Mechanistic and pharmacologic studies of selective mithramycin analogues targeting EWS-FLI1 in Ewing sarcoma

尤文肉瘤肿瘤家族(ESFT)是一类具有韧性的破坏性骨和软组织肿瘤家族 主要影响儿童和年轻人。目前高度细胞毒性的五种药物联合治疗提供了 总体存活率只有30%。仅存在于肿瘤细胞中的异常转录因子EWS-FLI1是致癌的 EWS的驱动程序。然而，转录因子被认为是“不能下药的”，直到最近的一次NCI筛查 发现米特拉霉素(MTM)作为一种有效的EWS-FLI1拮抗剂。MTM已被证明毒性太大，使用的是一种狭窄的 治疗窗口和较差的药代动力学(PK)特性。在这里，我们提出机械学和药理学 新型MTM类似物(MTM-SA)的研究具有显著降低毒性、提高靶向性和 极大地改善了PK属性。与其他地方报道的其他类似物形成对比的是，这些类似物仍在遭受贫穷 PK性质，MTM-SA类似物表现出优越的动力学和降低的毒性。这个项目的目标是 通过结构、生化和药理学研究获得对MTM作用模式的分子洞察 以产生一种高效和选择性的抗ESFT治疗。以协助Aim1的合成工作并确定 具有临床潜力的类似物，我们将在目标2中进行分子结构-功能水平研究，以确定 转录因子EWS-FLI1如何与DNA微卫星重复序列和转录因子Runx2(各自 肿瘤发生所必需的相互作用)，以及MTM-SA如何破坏这些致癌功能。在AIM 3将评估体外细胞毒性和靶向选择性，以确定将在 将评估人源化肝脏小鼠的毒性、PK和代谢以及疗效的药理学研究 在尤文肉瘤的异种移植和PDX模型中。该项目将由一个具有既定职责的团队执行 在基于片段的药物设计和天然药物的半合成路线方面拥有丰富经验的合作人员 产品、X射线结晶学、生物物理和分子生物学研究以及药理学评价。 我们预计，这些结构-功能研究将确定一种可能进入临床试验的主要候选者 ESFT的治疗。

Ewing sarcoma family of tumors (ESFT) is a family of resilient devastating cancers of bone and soft tissue affecting primarily children and young adults. Current highly cytotoxic combination therapy of five drugs provides only 30% overall survival. The aberrant transcription factor EWS-FLI1 present only in tumor cells is the oncogenic driver of EWS. However, transcription factors were believed to be “undruggable”, until a recent NCI screening found mithramycin (MTM) to act as a potent EWS-FLI1 antagonist. MTM has proven to be too toxic with a narrow therapeutic window and poor pharmacokinetic (PK) properties. Here we propose mechanistic and pharmacology studies of novel MTM analogues (MTM-SA) with significantly reduced toxicity, increased target specificity and greatly improved PK properties. In contrast to other analogues reported elsewhere, which still suffer from poor PK properties, the MTM-SA analogs display superior kinetics and reduced toxicity. The goal of this project is to gain molecular insights into the mode of action of MTM via structural, biochemical and pharmacological studies to generate a highly efficacious and selective anti-ESFT treatment. To aid synthetic efforts in Aim1 and identify analogues with clinical potential, we will perform molecular structure-function level studies in Aim 2 to determine how transcription factor EWS-FLI1 interacts with DNA microsatellite repeats and transcription factor Runx2 (each a necessary interaction for oncogenesis), and how these oncogenic functions are disrupted by MTM-SA. In Aim 3 will assess the in vitro cytotoxicity and target selectivity to identify analogues that will be evaluated in pharmacologic studies that will assess toxicity in humanized liver mice, PK and metabolism, as well as efficacy in xenograft and PDx models of Ewing Sarcoma. The project will be carried out by a team with an established collaboration who have extensive experience in fragment-based drug design and semi-synthetic routes of natural products, X-ray crystallographic, biophysical and molecular biology studies, and pharmacological evaluations. We expect that these structure-function studies will identify a lead-candidate that could enter a clinical trial for the treatment of ESFT.