Insights from somatic genomics lead to novel therapeutic targets in inherited bone marrow failure syndromes: Shwachman-Diamond Syndrome as a model.

项目摘要/摘要 遗传性骨髓衰竭综合征的特点是造血细胞再生不全和易患 骨髓发育不良。这些疾病被低估了，唯一有效的治疗方法是造血干细胞。 细胞移植。为了了解介导骨髓衰竭和克隆进化的机制， 这项建议的重点是Shwachman-Diamond综合征(SDS)，一种骨髓衰竭和MDS易感性 由SBDS基因的双等位基因突变引起的综合征。该基因编码一种调节核糖体的蛋白质。 成熟。SBDS蛋白特异性地促进成熟的、翻译上有活性的80S核糖体的形成 通过辅助催化将EIF6从60S核糖体亚基上移除。本报告中提供的初步数据 一项提案发现了骨骼中EIF6和TP53的反复和持续的体细胞错义突变 抑郁症患者的骨髓。最常见的EIF6突变会导致功能丧失并使EIF6失效 与60S核糖体亚基结合，改善了SDS细胞的适合性，为SDS治疗确定了一个新的靶点。 这一建议的中心假设是遗传性骨髓衰竭综合征已经减少 遗传性分子缺陷和体细胞突变决定的造血细胞适合性 潜在的健康缺陷或绕过细胞应激途径。该提案通过以下方式解决了这个假设 具体目标如下：(1)确定EIF6和EIF6的来源和造血后果 SDS中的TP53体细胞突变。(2)评估EIF6：60S相互作用的中断和EIF6的降低 蛋白质在十二烷基硫酸钠中的稳定性。在第一个目标中，单细胞DNA测序与DNA条形码相结合 免疫表型将用于确定克隆性构型和单个细胞的基因型-表型 十二烷基硫酸钠骨髓的体细胞突变。在第二个目标中，合理的从头设计的微型蛋白质将被 用于审问作为治疗策略的EIF6：60S结合。这项研究产生的数据将向 证候群相关骨髓衰竭的病理生理学和治疗，并可用于指导治疗 在其他骨髓衰竭综合征中，克隆性造血和核糖体成熟的研究。A K08 奖励将为候选人提供受保护的时间，以完成额外的计算方面的学术培训 生物学、体细胞基因组分析和高通量蛋白质组学方法。详细的职业发展 提出了一项计划，使候选人能够获得上述额外的科学专门知识和培训 在赠款撰写、领导力和管理技能方面。拟议的研究将在达纳- 法伯/波士顿儿童医院癌症和血液病中心，是购买 额外的培训。这位候选人保护了研究时间和强大的机构支持。此外， 候选人将有机会进入哈佛医学院附属的博德研究所和蛋白质研究所 创新。这些机构是世界知名的临床和研究中心，将使应聘者能够 开发更多的数据和培训，成为研究骨髓衰竭的独立内科医生兼科学家。

PROJECT SUMMARY/ABSTRACT Inherited bone marrow failure syndromes are characterized by hematopoietic aplasia and predisposition to myelodysplasia. These disorders are underdiagnosed, and the only curative treatment is hematopoietic stem cell transplant. In order to understand the mechanisms that mediate bone marrow failure and clonal evolution, this proposal focuses on Shwachman-Diamond Syndrome (SDS), a bone marrow failure and MDS predisposition syndrome caused by biallelic mutations in the SBDS gene. This gene encodes a protein that regulates ribosome maturation. The SBDS protein specifically promotes formation of the mature, translationally active 80S ribosome by aiding in catalysis of removal of EIF6 from the 60S ribosomal subunit. Preliminary data presented in this proposal has identified recurrent and persistent somatic missense mutations in EIF6 and TP53 within the bone marrow of patients with SDS. The most common EIF6 mutation leads to a loss of function and abolishes EIF6 binding with the 60S ribosomal subunit and improves SDS cell fitness, identifying a novel target for SDS therapy. The central hypothesis of this proposal is that inherited bone marrow failure syndromes have decreased hematopoietic cell fitness determined by the inherited molecular defect and somatic mutations specifically rescue the underlying fitness defect or bypass cell stress pathways. The proposal addresses this hypothesis with the following specific aims: (1) Identify the hematopoietic cell of origin and hematopoietic consequences of EIF6 and TP53 somatic mutations in SDS. (2) Evaluate the disruption of the EIF6:60S interaction and decreased EIF6 protein stability in SDS. In the first aim, single cell DNA sequencing combined with DNA-barcoded immunophenotyping will be used to determine clonal architecture and a single cell genotype-phenotype of somatic mutations in the SDS bone marrow. In the second aim, rational de novo designed miniproteins will be used to interrogate EIF6:60S binding as a therapeutic strategy. Data generated from this study will inform the pathophysiology and treatment of SDS-associated bone marrow failure and can be applied to inform treatment of other bone marrow failure syndromes, the study of clonal hematopoiesis and ribosome maturation. A K08 award will provide the candidate with protected time to complete additional academic training in computational biology, somatic genomic analysis and high-throughput proteomics approaches. A detailed career development plan is proposed which will enable the candidate to attain additional scientific expertise noted above and training in grant-writing, leadership and management skills. The proposed studies will take place at the Dana- Farber/Boston Children's Hospital Cancer and Blood disorders center, an exemplary location to acquire additional training. The candidate has protected research time and strong institutional support. In addition, the candidate will have access to the Harvard Medical School-affiliated Broad Institute and Institute for Protein Innovation. These institutions are world renowned clinical and research centers that will enable the candidate to develop additional data and training to become an independent physician-scientist studying bone marrow failure.