The role of oxidative stress in the pathogenesis of Reticular Dysgenesis and thetherapeutic potential of antioxidants

氧化应激在网状发育不全发病机制中的作用和抗氧化剂的治疗潜力

• 批准号: 9317208
• 负责人: Katja Gabriele Weinacht
• 金额: $ 4.94万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-14 至 2021-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9317208
• 关键词: Adenine Nucleotides; Adenosine Diphosphate; Adenosine Monophosphate; Advisory Committees; Affect; Aftercare; Antioxidants; Area; Award; Bacterial Infections; Biology; Boston; CRISPR/Cas technology; Cause of Death; Cell Differentiation process; Cell Line; Cell model; Cell physiology; Cessation of life; Child; Clinical; Complement; Constitution; Dana-Farber Cancer Institute; Data; Defect; Development; Development Plans; Diagnosis; Disease; Disease model; Effectiveness; Engraftment; Ensure; Enzymes; Generations; Genes; Glutathione; Goals; Health; Hematological Disease; Hematology; Hematopoiesis; Hematopoietic; Hematopoietic Stem Cell Transplantation; Immune; Immunologic Deficiency Syndromes; Immunology; In Situ; In Vitro; Institution; Lead; Life; Lymphopenia; Mediating; Mentors; Metabolic; Metabolism; Microbiology; Mitochondria; Modeling; Molecular; Mus; Mutate; Mutation; Mycoses; Myelogenous; Myeloid Cells; Natural Immunity; Nature; Neutropenia; Oncology; Outcome; Oxidation-Reduction; Oxidative Stress; Pathogenesis; Pathology; Patients; Pediatric Hematologist; Pediatric Hematology; Pediatric Hospitals; Physicians; Play; Positioning Attribute; Research; Research Personnel; Research Project Grants; Research Proposals; Research Training; Role; Scientist; Sensorineural Hearing Loss; Severe Combined Immunodeficiency; Specimen; Staging; Structure; Supportive care; Testing; Therapeutic; Therapeutic Human Experimentation; Time; Training; Translating; Translational Research; Transplantation; Universities; Work; Xenograft Model; Xenograft procedure; Zebrafish; adaptive immunity; adenylate kinase; base; career; career development; clinical care; congenital immunodeficiency; design; experience; gene correction; high risk; human disease; humanized mouse; improved; in vitro Model; in vivo; induced pluripotent stem cell; infancy; mitochondrial dysfunction; mitochondrial metabolism; mouse model; neutrophil; novel strategies; outcome forecast; progenitor; reconstitution; reticular dysgenesis; stem; stem cell biology; stem cell differentiation; stem cells; tandem mass spectrometry; targeted treatment; tenure track; therapeutic target; tissue culture

 DESCRIPTION (provided by applicant): Reticular Dysgenesis (RD) is one of the most serious forms of severe combined immunodeficiency (SCID) because it affects both innate and adaptive immunity. The disease is characterized by arrested neutrophil maturation, profound lymphopenia, and sensorineural hearing loss. It is invariably fatal early in life unless immune reconstitution is achieved by hematopoietic stem cell transplantation (HSCT). The simultaneous occurrence of severe neutropenia and lymphopenia is responsible for a high risk of death in infancy, and the predominance of bacterial and fungal infections supports neutropenia as the prevailing cause of death. RD is caused by mutations in the mitochondrial enzyme Adenylate Kinase 2 (AK2), however, how AK2 defects translate into disease pathology is largely unknown. Overall, transplant outcomes in RD are significantly worse compared to any other form of SCID, suggesting that the nature of the genetic defect may directly impact the poor prognosis. Therefore, elucidating the mechanistic basis of RD is critical in order to target the underlying problem and develop additional therapeutic options. Our prior work in induced pluripotent stem cell and zebrafish models of RD has shown that hematopoietic stem and progenitor cell differentiation is compromised but can be rescued by treatment with antioxidant agents (Rissone, Weinacht et al., J Exp Med, 2015, in press). These data led to the hypothesis that AK2 defects impair mitochondrial metabolism and increase oxidative stress, and that antioxidant agents improve mitochondrial function and represent a targeted supportive therapy to treat the neutropenia and overall constitution of patients with RD. To test this hypothesis I will dissect th molecular mechanisms underlying RD and their responsiveness to antioxidants by defining mitochondrial metabolism, oxidative stress and antioxidant reserve in different iPSC-models of RD before and after treatment with antioxidants. To demonstrate that the in vitro findings can be translated to patients, I will develop a xenograft model of RD based on transplantation of AK2-deficient iPSC-derived respecified multipotent hematopoietic progenitors into NSG-mice, and examine how antioxidant treatment affects engraftment, differentiation potential and mitochondrial function. If my preliminary findings that antioxidants improve mitochondrial function are supported by the proposed research, the therapeutic potential of these agents could be exploited in a much wider range of diseases, in which mitochondrial pathology and oxidative stress are at play. I am a pediatric hematologist with focus on immune and immune-mediated diseases and substantial prior research experience in molecular microbiology, stem cell differentiation and reprogramming, who is seeking K08 support for mentored research under the guidance of Dr. Luigi Notarangelo, Division of Immunology, Boston Children's Hospital, with Dr. George Daley, Division of Hematology/Oncology, Boston Children's Hospital/Dana-Farber Cancer Institute, as co-mentor. My long-term career objective is to obtain a tenure-track position as a physician-scientist at an academic center with special commitment to the study, diagnosis and treatment of primary immunodeficiencies. The K08 award will provide the protected time I need to advance my training in stem cell biology, metabolism, xenografting, in situ gene editing and translational research. I will devote a minimum of 80% of my time to a focused research project investigating the molecular mechanisms underlying RD and its responsiveness to antioxidants, and will complement this with 20% of my effort dedicated to clinical care of children with hematologic diseases. Boston Children's Hospital, Dana-Farber Cancer Institute and Harvard University are internationally recognized research institutions with renowned expert researchers in the areas of stem cell biology, hematopoiesis, metabolism, in situ gene editing, and development of xenograft-models. Furthermore, the Divisions of Pediatric Hematology/Oncology and Immunology have a distinguished record of training successful physician-scientists. I have assembled a mentoring and advisory committee, consisting of Drs. Raif Geha, David Williams, Alan Beggs, and Suneet Agarwal, who will guide my research and training experience. The expertise of my advisory committee will be complemented by a set of additional collaborators who are experts in their respective fields (Dr. Kiran Musunuru, gene editing with CRISPR/Cas9; Dr. Marcia Haigis, mitochondrial biology; and Dr. Giancarlo la Marca, Tandem Mass Spectrometry). This research proposal is part of a structured plan with scientific, technical, clinical training and career development components. The career development plan builds upon my prior research and clinical experiences with the goal of ensuring that I acquire the expertise required to become a successful, independent investigator whose focus is on translating disease mechanisms underlying immunodeficiencies into targeted therapies for patients.

 描述（由申请人提供）：网状发育不全（RD）是严重联合免疫缺陷（SCID）的最严重形式之一，因为它影响先天性和适应性免疫，该疾病的特征是中性粒细胞成熟停滞、严重淋巴细胞减少和感音神经性听力。除非通过造血干细胞移植（HSCT）实现免疫重建，否则它在生命早期总是致命的。淋巴细胞减少症是婴儿期死亡的高风险的原因，而细菌和真菌感染占主导地位，支持中性粒细胞减少症是主要的死亡原因，而 RD 是由线粒体酶腺苷酸激酶 2 (AK2) 突变引起的，然而，AK2 缺陷如何导致的。总体而言，与任何其他形式的 SCID 相比，RD 的移植结果明显较差，这表明遗传缺陷的性质可能直接影响不良预后。因此，阐明 RD 的机制基础对于解决根本问题并开发其他治疗方案至关重要。我们之前在 RD 的诱导多能干细胞和斑马鱼模型中的工作表明，造血干细胞和祖细胞的分化受到损害，但可以得到改善。通过抗氧化剂治疗可挽救（Rissone、Weinacht 等人，J Exp Med，2015 年，出版中）这些数据得出了 AK2 缺陷损害线粒体的假设。代谢并增加氧化应激，抗氧化剂可改善线粒体功能，是治疗 RD 患者中性粒细胞减少症和整体体质的靶向支持疗法。为了检验这一假设，我将通过定义来剖析 RD 背后的分子机制及其对抗氧化剂的反应。 RD 不同 iPSC 模型在抗氧化剂治疗前后的线粒体代谢、氧化应激和抗氧化剂储备 为了证明体外研究结果可以转化为患者，我将开发一个异种移植模型。 RD 基于将 AK2 缺陷的 iPSC 衍生的重新特异的多能造血祖细胞移植到 NSG 小鼠中，并研究抗氧化剂治疗如何影响植入、分化潜力和线粒体功能。如果我的初步发现抗氧化剂改善线粒体功能得到了拟议研究的支持，这些药物的治疗潜力可以用于更广泛的疾病，其中线粒体病理学和氧化应激在其中发挥作用。免疫和免疫介导的疾病以及分子微生物学、干细胞分化和重编程方面的丰富先前研究经验，他正在波士顿儿童医院免疫学部 Luigi Notarangelo 博士和 George 博士的指导下寻求 K08 支持以进行指导研究Daley，波士顿儿童医院/达纳法伯癌症研究所血液学/肿瘤学部的联合导师，我的长期职业目标是获得终身教授职位。 K08 奖将为我提供在干细胞生物学、新陈代谢、异种移植、原位基因编辑和转化方面的培训所需的受保护时间。我将投入至少 80% 的时间进行重点研究项目，调查 RD 的分子机制及其对抗氧化剂的反应，并将用 20% 的时间致力于儿童的临床护理来补充这一点。 波士顿儿童医院、达纳法伯癌症研究所和哈佛大学是国际公认的研究机构，在干细胞生物学、造血、代谢、原位基因编辑和异种移植模型开发领域拥有著名的专家研究人员。儿科血液学/肿瘤学和免疫学部门在培训成功的医师科学家方面有着杰出的记录，我组建了一个指导和咨询委员会，由 Raif Geha 博士、大卫·威廉姆斯 (David Williams)、艾伦·贝格斯 (Alan Beggs) 和苏尼特·阿加瓦尔 (Suneet Agarwal) 将指导我的研究和培训经验，我的咨询委员会的专业知识将得到一组其他合作者的补充，他们都是各自领域的专家（基兰·穆苏鲁 (Kiran Musunuru) 博士，基因编辑）。 CRISPR/Cas9 博士；Marcia Haigis 博士，线粒体生物学；Giancarlo la Marca 博士，串联质谱法）该研究计划是科学、技术、临床培训和职业发展的结构化计划的一部分。职业发展计划建立在我之前的研究和临床经验的基础上，目的是确保我获得成为一名成功的独立研究者所需的专业知识，其重点是将免疫缺陷的疾病机制转化为患者的靶向治疗。