Optimization and Characterization of "MYC Degraders" for Pediatric Medulloblastoma

项目摘要 髓母细胞瘤（MB）是一种高度恶性的小脑肿瘤，最常见于儿童。 尽管在治疗方面取得了进展-包括手术，放疗和化疗-大约三分之一的人 MB患者仍然死于这种疾病，幸存者由于治疗而遭受严重的副作用。基因组 对患者样本的分析和生物信息学分析已经确定了四个亚组的MB-WNT，Sonic 刺猬（SHH），第3组（MYC驱动）和第4组。这些亚组在突变、基因 表达谱和患者结果，并且患者可以使用相关的遗传和 免疫组织化学标记物。大约四分之一的MB是第3组（G3）肿瘤，其表现出 c-Myc（MYC）癌基因的过度表达或扩增。G3 MB患者更有可能出现 在诊断时有转移性疾病，复发率较高，生存率最差。 因此，迫切需要更有效的G3 MB疗法。 为了鉴定在G3 MB中有效的MYC抑制剂，我们开发了一种表型、靶标不可知的测定方法， 使用分离自G3 MB原位患者来源的异种移植物（PDX）的疾病相关细胞。该测定 旨在鉴定降低内源性MYC水平的小分子-MYC的标志性分子标志物， G3 MB -在4小时内，优先调节直接影响MYC稳定性的靶点，避免间接或关闭 在以后时间点的目标效果。我们应用该试验筛选了100，000种化合物， 小分子支架，其稳健地降低细胞MYC水平。命中率在严格的测试中得到验证 漏斗设计，以避免不希望的作用机制，并探讨了几个支架的初始SAR。 从这些研究中，最有希望的系列也优先考虑在100 nM效价范围内的活性 作为指示良好BBB渗透的化合物性质。我们进一步证实， 在暴露48小时后，一系列的细胞活力降低，并且这种效应与MYC中的效力相关 比色法该系列中最具活性的化合物SBI 1242的初步数据表明，这种细胞凋亡的减少， G3 MB患者细胞的存活率相对于iPSC衍生的神经元是选择性的，这表明可能的治疗窗口。 该提案的目标是进一步优化SBI 1242用于临床前体内测试，并使用最佳类似物 测试我们的假设，即在高度疾病相关的疾病中鉴定的G3 MB签名生物标志物MYC的抑制剂， 上下文可以安全有效地阻止或逆转我们的G3 MB特异性原位PDX小鼠中的肿瘤生长 模型我们预计，这些研究的成功完成将是我们朝着长远目标迈出的重要一步。 目标是为MB和其他MYC驱动的癌症确定新的，安全的和有效的治疗方法。

PROJECT SUMMARY Medulloblastoma (MB) is a highly malignant tumor of the cerebellum that occurs most frequently in children. Despite advances in treatment – including surgery, radiation, and chemotherapy – approximately one-third of MB patients still die from the disease and survivors suffer severe side effects as a result of treatment. Genomic profiling and bioinformatic analyses of patient samples have identified four subgroups of MB – WNT, Sonic hedgehog (SHH), Group 3 (MYC-driven) and Group 4. These subgroups differ in terms of mutations, gene expression profiles and patient outcomes, and patients can be stratified using relevant genetic and immunohistochemical markers. Approximately one-quarter of MBs are Group 3 (G3) tumors, which exhibit overexpression or amplification of the c-Myc (MYC) oncogene. Patients with G3 MB are more likely to present with metastatic disease at time of diagnosis, have a higher incidence of recurrence and the poorest survival rate. Thus, more effective therapies for G3 MB are critically needed. To identify MYC inhibitors that would be effective in G3 MB, we developed a phenotypic, target-agnostic assay using disease-relevant cells isolated from G3 MB orthotopic patient-derived xenografts (PDXs). The assay was designed to identify small molecules that reduce endogenous MYC levels – the signature molecular marker of G3 MB – in 4 hours, to preferentially modulate targets directly affecting MYC stability and avoid indirect or off- target effects at later time points. We applied this assay to screen a 100,000 compound collection and identified small molecule scaffolds that robustly decrease cellular MYC levels. Hits were validated in a rigorous testing funnel designed to avoid undesired mechanisms of action, and initial SAR was explored for several scaffolds. From these studies the most promising series was prioritized with activities in the 100 nM potency range as well as compound properties indicative of good BBB penetration. We further confirmed that the compounds in this series decrease cell viability after 48 hours exposure and that this effect correlates with potency in the MYC assay. Preliminary data for the most active compound of this series, SBI1242, indicate that this decrease in cell viability of G3 MB patient cells is selective over iPSC-derived neurons, suggesting a possible therapeutic window. The goal of this proposal is to further optimize SBI1242 for preclinical in vivo testing and use the best analog to test our hypothesis that inhibitors of the G3 MB signature biomarker MYC identified in a highly disease-relevant context can safely and effectively arrest or reverse tumor growth in our G3 MB-specific orthotopic PDX mouse model. We anticipate that successful completion of these studies will be a significant step towards our long-term goal of identifying novel, safe, and effective treatments for MB and other MYC-driven cancers.