Comprehensive analysis of m6A regulator-based methylation modification patterns characterized by distinct immune profiles in colon adenocarcinomas

Recent studies have highlighted the biological significance of RNA N6-methyladenosine (m6A) modification in tumorigenicity and progression. However, it remains unclear whether m6A modifications also have potential roles in immune regulation and tumor microenvironment (TME) formation.Methods: In this study, we curated 23 m6A regulators and performed consensus molecular subtyping with NMF algorithm to determine m6A modification patterns and the m6A-related gene signature in colon cancer (CC). The ssGSEA and CIBERSORT algorithms were employed to quantify the relative infiltration levels of various immune cell subsets. An PCA algorithm based m6Sig scoring scheme was used to evaluate the m6A modification patterns of individual tumors with an immune response.Results: Three distinct m6A modification patterns were identified among 1307 CC samples, which were also associated with different clinical outcomes and biological pathways. The TME characterization revealed that the identified m6A patterns were highly consistent with three known immune profiles: immune-inflamed, immune-excluded, and immune-desert, respectively. Based on the m6Sig score, which was extracted from the m6A-related signature genes, CC patients can be divided into high and low score subgroups. Patients with lower m6Sig score was characterized by prolonged survival time and enhanced immune infiltration. Further analysis indicated that lower m6Sig score also correlated with greater tumor mutation loads, PD-L1 expression, and higher mutation rates in SMGs (e.g., PIK3CA and SMAD4). In addition, patients with lower m6Sig scores showed a better immune responses and durable clinical benefits in three independent immunotherapy cohorts.Conclusions: This study highlights that m6A modification is significantly associated with TME diversity and complexity. Quantitatively evaluating the m6A modification patterns of individual tumors will strengthen our understanding of TME characteristics and promote more effective immunotherapy strategies.

Background: Recent studies demonstrate that N6-methyladenosine (m6A) methylation plays a crucial role in colorectal cancer (CRC). Therefore, we conducted a comprehensive analysis to assess the m6A modification patterns and identify m6A-modified genes in patients with CRC recurrence.Methods: The m6A modification patterns were comprehensively evaluated by the NMF algorithm based on the levels of 27 m6A regulators, and tumor microenvironment (TME) cell-infiltrating characteristics of these modification patterns were systematically assessed by ssGSEA and CIBERSORT algorithms. The principal component analysis algorithm based on the m6A scoring scheme was used to explore the m6A modification patterns of individual tumors with immune responses. The weighted correlation network analysis and univariable and multivariable Cox regression analyses were applied to identify m6A-modified gene signatures. The single-cell expression dataset of CRC samples was used to explore the tumor microenvironment affected by these signatures.Results: Three distinct m6A modification patterns with significant recurrence-free survival (RFS) were identified in 804 CRC patients. The TME characterization revealed that the m6A modification pattern with longer RFS exhibited robust immune responses. CRC patients were divided into high- and low-score subgroups according to the m6A score individually, which was obtained from the m6A-related signature genes. The patients with low m6A scores had both longer RFS and overall survival (OS) with altered immune cell infiltration. Notably, m6A-modified genes showed significant differences related to the prognosis of CRC patients in the meta-GEO cohort and TCGA cohort. Single-cell expression indicated that ALVRL1 was centrally distributed in endothelial tip cells and stromal cells.Conclusion: The m6A modification plays an indispensable role in the formation of TME diversity and complexity. Importantly, the signatures (TOP2A, LRRC58, HAUS6, SMC4, ACVRL1, and KPNB1) were identified as m6A-modified genes associated with CRC recurrence, thereby serving as a promising predictive biomarker or therapeutic target for patients with CRC recurrence.

<p>RNA N6-methyladenosine (m<sup>6</sup>A) modification in tumorigenesis and progression has been highlighted and discovered in recent years. However, the molecular and clinical implications of m<sup>6</sup>A modification in melanoma tumor microenvironment (TME) and immune infiltration remain largely unknown. Here, we utilized consensus molecular clustering with nonnegative matrix factorization based on the melanoma transcriptomic profiles of 23 m<sup>6</sup>A regulators to determine the m<sup>6</sup>A modification clusters and m<sup>6</sup>A-related gene signature. Three distinct m<sup>6</sup>A modification patterns (m<sup>6</sup>A-C1, C2, and C3), which are characterized by specific m<sup>6</sup>A regulator expression, survival outcomes, and biological pathways, were identified in more than 1,000 melanoma samples. The immune profile analyses showed that these three m<sup>6</sup>A modification subtypes were highly consistent with the three known immune phenotypes: immune-desert (C1), immune-excluded (C2), and immune-inflamed (C3). Tumor digital cytometry (CIBERSORT, ssGSEA) algorithm revealed an upregulated infiltration of CD8<sup>+</sup> T cell and NK cell in m<sup>6</sup>A-C3 subtype. An m<sup>6</sup>A scoring scheme calculated by principal component of m<sup>6</sup>A signatures stratified melanoma patients into high- and low-m<sup>6</sup>sig score subgroups; a high score was significantly associated with prolonged survival and enhanced immune infiltration. Furthermore, fewer somatic copy number alternations (SCNA) and PD-L1 expression were found in patients with high m<sup>6</sup>Sig score. In addition, patients with high m<sup>6</sup>Sig score demonstrated marked immune responses and durable clinical benefits in two independent immunotherapy cohorts. Overall, this study indicated that m<sup>6</sup>A modification is involved in melanoma tumor microenvironment immune regulation and contributes to formation of tumor immunogenicity. Comprehensive evaluation of the m<sup>6</sup>A modification pattern of individual tumors will provide more insights into molecular mechanisms of TME characterization and promote more effective personalized biotherapy strategies.</p>

Multiple roles of p53 in cancer development: Regulation of tumor microenvironment, m6A modification and diverse cell death mechanisms

Action of m6A-related gene signatures on the prognosis and immune microenvironment of colonic adenocarcinoma

BackgroundSeveral recent studies have confirmed epigenetic regulation of the immune response. However, the potential role of RNA N6-methyladenosine (m6A) modifications in cervical cancer and tumour microenvironment (TME) cell infiltration remain unclear.ResultsWe evaluated and analysed m6A modification patterns in 307 cervical cancer samples from The Cancer Genome Atlas (TCGA) dataset based on 13 m6A regulators. Pearson correlation analysis was used to identify lncRNAs associated with m6A, followed by univariate Cox regression analysis to screen their prognostic role in cervical cancer patients. We also correlated TME cell infiltration characteristics with modification patterns. We screened six m6A-associated lncRNAs as prognostic lncRNAs and established the prognostic profile of m6A-associated lncRNAs by least absolute shrinkage and choice of operator (LASSO) Cox regression. The corresponding risk scores of the patients were derived based on their prognostic features, and the correlation between this feature model and disease prognosis was analysed. The prognostic model constructed based on the TCGA-CESC (The Cancer Genome Cervical squamous cell carcinoma and endocervical adenocarcinoma) dataset showed strong prognostic power in the stratified analysis and was confirmed as an independent prognostic indicator for predicting the overall survival of patients with CESC. Enrichment analysis showed that biological processes, pathways, and markers associated with malignancy were more common in the high-risk subgroup. Risk scores were strongly correlated with the tumour grade. ECM receptor interactions and pathways in cancer were enriched in Cluster 2, while oxidative phosphorylation and other biological processes were enriched in Cluster 1. The expression of immune checkpoint molecules, including programmed death 1 (PD-1) and programmed death ligand 1 (PD-L1), was significantly increased in the high-risk subgroup, suggesting that this prognostic model could be a predictor of immunotherapy.ConclusionsThis study reveals that m6A modifications play an integral role in the diversity and complexity of TME formation. Assessing the m6A modification patterns of individual tumours will help improve our understanding of TME infiltration characteristics and thus guide immunotherapy more effectively. We also developed an independent prognostic model based on m6A-associated lncRNAs as a predictor of overall survival, which can also be used as a predictor of immunotherapy.

Objective: N6-methyladenosine (m6A) modification is involved in modulating various biological processes in human cancers. But the implication of m6A modification in lung adenocarcinoma (LUAD) is still unclear. Hence, this study conducted a comprehensive analysis of the expression and clinical implication of m6A regulators in LUAD.Methods: Consensus clustering analysis of 502 LUAD samples in the TCGA dataset was presented based on the expression profiles of 20 m6A regulators using ConsensusClusterPlus package. Overall survival (OS), activation of signaling pathways and tumor immunity (immune/stromal score, tumor purity, expression of HLA and immune checkpoints, and immune cell infiltration) were compared between m6A modification patterns. The m6A-related genes between patterns were identified and prognostic m6A-related genes were imported into LASSO-cox regression analysis. The m6A risk score was developed and its prognostic implication was evaluated and externally verified in the GSE30219 and GSE72094 dataset. Furthermore, a nomogram that contained independent prognostic indicators was established, followed by external verification.Results: Two m6A modification patterns were clustered across LUAD based on the expression similarity of the m6A regulators via consensus clustering analysis, with distinct OS, activation of signaling pathways and tumor immunity. Totally, 213 m6A-related genes that were identified by comparing two patterns were significantly related to LUAD prognosis. By LASSO method, we constructed the m6A risk score that was a reliable and independent prognostic factor for LUAD. Patients with low m6A risk score displayed a prominent survival advantage. After incorporating independent clinical features, we developed the prognostic nomogram that exhibited high predictive accuracy and the best clinical net benefit for OS.Conclusion: Collectively, our study may provide a clinically useful tool for precise prognostic management and optimization of immunotherapeutic strategies for LUAD patients.

Objective: N6-methyladenosine (m6A) modification is involved in modulating various biological processes in human cancers. But the implication of m6A modification in lung adenocarcinoma (LUAD) is still unclear. Hence, this study conducted a comprehensive analysis of the expression and clinical implication of m6A regulators in LUAD. Methods: Consensus clustering analysis of 502 LUAD samples in the TCGA dataset was presented based on the expression profiles of 20 m6A regulators using ConsensusClusterPlus package. Overall survival (OS), activation of signaling pathways and tumor immunity (immune / stromal score, tumor purity, expression of HLA and immune checkpoints, and immune cell infiltration) were compared between m6A modification patterns. The m6A-related genes between patterns were identified and prognostic m6A-related genes were imported into LASSO-cox regression analysis. The m6A risk score was developed and its prognostic implication was evaluated and externally verified in the GSE30219 and GSE72094 dataset. Furthermore, a nomogram that contained independent prognostic indicators was established, followed by external verification. Results: Two m6A modification patterns were clustered across LUAD based on the expression similarity of the m6A regulators via consensus clustering analysis, with distinct OS, activation of signaling pathways and tumor immunity. Totally, 213 m6A-related genes that were identified by comparing two patterns were significantly related to LUAD prognosis. By LASSO method, we constructed the m6A risk score that was a reliable and independent prognostic factor for LUAD. Patients with low m6A risk score displayed a prominent survival advantage. After incorporating independent clinical features, we developed the prognostic nomogram that exhibited high predictive accuracy and the best clinical net benefit for OS. Conclusion: Collectively, our study may provide a clinically useful tool for precise prognostic management and optimization of immunotherapeutic strategies for LUAD patients.

N6-methyladenosine (m6A) modification is the most prevalent chemical modification in eukaryotic mRNA and is associated with the development of various immune diseases. However, the role of m6A methylation in rheumatoid arthritis (RA) development is unclear. We preliminarily explored the role of m6A methylation-related mRNAs in RA for its clinical application. The discovery of m6A methylation-modifying genes in this study may provide a fresh perspective on the development of drugs for RA treatment. High-throughput sequencing combined with methylated RNA immunoprecipitation (MeRIP-seq) and RNA sequencing were used to assess whole-transcriptome m6A modifications in the synovium of patients with RA. The relationship between m6A-modified target genes and RA inflammation and macrophages was determined. The expression of the m6A-modified significant transcript-enriched inflammatory signaling pathway was assessed through animal experiments. Differentially expressed m6A genes were correlated with macrophage activation involved in immune response, vascular endothelium, MAPK signaling pathway, PI3K − Akt signaling pathway, and other inflammatory processes. Furthermore, combined analysis with m6A-seq and RNA-seq revealed 120 genes with significant changes in both m6A modification and mRNA expression. We selected the top 3 candidate mRNAs that were upregulated and downregulated simultaneously. The expression of phosphatase and tensin homolog deleted on chromosome ten (PTEN) mRNA and protein in RA patients was lower than that in healthy control (HC). SHC-binding protein 1 (SHCBP1) and neurexophilin-3 (NXPH3) mRNA expressions were increased in RA patients. The expression of M1 macrophages was increased in RA patients. RA markers are such as rheumatoid factor (RF) and peptide containing citrulline (CCP). Further animal experiments showed that the expression of synovial MAPK, PI3K, and Akt1 proteins in the RA model was increased, and the PTEN, p-PTEN protein expression was decreased. PI3K, Akt1, PTEN, and p-PTEN were correlated to RA joint inflammation. This study revealed a unique pattern of differential m6A methylation modifications in RA and concluded that m6A modification is related to the occurrence of RA synovial inflammation.

BackgroundThe association involving N6-methyladenosine (m6A) modification, molecular subtype and specific immune cell group in tumor microenvironment has been the focus of recent studies. The underlying function of m6A modification in thyroid cancer (TC) remains elusive.MethodsThe m6A modification regulations, molecular character and tumor immune profile of 461 TC patients were explored and then the correlation between them were comprehensively evaluated. The m6Ascore was established using principal component analysis (PCA) to quantify the m6A pattern of individual TC patients. The prognostic significance of the m6Ascore was evaluated by multivariate Cox regression analysis.ResultsFour m6Aclusters (mc1, 2, 3, 4)—characterized by differences in extent of aneuploidy, expression of immunomodulatory genes, mRNA or lncRNA expression pattern and prognosis were identified. T Preliminary validation of m6Ascore was a potential independent prognostic factor of TC involving in mc3. Finally, the prognostic value of the m6Ascore and its association with copy number variation (CNV) and tumor immune microenvironment (TIME) of TC in mc3 were verified.ConclusionsThe correlation between m6A modification, the copy number burden and tumor immune landscape in TC was demonstrated. A m6Acluster-mc3 with low m6Ascore and high CNV molecular subtype was identified with poor clinical prognosis, low infiltrating immunocyte and weak effector T cell. A three-gene clinical prognosis model for TC based on 4 m6a cluster expression was established. Understanding of TIME is enhanced by comprehensive assessment of m6A patterns in individual TC patients and gives a new insight toward improved immunotherapy strategies for TC cancer patients.

BackgroundThe role of epigenetic modulation in immunity is receiving increased recognition—particularly in the context of RNA N6-methyladenosine (m6A) modifications. Nevertheless, it is still uncertain whether m6A methylation plays a role in the onset and progression of intracranial aneurysms (IAs). This study aimed to establish the function of m6A RNA methylation in IA, as well as its correlation with the immunological microenvironment.MethodsOur study included a total of 97 samples (64 IA, 33 normal) in the training set and 60 samples (44 IA, 16 normal) in the validation set to systematically assess the pattern of RNA modifications mediated by 22 m6A regulators. The effects of m6A modifications on immune microenvironment features, i.e., immune response gene sets, human leukocyte antigen (HLA) genes, and infiltrating immune cells were explored. We employed Lasso, machine learning, and logistic regression for the purpose of identifying an m6A regulator gene signature of IA with external data validation. For the unsupervised clustering analysis of m6A modification patterns in IA, consensus clustering methods were employed. Enrichment analysis was used to assess immune response activity along with other functional pathways. The identification of m6A methylation markers was identified based on a protein–protein interaction network and weighted gene co-expression network analysis.ResultsWe identified an m6A regulator signature of IGFBP2, IGFBP1, IGF2BP2, YTHDF3, ALKBH5, RBM15B, LRPPRC, and ELAVL1, which could easily distinguish individuals with IA from healthy individuals. Unsupervised clustering revealed three m6A modification patterns. Gene enrichment analysis illustrated that the tight junction, p53 pathway, and NOTCH signaling pathway varied significantly in m6A modifier patterns. In addition, the three m6A modification patterns showed significant differences in m6A regulator expression, immune microenvironment, and bio-functional pathways. Furthermore, macrophages, activated T cells, and other immune cells were strongly correlated with m6A regulators. Eight m6A indicators were discovered—each with a statistically significant correlation with IA—suggesting their potential as prognostic biological markers.ConclusionOur study demonstrates that m6A RNA methylation and the immunological microenvironment are both intricately correlated with the onset and progression of IA. The novel insight into patterns of m6A modification offers a foundation for the development of innovative treatment approaches for IA.

The role of RNA N6-methyladenosine (m6A) modification in the regulation of the immune microenvironment in ischaemic cardiomyopathy (ICM) remains largely unclear. This study first identified differential m6A regulators between ICM and healthy samples, and then systematically evaluated the effects of m6A modification on the characteristics of the immune microenvironment in ICM, including the infiltration of immune cells, the human leukocyte antigen (HLA) gene, and HALLMARKS pathways. A total of seven key m6A regulators, including WTAP, ZCH3H13, YTHDC1, FMR1, FTO, RBM15 and YTHDF3, were identified using a random forest classifier. A diagnostic nomogram based on these seven key m6A regulators could effectively distinguish patients with ICM from healthy subjects. We further identified two distinct m6A modification patterns (m6A cluster-A and m6A cluster-B) that are mediated by these seven regulators. Meanwhile, we also noted that one m6A regulator, WTAP, was gradually upregulated, while the others were gradually downregulated in the m6A cluster-A vs. m6A cluster-B vs. healthy subjects. In addition, we observed that the degree of infiltration of the activated dendritic cells, macrophages, natural killer (NK) T cells, and type-17 T helper (Th17) cells gradually increased in m6A cluster-A vs. m6A cluster-B vs. healthy subjects. Furthermore, m6A regulators, including FTO, YTHDC1, YTHDF3, FMR1, ZC3H13, and RBM15 were significantly negatively correlated with the above-mentioned immune cells. Additionally, several differential HLA genes and HALLMARKS signalling pathways between the m6A cluster-A and m6A cluster-B groups were also identified. These results suggest that m6A modification plays a key role in the complexity and diversity of the immune microenvironment in ICM, and seven key m6A regulators, including WTAP, ZCH3H13, YTHDC1, FMR1, FTO, RBM15, and YTHDF3, may be novel biomarkers for the accurate diagnosis of ICM. Immunotyping of patients with ICM will help to develop immunotherapy strategies with a higher level of accuracy for patients with a significant immune response.

The specific functions of RNA N6-methyladenosine (m6A) modifications in the glioma tumor microenvironment (TME) and glioma patient prognosis and treatment have not been determined to date. The objective of the study was to determine the role of m6A modifications in glioma TME. Nonnegative matrix factorization (NMF) methods were used to determine m6A clusters and m6A gene signatures based on 21 genes relating to m6A modifications. TME characteristics for each m6A cluster and m6A gene signature were quantified by established m6A score. The utility of m6A score was validated in immunotherapy and other antiangiogenic treatment cohorts. Three m6A clusters were identified among 3,395 glioma samples, and they were linked to different biological activities and clinical outcomes. The m6A clusters were highly consistent with immune profiles known as immune-inflamed, immune-excluded, and immune-desert phenotypes. Clusters within individual tumors could predict glioma inflammation, molecular subtypes, TME stromal activity, genetic variation, alternative splicing, and prognosis. As for the m6A score and m6A gene signature, patients with low m6A scores exhibited an increased tumor mutation burden, immune activity, neoantigen load, and prolonged survival. A low m6A score indicated the potential for a low level of T-cell dysfunction, a considerably better treatment response, and durable clinical benefits from immunotherapy, bevacizumab and regorafenib. Glioma m6A clusters and gene signatures have distinctive TME features. The m6A gene signature may guide prognostic assessments and promote the use of effective strategies.

Maternal stress caused by the environmental factors varied in intrauterine and extrauterine during pregnancy may significantly affect placental and fetal development, as well as offspring health in adulthood. Epigenetic mechanisms are frequently invoked to elucidate these effects. RNA N6-methyladenosine (m6A) modification, one of the most prevalent and abundant post-transcriptional epigenetic modifications in eukaryotic mRNA, has recently garnered widespread attention in life sciences. RNA m6A modification plays critical roles in RNA splicing, translation, localization, stability, and has been implicated in various biological processes, including embryonic development, sex determination, and disease pathogenesis. In studies of placental developmental abnormalities induced by maternal stress during pregnancy, m6A modification has emerged as a key mechanism. This article initially introduces the impact of RNA m6A methylation modification on placental development, subsequently elaborates on recent advances in understanding how maternal stress induces placental abnormalities via m6A modification, and finally summarizes unresolved key questions in this field. This review aims to propose strategies for preventing and treating placental developmental abnormalities caused by maternal stress.

BackgroundThe regulatory role of N6-methyladenosine (m6A) modification in the onset and progression of cancer has garnered increasing attention in recent years. However, the specific role of m6A modification in pulmonary metastasis of colorectal cancer remains unclear.MethodsThis study identified differential m6A gene expression between primary colorectal cancer and its pulmonary metastases using transcriptome sequencing and immunohistochemistry. We investigated the biological function of METTL3 gene both in vitro and in vivo using assays such as CCK-8, colony formation, wound healing, EDU, transwell, and apoptosis, along with a BALB/c nude mouse model. The regulatory mechanisms of METTL3 in colorectal cancer pulmonary metastasis were studied using methods like methylated RNA immunoprecipitation quantitative reverse transcription PCR, RNA stability analysis, luciferase reporter gene assay, Enzyme-Linked Immunosorbent Assay, and quantitative reverse transcription PCR.ResultsThe study revealed high expression of METTL3 and YTHDF1 in the tumors of patients with pulmonary metastasis of colorectal cancer. METTL3 promotes epithelial-mesenchymal transition in colorectal cancer by m6A modification of SNAIL mRNA, where SNAIL enhances the secretion of CXCL2 through the NF-κB pathway. Additionally, colorectal cancer cells expressing METTL3 recruit M2-type macrophages by secreting CXCL2.ConclusionMETTL3 facilitates pulmonary metastasis of colorectal cancer by targeting the m6A-Snail-CXCL2 axis to recruit M2-type immunosuppressive macrophages. This finding offers new research directions and potential therapeutic targets for colorectal cancer treatment.