Characterization of Calmodulin and CHK2 inhibitors for the treatment of Diamond B

用于治疗 Diamond B 的钙调蛋白和 CHK2 抑制剂的表征

• 批准号: 8976079
• 负责人: Elizabeth R Macari
• 金额: $ 5.8万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-12-01 至 2016-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8976079
• 关键词: Adenocarcinoma; Adverse effects; Animal Model; Antipsychotic Agents; Binding; Binding Proteins; Biogenesis; Biology; Blood - brain barrier anatomy; Bone Marrow Transplantation; CD34 gene; CHEK2 gene; Calmodulin; Cell Culture Techniques; Cells; Chemicals; Child; Clinical Trials; Confocal Microscopy; Craniofacial Abnormalities; Data; Defect; Development; Diamond; Diamond-Blackfan anemia; Disease; Dysmyelopoietic Syndromes; Embryo; Erythrocytes; Erythroid; Erythropoiesis; FDA approved; Fluorescence; Genes; Goals; Health; Hematopoiesis; Hematopoietic; Hemoglobin concentration result; Human; In Vitro; Lead; Libraries; MDM2 gene; Malignant Neoplasms; Mediating; Messenger RNA; Modeling; Mutate; Mutation; Nuclear; Pancytopenia; Patients; Permeability; Phenotype; Phosphorylation; Phosphotransferases; Process; Protein Deficiency; Protein p53; RPS19 gene; Ribosomal Proteins; Ribosomes; Role; Syndrome; TP53 gene; Testing; Therapeutic Uses; Toxic effect; Transcript; Translating; Translations; Work; Zebrafish; cancer risk; chromosome 5q loss; effective therapy; high throughput screening; in vivo; ineffective therapies; inhibitor/antagonist; knock-down; leukemia/lymphoma; mortality; mutant; novel; osteogenic; prevent; scaffold; screening; small hairpin RNA; targeted treatment; transcriptome sequencing

DESCRIPTION (provided by applicant): Ribosomal protein mutations are found in many diseases, including Diamond Blackfan anemia (DBA), where defective erythropoiesis, craniofacial abnormalities and increased cancer risk are major complications. Ribosomal protein mutations cause p53 activation through accumulation of free ribosomal proteins that bind and sequester MDM2, the negative regulator of p53. Current DBA treatments are ineffective and associated with toxicity. To find effective therapies, we performed an in vivo chemical screen in a zebrafish DBA model and found that inhibitors of calmodulin (CaM), and CaM-dependent kinase CHK2, block p53 activity and rescue red blood cell development. Many CaM inhibitors are FDA approved antipsychotics, but cross the blood brain barrier (BBB), and have irreversible, negative side effects. Our goals are to elucidate the mechanism by which CaM and CHK2 inhibitors block p53 nuclear activation, and to find CaM or CHK2 inhibitors that do not cross the BBB, and can therefore be used in clinical trials for DBA patients. DBA cell culture models will be used do mutational analysis to determine the mechanism of CaM inhibition on p53 nuclear accumulation. Additionally, a zebrafish DBA model will be used to study the effects of ribosomal protein deficiency and the role of p53 on translation, in vivo. To find compounds that will not cross the BBB, we will use a novel chemical scaffold approach to generate 400 new CaM inhibitors to be tested for BBB permeability using a fluorescence confocal microscopy screening approach. Select chemicals will then be tested for erythroid rescue in zebrafish and in human/mammalian DBA models. This work will identify potential compounds to further our understanding of ribosome-p53 biology and develop for clinical trials for ribosomal protein disorders.

描述(申请人提供)：核糖体蛋白突变在许多疾病中被发现，包括钻石黑扇贫血(DBA)，红细胞生成缺陷、头面部异常和癌症风险增加是主要并发症。核糖体蛋白突变通过积累结合和隔离MDM2的游离核糖体蛋白来激活P53，MDM2是P53的负调控因子。目前的DBA治疗是无效的，并与毒性有关。为了找到有效的治疗方法，我们在斑马鱼DBA模型中进行了体内化学筛选，发现钙调蛋白(CaM)的抑制剂和CaM依赖的激酶Chk2可以阻断P53的活性并挽救红细胞的发育。许多钙调素抑制剂是FDA批准的抗精神病药物，但可以通过血脑屏障，具有不可逆转的负面副作用。我们的目标是阐明CaM和Chk2抑制剂阻断P53核激活的机制，并寻找不跨越血脑屏障的CaM或Chk2抑制剂，因此可以用于DBA患者的临床试验。采用DBA细胞培养模型进行突变分析，以确定CaM抑制P53核积聚的机制。此外，还将使用斑马鱼DBA模型在体内研究核糖体蛋白缺乏的影响和P53在翻译中的作用。为了寻找不会通过血脑屏障的化合物，我们将使用一种新的化学支架方法来产生400种新的CaM抑制剂，以使用荧光共聚焦显微镜筛选方法来测试血脑屏障的渗透性。然后，将在斑马鱼和人类/哺乳动物DBA模型中测试选定的化学物质对红血球的拯救作用。这项工作将确定潜在的化合物，以加深我们对核糖体-p53生物学的理解，并为核糖体蛋白紊乱的临床试验开发。