Functions, mechanisms, and therapeutic potential of fetal hemoglobin inducers

胎儿血红蛋白诱导剂的功能、机制和治疗潜力

基本信息

批准号：
10308676
负责人：
Gerd A Blobel
金额：
$ 65.29万
依托单位：
CHILDREN'S HOSP OF PHILADELPHIA
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-08-05 至 2022-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10308676
关键词：
5&apos Untranslated Regions Accounting Adult Animals Attenuated Biological Process Birth CRISPR screen CRISPR/Cas technology Cell Culture Techniques Cell Line Cells Cellular biology Chromatin Clone Cells Codon Nucleotides Collaborations Complement Custom Data Defect Disease Enzymes Erythroblasts Erythrocytes Erythroid Erythroid Cells Fetal Hemoglobin Foundations Genes Genetic Screening Genetic Transcription Genetic Translation Globin Goals Hematopoietic stem cells Hemoglobin Hemoglobin concentration result Hemoglobinopathies Human Knockout Mice Laboratories Libraries Light Mass Spectrum Analysis Measures Medical Messenger RNA Monitor Mus Open Reading Frames PRKR gene Panthera leo Patients Pharmacology Phosphoric Monoester Hydrolases Phosphorylation Phosphotransferases Pre-Clinical Model Process Production Protein Kinase Proteins Proteome Proteomics Regulation Repression Resolution Ribosomes Role Severity of illness Sickle Cell Anemia Sum Surveys Testing Thalassemia Therapeutic Therapeutic Index Tissues Translations Western Blotting Work base beta Globin beta Thalassemia combinatorial design detection limit druggable target experimental study fetal gamma Globin improved improved outcome insight mouse model mutant novel restoration sickling small molecule synergism tool transcription factor transcriptome

项目摘要

Abstract Sickle cell disease (SCD) and some types of b-thalassemia that are caused by defects in the adult form of hemoglobin manifest shortly after birth, when the switch from the fetal to the adult form of hemoglobin is complete. Even a partial reversal of this switch is associated with an improved course of these diseases. We employed a newly improved CRISPR-Cas9 platform to carry out a kinase domain-focused genetic screen to identify potentially druggable molecules that repress fetal hemoglobin (HbF) production. This screen uncovered HRI (also known as EIF2AK1), an erythroid-specific protein kinase that regulates protein translation. Depletion of HRI elevates HbF levels in human erythroid cells with few additional perturbations. HRI loss reduces the expression of the major HbF repressor BCL11A, and restoration of BCL11A expression partially restores HbF repression. Moreover, HRI depletion reduces sickling of SCD-derived human erythroid cells in culture. In Aim 1 we will comprehensively dissect HRI function by assessing the transcriptome and proteome of HRI-depleted cells. The goals of Aim 2 are to study the mechanism by which HRI regulates BCL11A, identify additional HRI regulated HbF repressors, and examine the global impact of HRI on protein translational control in primary human erythroid cells. Aim 3 will explore synergies with previously known HbF inducers both using a candidate approach, and by unbiased genetic screens for novel synergies. In Aim 4 we will examine the effects of HRI loss on SCD by generating HRI-deficient humanized SCD mouse models. In sum, these studies explore the role of HRI in human red cell biology and examine HRI as target for pharmacologic HbF induction alone or in combination with mechanistically distinct HbF inducers.

抽象的镰状细胞疾病（SCD）和某些由缺陷引起的某些类型的B-心理症出生后不久，当胎儿切换时，血红蛋白的成年形式表现出来成年血红蛋白形式的完整。即使是部分逆转此开关与这些疾病的改善过程有关。我们采用了新的改进 CRISPR-CAS9平台执行以激酶结构域为中心的遗传屏幕以识别抑制胎儿血红蛋白（HBF）产生的潜在可药物分子。这屏幕揭开的HRI（也称为EIF2AK1），一种红斑特异性蛋白激酶调节蛋白质翻译。 HRI的耗竭升高了人类的HBF水平红细胞细胞几乎没有扰动。 HRI损失降低了主要的HBF阻遏物BCL11A和BCL11A表达的恢复部分恢复 HBF抑制。此外，HRI耗尽减少了SCD衍生的人的病态培养中的红细胞细胞。在AIM 1中，我们将通过评估缺乏HRI的细胞的转录组和蛋白质组。目标2的目标是要研究HRI调节BCL11A的机制，请确定其他HRI 调节HBF阻遏物，并检查HRI对蛋白质翻译的全球影响控制原代人红细胞细胞。 AIM 3将探索以前的协同作用已知的HBF诱导剂都使用候选方法，又是通过公正的遗传筛选用于新颖的协同作用。在AIM 4中，我们将检查HRI损失对SCD的影响生成HRI缺陷的人源性SCD小鼠模型。总而言之，这些研究探索了 HRI在人类红细胞生物学中的作用并研究HRI作为药理的靶标 HBF诱导单独或与机械上不同的HBF诱导剂结合使用。