bims-rimeca Biomed News
on RNA methylation in cancer
Issue of 2021‒05‒02
twenty-one papers selected by
Sk Ramiz Islam
Saha Institute of Nuclear Physics
  1. Expression and clinical prognostic value of m6A RNA methylation modification in breast cancer.
  2. Characterization of the Prognostic m6A-Related lncRNA Signature in Gastric Cancer.
  3. N6-Methyladenosine, DNA Repair, and Genome Stability.
  4. The role of RNA m6A methylation in the regulation of postnatal hypoxia-induced pulmonary hypertension.
  5. Analysis of RNA m6A methylation regulators and tumour immune cell infiltration characterization in prostate cancer.
  6. N6-methyladenosine-dependent signalling in cancer progression and insights into cancer therapies.
  7. Main N6-Methyladenosine Readers: YTH Family Proteins in Cancers.
  8. Saikosaponin D exhibits anti-leukemic activity by targeting FTO/m6A signaling.
  9. Multiple Functions of RNA Methylation in T Cells: A Review.
  10. METTL3 involves the progression of osteoarthritis probably by affecting ECM degradation and regulating the inflammatory response.
  11. The RNA helicase DDX5 promotes viral infection via regulating N6-methyladenosine levels on the DHX58 and NFκB transcripts to dampen antiviral innate immunity.
  12. MTTL3 upregulates microRNA-1246 to promote occurrence and progression of NSCLC via targeting paternally expressed gene 3.
  13. Splice site m6A methylation prevents binding of U2AF35 to inhibit RNA splicing.
  14. M6ADD: a comprehensive database of m6A modifications in diseases.
  15. Identification of Expression Patterns and Potential Prognostic Significance of m5C-Related Regulators in Head and Neck Squamous Cell Carcinoma.
  16. Small-Molecule Inhibitors of the RNA M6A Demethylases FTO Potently Support the Survival of Dopamine Neurons.
  17. Small molecule inhibition of METTL3 as a strategy against myeloid leukaemia.
  18. Tumors exploit FTO-mediated regulation of glycolytic metabolism to evade immune surveillance.
  19. METTL3 Regulates Angiogenesis by Modulating let-7e-5p and miRNA-18a-5p Expression in Endothelial Cells.
  20. ATP-Independent Initiation during Cap-Independent Translation of m6A-Modified mRNA.
  21. Methylation of NRN1 is a Novel Synthetic Lethal Marker for Combined PI3K-Akt-mTOR and ATR Inhibitors.
  1. Biomark Res. 2021 Apr 29. 9(1): 28
    Expression and clinical prognostic value of m6A RNA methylation modification in breast cancer.
    Fangchao Zheng, Feng Du, Haili Qian, Jiuda Zhao, Xue Wang, Jian Yue, Nanlin Hu, Yiran Si, Binghe Xu, Peng Yuan.
      BACKGROUND: N6-methyladenosine(m6A) methylation modification affects the tumorigenesis, progression, and metastasis of breast cancer (BC). However, the expression characteristics and prognostic value of m6A modification in BC are still unclear. We aimed to evaluate the relationship between m6A modification and clinicopathological characteristics, and to explore the underlying mechanisms.METHODS: Three public cohorts and our clinical cohort were included: 1091 BC samples and 113 normal samples from the TCGA database, 1985 BC samples from the METABRIC database, 1764 BC samples from the KM Plotter website, and 134 BC samples of our clinical cohort. We collected date from these cohorts and analyzed the genetic expression, gene-gene interactions, gene mutations, copy number variations (CNVs), and clinicopathological and prognostic features of 28 m6A RNA regulators in BC.
    RESULTS: This study demonstrated that some m6A regulators were significantly differenially expressed in BCs and their adjacent tissues, and also different in various molecular types. All 28 studied m6A regulators exhibited interactions. KIAA1429 had the highest mutation frequency. CNVs of m6A regulators were observed in BC patients. The expression of the m6A regulators was differentially associated with survival of BC. Higher CBLL1 expression was associated with a better prognosis in BC than lower CBLL1 expression. Functional analysis showed that CBLL1 was related to the ESR1-related pathway, apoptosis-related pathway, cell cycle pathway and immune-related pathway in BC.
    CONCLUSIONS: m6A RNA modification modulated gene expression and thereby affected clinicopathological features and survival outcomes in BC. CBLL1 may be a promising prognostic biomarker for BC patients.
    Keywords:  Breast cancer; CBLL1; Methylation; N6-methyladenosine modification; Prognosis
    DOI:  https://doi.org/10.1186/s40364-021-00285-w
  2. Front Oncol. 2021 ;11 630260
    Characterization of the Prognostic m6A-Related lncRNA Signature in Gastric Cancer.
    Haixu Wang, Qingkai Meng, Bin Ma.
      N6-methyladenosine (m6A) is a common form of mRNA modification regulated by m6A RNA methylation regulators and play an important role in the progression of gastric cancer (GC). However, the prognostic role of m6A-related lncRNA in gastric cancer has not been fully explored. This study aims at exploring the biological function and prognostic roles of the m6A-related lncRNA signature in gastric cancer. A total of 800 m6A-related lncRNAs were identified through Pearson correlation analysis between m6A regulators and all lncRNAs. Eleven m6A-related lncRNA signatures were identified through a survival analysis and the Kaplan-Meier (KM) curve analysis results suggest that patients in the low-risk group have a better overall survival (OS) and disease-free survival (DFS) outcome than the high-risk group. Also, the lncRNA signature can serve as an independent prognostic factor for OS and DFS. The gene set enrichment analysis (GSEA) result suggests that patients in the high-risk group were mainly enriched in the ECM receptor interaction, focal adhesion, and cytokine-cytokine receptor interaction pathway, while the low-risk group was characterized by the base excision repair pathway. We further constructed an individualized prognostic prediction model via the nomogram based on the independent prognostic factor for the OS and DFS, respectively. In addition, some candidate drugs aimed at GC risk group differentiation were identified using the Connective Map (CMAP) database. Lastly, four subgroups (C1, C2, C3, and C4) were identified based on the m6A-related lncRNA expression, through a consensus clustering algorithm. Among them, C1 and C2 have a greater likelihood to respond to immune checkpoint inhibitor immunotherapy, suggesting that the C1 and C2 subgroup might benefit from immunotherapy. In conclusion, the m6A-related lncRNA signature can independently predict the OS and DFS of GC and may aid in development of personalized immunotherapy strategies.
    Keywords:  N6-methyladenosine; gastric cancer; immunotherapy; lncRNA signature; molecular subgroups; nomogram
    DOI:  https://doi.org/10.3389/fonc.2021.630260
  3. Front Mol Biosci. 2021 ;8 645823
    N6-Methyladenosine, DNA Repair, and Genome Stability.
    Fei Qu, Pawlos S Tsegay, Yuan Liu.
      N6-methyladenosine (m6A) modification in mRNAs and non-coding RNAs is a newly identified epitranscriptomic mark. It provides a fine-tuning of gene expression to serve as a cellular response to endogenous and exogenous stimuli. m6A is involved in regulating genes in multiple cellular pathways and functions, including circadian rhythm, cell renewal, differentiation, neurogenesis, immunity, among others. Disruption of m6A regulation is associated with cancer, obesity, and immune diseases. Recent studies have shown that m6A can be induced by oxidative stress and DNA damage to regulate DNA repair. Also, deficiency of the m6A eraser, fat mass obesity-associated protein (FTO) can increase cellular sensitivity to genotoxicants. These findings shed light on the novel roles of m6A in modulating DNA repair and genome integrity and stability through responding to DNA damage. In this mini-review, we discuss recent progress in the understanding of a unique role of m6As in mRNAs, lncRNAs, and microRNAs in DNA damage response and regulation of DNA repair and genome integrity and instability.
    Keywords:  DNA damage; DNA repair; N6-methyladenosine; genome stability; histone modifications
    DOI:  https://doi.org/10.3389/fmolb.2021.645823
  4. Respir Res. 2021 Apr 26. 22(1): 121
    The role of RNA m6A methylation in the regulation of postnatal hypoxia-induced pulmonary hypertension.
    Shanshan Xu, Xuefeng Xu, Ziming Zhang, Lingling Yan, Liyan Zhang, Lizhong Du.
      BACKGROUND: Pulmonary hypertension (PH) is a complex pulmonary vascular disease characterized by an imbalance in vasoconstrictor/vasodilator signaling within the pulmonary vasculature. Recent evidence suggests that exposure to hypoxia early in life can cause alterations in the pulmonary vasculature and lead to the development of PH. However, the long-term impact of postnatal hypoxia on lung development and pulmonary function remains unknown. N6-methyladenosine (m6A) regulates gene expression and governs many important biological processes. However, the function of m6A in the development of PH remains poorly characterized. Thus, the purpose of this investigation was to test the two-fold hypothesis that (1) postnatal exposure to hypoxia would alter lung development leading to PH in adult rats, and (2) m6A modification would change in rats exposed to hypoxia, suggesting it plays a role in the development of PH.METHODS: Twenty-four male Sprague-Dawley rats were exposed to a hypoxic environment (FiO2: 12%) within 24 h after birth for 2 weeks. PH was defined as an increased right ventricular pressure (RVP) and pathologic changes of pulmonary vasculature measured by α-SMA immunohistochemical staining. Methylated RNA immunoprecipitation sequencing (MeRIP-seq) was performed to analyze m6A modification changes in lung tissue in 2- and 9-week-old rats that were exposed to postnatal hypoxia.
    RESULTS: Mean pulmonary arterial pressure, lung/body weight ratio, and the Fulton index was significantly greater in rats exposed to hypoxia when compared to control and the difference persisted into adulthood. m6A methyltransferase and demethylase proteins were significantly downregulated in postnatal hypoxia-induced PH. Distinct m6A modification peak-related genes differed between the two groups, and these genes were associated with lung development.
    CONCLUSIONS: Our results indicate postnatal hypoxia can cause PH, which can persist into adulthood. The development and persistence of PH may be because of the continuous low expression of methyltransferase like 3 affecting the m6A level of PH-related genes. Our findings provide new insights into the impact of postnatal hypoxia and the role of m6A in the development of pulmonary vascular pathophysiology.
    Keywords:  N6-methyladenosine; Postnatal hypoxia; Pulmonary hypertension; RNA methylation
    DOI:  https://doi.org/10.1186/s12931-021-01728-6
  5. Artif Cells Nanomed Biotechnol. 2021 Dec;49(1): 407-435
    Analysis of RNA m6A methylation regulators and tumour immune cell infiltration characterization in prostate cancer.
    Yue Zhao, Huimin Sun, Jianzhong Zheng, Chen Shao.
      Potential roles of RNA N6-methyladenosine (m6A) modification in tumour microenvironment (TME) cell infiltration has been demonstrated in recent studies. Nonetheless, the mechanism of its regulation remains unknown and immunotherapy has been marginal in prostate cancer. We demonstrated the expression of different m6A regulators within prostate cancer related to genetic variation, alternative splicing (AS), tumour mutational burden (TMB) and TME. Unsupervised clustering and risk prediction model constructed by 24 m6A regulators could predict scores of TME and prostate cancer patients prognosis. T cells CD8 was the intersection of immune cells which are related to multiple biological processes, and the fraction of T cells CD8 strongly correlates with immune associated gene sets. m6A methylation modification and immune cells infiltration played a nonnegligible role in prostate cancer. Our study represents a step towards personalized immunotherapy for prostate cancer patients.
    Keywords:  Prostate cancer; immune cell infiltration; m6A RNA methylation; prognosis; tumour microenvironment
    DOI:  https://doi.org/10.1080/21691401.2021.1912759
  6. J Exp Clin Cancer Res. 2021 Apr 29. 40(1): 146
    N6-methyladenosine-dependent signalling in cancer progression and insights into cancer therapies.
    Fenghua Tan, Mengyao Zhao, Fang Xiong, Yumin Wang, Shanshan Zhang, Zhaojian Gong, Xiayu Li, Yi He, Lei Shi, Fuyan Wang, Bo Xiang, Ming Zhou, Xiaoling Li, Yong Li, Guiyuan Li, Zhaoyang Zeng, Wei Xiong, Can Guo.
      The N6-methyladenosine (m6A) modification is a dynamic and reversible epigenetic modification, which is co-transcriptionally deposited by a methyltransferase complex, removed by a demethylase, and recognized by reader proteins. Mechanistically, m6A modification regulates the expression levels of mRNA and nocoding RNA by modulating the fate of modified RNA molecules, such as RNA splicing, nuclear transport, translation, and stability. Several studies have shown that m6A modification is dysregulated in the progression of multiple diseases, especially human tumors. We emphasized that the dysregulation of m6A modification affects different signal transduction pathways and involves in the biological processes underlying tumor cell proliferation, apoptosis, invasion and migration, and metabolic reprogramming, and discuss the effects on different cancer treatment.
    Keywords:  Cancer progression; N6-methyladenosine; RNA fate; Signal transduction pathway; Therapy
    DOI:  https://doi.org/10.1186/s13046-021-01952-4
  7. Front Oncol. 2021 ;11 635329
    Main N6-Methyladenosine Readers: YTH Family Proteins in Cancers.
    Xin-Yuan Dai, Liang Shi, Zhi Li, Hai-Yan Yang, Ji-Fu Wei, Qiang Ding.
      Among the over 150 RNA modifications, N6-methyladenosine (m6A) is the most abundant internal modification in eukaryotic RNAs, not only in messenger RNAs, but also in microRNAs and long non-coding RNAs. It is a dynamic and reversible process in mammalian cells, which is installed by "writers," consisting of METTL3, METTL14, WTAP, RBM15/15B, and KIAA1429 and removed by "erasers," including FTO and ALKBH5. Moreover, m6A modification is recognized by "readers," which play the key role in executing m6A functions. IYT521-B homology (YTH) family proteins are the first identified m6A reader proteins. They were reported to participate in cancer tumorigenesis and development through regulating the metabolism of targeted RNAs, including RNA splicing, RNA export, translation, and degradation. There are many reviews about function of m6A and its role in various diseases. However, reviews only focusing on m6A readers, especially YTH family proteins are few. In this review, we systematically summarize the recent advances in structure and biological function of YTH family proteins, and their roles in human cancer and potential application in cancer therapy.
    Keywords:  IYT521-B homology domain proteins; N6-methyladenosine; RNA metabolism; cancer; cancer therapy
    DOI:  https://doi.org/10.3389/fonc.2021.635329
  8. Theranostics. 2021 ;11(12): 5831-5846
    Saikosaponin D exhibits anti-leukemic activity by targeting FTO/m6A signaling.
    Kaiju Sun, Yangyang Du, Yuzhu Hou, Mingyue Zhao, Jiajia Li, Yazhe Du, Lingxiao Zhang, Changbao Chen, Hongmei Yang, Fei Yan, Rui Su.
      Purpose: The implementation of targeted therapies for acute myeloid leukemia (AML) has been challenging. Fat mass and obesity associated protein (FTO), an mRNA N6-methyladenosine (m6A) demethylase, functions as an oncogene that promotes leukemic oncogene-mediated cell transformation and leukemogenesis. Here, we investigated the role of Saikosaponin-d (SsD) in broad anti-proliferation effects in AML and evaluated the m6A demethylation activity by targeting FTO of SsD. Methods: It was examined whether and how SsD regulates FTO/m6A signaling in AML. The pharmacologic activities and mechanisms of actions of SsD in vitro, in mice, primary patient cells, and tyrosine kinase inhibitors-resistant cells were determined. Results: SsD showed a broadly-suppressed AML cell proliferation and promoted apoptosis and cell-cycle arrest both in vitro and in vivo. Mechanistically, SsD directly targeted FTO, thereby increasing global m6A RNA methylation, which in turn decreased the stability of downstream gene transcripts, leading to the suppression of relevant pathways. Importantly, SsD also overcame FTO/m6A-mediated leukemia resistance to tyrosine kinase inhibitors. Conclusion: Our findings demonstrated that FTO-dependent m6A RNA methylation mediated the anti-leukemic actions of SsD, thereby opening a window to develop SsD as an epitranscriptome-base drug for leukemia therapy.
    Keywords:  FTO; N6-methyladenosine; Saikosaponin D; leukemia
    DOI:  https://doi.org/10.7150/thno.55574
  9. Front Immunol. 2021 ;12 627455
    Multiple Functions of RNA Methylation in T Cells: A Review.
    Yinong Chao, Hua-Bing Li, Jing Zhou.
      RNA modification represents one of the most ubiquitous mechanisms of epigenetic regulation and plays an essential role in modulating cell proliferation, differentiation, fate determination, and other biological activities. At present, over 170 types of RNA modification have been discovered in messenger RNA (mRNA) and noncoding RNA (ncRNA). RNA methylation, as an abundant and widely studied epigenetic modification, is crucial for regulating various physiological or pathological states, especially immune responses. Considering the biological significance of T cells as a defense against viral infection and tumor challenge, in this review, we will summarize recent findings of how RNA methylation regulates T cell homeostasis and function, discuss the open questions in this rapidly expanding field of RNA modification, and provide the theoretical basis and potential therapeutic strategies involving targeting of RNA methylation to orchestrate beneficial T cell immune responses.
    Keywords:  RNA methylation; T cell; epigenetics; immune function; m6A
    DOI:  https://doi.org/10.3389/fimmu.2021.627455
  10. Life Sci. 2021 Apr 21. pii: S0024-3205(21)00513-0. [Epub ahead of print] 119528
    METTL3 involves the progression of osteoarthritis probably by affecting ECM degradation and regulating the inflammatory response.
    Weilin Sang, Song Xue, Yafei Jiang, Haiming Lu, Libo Zhu, Cong Wang, Jinzhong Ma.
      We aimed to identify RNA N6-methyladenosine methylation associated genes in osteoarthritis (OA), and to explore possible regulatory mechanisms of these RNA methylation associated genes. Bioinformatics analyses, including differential expression analysis, functional enrichment analysis, verification analysis, and box plot analysis, were conducted based on different datasets from OA and non-OA patients. Gene expression at mRNA and protein levels was determined by quantitative reverse transcription PCR, western blot and immunofluorescence. Interleukin 1β (IL-1β)-treated SW1353 cells was used as cell model. Lentiviral vector was used for over-expression METTL3 in vitro. CCK-8 assay kit was used to determine cell viability and inflammatory cytokines (IL-1α, IL-6, IL-8, IL-10 and TNF-α) was detected using ELISA kits. Bioinformatics analysis showed that METTL3 expression was decreased in OA group, which was confirmed in clinical samples. Expression of METTL3 was also reduced in IL-1β-treated cells. Levels of inflammatory cytokines were obviously reduced in the METTL3 overexpression group, while IL-1β treatment reversed such decrease caused by METTL3 overexpression (p < 0.05). Both METTL3 overexpression and IL-1β treatment promoted expression of p65 protein and p-ERK (p < 0.01). Additionally, increased expression of MMP1 and MMP3, and decreased expression of MMP13, TIMP-1, and TIMP-2 at both mRNA and protein levels were observed in the METTL3 overexpression group when compared with the control group (p < 0.01). Expression of m6A methylation gene METTL3 was reduced in OA. METTL3 is involved in OA probably by regulating the inflammatory response. METTL3 overexpression may affect extracellular matrix degradation in OA by adjusting the balance between TIMPs and MMPs.
    Keywords:  Bioinformatics analysis; Immunofluorescence; Osteoarthritis; Quantitative RT-PCR; Western blot
    DOI:  https://doi.org/10.1016/j.lfs.2021.119528
  11. PLoS Pathog. 2021 Apr;17(4): e1009530
    The RNA helicase DDX5 promotes viral infection via regulating N6-methyladenosine levels on the DHX58 and NFκB transcripts to dampen antiviral innate immunity.
    Jian Xu, Yunhong Cai, ZhenBang Ma, Bo Jiang, Wenxiao Liu, Jing Cheng, Nannan Guo, Zishu Wang, Joshua E Sealy, Cuiping Song, Xiaojia Wang, Yongqing Li.
      Multi-functional DEAD-box helicase 5 (DDX5), which is important in transcriptional regulation, is hijacked by diverse viruses to facilitate viral replication. However, its regulatory effect in antiviral innate immunity remains unclear. We found that DDX5 interacts with the N6-methyladenosine (m6A) writer METTL3 to regulate methylation of mRNA through affecting the m6A writer METTL3-METTL14 heterodimer complex. Meanwhile, DDX5 promoted the m6A modification and nuclear export of transcripts DHX58, p65, and IKKγ by binding conserved UGCUGCAG element in innate response after viral infection. Stable IKKγ and p65 transcripts underwent YTHDF2-dependent mRNA decay, whereas DHX58 translation was promoted, resulting in inhibited antiviral innate response by DDX5 via blocking the p65 pathway and activating the DHX58-TBK1 pathway after infection with RNA virus. Furthermore, we found that DDX5 suppresses antiviral innate immunity in vivo. Our findings reveal that DDX5 serves as a negative regulator of innate immunity by promoting RNA methylation of antiviral transcripts and consequently facilitating viral propagation.
    DOI:  https://doi.org/10.1371/journal.ppat.1009530
  12. Mol Ther Nucleic Acids. 2021 Jun 04. 24 542-553
    MTTL3 upregulates microRNA-1246 to promote occurrence and progression of NSCLC via targeting paternally expressed gene 3.
    Shaohong Huang, Shaoning Luo, Chulian Gong, Limin Liang, Yi Xiao, Mingan Li, Jinyuan He.
      Non-small cell lung cancer (NSCLC) is one of the major causes of morbidity and mortality worldwide. We aimed to investigate the role of N6-methyladenosine (m6A) methyltransferase-like 3 (METTL3) regulating microRNA-1246 (miR-1246) in the progression of NSCLC by targeting paternally expressed gene 3 (PEG3). METTL3, miR-1246, and PEG3 expression in tissues was assessed, and the predictive role of METTL3 in prognosis of patients with NSCLC was detected. NSCLC cells were relatively treated with altered expression of METTL3, miR-1246, or PEG3 to measure their roles in the proliferation, migration, invasion, apoptosis, and in vivo growth of the NSCLC cells. The RNA m6A level was determined, and the targeting relationship between miR-1246 and PEG3 was confirmed. Our results revealed that METTL3 and miR-1246 were upregulated, whereas PEG3 was downregulated in NSCLC tissues. METTL3 knockdown or PEG3 overexpression in NSCLC cells suppressed malignant behaviors of NSCLC cells. METTL3 affected the m6A modification of miR-1246, thus upregulating miR-1246 and miR-1246-targeted PEG3. The elevation of PEG3 reversed the effects of miR-1246 upregulation on NSCLC cells. This study revealed that m6A methyltransferase METTL3 affects the m6A modification of miR-1246, thus upregulating miR-1246 to promote NSCLC progression by inhibiting PEG3.
    Keywords:  m6A methyltransferase-like 3; m6A modification; microRNA-1246; non-small cell lung cancer; paternally expressed gene 3; tumorigenesis
    DOI:  https://doi.org/10.1016/j.omtn.2021.02.020
  13. Cell. 2021 Apr 27. pii: S0092-8674(21)00435-9. [Epub ahead of print]
    Splice site m6A methylation prevents binding of U2AF35 to inhibit RNA splicing.
    Mateusz Mendel, Kamila Delaney, Radha Raman Pandey, Kuan-Ming Chen, Joanna M Wenda, Cathrine Broberg Vågbø, Florian A Steiner, David Homolka, Ramesh S Pillai.
      The N6-methyladenosine (m6A) RNA modification is used widely to alter the fate of mRNAs. Here we demonstrate that the C. elegans writer METT-10 (the ortholog of mouse METTL16) deposits an m6A mark on the 3' splice site (AG) of the S-adenosylmethionine (SAM) synthetase pre-mRNA, which inhibits its proper splicing and protein production. The mechanism is triggered by a rich diet and acts as an m6A-mediated switch to stop SAM production and regulate its homeostasis. Although the mammalian SAM synthetase pre-mRNA is not regulated via this mechanism, we show that splicing inhibition by 3' splice site m6A is conserved in mammals. The modification functions by physically preventing the essential splicing factor U2AF35 from recognizing the 3' splice site. We propose that use of splice-site m6A is an ancient mechanism for splicing regulation.
    Keywords:  3' splice site; METT-10; METTL16; SAM homeostasis; SAM synthetase; U2AF35/65; U6 snRNA; m(6)A; spermatogenesis; splicing
    DOI:  https://doi.org/10.1016/j.cell.2021.03.062
  14. RNA Biol. 2021 Apr 27. 1-9
    M6ADD: a comprehensive database of m6A modifications in diseases.
    Dianshuang Zhou, Hongli Wang, Fanqi Bi, Jie Xing, Yue Gu, Cong Wang, Menyan Zhang, Yan Huang, Jiaqi Zeng, Qiong Wu, Yan Zhang.
      N6-methyladenosine (m6A) modification is an important regulatory factor affecting diseases, including multiple cancers and it is a developing direction for targeted disease therapy. Here, we present the M6ADD (m6A-diseases database) database, a public data resource containing manually curated data on potential m6A-disease associations for which some experimental evidence is available; the related high-throughput sequencing data are also provided and analysed by using different computational methods. To give researchers a tool to query the m6A modification data, the M6ADD was designed as a web-based comprehensive resource focusing on the collection, storage and online analysis of m6A modifications, aimed at exploring the associations between m6A modification and gene disorders and diseases. The M6ADD includes 222 experimentally confirmed m6A-disease associations, involving 59 diseases from a review of more than 2000 published papers. The M6ADD also includes 409,229 m6A-disease associations obtained by computational and statistical methods from 30 high-throughput sequencing datasets. In addition, we provide data on 5239 potential m6A regulatory proteins related to 24 cancers based on network analysis prediction methods. In addition, we have developed a tool to explore the function of m6A-modified genes through the protein-protein interaction networks. The M6ADD can be accessed at http://m6add.edbc.org/.
    Keywords:  M6a modification; diseases; experimentally confirmed data; high-throughput sequencing data
    DOI:  https://doi.org/10.1080/15476286.2021.1913302
  15. Front Oncol. 2021 ;11 592107
    Identification of Expression Patterns and Potential Prognostic Significance of m5C-Related Regulators in Head and Neck Squamous Cell Carcinoma.
    Zhenyuan Han, Biao Yang, Yu Wang, Xiuxia Zeng, Zhen Tian.
      5-Methylcytosine (m5C) methylation is a major epigenetic technique of RNA modification and is dynamically mediated by m5C "writers," "erasers," and "readers." m5C RNA modification and its regulators are implicated in the onset and development of many tumors, but their roles in head and neck squamous cell carcinoma (HNSCC) have not yet been completely elucidated. In this study, we examined expression patterns of core m5C regulators in the publicly available HNSCC cohort via bioinformatic methods. The differentially expressed m5C regulators could divide the HNSCC cohort into four subgroups with distinct prognostic characteristics. Furthermore, a three-gene expression signature model, comprised of NSUN5, DNMT1, and DNMT3A, was established to identify individuals with a high or low risk of HNSCC. To explore the underlying mechanism in the prognosis of HNSCC, screening of differentially expressed genes, followed by the analysis of functional and pathway enrichment, from individuals with high- or low-risk HNSCC was performed. The results revealed a critical role for m5C RNA modification in two aspects of HNSCC: (1) dynamic m5C modification contributes to the regulation of HNSCC progression and (2) expression patterns of NSUN5, DNMT1, and DNMT3A help to predict the prognosis of HNSCC.
    Keywords:  TCGA; expression pattern; head and neck squamous cell carcinoma; m5C RNA methylation; prognostic signature
    DOI:  https://doi.org/10.3389/fonc.2021.592107
  16. Int J Mol Sci. 2021 Apr 26. pii: 4537. [Epub ahead of print]22(9):
    Small-Molecule Inhibitors of the RNA M6A Demethylases FTO Potently Support the Survival of Dopamine Neurons.
    Simona Selberg, Li-Ying Yu, Olesja Bondarenko, Esko Kankuri, Neinar Seli, Vera Kovaleva, Koit Herodes, Mart Saarma, Mati Karelson.
      The fat mass and obesity-associated protein (FTO), an RNA N6-methyladenosine (m6A) demethylase, is an important regulator of central nervous system development, neuronal signaling and disease. We present here the target-tailored development and biological characterization of small-molecule inhibitors of FTO. The active compounds were identified using high-throughput molecular docking and molecular dynamics screening of the ZINC compound library. In FTO binding and activity-inhibition assays the two best inhibitors demonstrated Kd = 185 nM; IC50 = 1.46 µM (compound 2) and Kd = 337 nM; IC50 = 28.9 µM (compound 3). Importantly, the treatment of mouse midbrain dopaminergic neurons with the compounds promoted cellular survival and rescued them from growth factor deprivation induced apoptosis already at nanomolar concentrations. Moreover, both the best inhibitors demonstrated good blood-brain-barrier penetration in the model system, 31.7% and 30.8%, respectively. The FTO inhibitors demonstrated increased potency as compared to our recently developed ALKBH5 m6A demethylase inhibitors in protecting dopamine neurons. Inhibition of m6A RNA demethylation by small-molecule drugs, as presented here, has therapeutic potential and provides tools for the identification of disease-modifying m6A RNAs in neurogenesis and neuroregeneration. Further refinement of the lead compounds identified in this study can also lead to unprecedented breakthroughs in the treatment of neurodegenerative diseases.
    Keywords:  ALKBH5; FTO; RNA; dopamine neurons; drug design; m6A
    DOI:  https://doi.org/10.3390/ijms22094537
  17. Nature. 2021 Apr 26.
    Small molecule inhibition of METTL3 as a strategy against myeloid leukaemia.
    Eliza Yankova, Wesley Blackaby, Mark Albertella, Justyna Rak, Etienne De Braekeleer, Georgia Tsagkogeorga, Ewa S Pilka, Demetrios Aspris, Dan Leggate, Alan G Hendrick, Natalie A Webster, Byron Andrews, Richard Fosbeary, Patrick Guest, Nerea Irigoyen, Maria Eleftheriou, Malgorzata Gozdecka, Joao M L Dias, Andrew J Bannister, Binje Vick, Irmela Jeremias, George S Vassiliou, Oliver Rausch, Konstantinos Tzelepis, Tony Kouzarides.
      The N6-methyladenosine (m6A) is an abundant internal RNA modification1,2 catalysed predominantly by the METTL3-METTL14 methyltransferase complex3,4. The m6A writer METTL3 has been linked to the initiation and maintenance of acute myeloid leukaemia (AML), but its true therapeutic importance is still unknown5-7. Here we present the identification and characterization of a highly potent and selective first-in-class catalytic inhibitor of METTL3 (STM2457) and its co-crystal structure bound to METTL3/METTL14. Treatment with STM2457 leads to reduced AML growth, and an increase in differentiation and apoptosis. These cellular effects are accompanied by selective reduction of m6A levels on known leukaemogenic mRNAs and a decrease in their expression consistent with a translational defect. We demonstrate that pharmacological inhibition of METTL3 in vivo leads to impaired engraftment and prolonged survival in various AML mouse models, specifically targeting key stem-cell subpopulations of AML. Collectively, these results reveal the inhibition of METTL3 as a potential therapeutic strategy against AML, and provide proof of concept that the targeting of RNA modifying enzymes represents a promising new avenue for anti-cancer therapy.
    DOI:  https://doi.org/10.1038/s41586-021-03536-w
  18. Cell Metab. 2021 Apr 21. pii: S1550-4131(21)00167-4. [Epub ahead of print]
    Tumors exploit FTO-mediated regulation of glycolytic metabolism to evade immune surveillance.
    Yi Liu, Guanghao Liang, Hongjiao Xu, Wenxin Dong, Ze Dong, Zhiwei Qiu, Zihao Zhang, Fangle Li, Yue Huang, Yilin Li, Jun Wu, Shenyi Yin, Yawei Zhang, Peijin Guo, Jun Liu, Jianzhong Jeff Xi, Peng Jiang, Dali Han, Cai-Guang Yang, Meng Michelle Xu.
      The ever-increasing understanding of the complexity of factors and regulatory layers that contribute to immune evasion facilitates the development of immunotherapies. However, the diversity of malignant tumors limits many known mechanisms in specific genetic and epigenetic contexts, manifesting the need to discover general driver genes. Here, we have identified the m6A demethylase FTO as an essential epitranscriptomic regulator utilized by tumors to escape immune surveillance through regulation of glycolytic metabolism. We show that FTO-mediated m6A demethylation in tumor cells elevates the transcription factors c-Jun, JunB, and C/EBPβ, which allows the rewiring of glycolytic metabolism. Fto knockdown impairs the glycolytic activity of tumor cells, which restores the function of CD8+ T cells, thereby inhibiting tumor growth. Furthermore, we developed a small-molecule compound, Dac51, that can inhibit the activity of FTO, block FTO-mediated immune evasion, and synergize with checkpoint blockade for better tumor control, suggesting reprogramming RNA epitranscriptome as a potential strategy for immunotherapy.
    Keywords:  RNA m6A modification; demethylase FTO; epitranscriptome; glycolytic metabolism; immune surveillance and immunotherapy
    DOI:  https://doi.org/10.1016/j.cmet.2021.04.001
  19. Arterioscler Thromb Vasc Biol. 2021 Apr 29. ATVBAHA121316180
    METTL3 Regulates Angiogenesis by Modulating let-7e-5p and miRNA-18a-5p Expression in Endothelial Cells.
    Aránzazu Chamorro-Jorganes, Walid K Sweaad, Rajesh Katare, Marie Besnier, Maryam Anwar, N Beazley-Long, Graciela Sala-Newby, Iñigo Ruiz-Polo, Dhananjie Chandrasekera, Alison A Ritchie, Andrew V Benest, Costanza Emanueli.
      OBJECTIVE: Postnatal angiogenesis is critical in vascular homeostasis and repair. m6A RNA methylation is emerging as a new layer for fine-tuning gene expression. Although the contribution of the m6A-catalyzing enzyme, METTL3 (methyltransferase-like 3), in cancer biology has been described, its role in endothelial cell (EC) function, particularly during angiogenesis, remains unclear. Approach and Results: To characterize the relevance of METTL3 in angiogenesis regulation, we performed gain- and loss-of-function studies in vitro. We demonstrated that depletion of METTL3 in ECs reduced the level of m6A and impaired EC function, whereas adenovirus-mediated METTL3 overexpression increased angiogenesis. Mechanistically, we showed that METTL3 depletion in ECs decreased mature angiogenic microRNAs let-7e-5p and the miR-17-92 cluster, and increased the expression of their common target, Tsp1 (thrombospondin 1). Conversely, Ad.METTL3 increased the expression of let-7e-5p and miR-17-92 cluster and reduced protein levels of Tsp1 in ECs. Moreover, overexpression of let-7e-5p and miR-18a-5p restored the angiogenic potential of METTL3-depleted ECs. We corroborated our data in vivo employing 3 mouse models. When tested in an in vivo Matrigel plug assay, METTL3-depleted ECs had diminished ability to vascularize the plug, whereas overexpression of METTL3 promoted angiogenesis. Local Ad.METTL3 gene transfer increased postischemic neovascularization in mice with either unilateral limb ischemia or myocardial infarction.CONCLUSIONS: METTL3 regulates m6A RNA methylation in ECs. Endogenous METTL3 is essential for EC function and angiogenesis, potentially through influencing let-7e and miR-17-92 cluster processing. Thus, the therapeutic modulation of METTL3 should be considered as a new approach for controlling angiogenic responses in the clinical setting.
    Keywords:  endothelial cell; gene expression; homeostasis; microRNA; thrombospondin
    DOI:  https://doi.org/10.1161/ATVBAHA.121.316180
  20. Int J Mol Sci. 2021 Apr 01. pii: 3662. [Epub ahead of print]22(7):
    ATP-Independent Initiation during Cap-Independent Translation of m6A-Modified mRNA.
    Pavel A Sakharov, Egor A Smolin, Dmitry N Lyabin, Sultan C Agalarov.
      The methylation of adenosine in the N6 position (m6A) is a widely used modification of eukaryotic mRNAs. Its importance for the regulation of mRNA translation was put forward recently, essentially due to the ability of methylated mRNA to be translated in conditions of inhibited cap-dependent translation initiation, e.g., under stress. However, the peculiarities of translation initiation on m6A-modified mRNAs are not fully known. In this study, we used toeprinting and translation in a cell-free system to confirm that m6A-modified mRNAs can be translated in conditions of suppressed cap-dependent translation. We show for the first time that m6A-modified mRNAs display not only decreased elongation, but also a lower efficiency of translation initiation. Additionally, we report relative resistance of m6A-mRNA translation initiation in the absence of ATP and inhibited eIF4A activity. Our novel findings indicate that the scanning of m6A-modified leader sequences is performed by a noncanonical mechanism.
    Keywords:  48S complex assembly; m6A-modified RNA; toeprinting; translation initiation
    DOI:  https://doi.org/10.3390/ijms22073662
  21. Cancer Sci. 2021 May 01.
    Methylation of NRN1 is a Novel Synthetic Lethal Marker for Combined PI3K-Akt-mTOR and ATR Inhibitors.
    Wushuang Du, Aiai Gao, James G Herman, Lidong Wang, Lirong Zhang, Shunchang Jiao, Mingzhou Guo.
      Wnt, PI3K-Akt-mTOR and NF-κB pathways were reported to be involved in DNA damage repair (DDR). DDR deficient cancers become critically dependent on backup DNA repair pathways. Neuritin 1 (NRN1) is reported to be involved in PI3K-Akt-mTOR, and its role in DDR remains unclear. Methylation-specific PCR, siRNA, flow cytometry, esophageal cancer cell lines and xenograft mouse models were used to examine the role of NRN1 in esophageal cancer. The expression of NRN1 is frequently repressed by promoter region methylation in human esophageal cancer cells. NRN1 was methylated in 50.4% (510/1012) of primary esophageal cancer samples. NRN1 methylation is associated significantly with age (p<0.001), tumor size (p<0.01), TNM stage (p<0.001), differentiation (p<0.001) and alcohol consumption (p<0.05). We found NRN1 methylation is an independent prognostic factor for poor 5-years OS (p<0.001). NRN1 inhibits colony formation, cell proliferation, migration and invasion, and induces apoptosis and G1/S arrest in esophageal cancer cells. NRN1 suppresses KYSE150 and KYSE30 cells xenografts growth in nude mice. PI3K signaling is reported to activate ATR signaling by targeting CHK1, the downstream component of ATR. By analyzing the synthetic efficiency of NVP-BEZ235 (PI3K inhibitor) and VE-822 (an ATR inhibitor), we found that combination of NVP-BEZ235 and VE-822 increased cytotoxicity in NRN1 methylated esophageal cancer cells, as well as KYSE150 cell xenografts. In conclusions, NRN1 suppresses esophageal cancer growth both in vitro and in vivo by inhibiting PI3K-Akt-mTOR signaling. Methylation of NRN1 is a novel synthetic lethal marker for PI3K-Akt-mTOR and ATR inhibitors in human esophageal cancer.
    Keywords:  ATR inhibitor; DNA Damage Repair; DNA methylation; NRN1; PI3K signaling
    DOI:  https://doi.org/10.1111/cas.14917
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