Role of RNA methylation in chemoresistant cancer cells

RNA 甲基化在化疗耐药癌细胞中的作用

• 批准号: 10083714
• 负责人: Shobha Vasudevan
• 金额: $ 19.26万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-15 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10083714
• 关键词: 3' Untranslated Regions; ATM Signaling Pathway; Acute Myelocytic Leukemia; Adenosine; Affect; Affinity Chromatography; Antibodies; Biological Assay; Blast Cell; Blood; Cancer Etiology; Cancer Relapse; Cell Cycle; Cell Line; Cell Survival; Cells; Chemicals; Chemoresistance; Clinical; Co-Immunoprecipitations; Complex; Coupled; Data; Disease remission; Gene Expression; Genes; Genetic Transcription; Goals; Heterogeneity; Immunoprecipitation; Lead; Malignant Neoplasms; Maps; Mediating; Messenger RNA; Methods; Methylation; Methyltransferase; Modification; N(6)-ribosyladenine; Normal Cell; Outcome; Patients; Population; Positioning Attribute; Post-Transcriptional Regulation; Proteome; Proteomics; RNA; RNA methylation; RNA-Binding Proteins; Reader; Recurrence; Regulation; Resistance; Role; Sampling; Signal Transduction; Site; Testing; Therapeutic; Translating; Translations; acute myeloid leukemia cell; base; biological heterogeneity; cancer cell; cancer stem cell; chemotherapy; crosslink; cytokine; immune resistance; in vivo; in vivo Model; inhibitor/antagonist; leukemic stem cell; mRNA Expression; new therapeutic target; protein degradation; stem cell biomarkers; targeted treatment; therapeutic biomarker; transcriptome; tumor

Summary Role of RNA methylation in chemoresistant cancer cells Goal Based on our data, we propose that transiently quiescent populations in acute myeloid leukemia (AML) are maintained in part by RNA methylation, permitting synthesis of survival and tumor promoting regulators necessary for chemoresistance and subsequent AML persistence. The primary objective is to characterize the role of RNA methylation in regulating gene expression in resistant AML, which contributes to chemosurvival. Significance AML is a serious malignancy that displays clinical resistance due to heterogeneity. Resistant AML cells include quiescent (G0) cells that are transiently arrested states, and thus resist clinical therapy that targets only cycling cells; these cells re-enter proliferation to cause AML persistence. A primary issue is that resistance regulators remain to be uncovered. A second need is to identify markers to detect resistant cells. Such cells show distinct gene expression that permits chemosurvival (PNAS 2014, Mol. Cell 2016, Biorxiv/ 418715). Canonical translation is decreased; yet specific mRNAs are expressed by unknown mechanisms. Regulation of specific gene expression in resistant AML needs to be uncovered & targeted to curtail AML. Premise Profiling in resistant cells revealed expression of factors that affect cell survival. Expressed mRNAs in G0 resistant cells have extended 3? untranslated regions (UTRs) to include regulatory sites, such as for RNA methylation. Methylation at N6 position of adenosines (m6A) are unique marks on RNAs that regulate gene expression via RNA binding proteins called readers, to control distinct cell states. Role of m6A in resistant AML remains to be uncovered. We find the m6A methyltransferase increases due to therapy induced signals. Our data reveal that m6A is needed for specific gene expression & resistance. We developed inhibitors to block therapy induced signals & antisense to block unique m6A sites on extended 3’UTRs, which reduce resistance. These data suggest that m6A regulates specific gene expression to enable therapy survival. Characterization of mRNAs that are modified & expressed in resistant cells, in vivo & in patient samples by m6A, their readers, their role in resistance, & of inhibitors that block m6A, will provide markers & therapeutics to curb resistance. Method Aim I will characterize m6A readers that regulate known modified mRNAs, by in vivo crosslinking coupled RNA affinity purification in resistant AML cells & in vivo models. Chemical inhibitors to therapy induced signaling that promotes m6A and antisense to block mapped m6A sites will be tested to curb resistance. Aim II will globally identify m6A targets that contribute to resistance in cell lines & in vivo models, using m6A antibody & methyltransferase immunoprecipitation. Role of readers & m6A mRNAs in resistant cells & in vivo is tested by depletions coupled with chemosurvival assays; their expression will be verified in resistant patient samples. Outcomes mRNAs expressed by m6A, their readers, their role in resistance in AML cells & in vivo, & inhibitors of m6A to curb resistance will be identified. Their expression & efficacy in patient samples will be uncovered.

RNA甲基化在化疗耐药癌细胞中的作用 根据我们的数据，我们提出急性髓系白血病（AML）中短暂静止的人群， 部分由RNA甲基化维持，允许合成存活和肿瘤促进调节剂 化疗耐药性和随后的AML持续性所必需的。主要目的是描述 RNA甲基化在耐药AML中调节基因表达的作用，这有助于化疗存活。 AML是一种严重的恶性肿瘤，由于异质性而表现出临床耐药性。耐 AML细胞包括处于短暂停滞状态的静止（G 0）细胞，因此抵抗临床治疗， 仅针对周期细胞;这些细胞重新进入增殖，导致AML持续存在。一个主要问题是， 抗性调节剂仍有待发现。第二个需要是鉴定标记物以检测抗性细胞。 这样的细胞显示出允许化学存活的不同基因表达（PNAS 2014，Mol.细胞2016，Biorxiv/ 418715）。典型的翻译减少，但特定的mRNA表达的未知机制。 耐药AML中特定基因表达的调控需要被发现和靶向以减少AML。 在抗性细胞中的预分析揭示了影响细胞存活的因子的表达。表达的mRNA G 0耐药细胞已延长3？非翻译区（UTR）包括调控位点，如RNA 甲基化腺苷N6位甲基化（m6 A）是调控基因表达的RNA上的独特标记 通过称为阅读器的RNA结合蛋白表达，以控制不同的细胞状态。m6 A在耐药AML中的作用 仍有待发现。我们发现由于治疗诱导的信号，m6 A甲基转移酶增加。我们 数据显示，m6 A是特定基因表达和抗性所必需的。我们开发了抑制剂 治疗诱导的信号和反义阻断延长的3 'UTR上的独特m6 A位点，从而降低耐药性。 这些数据表明，m6 A调节特定的基因表达，使治疗生存。表征 在抗性细胞中、体内和患者样品中被m6 A修饰和表达的mRNA，它们的阅读器， 它们在耐药性中的作用，以及阻断m6 A的抑制剂，将提供抑制耐药性的标志物和治疗剂。 方法目的我将通过体内交联来表征调节已知修饰的mRNA的m6 A阅读器 在耐药AML细胞和体内模型中偶联RNA亲和纯化。治疗诱导的化学抑制剂 将测试促进m6 A的信号传导和反义阻断定位的m6 A位点以抑制抗性。Aim II 将使用m6 A抗体，在细胞系和体内模型中全面鉴定导致耐药性的m6 A靶标 和甲基转移酶免疫沉淀。测试了阅读器和m6 A mRNA在抗性细胞和体内的作用 通过消耗结合化疗存活测定;将在耐药患者样品中验证它们的表达。 结果m6 A表达的mRNAs，它们的阅读器，它们在AML细胞和体内耐药中的作用，以及抑制剂 m6 A的抗阻力能力。它们在患者样本中的表达和功效将被发现。

项目成果

NR4A1 regulates expression of immediate early genes, suppressing replication stress in cancer.

• DOI: 10.1016/j.molcel.2021.09.016
• 发表时间: 2021-10-07
• 期刊: Molecular cell
• 影响因子: 16
• 作者: Guo H;Golczer G;Wittner BS;Langenbucher A;Zachariah M;Dubash TD;Hong X;Comaills V;Burr R;Ebright RY;Horwitz E;Vuille JA;Hajizadeh S;Wiley DF;Reeves BA;Zhang JM;Niederhoffer KL;Lu C;Wesley B;Ho U;Nieman LT;Toner M;Vasudevan S;Zou L;Mostoslavsky R;Maheswaran S;Lawrence MS;Haber DA
• 通讯作者: Haber DA

Long-read sequencing reveals complex patterns of wraparound transcription in polyomaviruses.

• DOI: 10.1371/journal.ppat.1010401
• 发表时间: 2022-04
• 期刊: PLoS pathogens
• 影响因子: 6.7