Novel therapeutic targets for Diamond Blackfan anemia and other ribosomopathies

描述(由申请人提供)：应聘者Anupama Narla提出了一项为期5年的职业发展计划，旨在利用三种不同的方法，寻找治疗钻石黑扇贫血(DBA)和其他核糖体功能障碍疾病的新型治疗药物，同时建立转化性儿科血液学的学术生涯。这项建议的具体目的是：(1)确定来那度胺对红系分化的作用机制，并研究其在钻石黑扇贫血治疗中的潜在作用；(2)培养和鉴定来自钻石黑扇贫血和5q综合征患者的iPS细胞；以及(3)在核糖体单倍体不足的细胞中进行RNAi筛选，以确定新的治疗药物。她的主要导师本杰明·埃伯特博士发现RPS14是5q综合征红系表型的候选基因，并迅速确立了自己在新兴核糖体疾病领域的领导者地位。核糖体病最近才在人类中被发现，它包括一系列疾病，其中遗传异常导致核糖体生物发生和功能受损。除了DBA和5q综合征外，正常核糖体生物发生所需的其他基因突变也与几种罕见的先天性综合征有关，包括Shwachman-Diamond综合征、先天性角化不良、软骨毛发发育不良和Treacher Collins综合征。在相对较短的时间内，候选人在Ebert博士的直接监督下，熟练掌握了成功实现其提案目标所需的所有技术和方法技能。她描述了地塞米松和来那度胺单独和联合使用对体外培养的原代人骨髓祖细胞分化的影响。她发现，这两种药物都促进了正常CD34+细胞和CD34+细胞的红细胞生成，这些细胞的核糖体功能障碍类型在DBA和del(5q)骨髓增生异常综合征中发现。她还与布罗德研究所合作，使用细胞培养中氨基酸的稳定同位素标记(SILAC)将CBR1确定为来那度胺的靶标。她还帮助产生了一株红细胞生成缺陷的iPS细胞系，该细胞系来自一名DBA患者。她的第一篇作者论文关于地塞米松和来那度胺对红细胞生成的影响，在第一轮积极评价后，将于2011年3月重新提交给血液。她是核糖病领域三篇综述论文的第一作者，这三篇论文已经或正在发表在《血液》、《国际血液学杂志》和《血液学研讨会》上。她是一篇描述核糖体单倍体功能不全中P53激活对红系特异性影响的论文的第二作者；这篇论文作为一篇全体文章发表在《血液》印刷版上。此外，另一篇确定了microRNA miR-145在5q综合征病理生理学中的作用的第二作者论文将于2011年5月重新提交给血液。2009年12月，她在美国血液病学会上以口头形式介绍了这项研究。基于她在埃伯特实验室的科学成就和她在儿科血液学方面的背景，这位候选人概述了她的短期和长期职业目标，在这些目标中，她将继续将80%的精力投入到翻译血液学的研究中，将20%的时间投入到骨髓衰竭综合征和教育的临床实践中。为了实现这些目标，她组建了一支由导师、顾问和合作者组成的才华横溢的团队。除了她的主要导师埃伯特博士，候选人还将由伦纳德·佐恩共同指导，后者在成功指导内科科学家方面有着既定的记录。她的导师将至少每月与候选人会面，以监督和帮助她过渡到独立。戴维·威廉姆斯(David Williams)和南希·伯林纳(Nancy Berliner)都是血液学部门的负责人和国际领导者，他们将担任候选人的研究委员会，该委员会至少每六个月举行一次会议，以确保科学研究计划取得成功。这位候选人还招募了iPS技术领域的领先者George Daley、波士顿儿童医院骨髓衰竭诊所主任Colin Sief和领导基于SILAC的靶标识别开发的Steven Carr作为她的项目的合作者。SILAC靶标识别允许无偏见地识别与药物结合的蛋白质。最后，候选人的好处是在波士顿儿童医院的卡普研究大楼进行她提出的研究，那里也是大卫·威廉姆斯、斯科特·阿姆斯特朗、斯图尔特·奥尔金、伦纳德·宗、南希·伯林纳和乔治·戴利的实验室。这些集中在造血方面的临床和科学专业知识将成为纳拉博士的额外资源。执行本提案中概述的具体目标和研究职业发展计划将使候选人能够成功地过渡到职业生涯，成为一名专注于开发核糖体生物发生障碍新疗法的翻译研究的独立研究员。

DESCRIPTION (provided by applicant): The candidate, Anupama Narla, presents a 5-year career development plan that seeks to identify novel therapeutic agents for Diamond Blackfan anemia (DBA) and other disorders of ribosome dysfunction, using three different approaches, while establishing an academic career in translational pediatric hematology. The specific aims of this proposal are to: (1) determine the mechanism of action of lenalidomide on erythroid differentiation and study its potential role in the treatment of Diamond Blackfan anemia; (2) generate and characterize iPS cells derived from patients with Diamond Blackfan anemia and the 5q- syndrome; and (3) perform an RNAi screen in cells with ribosomal haploinsufficiency in order to identify novel therapeutic agents. Her primary mentor, Dr. Benjamin Ebert, identified RPS14 as the candidate gene for the erythroid phenotype of the 5q- syndrome, and has quickly established himself as a leader in the emerging field of ribosomopathies. Ribosomopathies, which have only very recently been identified in humans, comprise a collection of disorders in which genetic abnormalities cause impaired ribosome biogenesis and function. In addition to DBA and the 5q- syndrome, mutations in other genes required for normal ribosome biogenesis have been implicated in several rare congenital syndromes including Shwachman-Diamond Syndrome, Dyskeratosis Congenita, Cartilage Hair Hypoplasia, and Treacher Collins Syndrome. In a relatively short period of time, the candidate, under the direct supervision of Dr. Ebert, has gained proficiency in all of the technical and methodological skills needed to successfully carry out the objectives of her proposal. She has characterized the effects of dexamethasone and lenalidomide, individually and in combination, on the differentiation of primary human bone marrow progenitor cells in vitro. She found that both agents promote erythropoiesis in normal CD34+ cells and in CD34+ cells with the types of ribosome dysfunction found in DBA and del(5q) myelodysplastic syndrome. She has also identified CBR1 as a target of lenalidomide using stable isotope labeling with amino acids in cell culture (SILAC) in collaboration with the Broad Institute. And she has been instrumental in the generation of an iPS cell line with defective erythropoiesis, derived from a patient with DBA. Her first author paper on the effects of dexamethasone and lenalidomide on erythropoiesis is being re-submitted to Blood in March 2011 after an initial round of positive reviews. She is the first author on three review papers in the field of ribosomopathies which have been or are being published in Blood, International Journal of Hematology, and Seminars in Hematology. She is the second author on a paper that described the erythroid specific effects of p53 activation in ribosomal haploinsufficiency; this paper was featured as a plenary article in the print edition of Blood. In addition, another second author paper which identified the role of a microRNA, miR-145, in the pathophysiology of the 5q- syndrome will be resubmitted to Blood in May 2011. She presented this work as an oral session at the American Society of Hematology in December 2009. Building on her scientific accomplishments in the Ebert lab and her background in pediatric hematology, the candidate has outlined both short and long-term career goals in which she would continue to devote 80% of her effort to research in translational hematology and 20% of her time to clinical practice in bone marrow failure syndromes and education. To accomplish these goals, she has assembled a highly talented team of mentors, advisors, and collaborators. In addition to her primary mentor, Dr. Ebert, the candidate will be co-mentored by Leonard Zon who has an established record of successfully mentoring physician-scientists. Her mentors will meet with the candidate at least monthly to supervise and assist in her transition to independence. David Williams and Nancy Berliner, both division heads and international leaders in hematology, are serving as the candidate's research committee which will meet, at minimum, every six months to ensure a successful scientific research program. The candidate has also recruited George Daley, a leader in the field of iPS technology; Colin Sieff, Director of the Bone Marrow Failure Clinic at Children's Hospital Boston; and Steven Carr, who led the development of SILAC-based target identification which allows for the unbiased identification of proteins that bind to a drug, as collaborators for her project. Finally, the candidate has the benefit of carrying out her proposed research in the Karp Research Building at Children's Hospital Boston which also houses the labs of David Williams, Scott Armstrong, Stuart Orkin, Leonard Zon, Nancy Berliner, and George Daley. This concentration of clinical and scientific expertise on hematopoiesis will be an additional resource to Dr. Narla. Execution of the specific aims and research career development plan outlined in this proposal will allow the candidate to successfully transition to a career as an independent investigator in translational research focused on developing new therapies for disorders of ribosome biogenesis.