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MicroRNA-based epigenetic approach to induce fetal hemoglobin

基于 MicroRNA 的表观遗传学方法诱导胎儿血红蛋白

基本信息

批准号：
10212449
负责人：
Athena Starlard-Davenport
金额：
$ 30.53万
依托单位：
UNIVERSITY OF TENNESSEE HEALTH SCI CTR
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-07-13 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10212449
关键词：
3&apos Untranslated Regions ANXA5 gene Address Adult Biological Assay Blood Cell Count CD34 gene Cell Proliferation Cells Child Cholesterol Clinical Complete Blood Count Continuous Infusion Cytosine DNA DNA Damage DNA Methylation DNA Methyltransferase Inhibitor DNA Modification Methylases DNA Sequence Alteration Data Development Differentiation Antigens Disease Dose Enzymes Epigenetic Process Erythroid Erythropoiesis FDA approved Fertility Fetal Hemoglobin Flow Cytometry Fluorescence Functional disorder GYPA gene Gene Expression Gene Expression Profiling Gene Expression Regulation Gene Silencing Genetic Transcription Genome Genotype Globin Goals Hematopoiesis Hemoglobinopathies Human Hypermethylation Hypoxia In Vitro Individual Infant Infusion procedures Knowledge Liquid substance MYB gene Malignant Neoplasms Measures Mediating Messenger RNA Methylation MicroRNAs Molecular Monitor Mus Mutation Oncogenes Patients Peripheral Blood Mononuclear Cell Pharmaceutical Preparations Pharmacotherapy Phenotype Physiological Plasmids Play Population Propidium Diiodide Publishing Pump Regulation Research Reticulocyte count Role Severity of illness Sickle Cell Sickle Cell Anemia Spleen Subcutaneous Injections Switch Genes Symptoms System TFRC gene Testing Therapeutic Therapeutic Intervention Tissues Transfection Transferase Treatment Efficacy Untranslated RNA Work base bench to bedside beta Globin cancer cell chromatin modification drug development efficacy testing epigenetic silencing erythroid differentiation gamma Globin hydroxyurea in vivo innovation methylation pattern mouse model novel novel therapeutics overexpression pre-clinical prevent progenitor promoter sickling side effect small molecule stem cells subcutaneous transcriptome sequencing

项目摘要

Project Abstract This SHINE II application seeks to explore epigenetic mechanisms that control globin gene regulation during erythropoiesis. The central hypothesis of this project is that that miR-29b reverses the γ-globin to β-globin gene switch and induces HbF via modulation of DNA methylation. Therapeutic interventions aimed at inducing HbF expression is an effective approach for ameliorating the clinical symptoms of sickle cell disease (SCD) in adults and children. Hydroxyurea is the only FDA-approved drug with proven efficacy for inducing HbF in patients with SCD, but DNA methyltransferase (DNMT) inhibitors have shown promise as HbF inducers by producing proximal γ-globin promoter DNA hypomethylation. However, DNMT inhibitors can produce off-target side effects. Small non-coding microRNAs (miR) are attractive molecules for targeting repressors of γ-globin gene expression and studies show that miR-29b inhibits DNA methylation through direct targeting of the 3’ untranslated region of DNMT3A and DNMT3B. The objective of this proposal is to test the efficacy of miR-29b as an HbF inducer. Our published work shows that miR-29b is important for reactivating γ-globin transcription and HbF expression by targeting MYB, a known repressor of γ-globin involved in mediating DNA methylation and gene silencing. To test our central hypothesis, we will accomplish one specific aim to determine the ability of miR-29b to mediate epigenetic changes in DNA methylation in the HBB locus and reactivate γ-globin transcription and HbF expression during adult erythropoiesis. Sub-aim A will explore the molecular effects of miR-29b on γ-globin regulation using an in vitro primary liquid culture system to generate erythroid progenitors from peripheral blood mononuclear cells isolated from individuals with SCD. These findings will compare to normal erythroid progenitors to validate the ability of miR-29b to reactivate γ-globin transcription through proximal promoter DNA methylation. Off-target effects mediated by miR-29b will be investigated by DNA mutation and RNA-seq analyses. To define the role of miR-29b in reversing phenotype, erythroid progenitor sickling under hypoxic conditions will be evaluated. Sub-aim B will establish the optimal dose of miR-29b that mediates γ-globin reactivation and HbF induction in vivo using a preclinical Townes SCD mouse model. This research highlights a novel miRNA-based epigenetic approach to induce HbF to impact the discovery of new drugs to expand treatment options for SCD.

项目摘要该 SHINE II 应用程序旨在探索控制珠蛋白基因的表观遗传机制红细胞生成过程中的调节。该项目的中心假设是 miR-29b 逆转 γ-珠蛋白至 β-珠蛋白基因开关并通过 DNA 调节诱导 HbF 甲基化。旨在诱导 HbF 表达的治疗干预是一种有效的方法用于改善成人和儿童镰状细胞病（SCD）的临床症状。羟基脲是 FDA 批准的唯一一种经证实可有效诱导 HbF 的药物 SCD，但 DNA 甲基转移酶 (DNMT) 抑制剂已显示出作为 HbF 诱导剂的前景产生近端 γ-珠蛋白启动子 DNA 低甲基化。然而，DNMT 抑制剂可以产生脱靶副作用。小的非编码 microRNA (miR) 是有吸引力的分子靶向 γ-珠蛋白基因表达的阻遏物，研究表明 miR-29b 抑制 DNA 通过直接靶向 DNMT3A 和 DNMT3B 的 3' 非翻译区进行甲基化。这该提案的目的是测试 miR-29b 作为 HbF 诱导剂的功效。我们发表的作品显示 miR-29b 对于重新激活 γ-珠蛋白转录和 HbF 表达非常重要靶向 MYB，一种已知的 γ-珠蛋白抑制因子，参与介导 DNA 甲基化和基因沉默。为了检验我们的中心假设，我们将实现一个特定目标来确定 miR-29b 介导 HBB 基因座 DNA 甲基化表观遗传变化的能力在成人红细胞生成过程中重新激活 γ-珠蛋白转录和 HbF 表达。子目标A将使用体外原液探索 miR-29b 对 γ-珠蛋白调节的分子效应从分离的外周血单核细胞中产生红系祖细胞的培养系统来自 SCD 患者。这些发现将与正常红系祖细胞进行比较以验证 miR-29b 通过近端启动子 DNA 重新激活 γ-球蛋白转录的能力甲基化。 miR-29b 介导的脱靶效应将通过 DNA 突变和 RNA-seq 分析。确定 miR-29b 在逆转红系祖细胞表型中的作用将评估缺氧条件下的镰状细胞。子目标 B 将确定最佳剂量使用临床前 Townes 介导体内 γ-球蛋白再激活和 HbF 诱导的 miR-29b SCD小鼠模型。这项研究强调了一种新的基于 miRNA 的表观遗传学方法来诱导 HbF 影响新药的发现，以扩大 SCD 的治疗选择。