Analysis

Pan-cancer analysis of m1A writer gene RRP8: implications for immune infiltration and prognosis in human cancers

Zhihui Huang 1,2,8 . Koo Han Yoo3 . Duohui Li4 . Qingxin Yu5,9 . Luxia Ye6 . Wuran Wei7

Received: 12 June 2024 / Accepted: 2 September 2024

Published online: 12 September 2024

@ The Author(s) 2024 OPEN

Abstract

Background Ribosomal RNA Processing 8 (RRP8) is a gene associated with RNA modification and has been implicated in the development of several types of tumors in recent research. Nevertheless, the biological importance of RRP8 in pan-cancer has not yet been thoroughly and comprehensively investigated.

Methods In this study, we conducted an analysis of various public databases to investigate the biological functions of RRP8. Our analysis included examining its correlation with pan-cancer prognosis, heterogeneity, stemness, immune checkpoint genes, and immune cell infiltration. Furthermore, we utilized the GDSC and CTRP databases to assess the sensitivity of RRP8 to small molecule drugs.

Results Our findings indicate that RRP8 exhibits differential expression between tumor and normal samples, particularly impacting the prognosis of various cancers such as Adrenocortical carcinoma (ACC) and Kidney Chromophobe (KICH). The expression of RRP8 is intricately linked to tumor heterogeneity and stemness markers. Additionally, RRP8 shows a positive correlation with the presence of tumor-infiltrating cells, with TP53 being the predominant mutated gene in these malignancies.

Conclusion Our findings suggest that RRP8 may serve as a potential prognostic marker and therapeutic target in a variety of cancer types.

Keywords Pan cancer . RNA 1-methylcytosine . Ribosomal RNA Processing 8 . Tumor-infiltrating cells

Abbreviations

m6AN6-methyladenosine
m5C5-Methylcytosine
m1AN1-methyladenosine
RRP8Ribosomal RNA Processing 8
TCGAThe Cancer Genome Atlas

Supplementary Information The online version contains supplementary material available at https://doi.org/10.1007/s12672-024- 01299-0.

☒ Duohui Li, 2860263085@qq.com; ☒ Qingxin Yu, qingxinyu0220@163.com; ☒ Luxia Ye, ylx941016@163.com; ☒ Wuran Wei, weiwuranwch@126.com; Zhihui Huang, 624539501@qq.com; Koo Han Yoo, yookoohan@khu.ac.kr | 1Operating Room, West China Hospital, Sichuan University, Chengdu, China. 2West China School of Nursing, Sichuan University, Chengdu, China. 3Department of Urology, Kyung Hee University, Seoul, South Korea. 4Department of Pharmacy Management, Anqing Municipal Hospital, Anqing 246000, Anhui, China. 5Department of Pathology, Ningbo Clinical Pathology Diagnosis Center, Ningbo 315211, Zhejiang, China. 6Department of Public Research Platform, Taizhou Hospital of Zhejiang Province Affiliated to Wenzhou Medical University, Linhai, China. 7Department of Urology, Institute of Urology, West China Hospital, Sichuan University, Chengdu 610041, China. 8West China Tianfu Hospital, Sichuan University, Chengdu, China. 9Department of pathology, Ningbo Medical Centre Lihuili Hospital, Ningbo, China.

Check for updates

Discover Oncology

(2024) 15:437

| https://doi.org/10.1007/s12672-024-01299-0

Discover

OSOverall survival
PFIProgression-free interval
DFSDisease-free survival
DSSDisease-specific survival
DNAssDNA methylation based
DMPssDifferentially methylated probes-based
EHNssEnhancer elements/DNA methylation-based
RNASSRNA expression-based
EREG-METHssEpigenetically regulated DNA methylation-based
EREG-METHssEpigenetically regulated RNA methylation-based
TMBTumor mutation burden
MATHMutant-allele tumor heterogeneity
LOHLoss of heterozygosity
NEONeoantigen
HRDHomologous recombination deficiency
MSIMicrosatellite instability
ACCAdrenocortical carcinoma
KICHKidney Chromophobe
LGGBrain Lower Grade Glioma
LIHCLiver hepatocellular carcinoma
KIPANPan-kidney cohort carcinoma
KIRPRenal papillary cell carcinoma
GBMLGGGlioma
LUSCLung squamous cell carcinoma
COADColon adenocarcinoma
PCPGPheochromocytoma and paraganglioma
THYMThymoma
NKNatural killer
STADStomach adenocarcinoma
GDSCGenomics of Cancer Drug Sensitivity
CTRPPan-Cancer Cancer Therapy Response Portal
PVRPoliovirus receptor
GSEAGene set enrichment analysis
KEGGKyoto encyclopedia of genes and genomes
MAPKMitogen-activated protein kinase
ECM-receptorExtracellular Matrix-receptor

1 Introduction

A central dogma of molecular biology is to elucidate the fundamental principles of the flow of genetic information within biological systems, contributing to our understanding of cellular processes [1, 2]. This principle assumes that genetic information passes from DNA to RNA to protein, involving transcription, translation, and replication processes. It has also been expanded to include RNA self-replication in certain viruses (e.g., borna disease virus), a process that challenges the conventional flow of information and offers insights into the origins of life and molecular evolution [3]. The historical development of RNA self-replication can be traced back to the discovery of ribozymes by Thomas Cech and Sidney Alt- man in the 1980s [4]. Their work led to the award of the Nobel Prize in Chemistry in 1989. Ribozymes are RNA molecules that can catalyze specific biochemical reactions, such as RNA splicing, showcasing the dual genetic and catalytic roles of RNA. The role of epigenetic modifications in cancer has been a major focus of research since the late twentieth century. Abnormal DNA methylation patterns are one of the earliest epigenetic changes associated with cancer, characterized by hypomethylation of oncogenes and hypermethylation of tumor suppressor genes [5]. Additionally, the field of epigenom- ics has revealed the intricate layers of regulation in eukaryotic cells. This encompasses various covalent modifications to histones and nucleic acids, changes in nucleosome arrangement, three-dimensional chromatin conformation, RNA

Discover

splicing machinery, and the functional roles of non-coding genomic elements [6, 7]. Epigenetic mechanisms dynami- cally regulate chromatin structure and fine-tune gene expression, significantly influencing various biological properties. These mechanisms are integral to epigenetic-based transgenic technologies and play a crucial role in the pathogenesis and intervention of diseases, particularly cancer [8, 9].

Recent technological advances have significantly propelled the field of epigenomics, enabling more in-depth and thorough analysis of epigenetic modifications. High-throughput sequencing technology has facilitated detailed mapping of DNA methylation and chromatin immunoprecipitation sequencing, allowing for rapid and cost-effective sequencing of entire genomes and epigenomes [10, 11]. Furthermore, single-cell RNA sequencing at an unprecedented resolution has provided valuable insights into cellular heterogeneity and chromatin accessibility [12, 13]. In addition, CRISPR-based epigenome editing technologies have made targeted modification of epigenetic marks possible, enabling functional studies of epigenetic regulation [14, 15]. These advancements have played a crucial role in identifying numerous chemical modifications in DNA and RNA, thereby enhancing our understanding of epigenetic regulation. Epigenetic modifica- tions are mediated by specialized enzymes known as ‘writers’, ‘erasers’, and ‘readers’, each playing a distinct role in the attachment, removal, and recognition of chemical groups [16]. Since the 1960s, over 100 RNA modifications have been discovered, which have diverse functions in determining cell fate [17]. As research progresses, it has become evident that RNA not only participates in protein synthesis but also has a direct impact on gene expression through microRNAs and long non-coding RNAs [18]. RNA modifications, which are chemical alterations made to RNA molecules post-tran- scriptionally, have significant impacts on RNA metabolism, including processes such as splicing, stability, translation, and decay [19, 20]. The most common internal modification in eukaryotic mRNA, N6-methyladenosine (m6A), plays a critical role in regulating embryonic development, stem cell differentiation, circadian rhythms, and stress responses [21]. Other important modifications include 5-methylcytosine (m5C), which stabilizes RNA and improves translation efficiency, and N1-methyladenosine (m1A), which promotes RNA stability and proper folding [22, 23]. In the context of tumors, aberrant RNA modifications are associated with tumorigenesis. For instance, inhibiting the expression of m6A demethylase ALKBH5 can suppress the proliferation and invasion of neuroblastoma, while alterations in m5C levels regulated by NSUN2 are linked to poor prognosis in colorectal cancer and bladder cancer [24]. Targeting these pathways holds therapeutic promise: Specific enzyme inhibitors have demonstrated efficacy in preclinical models, highlighting the importance of understanding RNA modifications in cancer biology and the potential for new therapeutic interventions. This study aimed to investigate the immuno-oncology role of ribosomal RNA processing 8 (RRP8) in human cancers through a pan-cancer analysis. Recent studies have indicated differential expression of RRP8 in tumors, suggesting its significance in tumorigenesis and its potential as a prognostic indicator for liver cancer [25]. RRP8, the yeast ortholog of mammalian nuclear methyl protein (NML), is linked to the 1m1A modification of 25S rRNA [26]. Additionally, it plays a role in energy-dependent silencing of ribosomal DNA, histone recruitment, and DNA repair processes [27, 28]. DNA damage triggers the generation of NML complexes, leading to the formation of rDNA isochromes and suppression of rRNA transcription. Immunohistochemistry findings indicate that breast tumors lacking detectable nucleosomal NML expression are associated with a lower survival rate [28]. Exploring the influence of RRP8 on immune regulatory genes and immune checkpoints can offer valuable insights into its potential as a therapeutic target and biomarker for immuno- therapy response. Additionally, conducting pan-cancer analyses can provide a comprehensive understanding of RRP8’s role across various cancer types, which is crucial for assessing its broader relevance and potential applications in preci- sion oncology.

2 Materials and methods

2.1 Data acquisition and processing

In accordance with our preceding research, we obtained the Cancer Genome Atlas (TCGA) pan-cancer dataset from the USCS database [29, 30]. We extracted the expression data of RRP8 in each sample by integrating the TCGA prognostic dataset from previous studies [31]. We screened samples with an expression level of 0, starting from normal solid tis- sue, primary tumor, and primary cancer-derived blood-peripheral blood. To enhance the robustness of our analysis, we subjected each expression value to a log2 (x+0.001) transformation. Cancer types represented by a sample size of fewer than 3 were systematically excluded. For identifying significant variances, we employed the unpaired Wilcoxon rank-sum test in conjunction with the sign test.

Discover

2.2 Pan-cancer survival analysis and relationship with clinical features

Metastatic samples from Primary Blood Derived Cancer-Peripheral Blood, primary tumor, and TCGA-SKCM databases. Expression data for 39 cancer types were obtained by excluding samples with an expression level of 0 or a follow-up period of less than 30 days. We stratified patients into either high- or low-expression cohort, predicated on the median expression value corresponding to each gene. The prognostic value of RRP8 was analyzed using the Cox proportional hazards regression model, considering overall survival (OS), disease-specific survival (DSS), disease-free survival (DFS) and progression-free interval (PFI) as prognostic analysis indicators [32, 33]. Furthermore, the correlation between gene expression and clinical stage, gender, and other clinical characteristics was evaluated using the unpaired Wilcoxon rank sum test, sign test, and Kruskal test. A dedicated exploration was also undertaken to discern the potential correlation between RRP8 expression and patient age.

2.3 Analysis of tumor heterogeneity, stemness and mutation landscape

Tumor stemness indicators were calculated by analyzing tumor methylation and mRNA expression signatures. These indi- cators include six categories: DNA methylation-based (DNAss), differentially methylated probe-based (DMPss), enhancer element/DNA methylation-based (ENHss), RNA expression-based (RNAss), appearance-based genetically regulated DNA methylation (EREG-METHss), and RNA methylation based on epigenetic regulation (EREG-METHss). Additionally, Spear- man analysis was performed to determine the correlation between tumor stemness characteristics and RRP8 expression. Tumor mutation burden (TMB), mutant allelic tumor heterogeneity (MATH), tumor ploidy, tumor purity, loss of heterozy- gosity (LOH), neoantigens (NEO), Microsatellite instability (MSI) and homologous recombination deficiency (HRD) serve as reflective indicators of tumor heterogeneity, using Spearman’s rank correlation coefficient [33, 34]. We analyzed gene expression and mutations in Adrenocortical carcinoma (ACC), Kidney Chromophobe (KICH), Brain Lower Grade Glioma (LGG), and Liver hepatocellular carcinoma (LIHC). The mutation frequency between samples in each group was evalu- ated using the Chi-square test [33].

2.4 Analysis of RNA modifications, checkpoints, tumor immune microenvironment (TME) and drug sensitivity

We undertook a comprehensive analysis to discern the potential correlations between the expression levels of RRP8 mRNA and an array of immune-related genes. These genes span several categories, including stimulatory checkpoints, heterogeneous checkpoints, and an extensive array of immunomodulatory genes (encompassing receptors, major his- tocompatibility complex molecules, chemokines, immunosuppressive, and immunostimulatory factors). Employing a structured data matrix, we investigated the relationship between RRP8 expression and 44 genes distributed across three RNA modification subcategories: m1A (containing 10 genes), m5C (containing 13 genes), and m6A (containing 21 genes). To gain insights into the tumor microenvironment, the Timer tool was utilized [35]. Concurrently, an exploration of drug sensitivities was conducted using datasets from the Genomics of Cancer Drug Sensitivity (GDSC) and the Pan-Cancer Cancer Therapy Response Portal (CTRP) via the GSCALite platform [36].

2.5 Gene enrichment analysis and nomogram

The LIHC cohort of RNAseq data type from the LinkedOmics database (http://www.linkedomics.org/login.php) was uti- lized as the research subject [37]. Gene Set Enrichment Analysis (GSEA) tool was employed to conduct Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis on RRP8-related genes. Additionally, nomogram analysis and visualization were based on survival data of LIHC from TCGA.

2.6 Statistical analysis

Based on the normality and homogeneity of variance within the data, either a one-way ANOVA or the Mann-Whitney U test was employed for the statistical analysis of continuous variables across three or more groups. For quantitative data comparisons between two groups, the Student’s t-test was utilized. All data presented are expressed as the standard

Discover

deviation. All analyses were conducted using Sanger platform [33]. A p-value below 0.05 was considered statistical sig- nificance. ns, P≥0.05; * , P< 0.05; ** , P< 0.01; *** , P < 0.001.

3 Results

3.1 Differential expression and clinical value

Our study revealed notable variations in the expression levels of RRP8 in different types of human cancers as compared to normal samples. Specifically, we found high expression of RRP8 in 15 tumor tissues, while 2 tumor tissues showed low expression (Fig. 1A). Furthermore, our analysis demonstrated a strong correlation between this gene and OS (Fig. 1B), DFS (Fig. 1C), DSS (Fig. 1D), and PFI (Fig. 1E) in numerous cancer types. Notably, these included ACC, KICH, pan-kidney cohort carcinoma (KIPAN), renal papillary cell carcinoma (KIRP), LIHC, glioma (GBMLGG), LGG, lung squamous cell carci- noma (LUSC), Colon adenocarcinoma (COAD), and pheochromocytoma and paraganglioma (PCPG). Moreover, our find- ings revealed a significant association between RRP8 and ACC as well as LIHC across all the aforementioned prognostic indicators (Fig. 2B-E). Among the 9 types of cancer mentioned above, RRP8 mRNA expression exhibited a significant correlation with age. Specifically, there were 3 positive correlations and 6 negative correlations (Fig. 2B). Additionally, this gene displayed varying degrees of correlation with clinical characteristics (Figure S1).

3.2 Relationship of RRP8 with tumor heterogeneity, stemness and gene mutation

The correlation between RRP8 expression levels and tumor heterogeneity and stemness was further investigated in our study. We discovered a significant correlation between RRP8 expression levels and HRD status in 15 tumors (Fig. 2A). Addi- tionally, a positive correlation between RRP8 expression and LOH was observed in 6 tumors (Fig. 2B). Regarding MATH, we found a negative correlation between RRP8 mRNA expression and 10 tumors (Fig. 2C). Our results demonstrated that RRP8 expression was significantly correlated with MSI in 11 tumors, including GBMLGG, LUSC, and KIRC (Fig. 2D). However, NEO was only associated with RRP8 expression in 6 tumors (Fig. 2E). RRP8 expression was found to be correlated with TMB in 13 tumors (Fig. 2F). In our analysis of tumor stemness, we observed a correlation between the expression level of RRP8 in Thymoma (THYM) and all six tumor stemness (Fig. 3A-F).

Tumor gene mutations play a crucial role in determining their biological behavior. In this study, we focused on analyz- ing the mutation patterns of RRP8, a gene known for its prominent role in tumors. We compared the mutation profiles of the RRP8 high-expression group with the low-expression group and identified the significantly mutated genes. Our findings revealed that TP53 was the most mutated gene. Additionally, we observed mutations in CTNBI, MUC, TTN and HMCN in ACC, TP53, PTEN, ZAN, TTN and CFAP47 in KICH, IDH1, TP53, ATRX.CIC and EGFR in LGG, TP53, ARID1A, JMUC17 and PCDH7 in LIHC, were significant mutated between the two groups (Fig. 4A-E).

3.3 Relationship between RRP8 expression with immune regulation, checkpoints, RNA modification and drug sensitivity

Our findings indicate that RRP8 exhibits a positive correlation with immune regulatory genes in most urinary system tumors, including ACC, KIRP, KICH, KIPAN, and BLCA (Fig. 5A). Notably, RRP8 expression levels are largely negatively cor- related with immune regulatory genes (Fig. 5A). Similarly, we observed a positive correlation between RRP8 and several pre-immune checkpoints in various urological tumors (BLCA, KICH, KIRP) as well as UVM, OV, and LGG (Fig. 5B). Consist- ent with the aforementioned results, RRP8 displays a negative correlation with the majority of tumor infiltrating cells in THYM and THCA, while OV, KIRC, and KIRP exhibit a positive correlation with most tumor infiltrating cells. Of particular interest, BLCA demonstrates a strong correlation with CD4 T+ cells, CD8+T cells, neutrophils, macrophages, and dendritic cells (Fig. 6A, B). Furthermore, we identified a relationship between RRP8 expression and drug sensitivity, as depicted in Fig. 6C and D. Notably, lapatinib, erlotinib, Saracatinib and gefitinib exhibited relatively promising efficacy among the experimental drugs.

Discover

A

**

*

*

*





*





*

*



7

6

Expression

5

T

T

T

Group

4

1

I

T

T

T

·

1

I

O

.

1

I

T

Tumor Normal

T

P

L

E

3

I

O 1

I

2

1

0

GBM(T=153,N=5)

GBMLGG(T=662,N=5)

LGG(T=509,N=5)

CESC(T=304,N=3)

LUAD(T=513,N=109)

COAD(T=288,N=41)

COADREAD(T=380,N=51)

BRCA(T=1092,N=113)

ESCA(T=181,N=13)

STES(T=595,N=49)

STAD(T=414,N=36)

HNSC(T=518,N=44)

LIHC(T=369,N=50)

READ(T=92,N=10)

PCPG(T=177,N=3)

KICH(T=66,N=129)

CHOL(T=36,N=9)

B

C

CancerCode

pvalue

Hazard Ratio(95%C1)

TOGA-LIHC(N-294)

3.0e-3

1.87(1 24,2.83)

TOGA-ACC(N=44)

3.8c-3

6.16(1.76,21.61)

TOGA-COAD(N=103)

0.01

TOGA-KIRP(N-177)

9.92(2.02,48.78)

0,06

2.98(0.95,9.31)

TOGA-LGG(N=126)

0.10

2.60(0.84,8.06)

TOGA-COADREAD(N-132)

0.11

2.99(0.82,10.94)

TOGA-GBMLOG(N-127)

0.12

2.46(0.80,7.58)

TOGA-KICH(N-29)

0,14

98.60(0.16,59648.13)

TCGA-LUSC(N-292)

0.26

1.43(0.77,2.68)

TOGA-KIPAN(N-319)

0.30

1.49(0.71,3.11)

TCGA-HNSC(N-128)

0.31

1.57(0.66,3.71)

TCGA-PRAD(N-337)

0.37

1.74(0.52,5.84)

TCGA-SARC(N-149)

1.20(0.69,2.08)

TOGA-PCPG(N=152)

0.52 0.53

2.02(0.22,18.50)

TCGA-TGCT(N=101)

0.59

1.32(0.47,3.69)

TCGA-CESC(N=171)

0.67

1.25(0.44,3.54)

TCGA-DLBC(N=26)

0.91

1.16(0.10,13.81)

TOGA-LUAD(N=294)

0.93

TOGA-BLCA(N=184)

1.02(0.64,1.63)

0.97

1.02(0.45,2.27)

TOGA-UCEC(N-115)

0,97

1-0-1

1.01(0.52,1.98)

TOGA-ESCA(N-84)

5.2c-3

TCGA-STES(N-316)

0.18(0.06,0.60)

0.03

0.50(0.27,0.93)

TOGA-OV(N-203)

0.05

0.74(0.54,1.00)

TOGA-READ(N-29)

0,07

TOGA-THCA(N=352)

0.06(2.0e-3,1.56)

0.19

0.47(0.15,1.46)

TCGA-STAD(N-232)

0.42

1

TCGA-BRCA(N-905)

0.73(0.34,1.57)

0.44

0.84(0.54,1.30)

TOGA-MESO(N=14)

0.52

0.48(0.05,4.62)

TCGA-PAAD(N=68)

0.57

TCGA-UCS(N=26)

0.77(0.30,1.94)

0.64

TCGA-CHOL(N=23)

0.71(0.17,3.04)

0.78

0.81(0.18,3.59)

TCGA-KIRC(N-113)

0.92

0.94(0 27.3.32)

6 log2(Hazard Ratio(95%CI))

10 12 14

D

E

log2(Hazard Ratio(95%CI))

-4

4

log2(Hazard Ratio(95%CI))

log2(Hazard Ratio(95%CI))

CancerCodepvalueHazard Ratio(95%(1)
TOGA-GBMLGG(N=619)2.2e-163.49(2.59,4.71)
TOGA-LIHC(N=341)2.0042.81(1.84,4.30)
TOGA-ACC(N=77)1.2e-33.45(1.63,7.28)
TOGA-KICH(N=64)0.017.24(1.59,32.91)
TOGA-PCPG(N=170)0.0210.67(1.32,86.04)
TOGA-LGG(N-474)0,031/64(1.04,2 58)
TOGA-LUSC(N=468)0.161.28(0.91,1.80)
TOGA-HNSC(N=509)0.181.24(0,90,1.70)
TOGA-GBM(N=144)0.211.32(0.86,2.02)
TOGA-KIRC(N-515)0.211.31(0.86,1.99)
TOGA-MESO(N=84)0.231.58(0.75,3.31)
TOGA-COAD(N=278)0.271.43(0.76,2.70)
TOGA-SKCM-M(N=347)0.271.18(0.88,1.58)
TOGA-KIPAN(N=855)0.281.21(0.85,1.72)
TOGA-THCA(N=501)0.391.85(0.46,7.37)
TOGA-SKCM(N-444)0.401.12(0.85,1.48)
TOGA-LAML(N=144)0.431.24(0.73,2.13)
TOGA-BLCA(N-398)0.481.12(0.82,1.54)
TOGA-UVM(N=74)0.521.46(0.46,4.70)
TOGA-ESCA(N-175)0.551.17(0.70,1.95)
TOGA-STES(N=547)0.561.10(0.80,1.52)
TOGA-BRCA(N-1044)0.591.30(0.78,1.56)
TOGA-COADREAD(N=368)0.591.17(0.67,2.02)
TOGA-LUAD(N-490)0.651.08(0.77,1.53)
TOGA-PAAD(N=172)0.741.09(0.66,1.79)
TOGA-PRAD(N=492)1.2%(0.14,12.11)
TOGA-STAD(N=372)0.931.02(0.68,1.53)
TOGA-THYM(N=117)0.150.19(0.02,1.81)
TOGA-UCEC(N=166)0.230.74(0.45,1.21)
TOGA-READ(N-90)0.260.47(0.12,1.76)
TOGA-DLBC(N=44)0.260.40(0.08,1.99)
TOGA-CESC(N=273)0.420.78(0.43,1.43)
TOGA-CHOL(N=33)0.430.64(0.21,1.94)
TOGA-SARCIN-254)0.580.88(0.57,1.37)
TOGA-UCS(N=55)0.59-- 40.82(0.40,1.70)
TOGA-TOCT(N=128)0.650.49(0.02,11.09)
TOGA-OV(N=406)0.710.96(0.77,1.20)
TOGA-KIRIN-276)0.840.91(0.37,2.24)
TOGA-SK.CM-P(N=97)1.001.00(0.45,2.20)
CancerCodepvalueHazard Ratio(95%(T)
TOGA-GBMLGG(N-598)3.40-173.97(2.88,5.48)
TOGA-ACC(N=75)2.6c-33.27(1 51,7.07)
TOGA-LIHC(N-333)3.60-32.26(1 30,3,91)
TOGA-LGG(N=466)7.2e-31.95(1 20,3.17)
TOGA-LUSCIN-418)8.4c-32.11(1.22,3.64)
TOGA-KIPAN(N-840)0.021.66(1.09,2.52)
TOGA-KIRC(N=504)0.031.77(1.08,2.90)
TOGA-KICH(N=64)0.037.54(1.32,42.98)
TOGA-PCPG(N=170)0.068.55(0.76,95.95)
TOGA-COAD(N=263)0.112.15(0.85,5.46)
TOGA-COADREAD(N-347)0.141.92(0.82,4.50)
TOGA-HNSC(N-485)0.201.31(0.87,1.98)
TOGA-THCA(N-495)0.213.85(0.48,30.71)
TOGA-MESO(N-64)0.281.70(0.65,4.43)
TOGA-GBM(1-131)0.301.29(0.80,2.08)
TOGA-UVM(N-74)0.361.83(0.51,641)
TOGA-SKCM-M(N=341)0.441.13(0.83,1.54)
TOGA.BRCA(N=1025)0.471.19(0.74,1.89)
TOGA-SKCM(N-438)0.47L.11(0.83,1.49)
TOGA-PAAD(N=166)0.581.18(0.66,2.10)
TOGA-BLCA(N=385)0.631-0-11.10(0.75,1.61)
TOGA-KIRP(N=272)0.651.28(0.44,3.75)
TOGA-SKCM-P(N=97)0.651.25(0.47,3.36)
TOGA-THYM(N=117)0.731.68(0.09,29.76)
TOGA-STES(N-524)0.831.01 1-9-11.05(0.70,1.57)
TOGA-ESCAIN-173)1.06(0.56,2.00)
TOGA-DLBC(N=44)0.380.37(0.04,3.48)
TOGA-CESC(N-269)0.450.77(0.39,1.52)
TOGA-UCEC(N=164)0.450.79(0.44,1.44)
TOGA-CHOL(N-12)0.460.65(0.21,2.04)
TOGA-TGCT(N=128)0.520.29(6.8e-3,12.43)
TOGA-SARC(N=248)0.580.87(0.54,1.41)
TOGA-PRAD(N=490)0.620.45(0.02.10.63)
TOGA-OV(N=377)0.680.95(0.75,1.21)
TOGA-LUAD(N=457)0.790.94(0.61,1.45)
TOGA-READ(N-84)0.930.89(0.08,9.91)
TOGA-UCS(N=53)0.950.98(0.45,2.10)
TOGA-STAD(N-151)0.99[0 59.1.67)
CancerCodepvalueHazard Ratio(95%CT)
TOGA-GBMLGG(N-616)4.4c-142.75(2.12,3.58)
TOGA-ACC(N=76)1.le-43.50(1.86,6.60)
TOGA-LIHC(N-340)2.10-42.00(1.39,2.89)
TOGA-PCPG(N=168)2.46-35.04(1.73,14.65)
TOGA-KIPAN(N-845)5.3e-31.62(1.16,2.27)
TOGA-LUSC(N=467)0.021.66(1.11,2 50)
TOGA-KICH(N-64)0.024.86(1.33,17.74)
TOGA-LGG(N=472)0.031.48(1.04,2.12)
TOGA-GBM(N=143)0.031.63(1.06,2.50)
TOGA-KIRC(N=508)0.041.57(1.03,2.39)
TOGA-KIRP(N=273)0.052.19(1.01,4.74)
TOGA-COAD(N=275)0.081.71(0.95,3.08)
TOGA-HNSC(N=508)0.121.30(0.93,1.80)
TOGA-UVM(N=73)0.291.73(0.62,4.85)
TOGA-PRAD(N-492)0.421.33(0.66,2.69)
TOGA-COADREAD(N=363)0.431.23(0.74,2.03)
TOGA-MESO(N=82)0.511.31(0.59,2.90)
TOGA-THYM(N-117)0.531.53(0.41,5.75)
TOGA-TGCT(N-126)0.601.29(0.50,3.32)
TOGA-PAAD(N=171)0.771.07(0.67,1.71)
TOGA-THCA(N-499)0.831.09(0.50,2.39)
TOGA-BLCA(N-397)0.901.02(0.74,1.41)
TOGA-READ(N-88)0.010.19(0.05,0.71)
TOGA-STES(N=548)0.060.72(0.52,1.01)
TOGA-STAD(N-375)0.110.70(0.45,1.08)
TOGA-CHOL(N-33)0.210.48(0.15,1.50)
TOGA-ESCA(N-173)0.250.74(0.44,1.24)
TOGA-OV(N-406)0.320.90(0.73,1.11)
TOGA-UCEC(N-166)0.360.82(0.54,1.25)
TOGA-LUAD(N-486)0.400.87(0.63,1.20)
TOGA-CESC(N-273)0.510.82(0.46,1.48)
TOGA-SARC(N=250)0.570.90(0.61.1.31)
TOGA-SKCM(N-434)0.580.94(0.74,1.18)
TOGA-SKCM-M(N-338)0.641-0-10.94(0.74,1.21)
TOGA-DLBC(N-43)0.730.78(0.19,3.23)
TOGA-UCS(N=55)0.750.89(0.42,1.87)
TOGA-SKCM-P(N-96)0.790.91(0.47,1.78)
TOGA-BRCA(N-1043)0.980.99(0.71,1.40)

Fig. 1 Differential expression and prognosis analysis of RRP8. A Pan-cancer analysis of RRP8 for differential expression between tumor and normal tissues; B pan-cancer analysis of RRP8 for OS; C pan-cancer analysis of RRP8 for DFS; D pan-cancer analysis of RRP8 for DSS; E pan- cancer analysis of RRP8 for PFI; OS: overall survival; DFS: disease-free survival; DSS: disease-specific survival; PFI: progression-free interval

3.4 Gene enrichment analysis and nomogram

GSEA analysis showed that enrichment of mitogen-activated protein kinase and extracellular matrix-receptor path- ways in LIHC in relation to RRP8 (Fig. 7A, C, D). Additionally, a nomogram for LIHC was constructed incorporating survival data and clinical characteristics (Fig. 7B).

Discover

Fig. 2 The pan-cancer Spearman analysis of tumor heterogeneity and RRP8 expression. A The correlation between HRD and RRP8 level; B the correlation between LOH and RRP8 level; C the correlation between MATH and RRP8 level; D the correlation between MSI and RRP8 level; E the correlation between NEO and RRP8 level; F the correlation between TMB and RRP8 level. HRD: homologous recombination defi- ciency; LOH: loss of heterozygosity; MATH: mutant-allele tumor heterogeneity; MSI: microsatellite instability; NEO: neoantigen; TMB: tumor mutation burden

A

B

UCEC(N=178)

pValue

pValue

BRCA(N=1044)

0.0

ACC(N=74)

READ(N=87)

0.0

ESCA(N=158)

UCEC(N=175)

LUAD(N=500)

-0.2

CHOL(N=36)

-0.2

CHOL(N=36)

-0.4

CESC(N=291

CESC(N=291)

BRCA(N=1035

-0.4

OV(N=406)

UVM(N=79)

-0.6

STAD(N=400)

LUAD(N=495

-0.6

STES(N=563)

STAD(N=405)

-0.8

STES(N=558)

ESCA(N=158)

-0.8

READ(N-90)

-1.0

THYM(N=102)

KICH(N-66)

LUSC(N=476)

1.0

SKCM(N=102)

TGCT(N=147)

LGG(N=503)

DLBC(N=46)

OV(N=400)

UCS(N=56)

HNSC(N=505)

HNSC(N=500)

THYM(N-103)

PAAD(N=158)

BLCA(N-397

PRAD(N=469)

SARC(N=241)

GBMLGG(N=646)

MESO(N=81)

COADREAD(N=362)

PAAD(N=158)

BLCA(N=397

PRAD(N=470)

LAML(N=111)

MESO(N=81)

PCPG(N-160)

SARC(N=241)

SKCM(N=102)

LUSC(N=490)

KIRC(N=478)

LAML(N=111)

KIRC(N=495)

GBM(N=143)

COAD(N=275

COADREAD(N=373)

PCPG(N=159)

TGCT(N=147)-

KIPAN(N=844)

UVM(N=79)

KIPAN(N=821)

COAD(N=283)

KIRP(N=278)

LIHC(N=356)

LGG(N=501)

UCS(N=56)

LIHC(N=353

GBM(N=143)

KICH(N=65

KIRP(N=283)

THCA(N=462

THCA(N=461

DLBC(N=46

ACC(N=75)

GBMLGG(N=644)

-0.4

-0.2

0.0

0.2

0,4

-0.2

0.0

0.2

Correlation coefficient(spearman)

Correlation coefficient(spearman)

C

D

GBM(N=149)

pValue

UCEC(N=175)

0.0

GBMLGG(N=657)

pValue

GBMLGG(N=649)

ACC(N=77

0.0

ACC(N=77)

-0.2

CESC(N=302)

SKCM(N=102)

PCPG(N=177

-0.2

SARC(N=234)

-0.4

THYM(N=118)

THYM(N=118)

READ(N=89)

-0.4

BLCA(N=407)

-0.6

GBM(N=151

LGG(N=506)

-0.6

LGG(N=500)

LAML(N=123)

-0.8

PRAD(N=495)

MESO(N=83

-0.8

BRCA(N=980)

HNSC(N=498)

-1.0

SARC(N=252)

1.0

DLBC(N-37

COADREAD(N=374)

PAAD(N=168)

OV(N=303

COAD(N=285

PRAD(N=492)

THCA(N=487

BRCA(N=1039)

STAD(N=409)

BLCA(N=407

PAAD(N=176)

PCPG(N=176)

KIRC(N=334)

LAML(N=129)

CESC(N=286

KIRP(N=285)

UCS(N-57

READ(N-90)

OV(N=303)

THCA(N=493)

CHOL(N-35

LUAD(N=511)

LIHC(N=367

STES(N=589)

LUSC(N=485)

CHOL(N=36)

LUAD(N=508)

ESCA(N=180)

COADREAD(N=372)

TGCT(N=148)

LIHC(N=356)

STES(N=592)

COAD(N=282)

STAD(N=412)

LUSC(N=490

KIRP(N=279)

KIPAN(N=679)

HNSC(N=500)

MESO(N=81

KIPAN(N=688)

KICH(N=66

UCS(N=57

ESCA(N=180)

UCEC(N=180)

UVM(N=79

KIRC(N=337

UVM(N=79

TGCT(N=143)

SKCM(N=102)

KICH(N=66

DLBC(N=47

-0.3

-0.2

-0.1

0.0

0.1

0.2

0.3

-0.3

-0.2

-0.1

0.0

0.1

0.2

0.3

Correlation coefficient(spearman)

Correlation coefficient(spearman)

E

F

DLBC(N=33)

pValue

0.0

THYM(N=118)

pValue

THCA(N=175)

BRCA(N=981)

0.0

CHOL(N=30)

-0.2

LAML(N=126)

KICH(N=39)

UVM(N=79)

-0.2

LUAD(N=462)

-0.4

CHOL(N=36)

LUAD(N=509)

-0.4

MESO(N=65)

LGG(N=403)

-0.6

PRAD(N=492)

KIRP(N=279)

-0.6

SKCM(N=83)

-0.8

KIPAN(N=679)

PRAD(N=341)

DLBC(N=37

-0.8

BRCA(N=857)

1.0

CESC(N=286)

GBMLGG(N=520)

BLCA(N=407)

-1.0

THCA(N=489

SARC(N=177)

LUSC(N=486)

LUSC(N=447)

COAD(N=282)

ACC(N=57)

OV(N=303

READ(N=81)

COADREAD(N-372)

CESC(N-244)

SARC(N=234)

HNSC(N=498)

HNSC(N=446)

READ(N=90

TGCT(N=94)

UCS(N=57)

UVM(N=38)

TGCT(N=143)

GBM(N=117

KIRC(N=334)

KIPAN(N=636)

KICH(N-66)

THYM(N=64)

PCPG(N=177)

SKCM(N=102)

KIRP(N=260)

MESO(N=82)

UCS(N=50)

LIHC(N=357)

COADREAD(N=336)

ESCA(N=180)

KIRC(N=337)

GBM(N=149)

LIHC(N=337)

STES(N=589)

STAD(N=409)

BLCA(N=375)

PAAD(N=171)

COAD(N=255)

LGG(N-501)

PAAD(N=113)

UCEC(N=175)

PCPG(N=60)

ACC(N=77)

UCEC(N=166)

GBMLGG(N=650)

-0.2

-0.2

0.0

0.2

0.0

0.2

0.4

Correlation coefficient(spearman)

Correlation coefficient(spearman)

Discover

Fig. 3 The pan-cancer Spearman analysis of tumor stemness and RRP8 expression. A The correlation between tumor stemness and RRP8 level using DMPss; B the correlation between tumor stemness and RRP8 level using DNAss; C the correlation between tumor stemness and RRP8 level using ENHss; D the correlation between tumor stemness and RRP8 level using EREG.EXPss; E the correlation between tumor stemness and RRP8 level using EREG-METHss; F the correlation between tumor stemness and RRP8 level using RNAss. DNAss: DNA meth- ylation based; DMPss: differentially methylated probes-based; EHNss: enhancer elements/DNA methylation-based; RNAss: RNA expression- based; EREG-METHss: epigenetically regulated DNA methylation-based; EREG-METHss: epigenetically regulated RNA methylation-based

A

B

THYM(N=119)

pValue

THCA(N=499)

O.C

THYM(N=119)

pValue

BLCA(N=403)

THCA(N=499

0.0

LUAD(N=451)

-0.2

BLCA(N=403)

OV(N-9

-0.2

CESC(N=301

OV(N=9)

-0.4

CESC(N=301)

MESO(N=87)

LUAD(N=451)

-0.4

LAML(N=170)

- 0.€

BRCA(N=774)

LAML(N=170)

-0.6

BRCA(N=774)

SARC(N-253)

-0.8

CHOL(N=36)

DLBC(N=47)

SARC(N=253)

-0.8

TGCT(N=147)

- 1.0

KICH(N-65

LIHC(N-366)

1.0

LIHC(N=366)

KICH(N=65)

LUSC(N=361

MESO(N=87

CHOL(N=36)

DLBC(N=47

LUSC(N=361)

PRAD(N=491

TGCT(N=147)

COAD(N=271

READ(N=87)

COAD(N=271)

COADREAD(N=358)

COADREAD(N=358)

UCEC(N=173)

PCPG(N=176

UCEC(N=173)

STAD(N=369)

KIRP(N=268)

PCPG(N=176)

GBM(N=51)

KIRP(N=268)

GBM(N=51)

PRAD(N=491

STAD(N=369)

STES(N=548)

STES(N=548)

HNSC(N=512

READ(N=87

PAAD(N=156)

PAAD(N=156)

ESCA(N=179)

HNSC(N=512)

KIRC(N-309

KIRC(N=309)

LGG(N=507

KIPAN(N=642)

UCS(N=57

KIPAN(N=642)

LGG(N=507

ESCA(N=179

ACC(N=76

SKCM(N=102)

ACC(N=76

SKCM(N=102)

GBMLGG(N=558)

UCS(N=57

UVM(N=79)

GBMLGG(N=558)

UVM(N=79)

-0.4

-0.2

0.0

0.2

0.4

-0.4

-0.2

0.0

0.2

0.4

Correlation coefficient(spearman)

Correlation coefficient(spearman)

C

D

THYM(N=119)

pValue

KICH(N=65

0

TGCT(N=147)

pValue

CHOL(N-36)

0.0

BLCA(N=403)

OV(N-9

-0

BRCA(N=1080)

SARC(N=253)

-0.2

SARC(N=253)

LUAD(N=451)

-0

LGG(N=507

CESC(N=301

OV(N=297)

-0.4

CHOL(N-36

-0

GBMLGG(N=659)

PAAD(N=156)

-0.6

BRCA(N=774)

LAML(N=170)

-0

BLCA(N=403)

LIHC(N-366

UCEC(N=177)

0.8

THCA(N=499

-1

KIRP(N=283)

-1.0

LUSC(N=361

HNSC(N=512)

MESO(N=87

UCS(N=57)

DLBC(N=47

THCA(N=499)

STAD(N-369

KIRC(N=512)

PAAD(N=156

ESCA(N=179)

COAD(N-271

LUSC(N=483)

KIRP(N=268

KIPAN(N=860)

KIRC(N=309

LUAD(N=507)

PCPG(N=176

CESC(N=301

STES(N=548

PRAD(N-491)

COADREAD(N=358)

UVM(N=79)

KIPAN(N=642)

MESO(N-87)

TGCT(N=147

STES(N=578)

PRAD(N=491

LAML(N=167)

HNSC(N=512)

GBM(N=152)

READ(N=87)

KICH(N=65)

UCEC(N=173

STAD(N=399)

ESCA(N=179)

LIHC(N-366)

ACC(N=76

SKCM(N=102)

LGG(N=507

COAD(N-281)

GBM(N=51)

PCPG(N=176)

UCS(N=57

THYM(N-119)

SKCM(N=102

COADREAD(N=369)

GBMLGG(N=558)

ACC(N-76)

UVM(N=79

DLBC(N=47)

READ(N=88)

-0.4

-0.2

0.0

0.2

0.4

-0.4

-0.2

0.0

0.2

0.4

Correlation coefficient(spearman)

Correlation coefficient(spearman)

E

F

THYM(N=119)

pValue

O.C

GBMLGG(N=659)

pValue

THCA(N=499)

0.0

BLCA(N=403)

BRCA(N=1080)

-0.2

KIRP(N=283

LAML(N=170)

PRADIN-491

-0.2

CHOL(N=36)

BRCA(N=774)

-0.4

LGG(N-507

TGCT(N=147

-0.4

CESC(N=301

OV(N=9

- 0.€

UVM(N=79

SKCM(N=102

-0.6

LUAD(N=451

SARCIN-253

-0.8

BLCA(N=403

UCS(N=57

-0.8

LUSC(N=361

KICH(N=65

- 1.0

KICH(N-65)

KIRC(N=512)

-1.0

LIHC(N=366

CESC(N=301

DLBC(N=47

LAML(N=167

COAD(N=271)

GBM(N-152)

MESO(N=87)

PCPG(N=176

GBM(N=51)

LUAD(N=507

TGCT(N=147

KIPAN(N=860)

UCEC(N=173)

THCA(N=499

COADREAD(N=358)

ACC(N=76

PCPG(N=176

OV(N-297

STAD(N=369

CHOL(N=36

KIRP(N-268

LUSC(N=483

READ(N=87

PAAD(N=156)

STES(N=548)

STADIN-399

PRAD(N=491

STES(N=578

PAAD(N=156)

HNSC(N=512

LGG(N=507

DLBC(N=47

HNSC(N=512)

SARC(N=253

KIRC(N=309

COAD(N=281

KIPAN(N=642 UCS(N=57)

ESCA(N=179)

LIHC(N-366

SKCM(N=102

COADREAD(N=369)

ESCA(N=179

MESO(N=87

ACC(N=76)

UCEC(N=177

GBMLGG(N=558)

READ(N=88

UVM(N=79)

THYM(N=119)

-0.4

-0.2

0.0

0.2

0.4

-0.6

-0.4

-0.2

0.0

0.2

0.4

0.6

Correlation coefficient(spearman)

Correlation coefficient(spearman)

Discover

Fig. 4 Mutation landscape of RRP8. A Mutation landscapes of RRP8 for pan-cancer; B the top 5 mutation genes between high and low- expression of RRP8 in ACC patients; C the top 5 mutation genes between high and low- expression of RRP8 in KICH patients; D the top 5 mutation genes between high and low-expression of RRP8 in LGG patients; E the top 5 mutation genes between high and low- expression of RRP8 in LIHC patients. ACC: Adrenocortical carcinoma; KICH: Kidney Chro- mophobe; LGG: Brain Lower Grade Glioma; LIHC: Liver hepatocellular carcinoma

A

GBM(N=149,0.7%)-

Missense_Mutation

GBMLGG(N=649,0.2%)

Splice_Site

CESC(N-286,0.3%)

Frame_Shift_Ins

LUAD(N-508,1.8%)

Frame Shift_Del

Nonsense_Mutation

COAD(N=282,0.7%)

Nonstop_Mutation

COADREAD(N-372,1.3%)

-2.0

LAML(N-123,0.8%)

STES(N=589,0.7%)

SARC(N-234,0.4%)

KIPAN(N-679,0.1%)

STAD(N-409,1.0%)

UCEC(N=175,5.7%)

1.5

HNSC(N-498,0.4%)-

KIRC(N-334,0.3%)

LUSC(N-485,1.4%)

LIHC(N-356,0.3%)

THCA(N-487,0.2%)

READ(N-90,3.3%)

-1.0

PAAD(N=168,1.2%)

OV(N-303,1.3%)

SKCM(N-102,2.0%)

BLCA(N=407,0.5%)-

B

457aa

AdoMet MTases

47

MutCount

Missense_Mutation Frame Shift Del Nonsense_Mutation

MutCount2.

0

SampleGroup

0

3

10

Splice_Site

In_Frame_Del Frame_Shift_Ins

TP53(0.20)

35.1%

SampleGroup: HighExp LowExp

CTNNB1(0.32)

32.4%

MUC16(0.21)

29.7%

TTN(0.14)

24.3%

HMCNI(0.97)

18.9%

C

MutCount

Missense_Mutation Nonsense_Mutation Splice_Site

MutCount2

-

J

SampleGroup

0

5

15 20

Frame_Shift_Del

SampleGroup: HighExp LowExp

TP53(0.21)

67.9%

PTEN(1.00)

17.9%

ZAN(0.94)

17.9%

TTΝ(0.65)

14,3%

CFAP47(1.00)

10.7%

D

Mut Count

Missense Mutation Framo_Shill_Del

MulCount

-

A

Nonsense_Mutatice

SampleGroup

0

100 200 300

Frame Shift_Irs

In_Frame_Del Splice_Site

IDH 1(5.93-4)

86.8%

SampleGroup:

Low Exp HighExp

TPS3(1.40-3)

51.5%

ATRX(3.2e-5)

37.2%

CIC(4.55-3)

22.7%

EGFR(0.01)

7.0%

E

-& MutCount

Frame_Shift_Del Missense_Mutation Nonsense Mutation

MutCount

/

SampleGroup

0

100

In_Frame_Del Frame_Shift_Ins Splice_Site In_Frame_Ins

TP53(2.4c-4)

71.8%

SampleGroup: HighExp LowExp

ARIDIA(9.2e-3)

17.6%

MUCI7(0.03)

10.6%

PCDH7(0.05)

9.9%

ANKRDI2(0.03)

8.5%

Discover

Fig. 5 The Spearman analysis of RRP8 expression and regulatory genes and immune checkpoints. A The correlation of RRP8 expression with immune regulatory genes; B the correlation of RRP8 expression with immune checkpoint genes

A

3

B

Type

correlation coefficient

IL13

correlation coefficient

-1.0-0.5 0.0 0.5 1

1.0

VTCNI

pValue

C10orf54

-1.0-0.5 0.0 0.5 1.0

05

LAG3

pValue

0.0

1.0

Type: chemokine

PDCD1

SLAMF7

0,0

0.5

1.0

receptor

MHC

CTLA4

Type:

Immunoinhibitor

TIGIT

Inhibitory Stimulaotry

Immunostimulator

CD274

HAVCR2

IL10

16

KIR2DL1

[13

KIR2DL3

BTLA

IDO1

ARGI

EDNRB

ADORA2A

IL4

VEGFB

VEGFA

IL12A

CD276

TGFB1

HMGBI

CD70

TNFRSF18

ICOSLG

TNFSF9

TNFRSF14

TNFRSF4

CD27

A2A

CD28

LG2

CD40LG

ICOS

ITGB2

PRFI

AB

GZMA

CCL5

BTN3A1

BTN3A2

CD80

ICAMI

13C

TNFSF4

CXCL10

$138

CXCL9

IFNG

E14

IL2RA

TNFRSF9

RSF18

ENTPDI

CD40

ILIA

CX3CLI

TLR4

ILIB

TNF

SF18

IFNAI

IL2

IFNA2

SELP

UVM(N=19) THYMIN HY)

DLBC(N=41) THCA(N-504)

CHOLIN-36

LAMLON-173

PAAD(N=178) PRADIN 495)

KIPAN(N-884)

KIRCIN- 530 GBMLGG(N=662)

BLCA(N=407

KICHEN

PCPO(N=197

KIRPIN ORS

ACC(N=77)

SKCM(N-102)

READ(N=92)

COAD(N=)28

COADREAL(N”380)

ESCAIN-181)

BRCA(N=1092)

LUADIN=513

LUSCO

STAD(NASA)

STESIN-SOS

SARCIN-258)

MESOINS

UCECIN=180)

CESC(N=304)

INFRSP25

HNSC(N=518)

OADRI

GBM

4 Discussion

RNA modifications play a crucial role in the epigenomic machinery, offering a unique perspective to comprehend cancer biology. Unlike static DNA changes, RNA modifications provide a dynamic and reversible mechanism for regulating gene expression [38, 39]. This adaptability is particularly important in cancer, where rapid and adaptive changes in gene expression are vital for survival and proliferation in diverse microenvironments [40]. One of the primary functions of RNA modifications in cancer is their influence on the destiny of mRNA molecules. Modifications like m6A methylation have been observed to affect mRNA stability, decay, and translation efficiency [41-43]. These post-transcriptional modifications can result in altered expression of key oncogenes and tumor suppressors, thereby driving the oncogenic process. For instance, modified mRNAs may evade standard degradation pathways, leading to sustained expression of growth-promoting genes. The overexpression of the methyltransferase METTL3 in BCLA results in the downregulation of PTEN in an m6A-dependent manner, leading to a poor response to treatment in patients [44]. Moreover, elevated METTL3 is found to be an independent factor for poor prognosis in patients with LIHC and gastric cancer [45, 46]. The impact of RNA modification extends beyond mRNA to include various types of non-coding RNA, such as microRNA and long non-coding RNA. These molecules play crucial roles in regulating gene expression and cell signaling pathways associated with cancer. RNA modifications could influence the production,

Discover

Fig. 6 The Spearman analysis of RRP8 expression and RNA modification; Tumor immune environment and its correlation with RRP8 expres- sion and drug sensitivity analysis. A The correlation of RRP8 expression with genes of RNA modification; B the correlation of RRP8 expression with immune infiltrating cells using TIMER; C the correlation between gene expression and the sensitivity of GDSC drugs (top 10) in pan- cancer; D The correlation between gene expression and the sensitivity of CTRP drugs (top 3) in pan-cancer

A

Modification

Type

B

TRMTGIA

TRMT6IB

correlation coefficient

TRMTIOC

TRMT6

-1.0-0.5 0.0 0.5 1.0

0.38

0.27

0.21

0.27

YTHDF3

pValue

0.35

TCGA-PCPG(N=177)

correlation coefficient

YTHDCI

-0.16

0.21

0.15

0.08

YTHDF1




·

TCGA-GBMLGG(N=656)

YTHDF2

0,0

0.5

M

1.0

-0.16

-0.24

0.14

-0.31

0.11

Modification: mlA




·

-0.33


TCGA-THCA(N-503)

0.2 0.0 0.2

ALKBHI

ALKBH3

0.15

0.18

0.14

0.22

0.15

0.22

pValue



TCGA-KIRC(N=528)

NSUNS

msc

.**


0.20

0.24

NOP2

mbA

0.20

0.17

0.30 TCGA-KIRP(N=285)

DNMTI

Type:




**


0.0

-0.20

0.5

1.5

2.0

NSUN2

writer

*

TCGA-GBM(N=152)

NSUN4

reader

DNMT3A

eraser

0.17

·

).17

TCGA-PAAD(N=177)

DNMT3B

NSUN7

-0.10

0.12

TCGA-LGG(N=504)

NSUN6

NSUN3

0.11

e

0.13

0.14

0.16

TCGA-LIHC(N=363)

TRDMTI

0.10

0.21

0.26

0.19

0.26

TCGA-OV(N=416)

ALYREF

0.30

0.31

ZC3H13

**


-0.35

0.32



TCGA-THYM(N=118)

METTL14

CBLLI

TCGA-STAD(N=388)

KIAA1429

METTL3

TCGA-SKCM-M(N=351)

RBMISB

RBM15

TCGA-UVM(N=79)

WTAP

0.30

0.23

ALKBHS

**

TCGA-SKCM-P(N=101)

FTO

0.29

IGF2BP1

TCGA-KICH(N=65)

YTHDF3

YTHDC2

TCGA-UCEC(N=178)

YTHDCI

FMRI

TCGA-HNSC(N=517)

LRPPRC

0.25

0.30

HNRNPA2BI

*

**

TCGA-ACC(N=77)

HNRNPC

YTHDF2

TCGA-READ(N=91)

YTHDF1

ELAVLI

TCGA-MESO(N=85)

THYM(N-119)

0.20

*

TCGA-TGCT(N=132)

0.13

0.13

..

0.10

+

TCGA-PRAD(N=495)

0.27

0.13

0.30

C


**

0.17

0.38

D



TCGA-BLCA(N=405)

-0.17

-0.22

**


TCGA-SARC(N=258)

Lapatinib

TCGA-LUSC(N=491)

Erlotinib

TCGA-CHOL(N-36)

austocystin D

0.22

-0.18

Saracatinib

TCGA-ESCA(N=181)

TCGA-SKCM(N=452)

17-AAG

P

TCGA-CESC(N=291)

Docetaxel

0

0.0025

0.0050

TCGA-UCS(N=56)

0.0075

erlotinib

Gefitinib

0.06

0.0100

TCGA-BRCA(N=1077)

GSK1904529A

TCGA-COADREAD(N=373)

AKT inhibitor Vill

-0.14

TCGA-LUAD(N=500)

Cetuximab

vandetanib

TCGA-COAD(N=282)

-0.35

JNK Inhibitor VIII

.

TCGA-DLBC(N=46)

B cell

T cell CD4

T cell CD8

Neutrophil

Macrophage

DC

0.00

0.05

0.10

15

0.20

0.00

0.05

0.10

0.15

Correlation

Correlation

stability, and function of these noncoding RNAs, thereby affecting important cellular processes like apoptosis, angio- genesis, and metastasis [47, 48]. RNA methylation has been shown to play a role in promoting tumorigenesis through the regulation of metabolic pathways. Wang et al. discovered that TRMT8 and TRMT61A can combine to form an m1A methyltransferase complex, leading to an increase in m1A methylation. This increase in methylation further enhances the expression of PPAR8, which in turn triggers cholesterol synthesis and ultimately activates Hedgehog signaling, thereby driving tumorigenesis [49]. Cancer cells exploit the flexibility provided by RNA modifications to adapt to environmental stresses like hypoxia or nutrient deprivation, and to develop resistance to therapeutic interventions. Changes in RNA modification patterns can confer drug resistance through metabolic enzymes. Overexpression of METTL3 in BRCA cell lines has been observed to result in an increased rate of fatty acid beta oxidation, which is a key enzyme leading to chemotherapy resistance [50, 51]. This resistance mechanism enables tumor cells to become resistant to multiple drugs.

The discovery of RNA m1A modifications dates to the second half of the twentieth century [51]. This is a reversible methylation process that involves adding a methyl group to the N1 position of adenosine in cellular transcripts [52]. The modification of m1A in RNA can also alter the secondary structure of the RNA and its interactions with proteins, conse- quently impacting RNA metabolism, structure, stability, and ultimately regulating gene expression and various cellular processes. Specific methyltransferases primarily mediate this process, and it has been observed in various RNA types, including coding and non-coding RNAs [53, 54]. The significance of m1A modifications is particularly notable in cancer research because it can regulate the expression of tumor suppressor genes in response to changes in cellular conditions, making it a key factor in cancer cell adaptability and resilience. In hepatocellular carcinoma, elevated m1A scores are associated with poorer prognosis and increased immune cell infiltration in tumor tissues, underscoring their significance within the tumor immune microenvironment [55]. The m1A demethylase ALKBH3 regulates glycolysis in cancer cells in a manner dependent on its demethylation activity, highlighting its role in the metabolic reprogramming of cancer cells [56]. In colorectal cancer, m1A modification patterns markedly influence tumor progression, invasion, and metastasis,

Discover

Fig. 7 GSEA of RRP8 in the TCGA LIHC cohort and nomogram. A KEGG enrichment analysis of RRP8 in LIHC; B the nomogram of TCGA LIHC cohort; C enrichment plot of RRP8 in MAPK signaling pathway; D enrichment plot of RRP8 in ECM-receptor interaction. GSEA: Gene set enrichment analysis; LIHC: Liver hepatocellular carcinoma; KEGG: Kyoto encyclopedia of genes and genomes. MAPK: Mitogen-activated pro- tein kinase: ECM-receptor: Extracellular Matrix-receptor

A

B

FOR $ 0.05

FOR > 0.05

20

1.5

1.0

0.5

0.0

0.5

1.0

1,5

20

2,5

3.0

Ribosome

Oxidative phosphorylation

Points

0

20

40

60

80

100

Parkinson disease

Proteasome

Huntington disease

Non-alcoholic fatty liver disease (NAFLD)

Pathologic T stage

T2

Thermogenesis

Alzheimer disease

T1

T3&T4

Spliceosome

Retrograde endocannabinoid signaling

Female

Pyrimidine metabolism

Gender

Cardiac muscle contraction

Drug metabolism

Male

RNA polymerase

Ribosome biogenesis in eukaryotes

Age

> 60

Purine metabolism

Glutathione metabolism

60

Cytosolic DNA-sensing pathway

400

Base excision repair

AFP(ng/ml)

Protein export

Glyoxylate and dicarboxylate metabolism

Sulfur relay system Metabolic pathways

> 400

DNA replication

RRP8

High

Aminoacyl-tRNA biosynthesis

Low

Renin secretion

Wnt signaling pathway

Ras signaling pathway

Total Points

MAPK signaling pathway

Prolactin signaling pathway

0

100

200

300

Vascular smooth muscle contraction

Hippo signaling pathway

Linear Predictor

Rap1 signaling pathway

Hedgehog signaling pathway Neurotrophin signaling pathway

-1

-0.6

-0.2

0.2

0.6

1

1.4

TNF signaling pathway

1-year Survival Probability

Osteoclast differentiation

Focal adhesion

0.95

0.9

0.85

0.8 0.75 0.

7

CGMP-PKG signaling pathway

Inositol phosphate metabolism

3-year Survival Probability

Platelet activation

JAK-STAT signaling pathway

0.8

0.7

0.6

0.5

0.4

0.3

ECM-receptor interaction

TGF-beta signaling pathway

5-year Survival Probability

Signaling pathways regulating pluripotency of stem cells

MicroRNAs in cancer

0.7

0.6

0.5

0.4

0.3

0.2

2.0

-15

-1.0

0.5

Normalized Enrichment Score

0.0

0.5

1.0

1.5

20

2.5

3.0

C

D

Enrichment plot: MAPK signaling pathway

Enrichment plot: ECM-receptor interaction

0

0

a

à

Enrichment Score

Enrichment Score

0

O

10

8

2

:

0

¥

Ranked list metric

20

Ranked list metric

R

8

9

0

5000

10000

15000

0

5000

Rank in Ordered Dataset

10000

Rank in Ordered Dataset

15000

with high m1A levels correlating with worse prognosis and greater tumor burdens [57]. Furthermore, TRMT6-mediated m1A modification in colorectal cancer enhances cancer stem cell self-renewal and activates the EGFR/ERK signaling path- way, contributing to tumorigenesis [58]. In gynecological cancers, the m1A regulator TRMT10C is a predictor of poorer survival and promotes malignant behaviors, while its silencing results in reduced cancer cell proliferation and migration [59]. Studies have found that m1A regulatory factors can promote the proliferation of cancer cells in gastric tumor [60] and LIHC [61] by regulating the PI3K/AKT pathway [62]. Additionally, ALKBH3, which acts as an eraser for m1A, can also contribute to cancer cell invasion by destabilizing tRNA [63]. Detecting and analyzing m1A modifications primarily rely on advanced sequencing technology and immunoprecipitation methods [64, 65]. However, techniques for accurately identifying and mapping m1A modifications still require further refinement and development. The emergence of new computational methods and improved sequencing technologies holds promise for deepening our understanding of m1A modifications and their role in various biological contexts, particularly in tumorigenesis.

The RRP8 gene is 8.5 base pairs long and is located on chromosome 11p15.4 [54]. It is a protein-coding gene found in the cytoplasm and nucleus. Its functions include RNA polymerase 1 promoter opening and gene expression [26]. Additionally, it can bind to methylated histones and act as a methyltransferase. In vivo experiments have revealed that deficiency of RRP8 affects the translation of proteins involved in carbohydrate metabolism, making it a gene

Discover

associated with metabolic diseases and obesity [66]. While research in the field of cancer is limited, some studies suggest that overexpression of RRP8 is a poor prognostic marker in LIHC [67]. Furthermore, research by Han et al. demonstrated the correlation between RRP8 expression and the effectiveness of neoadjuvant chemotherapy in triple- negative breast cancer [68]. Our study discovered that RRP8 exhibits differential expression in many tumors, including GMBLGG, LIHC, ACC, KICH, LGG, and LUSC, suggesting a correlation with solid tumors. Additionally, we observed a significant correlation between the expression of RRP8 and advanced age in multiple tumor types. Growing evidence supports the notion that alterations in the epigenetic landscape during aging contribute to tumorigenesis [69, 70]. The substantial association between RRP8 expression and advanced age in tumors underscores the importance of investigating the genetic overlap between aging and tumorigenesis, providing insights into the genomic mechanisms underlying tumor initiation and progression.

This study examined the correlation between RRP8 expression level and immune regulatory genes, immune check- points, and tumor infiltrating cells. The results consistently showed a strong correlation between RRP8 and tumor infiltrating cells in urological tumors (KIRC, KIRP, BLCA), as well as a positive correlation with poliovirus receptor (PVR). PVR, also known as CD155, is a transmembrane glycoprotein involved in cell adhesion, contact inhibition, and proliferation [71, 72]. It plays a crucial role in mediating natural killer cell adhesion and triggering natural killer (NK) cell effector functions [73]. PVR forms an immune synapse between NK cells and target cells by binding to CD96 and CD226, activating NK cell cytotoxicity [74]. However, when its expression increases, its isomers compete with membrane-bound PVR for the binding of DNAM-1, allowing tumors to evade detection and elimination by NK cells [75]. While PVR is constitutively expressed at low levels in various tissues, studies have shown that its overexpression is associated with poor prognosis in different malignant tumors, promoting tumor progression and metastasis [76, 77]. Based on these findings, we hypothesize that RRP8 may induce tumor cells to express PVR, thereby inhibiting the function of NK cells.

Tumor heterogeneity arises from variations in epigenetics and the tumor microenvironment, which play crucial roles in tumor growth, metastasis, and response to treatment [78, 79]. In our study, we investigated the association between RRP8 expression and tumor heterogeneity, specifically focusing on TMB (total number of mutations in the coding region of an exon) and MSI. As a marker for predicting immunotherapy response, TMB has shown a good correlation in melanoma [80]. Stomach adenocarcinoma (STAD) accounts for over 1 million new cases annually and ranks as the fifth most prevalent malignant tumor worldwide [81, 82]. Unfortunately, it is often diagnosed at an advanced stage, limiting the efficacy of combination chemotherapy in improving patient outcomes. As a result, immunotherapy is emerging as a promising first-line treatment for advanced gastric cancer patients [83, 84]. Our findings indicate a positive correlation between RRP8 expression and TMB as well as MSI in STAD, suggesting that patients with high RRP8 expression may benefit more from immunotherapy. Cellular stemness refers to the capacity of primitive cells to undergo self-renewal and differentiation. During tumor progression, epigenetic dysregulation of tumor cells can cause cancerous dedifferentiation and acquisition of stemness traits. Undifferentiated tumors are more prone to metastasis, resulting in disease progression and a poor prognosis. KIPAN cancer is a prevalent form of malignant tumors [85]. The survival rates for kidney cancer are notably high (90%) when the tumor remains local- ized in the kidney. However, these rates drastically decrease to 12% when metastasis occurs. The primary organs affected by metastasis are the lungs, bones, liver, and brain, all of which exhibit limited response to treatment [86, 87]. In this study, we observed a positive correlation between the expression of RRP8 in KIPAN and DMPss and EREG. These findings suggest that patients with higher RRP8 expression may have an increased susceptibility to tumor progression and metastasis.

TP53 is the most frequently altered tumor suppressor gene in solid tumors [88]. As a transcription gene, the P53 protein participates in specific physiological activities depending on the type of cellular stress signal received. These signals can include oncogene activation, DNA damage, and repair [89]. Consistent with this, our findings indicate that the TP53 gene is the most mutated gene in ACC, KICH, and LIHC. Furthermore, we observed that patients with higher RRP8 expression also had a higher frequency of TP53 mutations, highlighting the importance of this gene in tumor development. Our study reveals the potential functions and clinical significance of RRP8 in various solid tumors.

This study has several limitations. The data utilized are exclusively sourced from TCGA and various external databases, which may introduce selection bias. The samples in these databases are often collected under specific conditions and may not comprehensively represent the general oncology patient population. Furthermore, the quality and completeness of the data in public databases may impact the results. Additionally, the role of RRP8 in tumors requires further verification through in vivo and in vitro experiments. Nevertheless, our pan-cancer analysis of RRP8 establishes a solid foundation and offers novel insights for future research.

Discover

5 Conclusions

Our findings suggested that RRP8 could serves a biomarker in many cancers and should deserve more attention of researchers.

Acknowledgements We appreciated the Figdraw (www.figdraw.com) and Chengdu Basebiotech Co, Ltd for their assistance in drawing and data process.

Author contributions ZHH proposed the project, conducted data analysis, interpreted the data, and wrote the manuscript; KHY and QXY ducted data analysis, interpreted the data; DHL, LXY and WRW supervised the project, and interpreted the data. All authors reviewed and edited the manuscript.

Funding This research was funded by a regional innovation cooperation project of Sichuan Province (Grant No. 23QYCX0136).

Data availability The data sets presented in this study are available in online repositories. The name and join number of the repository can be found in the article/supplement.

Declarations

Ethics approval and consent to participate Not available.

Consent for publication Not available.

Competing interests The authors declare no competing interests.

Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License, which permits any non-commercial use, sharing, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if you modified the licensed material. You do not have permission under this licence to share adapted material derived from this article or parts of it. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativeco mmons.org/licenses/by-nc-nd/4.0/.

References

1. Crick F. Central dogma of molecular biology. Nature. 1970;227(5258):561-3.

2. Li GW, Xie XS. Central dogma at the single-molecule level in living cells. Nature. 2011;475(7356):308-15.

3. Jordan I, Lipkin WI. Borna disease virus. Rev Med Virol. 2001;11(1):37-57.

4. Kruger K, et al. Self-splicing RNA: autoexcision and autocyclization of the ribosomal RNA intervening sequence of tetrahymena. Cell. 1982;31(1):147-57.

5. Esteller M. Epigenetics in cancer. N Engl J Med. 2008;358(11):1148-59.

6. Parmar JJ, Padinhateeri R. Nucleosome positioning and chromatin organization. Curr Opin Struct Biol. 2020;64:111-8.

7. Casado-Pelaez M, Bueno-Costa A, Esteller M. Single cell cancer epigenetics. Trends Cancer. 2022;8(10):820-38.

8. Dawson MA, Kouzarides T. Cancer epigenetics: from mechanism to therapy. Cell. 2012;150(1):12-27.

9. Xiao K, et al. mRNA-based chimeric antigen receptor T cell therapy: basic principles, recent advances and future directions. Interdiscipl Med. 2024;2(1): e20230036.

10. Reuter JA, Spacek DV, Snyder MP. High-throughput sequencing technologies. Mol Cell. 2015;58(4):586-97.

11. Wu Z, et al. Genomic characterization of peritoneal lavage cytology-positive gastric cancer. Chin J Cancer Res. 2024;36(1):66-77.

12. Feng DC, Zhu WZ, Wang J, Li DX, Shi X, Xiong Q, You J, Han P, Qiu S, Wei Q, Yang L. The implications of single-cell RNA-seq analysis in prostate cancer: unraveling tumor heterogeneity, therapeutic implications and pathways towards personalized therapy. Mil Med Res. 2024;11(1):21. https://doi.org/10.1186/s40779-024-00526-7.

13. Du H, et al. Single-cell RNA-seq and bulk-seq identify RAB17 as a potential regulator of angiogenesis by human dermal microvascular endothelial cells in diabetic foot ulcers. Burns & Trauma. 2023;11: tkad020.

14. Wang S-W, et al. Current applications and future perspective of CRISPR/Cas9 gene editing in cancer. Mol Cancer. 2022;21(1):57.

15. Dong M, et al. CRISPR/CAS9: a promising approach for the research and treatment of cardiovascular diseases. Pharmacol Res. 2022;185: 106480.

16. Zhao LY, et al. Mapping the epigenetic modifications of DNA and RNA. Protein Cell. 2020;11(11):792-808.

17. Roundtree IA, et al. Dynamic RNA modifications in gene expression regulation. Cell. 2017;169(7):1187-200.

18. Ghidotti P, Petraroia I, Fortunato O, Pontis F. Immunomodulatory role of EV-derived non-coding RNA in lung cancer. Extracell Vesicles Circ Nucleic Acids. 2023;4(1):59-71. https://doi.org/10.20517/evcna.2022.42.

Discover

| https://doi.org/10.1007/s12672-024-01299-0

(2024) 15:437

Discover Oncology

19. Li C, et al. N6-Methyladenosine in vascular aging and related diseases: clinical perspectives. Aging Dis. 2023. https://doi.org/10.14336/ AD.2023.0924-1.

20. Thompson MG, Sacco MT, Horner SM. How RNA modifications regulate the antiviral response. Immunol Rev. 2021;304(1):169-80.

21. Zou D, et al. Single-cell and spatial transcriptomics reveals that PTPRG activates the m6A methyltransferase VIRMA to block mitophagy- mediated neuronal death in Alzheimer’s disease. Pharmacol Res. 2024;201: 107098.

22. Jin H, et al. m(1)A RNA modification in gene expression regulation. Genes. 2022;13(5):910.

23. Yuan L, Mao L-H, Li J-Y. CAG repeat expansions increase N1-methyladenine to Alter TDP-43 phase separation: lights up therapeutic inter- vention for neurodegeneration. Aging Dis. 2024. https://doi.org/10.14336/AD.2024.0110.

24. Guan Q, et al. Variant rs8400 enhances ALKBH5 expression through disrupting miR-186 binding and promotes neuroblastoma progres- sion. Chin J Cancer Res. 2023;35(2):140-62.

25. You K, et al. RRP8, associated with immune infiltration, is a prospective therapeutic target in hepatocellular carcinoma. J Cancer Res Clin Oncol. 2024;150(5):245.

26. Peifer C, et al. Yeast Rrp8p, a novel methyltransferase responsible for m1A 645 base modification of 25S rRNA. Nucleic Acids Res. 2013;41(2):1151-63.

27. Zhu C, et al. Erroneous ribosomal RNAs promote the generation of antisense ribosomal siRNA. Proc Natl Acad Sci USA. 2018;115(40):10082-7.

28. Yang L, et al. Nucleolar repression facilitates initiation and maintenance of senescence. Cell Cycle. 2015;14(22):3613-23.

29. Feng D, et al. A pan-cancer analysis of the oncogenic role of leucine zipper protein 2 in human cancer. Exp Hematol Oncol. 2022;11(1):55.

30. Goldman MJ, et al. Visualizing and interpreting cancer genomics data via the Xena platform. Nat Biotechnol. 2020;38(6):675-8.

31. Liu, J., et al., An Integrated TCGA Pan-Cancer Clinical Data Resource to Drive High-Quality Survival Outcome Analytics. Cell, 2018. 173(2): p. 400-416 e11.

32. Cortese G, Scheike TH, Martinussen T. Flexible survival regression modelling. Stat Methods Med Res. 2010;19(1):5-28.

33. Shen W, et al. Sangerbox: a comprehensive, interaction-friendly clinical bioinformatics analysis platform. iMeta. 2022. https://doi.org/10. 1002/imt2.36.

34. Ozga AJ, Chow MT, Luster AD. Chemokines and the immune response to cancer. Immunity. 2021;54(5):859-74.

35. Li T, et al. TIMER: a web server for comprehensive analysis of tumor-infiltrating immune cells. Cancer Res. 2017;77(21):e108-10.

36. Liu CJ, et al. GSCALite: a web server for gene set cancer analysis. Bioinformatics. 2018;34(21):3771-2.

37. Vasaikar SV, et al. LinkedOmics: analyzing multi-omics data within and across 32 cancer types. Nucleic Acids Res. 2018;46(D1):D956-63.

38. Bi Z, et al. A dynamic reversible RNA N(6)-methyladenosine modification: current status and perspectives. J Cell Physiol. 2019;234(6):7948-56.

39. Zhao BS, Roundtree IA, He C. Post-transcriptional gene regulation by mRNA modifications. Nat Rev Mol Cell Biol. 2017;18(1):31-42.

40. Hanahan D, Robert A. Weinberg, Hallmarks of cancer: the next generation. Cell. 2011;144(5):646-74.

41. Zeng Z, et al. The m6A reader YTHDF2 alleviates the inflammatory response by inhibiting IL-6R/JAK2/STAT1 pathway-mediated high- mobility group box-1 release. Burns & Trauma. 2023;11: tkad023.

42. Chen L, et al. m6A methylation-induced NR1D1 ablation disrupts the HSC circadian clock and promotes hepatic fibrosis. Pharmacol Res. 2023;189: 106704.

43. Zhao J, et al. Emerging regulatory mechanisms of N6-methyladenosine modification in cancer metastasis. Phenomics. 2023;3(1):83-100.

44. Han J, et al. METTL3 promote tumor proliferation of bladder cancer by accelerating pri-miR221/222 maturation in m6A-dependent man- ner. Mol Cancer. 2019;18(1):110.

45. Zhou Y, et al. Expression profiles and prognostic significance of RNA N6-methyladenosine-related genes in patients with hepatocellular carcinoma: evidence from independent datasets. Cancer Manag Res. 2019;11:3921-31.

46. Yue B, et al. METTL3-mediated N6-methyladenosine modification is critical for epithelial-mesenchymal transition and metastasis of gastric cancer. Mol Cancer. 2019;18(1):142.

47. Ma S, et al. The interplay between m6A RNA methylation and noncoding RNA in cancer. J Hematol Oncol. 2019;12(1):121.

48. Tu B, et al. METTL3 boosts mitochondrial fission and induces cardiac fibrosis by enhancing LncRNA GAS5 methylation. Pharmacol Res. 2023;194: 106840.

49. Wang Y, et al. N(1)-methyladenosine methylation in tRNA drives liver tumourigenesis by regulating cholesterol metabolism. Nat Commun. 2021;12(1):6314.

50. Singh B, et al. Important role of FTO in the survival of rare panresistant triple-negative inflammatory breast cancer cells facing a severe metabolic challenge. PLoS ONE. 2016;11(7): e0159072.

51. Chen Z, et al. N6-methyladenosine-induced ERRgamma triggers chemoresistance of cancer cells through upregulation of ABCB1 and metabolic reprogramming. Theranostics. 2020;10(8):3382-96.

52. Dominissini D, et al. The dynamic N1-methyladenosine methylome in eukaryotic messenger RNA. Nature. 2016;530(7591):441-6.

53. El Yacoubi B, Bailly M, de Crécy-Lagard V. Biosynthesis and function of posttranscriptional modifications of transfer RNAs. Annu Rev Genet. 2012;46(1):69-95.

54. Sharma S, et al. Identification of a novel methyltransferase, Bmt2, responsible for the N-1-methyl-adenosine base modification of 25S rRNA in Saccharomyces cerevisiae. Nucleic Acids Res. 2013;41(10):5428-43.

55. Zhao M, Shen S, Xue C. A novel m1A-score model correlated with the immune microenvironment predicts prognosis in hepatocellular carcinoma. Front Immunol. 2022;13: 805967.

56. Wu Y, et al. RNA m1A methylation regulates glycolysis of cancer cells through modulating ATP5D. Proc Natl Acad Sci USA. 2022;119(28): e2119038119.

57. Jiang C, et al. Landscape of N1-methyladenosin (m1A) modification pattern in colorectal cancer. Cancer Rep. 2024;7(2): e1965.

58. Sui S, et al. Abstract 1713: TRMT6-mediated N1-methyladenosine methylation promotes tumorigenesis in colorectal cancer. Cancer Res. 2023;83(7_Supplement):1713-1713.

59. Wang Q, et al. m1A regulator TRMT10C predicts poorer survival and contributes to malignant behavior in gynecological cancers. DNA Cell Biol. 2020;39(10):1767-78.

Discover

60. Li J, et al. Differential analysis of RNA methylation regulators in gastric cancer based on TCGA data set and construction of a prognostic model. J Gastrointest Oncol. 2021;12(4):1384-97.

61. Shi Q, et al. Gene signatures and prognostic values of m1A-related regulatory genes in hepatocellular carcinoma. Sci Rep. 2020;10(1):15083.

62. Macari F, et al. TRM6/61 connects PKCalpha with translational control through tRNAi(Met) stabilization: impact on tumorigenesis. Onco- gene. 2016;35(14):1785-96.

63. Woo HH, Chambers SK. Human ALKBH3-induced m(1)A demethylation increases the CSF-1 mRNA stability in breast and ovarian cancer cells. Biochim Biophys Acta Gene Regul Mech. 2019;1862(1):35-46.

64. Thuring K, et al. Analysis of RNA modifications by liquid chromatography-tandem mass spectrometry. Methods. 2016;107:48-56.

65. Araujo Tavares RC, et al. MRT-ModSeq - rapid detection of RNA modifications with MarathonRT. J Mol Biol. 2023;435(22): 168299.

66. Sharma S, et al. A single N(1)-methyladenosine on the large ribosomal subunit rRNA impacts locally its structure and the translation of key metabolic enzymes. Sci Rep. 2018;8(1):11904.

67. Li D, et al. The m6A/m5C/m1A regulated gene signature predicts the prognosis and correlates with the immune status of hepatocellular carcinoma. Front Immunol. 2022;13: 918140.

68. Han Y, Wang J, Xu B. Novel biomarkers and prediction model for the pathological complete response to neoadjuvant treatment of triple- negative breast cancer. J Cancer. 2021;12(3):936-45.

69. Martin-Herranz DE, et al. Screening for genes that accelerate the epigenetic aging clock in humans reveals a role for the H3K36 methyl- transferase NSD1. Genome Biol. 2019;20(1):146.

70. Feng D, et al. Unraveling links between aging, circadian rhythm and cancer: Insights from evidence-based analysis. Chin J Cancer Res. 2024;36(3):341-50.

71. Mendelsohn CL, Wimmer E, Racaniello VR. Cellular receptor for poliovirus: molecular cloning, nucleotide sequence, and expression of a new member of the immunoglobulin superfamily. Cell. 1989;56(5):855-65.

72. O’Donnell JS, et al. Tumor intrinsic and extrinsic immune functions of CD155. Semin Cancer Biol. 2020;65:189-96.

73. de Andrade LF, Smyth MJ, Martinet L. DNAM-1 control of natural killer cells functions through nectin and nectin-like proteins. Immunol Cell Biol. 2014;92(3):237-44.

74. Martinet L, Smyth MJ. Balancing natural killer cell activation through paired receptors. Nat Rev Immunol. 2015;15(4):243-54.

75. Briukhovetska D, et al. T cell-derived interleukin-22 drives the expression of CD155 by cancer cells to suppress NK cell function and pro- mote metastasis. Immunity. 2023;56(1):143-61.

76. Chen J. Expression of CD155 protein in pancreatic cancer and its clinical significance. J Am Coll Surg. 2020;231(4):S158-9.

77. Li YC, et al. Overexpression of an immune checkpoint (CD155) in breast cancer associated with prognostic significance and exhausted tumor-infiltrating lymphocytes: a cohort study. J Immunol Res. 2020;2020:3948928.

78. Jardim DL, et al. The challenges of tumor mutational burden as an immunotherapy biomarker. Cancer Cell. 2021;39(2):154-73.

79. Zhang X, et al. Radiomics under 2D regions, 3D regions, and peritumoral regions reveal tumor heterogeneity in non-small cell lung cancer: a multicenter study. Radiol Med. 2023;128(9):1079-92.

80. Chan TA, Wolchok JD, Snyder A. Genetic basis for clinical response to CTLA-4 blockade in melanoma. N Engl J Med. 2015;373(20):1984.

81. Smyth EC, et al. Gastric cancer. Lancet. 2020;396(10251):635-48.

82. Yan X, et al. Stomach cancer burden in China: epidemiology and prevention. Chin J Cancer Res. 2023;35(2):81-91.

83. Fuchs CS, et al. Ramucirumab with cisplatin and fluoropyrimidine as first-line therapy in patients with metastatic gastric or junctional adenocarcinoma (RAINFALL): a double-blind, randomised, placebo-controlled, phase 3 trial. Lancet Oncol. 2019;20(3):420-35.

84. Ohtsu A, et al. Bevacizumab in combination with chemotherapy as first-line therapy in advanced gastric cancer: a randomized, double- blind, placebo-controlled phase III study. J Clin Oncol. 2011;29(30):3968-76.

85. Qi J, et al. National and subnational trends in cancer burden in China, 2005-20: an analysis of national mortality surveillance data. Lancet Public Health. 2023;8(12):e943-55.

86. Ge W, et al. Review and prospect of immune checkpoint blockade therapy represented by PD-1/PD-L1 in the treatment of clear cell renal cell carcinoma. Oncol Res. 2023;31(3):255-70.

87. Capitanio U, Montorsi F. Renal cancer. Lancet. 2016;387(10021):894-906.

88. Kandoth C, et al. Mutational landscape and significance across 12 major cancer types. Nature. 2013;502(7471):333-9.

89. Kastenhuber ER, Lowe SW. Putting p53 in context. Cell. 2017;170(6):1062-78.

Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Discover