Design of the First Mdm2 Targeting PROTACs for treatment of p53 Mutant or Deficient Cancers

项目总结/摘要 我们的建议集中在新的Mdm 2靶向蛋白水解靶向嵌合体（PROTAC）的发展 其在p53突变和缺陷癌细胞中有效降解Mdm 2，并预期靶向p53- MDM 2的独立功能。我们的先导化合物以高亲和力结合，降解Mdm 2，杀死癌细胞 其缺乏功能性p53，并且在体内有效。我们还证明了体内安全性和耐受性， 在小鼠药代动力学研究中，合成易处理性、代谢稳定性和适当的体内暴露 体内功效评价。肿瘤抑制因子p53是一种转录因子，在预防肿瘤的发生中起重要作用。 人类癌症在没有细胞应激的情况下，p53蛋白由于其结合而维持在低水平 Mdm 2，一种E3泛素连接酶。然而，一半的人类癌症通过突变或缺失使p53失活。 为了测试缺乏p53的癌细胞的存活是否需要Mdm 2，我们在原发性肝癌细胞中诱导缺失Mdm 2。 鼠p53-无效淋巴瘤和肉瘤细胞。mdm 2缺失导致体外和体内p53-null细胞凋亡 癌症，具有显著降低的肿瘤负荷和增加的存活益处。我们和其他人报告说 Mdm 2通过结合和调节其他蛋白质，如p53家族，具有p53非依赖性功能 Mre 11-Rad 50-Nbs 1 DNA断裂修复复合物中的p73和Nbs 1。p73具有同源的N-末端 在Mdm 2上的相同位点结合的p53的反式激活结构域。另外，与p53不同，p73很少 在人类癌症中失活。由于Mdm 2-p53抑制剂对p53失活的肿瘤无反应，我们 研究了PROTAC方法，并提供了初步数据，以证实其杀死突变或缺陷的p53的能力 癌细胞我们的假设是，通过Mdm 2靶向PROTAC降解Mdm 2将杀死癌症， 通过激活Mdm 2的p53非依赖性活性来突变或缺失p53。我们将用两个例子来验证这个假设。 具体目标。目标1将专注于扩展我们的领先Mdm 2靶向化合物的表征， 在体内异种移植模型中评估这些。目标2将是两种Mdm 2 PROTAC的电极导线优化， p53突变和缺失的癌症。这些目标的目标是具有特征化的、有效的和有力的Mdm 2 PROTAC用于治疗p53失活癌症的临床评价。

Project Summary / Abstract Our proposal focuses on the development of novel Mdm2-targeted proteolysis targeting chimeras (PROTACs) that efficiently degrade Mdm2 in p53 mutant and deficient cancer cells and are expected to target p53- independent functions of Mdm2. Our lead compounds bind with high affinity, degrade Mdm2, kill cancer cells that lack functional p53, and are efficacious in vivo. We have also demonstrated safety and tolerability in vivo, synthetic tractability, metabolic stability, and suitable in vivo exposure in mouse pharmacokinetic studies for in vivo efficacy evaluation. The tumor suppressor p53, a transcription factor, has an essential role in the prevention of human cancer. In the absence of cellular stress, the p53 protein is maintained at low levels due to its binding to Mdm2, an E3 ubiquitin ligase. However, half of all human cancers have inactivated p53 by mutation or deletion. To test if Mdm2 is required for the survival of cancer cells that lack p53 we inducibly deleted Mdm2 in primary murine p53-null lymphoma and sarcoma cells. Mdm2 loss resulted in apoptosis in vitro and in vivo in p53-null cancers, with significantly reduced tumor burden and increased survival benefit. We and others have reported that Mdm2 has p53-independent functions by binding and regulating other proteins, such as the p53 family member, p73, and Nbs1 in the Mre11-Rad50-Nbs1 DNA break repair complex. p73 has a homologous N-terminal transactivation domain to p53 which binds in the same site on Mdm2. Additionally, unlike p53, p73 is rarely inactivated in human cancers. Since Mdm2-p53 inhibitors are not responsive to tumors with inactivated p53, we pursued a PROTAC approach and provide preliminary data to confirm their ability to kill p53 mutated or deficient cancer cells. Our hypothesis is that the degradation of Mdm2 via Mdm2-targeted PROTACs will kill cancers with mutant or deleted p53 by activating the p53-independent activities of Mdm2. We will test this hypothesis with two Specific Aims. Aim 1 will focus on expanding the characterization of our lead Mdm2 targeting compounds and evaluate these in in vivo xenograft models. Aim 2 will be lead optimization of two Mdm2 PROTACs that killed p53 mutant and deleted cancers. The goal of these aims is to have a characterized, effective, and potent Mdm2 PROTAC for clinical evaluation for the treatment of p53-inactivated cancers.