Developing differentiation therapy for the treatment of patients with acute myeloid leukemia

项目摘要 本项目的目标是开发新的，有效的，耐受性良好的分化治疗的治疗 急性髓细胞白血病（AML）患者的治疗。 尽管在理解AML的基因组基础方面取得了显着进展，但患者仍然接受 和四十年前一样的化疗建立阿糖胞苷联合治疗的研究 1973年发表了蒽环类抗生素作为标准化疗（！）。免疫疗法和新靶点 突变（例如IDH，Flt 3-ITD，Dot 1 L）提高了对明确定义的亚组患者更好治疗的可能性。 人口数量。然而，分化治疗具有更全球适用于 有多种潜在突变的白血病HoxA 9在70%的AML中过表达 促使我们建立了一个表型筛选系统，以确定小分子，可以克服 通过HoxA 9建立的分化阻断。 高通量流式细胞术分化筛选，随后进行化合物的分子分析- 耐药细胞系，导致二氢乳清酸脱氢酶（DHODH）的鉴定为 可以触发骨髓分化的特异性抑制剂的靶点。DHODH及其在调节游泳池中的作用 细胞内尿苷的释放是AML的新靶点。在我们的工程细胞系中的这一意外的体外发现是 在人细胞系和AML的离体PDX模型中证实。我们已经证明DHODH 抑制剂在鼠同系白血病模型、人异种移植模型和PDX中具有高度体内活性 AML模型 本申请提出了确定细胞内蛋白质消耗的分子机制。 尿苷导致髓样分化。我们已经概述了一系列基于初步数据的研究 表明在正常造血干细胞和白血病细胞之间存在明确的治疗窗口， 细胞在尿苷饥饿期的生存能力。虽然可以说是雄心勃勃， 实验利用已完成的初步研究，使我们能够查询DHODH的效果 多模型系统中的抑制。特别地，对300+组细胞系的分析已经产生了细胞系 敏感的品系和对DHODH抑制的抗性高1000倍的品系。理解这一点的基础 引人注目的表型在患者选择方面具有基础科学和直接临床意义， 疾病指征。 了解嘧啶生物合成和髓样分化之间的联系将促进我们的研究。 了解正常骨髓生成和白血病背景下分化是如何失调的。的 具有良好药代动力学特性的DHODH有效抑制剂的现成可用性提高了令人兴奋的 快速临床翻译的可能性。 大卫赛克斯是马萨诸塞州总医院大卫斯卡登博士实验室的博士后研究员 (MGH)再生医学中心。大卫斯卡登是一个正式的教授， 指导研究员向独立过渡。再生医学中心是最先进的 研究实验室，包括显微镜和流式细胞术的核心设施，以及自己的动物设施。 大卫与布罗德研究所的研究人员建立了密切的合作关系，丹娜-法伯癌症研究所 研究所，波士顿儿童医院和拜耳医疗保健。这些合作者带来了丰富的 该项目的共同导师以及癌症表观遗传学，代谢物分析，药物 化学和临床前药物开发。 大卫的简历说明了他在学术上的成功。他做了一年的住院总医师， 作为一名教师和领导者的重要经验。他的初步研究得到了 美国国立卫生研究院（R 03）以及美国血液学会，亚历克斯的柠檬水站基金会，和 白血病和淋巴瘤协会。他致力于作为一个物理学家的职业生涯， 门诊和住院病人的临床责任（20%）与他的实验室研究（80%）。大卫是一家公司 相信患者接触有助于提供信息和指导基础研究。为此，他确定了一个 提出了一种全新的临床综合征（TEMPI综合征），并撰写了多篇临床病例报告。 大卫期待着继续指导，因为他学习进行和解释实验， 表观遗传和代谢产物分析，并与化学家在小分子的合理设计， 靶向分化停滞中的关键途径。今后几年将在初步研究的基础上取得丰硕成果 准备过渡到独立调查员职位。

Project Summary The goal of this project is to develop new, effective, and well-tolerated differentiation therapy for the treatment of patients with acute myeloid leukemia (AML). Despite remarkable advances in understanding the genomic underpinnings of AML, patients still receive the same chemotherapy that they did forty years ago. The study that established the combination of cytarabine and an anthracycline as standard chemotherapy was published in 1973(!). Immunotherapy, and novel target mutations (e.g. IDH, Flt3-ITD, Dot1L) raise the possibility of better therapies for well-defined patient sub- populations. However, differentiation therapy holds the potential of being more globally applicable for leukemias with a wide variety of underlying mutations. The finding that HoxA9 is overexpressed in 70% of AML prompted us to establish a phenotypic screening system to identify small molecules that could overcome the differentiation block established by HoxA9. A high-throughput flow-cytometry differentiation screen, followed by the molecular analysis of compound- resistant cell lines, led to the identification of the enzyme dihydroorotate dehydrogenase (DHODH) as the target of specific inhibitors that could trigger myeloid differentiation. DHODH, and its role in regulating the pool of intracellular uridine, is a novel target in AML. This unexpected in vitro finding in our engineered cell line was confirmed in human cell lines and in ex vivo PDX models of AML. We have since demonstrated that DHODH inhibitors are highly active in vivo in murine syngeneic leukemia models, human xenograft models, and PDX models of AML. This application proposes to determine the molecular mechanism through which the depletion of intracellular uridine results in myeloid differentiation. We have outlined a series of studies that build on preliminary data showing the existence of a clear therapeutic window between normal hematopoietic stem cells and leukemic cells in terms of their ability to survive periods of uridine starvation. While arguably ambitious, the proposed experiments take advantage of completed preliminary studies, allowing us to query the effect of DHODH inhibition in multiple model systems. In particular, the profiling of a 300+ panel of cell lines has yielded lines that are sensitive and lines that are 1000x more resistant to DHODH inhibition. Understanding the basis of this striking phenotype has both basic science and immediate clinical implications in terms of patient selection and disease indication. Understanding the link between pyrimidine biosynthesis and myeloid differentiation will advance our understanding of normal myelopoiesis and of how differentiation is dysregulated in the setting of leukemia. The ready availability of potent inhibitors of DHODH with favorable pharmacokinetic properties raises the exciting possibility of rapid clinical translation. David Sykes is a post-doctoral fellow in Dr. David Scadden’s laboratory at the Massachusetts General Hospital (MGH) Center for Regenerative Medicine. David Scadden is a full professor with a proven track-record of mentoring fellows in the transition to independence. The Center for Regenerative Medicine is a state-of-the-art research laboratory including microscopy and flow-cytometry core facilities as well as its own animal facility. David has established close collaborations with investigators at the Broad Institute, the Dana-Farber Cancer Institute, Boston’s Children’s Hospital, and Bayer Healthcare. These collaborators bring a wealth of co-mentorship to the project as well as scientific expertise in cancer epigenetics, metabolite profiling, medicinal chemistry, and pre-clinical drug development. David’s biosketch speaks to his academic successes. He spent a year as medical chief resident, gaining critical experience as a teacher and a leader. His preliminary research has been recognized with grants from the NIH (R03) as well as the American Society of Hematology, Alex’s Lemonade Stand Foundation, and the Leukemia and Lymphoma Society. He is dedicated to a career as a physician-scientist, and balances outpatient and inpatient clinical responsibilities (20%) with his laboratory research (80%). David is a firm believer that patient encounters help to inform and to guide basic research. To this end, he has identified an entirely new clinical syndrome (the TEMPI syndrome), and written a number of clinical case reports. David looks forward to continued mentorship as he learns to carry out and to interpret experiments in epigenetic and metabolite profiling, and to work with chemists in the rational design of small molecules around targeting critical pathways in differentiation arrest. The next years will build productively on preliminary studies in preparation for transition to an independent investigator position.