Engineering an HIV-resistant immune system for HIV cure

 描述（由申请人提供）：候选人，Hind Fadel，医学博士，哲学博士、是马约临床医学院的助理教授。她是医学系传染病科和马约诊所分子医学科的成员。Fadel博士是一位致力于基础和转化HIV-1研究和基因治疗的医学科学家。她已经聚集了一个经验丰富的导师，合作者和顾问团队，并提出了一个全面的研究，课程工作和职业发展计划，这将使她成为一个成功的独立调查员。马约诊所提供的环境非常适合该奖项的研究和职业发展部分的成功。她还维护和确定了马约诊所以外的顾问和合作者。职业发展计划将使她能够磨练她在基因组编辑与工程位点特异性核酸酶的技能，深入了解宿主因素在HIV-1生物学中的作用，并开发新的技能来操纵人类基因组用于HIV-1治疗应用和基因治疗。该研究计划的重点是开发艾滋病毒治疗策略，使用新型工程核酸酶靶向/编辑HIV-1依赖性和限制因子基因，以产生HIV-1抗性。该方法将应用于原代CD 34+造血干细胞（HSC），并在人源化小鼠中进行研究。 虽然抗逆转录病毒药物使治疗艾滋病成为可能，但在许多方面，它们是一种“半途而废的技术”。“它们使现有患者容易受到主要问题的影响，包括抵抗力，持续的免疫功能障碍和免疫衰老，复杂的代谢紊乱和加速衰老现象。第二个问题是，虽然药物治疗在理论上可以防止进一步传播，但与终身提供药丸组合和保持坚持有关的许多问题意味着这种流行病继续扩大。一种彻底的治疗方法可以解决这两个问题。直到最近，治愈被认为是未来主义的，但现在已经改变了。最近，一名艾滋病毒和白血病患者-所谓的“柏林患者”-通过骨髓移植治愈了一名缺乏艾滋病毒主要细胞辅因子（进入辅受体CCR 5）的捐赠者，这给该领域注入了活力。特定的方法（同种异体BMT）对没有癌症的人来说毒性太大。然而，靶向HIV需要的基因（宿主辅因子）的价值已经很清楚了。沿着最近出现的非常有前途的基因靶向技术，以及我们对疾病过程本身的理解的进步，柏林患者有助于催化使治愈成为该领域的中心优先事项。对患者自身的HSC进行修饰以在患有HIV/AIDS的受试者中再生抗HIV的免疫系统，再加上适当的温和调节，可能是有效的治疗策略。必须征聘和培养年轻的调查人员，以充实这一肯定是重要的、不断扩大的领域。 Fadel博士建议在开发治疗性基因治疗方法时针对HIV-1的两个弱点：（i）消除HIV-1所需的宿主依赖性因素，以及（ii）编辑HIV-1进化逃避的宿主限制因素，以恢复其限制HV-1的能力。她的中心假设是，工程核酸酶将有效地靶向HIV-1依赖性和限制性因子，以产生对HIV-1感染的抗性，用于治疗应用，并进一步加深我们对HIV-1发病机制中这些因素的理解。主要目标是：1）敲除人CD 4 + T细胞系中的HIV-1依赖性因子，以同时确定它们的病毒学作用并建立它们作为HIV治疗应用的治疗靶点的潜力; 3）在离体CD 34 + HSC细胞中产生依赖性因子敲除和限制性因子敲入，并在人源化小鼠的临床前动物研究中表征它们。 拟议的研究有可能深入了解多种宿主因子在HIV-1生物学中的作用，并导致HIV-1治疗的转化应用。在这种疾病中开发HSC介导的治疗方法也可以推进其他感染性，自身免疫性和血液学疾病的相关应用。

 DESCRIPTION (provided by applicant): The candidate, Hind Fadel, M.D., Ph.D., is an Assistant Professor at the Mayo Clinic College of Medicine. She is a member of the Division of Infectious Diseases in the Department of Medicine, and of the Department of Molecular Medicine at the Mayo Clinic. Dr. Fadel is a physician-scientist committed to a career in basic and translational HIV-1 research and gene therapy. She has gathered an experienced team of mentors, collaborators and advisors and is proposing a comprehensive research, course work, and career development plan that will position her to be a successful independent investigator. The environment provided by the Mayo Clinic is ideally suited for the success of both the research and career development components of the award. She has also maintained and identified advisors and collaborators outside Mayo Clinic. The career development plan will allow her to hone her skills in genome editing with engineered site-specific nucleases, to gain insights into host factors role in HIV-1 biology, and to develop new skills to manipulate human genome for HIV-1 cure applications and gene therapy. The research plan focuses on developing HIV cure strategies that use targeting/editing of HIV-1 dependency and restriction factor genes with novel engineered nucleases in order to create HIV-1 resistance. The methods will be applied to primary CD34+ hematopoietic stem cells (HSCs) and studied in humanized mice. While antiretroviral drugs have made treatment of HIV disease possible, they in many ways represent a "halfway technology." They leave existing patients vulnerable to major problems that include resistance, persistent immune dysfunction and immunosenescence, complex metabolic disturbances, and accelerated aging phenomena. The second problem is that while drug treatment can in theory prevent further transmission, the many problems associated with supplying the pill combinations life-long and with maintaining adherence have meant that the epidemic continues to expand. A definitive cure could solve both problems. Cure was until recently considered futuristic, but this has now changed. The recent cure of a patient with both HIV and leukemia -- the so-called "Berlin patient" -- by bone marrow transplantation from a donor lacking a main HIV cellular cofactor (the entry co-receptor CCR5) was energizing to the field. The specific approach (allogeneic BMT) is far too toxic for anyone without cancer. However, the value of targeting a gene that HIV needs (a host cofactor) has been made clear. Along with the very recent emergence of highly promising gene targeting technologies, and advances in our understanding of the disease process itself, the Berlin patient helped catalyze making cure a central priority in the field. Modification of a patient's own HSCs to regenerate an HIV- resistant immune system in subjects with HIV/AIDS, coupled with appropriately mild conditioning, could be an effective cure strategy. It is essential to recruit and develop young investigators to populate what is certain to be an important, expanding field. Dr. Fadel is proposing to target two vulnerabilities of HIV-1 as she develops curative gene therapy approaches: (i) eliminating host dependency factors HIV-1 requires, and (ii) editing host restriction factors that HIV-1 has evolved to evade in order to restore their ability to restrict HV-1. Her central hypothesis is that engineered nucleases will efficiently target HIV-1 dependency and restriction factors to generate resistance to HIV-1 infection for therapeutic application and t further our understanding of these factors in HIV-1 pathogenesis. The main aims are: 1) Knockout HIV-1 dependency factors in human CD4+ T-cell lines to simultaneously determine their virological roles and establish their potential for therapeutic targets for HIV cure applications; 2) Edit human restriction factors to restore HIV-1 resistance and determine effects on viral replication; 3) Create dependency factor knockouts and restriction factor knock-ins in CD34+ HSC cells ex-vivo and characterize them in pre-clinical animal studies in humanized mice. The proposed research has the potential to provide insights into the roles of multiple host factors in HIV-1 biology and to lead to translational applications for curative HIV-1 therapies. Developing HSC-mediated therapeutics in this disease can also advance related applications to other infectious, autoimmune and hematologic diseases.