RRM2B expression was positively associated with immune score and stromal score. Chemosensitivity analysis revealed that sensitivity of nelarabine was positively associated with high expressions of RRM1 and RRM2. The sensitivity of rapamycin was positively associated with high expressions of RRM2B.

Conclusion: Our findings demonstrated high expression profiles of RR subunits in liver cancer, which may provide novel insights for predicting the poor prognosis and increased chemosensitivity of liver cancer in clinic.

Figure 1

Expression of RR subunits in liver cancer and cancer cell lines. (a) mRNA expression levels of RRM1, RRM2, and RRM2B between liver cancer (tumor) and adjacent nonmalignant liver tissue (normal) in The Cancer Genome Atlas Liver Hepatocellular Carcinoma (TCGA-LIHC) cohort. (b) mRNA expression levels of RRM1, RRM2, and RRM2B in paired tumor and adjacent nonmalignant (normal) tissues from TCGA-LIHC cohort. (c) mRNA expression levels of RRM1 among various liver cancer cohorts in the Hepatocellular Carcinoma Expression Atlas Database (HCCDB). (d) mRNA expression levels of RRM2 among various liver cancer cohorts in the HCCDB. (e) mRNA expression levels of RRM2B among various liver cancer cohorts in the HCCDB. (f) mRNA expression levels of RRM1 in a variety of liver cancer cell lines from Cancer Cell Line Encyclopedia (CCLE). (g) mRNA expression levels of RRM2 in a variety of liver cancer cell lines from CCLE. (h) mRNA expression levels of RRM2B in a variety of liver cancer cell lines from CCLE. ^∗∗p < 0.01 and ^∗∗∗p < 0.001. The “ns” stands for “not significant”.

Figure 2

Correlation analysis of clinicopathological characteristics and expressions of RR subunits in liver cancer tissues. (a) Expressions of RRM1, RRM2, and RRM2B in different age groups (number of age < = 60 years: 177 and number of age > 60 years: 196). (b) Expressions of RRM1, RRM2, and RRM2B in different ethnic groups (number of Asian:160, number of Black or African American: 17, and number of White: 185). (c) Expressions of RRM1, RRM2, and RRM2B in different gender groups (number of female: 121 and number of male: 253). (d) Expressions of RRM1, RRM2, and RRM2B in different T stage groups (number of T1: 183, number of T2: 95, number of T3: 80, and number of T4: 13). (e) Expressions of RRM1, RRM2, and RRM2B in different grade groups (number of G1: 55, number of G2: 178, and number of G3 and G4: 136). (f) Expressions of RRM1, RRM2, and RRM2B in different stage groups (number of stage I: 173, number of stage II: 87, and number of stages III and IV: 90). (g) Expressions of RRM1, RRM2, and RRM2B in different tumor status groups (number of tumor-free: 202 and number of with tumor: 153). (h) Expressions of RRM1, RRM2, and RRM2B in different alpha-fetoprotein (AFP) protein level groups (number of AFP < = 400 ng/ml: 215 and number of AFP > 400 ng/ml: 65). ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001. The “ns” stands for “not significant.” “Tumor-free” means that liver cancer does not continue to be present, indicating no progression of the original liver cancer. “With tumor” means the progression of the original disease.

Figure 3

Prognostic impact of RRM1, RRM2, and RRM2B in TCGA-LIHC. (a) Kaplan-Meier overall survival analysis between low (N = 187) and high (N = 186) expression of RRM1. (b) Kaplan-Meier overall survival analysis between low (N = 187) and high (N = 186) expression of RRM2. (c) Kaplan-Meier overall survival analysis between low (N = 187) and high (N = 186) expression of RRM2B. (d) Kaplan-Meier disease-specific survival analysis between low (N = 184) and high (N = 181) expression of RRM1. (e) Kaplan-Meier disease-specific survival analysis between low (N = 183) and high (N = 182) expression of RRM2. (f) Kaplan-Meier disease-specific survival analysis between low (N = 185) and high (N = 180) expression of RRM2B. (g) The receiver operating characteristic (ROC) curve for the diagnosis of liver cancer based on RRM1. (h) The ROC curve for the diagnosis of liver cancer based on RRM2. (i) The ROC curve for the diagnosis of liver cancer based on RRM2B.

Figure 4

Mutation patterns and coexpression analyses of RR subunits. (a) Summary genetic alterations of RRM1, RRM2, and RRM2B in TCGA-LIHC (N = 370). (b) Frequency of different somatic alterations of RRM1, RRM2, and RRM2B in TCGA-LIHC. (c) Coexpression matrix of RRM1, RRM2, and RRM2B. (d) Interaction gene networks of RRM1, RRM2, and RRM2B, such as glutaredoxin (GLRX), thioredoxin (TXN), chromosome segregation 1-like (CSE1L), transcription factor binding to IGHM enhancer 3 (TFE3), adenylate kinase 1 (AK1), E2F transcription factor 3 (E2F3), cytidine/uridine monophosphate kinase 1 (CMPK1), isocitrate dehydrogenase (NAD(+)) 3 catalytic subunit alpha (IDH3A), thioredoxin reductase 1 (TXNRD1), guanylate kinase 1 (GUK1), E2F transcription factor 6 (E2F6), peptidylprolyl isomerase B (PPIB), coenzyme Q7, hydroxylase (COQ7), diphthamide biosynthesis 1 (DPH1), (polo-like kinase 1) PLK1, AT-rich interaction domain 2 (ARID2), MCM4, minichromosome maintenance complex component 5 (MCM5), WD repeat domain 43 (WDR43), and cyclin-dependent kinase inhibitor 2A (CDKN2A).

Figure 5

Functional enrichment analysis of RR subunits in liver cancer. (a) Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment results of RRM1-related genes (N = 374). (b) Ridge plot of GSEA results for high RRM1 expression. (c) GO and KEGG enrichment results of RRM2-related genes. (d) Ridge plot of GSEA results for high RRM2 expression. “NES,” normalized enrichment score (a significant positive NES value indicates that members of the gene set tend to appear at the top of the ranked transcriptome data.).

Figure 6

Immune analysis of RR subunits in liver cancer. (a) The correlation between RRM1 expression and profiles of immune infiltrating cells. (b) The correlation between RRM2 expression and profiles of immune infiltrating cells. (c) The correlation between RRM2B expression and profiles of immune infiltrating cells. (d) The correlation between RRM1 expression and immune score. (e) The correlation between RRM2 expression and stromal score. (f) The correlation between RRM2B expression and immune and stromal score. Abbreviation: TH2: T helper 2; TFH: T follicular helper cells; aDC: activated dendritic cell; Tcm: central memory T cell; TH1: T helper 1; TH17: T helper 17; NK: natural killer cells; Tem: effector memory T cell; pDC: plasmacytoid dendritic cell; DC: dendritic cell.