Y-Box 1 and normoxic HIF1 in Sonic hedgehog medulloblastoma tumor stem cell radiation resistance

项目摘要 髓母细胞瘤是儿科最常见的中枢神经系统恶性肿瘤。海流 髓母细胞瘤的治疗模式包括手术切除，肿瘤部位放射治疗和 脑脊液放射治疗和化疗。虽然这种疗法的治愈率约为70%， 幸存者受到长期神经副作用的困扰，肿瘤复发是致命的。因此，有必要 开发治疗方法，减少照射需求，减少复发， 并可成功应用于复发患者。髓母细胞瘤可分为4个分子 其中Sonic Hedgehog(SHH)亚组进一步分为带有野生型P53的SHH或 突变型P53，预后最差。SHH亚组的特点是局部复发最频繁。 然而，人们对导致复发的机制的了解有限，最近的研究表明 复发的遗传驱动因素可能与最初的肿瘤驱动因素完全不同，呈现靶向 针对主要肿瘤驱动者的治疗毫无意义。应用小鼠Shh髓母细胞瘤模型和 原代培养小脑颗粒神经前体细胞(CGNPs)，拟为Shh髓母细胞瘤 细胞起源，我们观察到致癌的DNA和RNA结合蛋白YB1促进 辐射后的DNA修复。我们还观察到YB1定位于血管周围窝瘤。 小鼠和人髓母细胞瘤的细胞。这些电池以其抗辐射性能而闻名。我们 也观察到其mRNA翻译被YB1促进的HIF1a同样在PVN中被发现， 我们假设它促进干细胞样表型的维持，而干细胞样表型也提供辐射 抵抗。HIF1a通常在常氧条件下被降解，但我们的初步研究表明 它是以活性氧物种(ROS)和NADPH氧化酶(NOX4)依赖的方式稳定的。这里, 我们建议使用小鼠Shh髓母细胞瘤模型来检验药物靶向的假设 YB1体内可诱导血管周围窝区肿瘤细胞死亡，对小鼠具有体内外治疗作用 带有ROS或NADPH氧化酶抑制剂的髓母细胞瘤将破坏HIF1a的稳定，并同样增加辐射 回应。最后，我们提出了一个翻译目标，其中我们将利用一种新的微流控药物递送 YB1和ROS抑制对新鲜切除的人细胞活力联合作用的平台 髓母细胞瘤原发和复发组织切片及HIF1a、YB1和NOX4定位分析 来自亚特兰大儿童医院新建立的生物库的髓母细胞瘤标本。我们的 研究有可能验证YB1活性和HIF1a稳定性作为新的治疗靶点。 髓母细胞瘤，并开创了筛选患者样本以预测对可能的药物的反应的先例 在复发的情况下给予，或改善原发肿瘤的放射反应。

Project Summary Medulloblastomas are the most common malignant pediatric tumor of the central nervous system. The current treatment paradigm for medulloblastomas includes surgical resection, irradiation of the tumor site and craniospinal irradiation as well as chemotherapy. While this regimen is associated with a ~70% cure rate, survivors are beset with long-term neurological side effects, and tumor recurrence is fatal. Thus, there is a need to develop therapeutic approaches that reduce the requirement for irradiation, reduce the incidence of recurrence, and may be successfully applied to patients that relapse. Medulloblastomas can be divided into 4 molecular subgroups, one of which, the Sonic hedgehog (SHH) subgroup, is further divided into SHH with wild type p53 or mutant p53, which bears the worst prognosis. The SHH subgroup features the most frequent local recurrence. However, there is a limited understanding of mechanisms causing recurrence, and recent studies have shown that genetic drivers of recurrence may be completely different than the original tumor drivers, rendering targeted therapies against the primary tumor drivers pointless. Using mouse models for Shh medulloblastoma and primary cultures of cerebellar granule neuron progenitor (CGNPs) cells, proposed to be Shh medulloblastoma cells-of-origin, we have made the observation that the oncogenic DNA- and RNA-binding protein YB1 promotes DNA repair after radiation. We have also observed that YB1 localizes to the perivascular niche (PVN) tumor cells in mouse and human medulloblastoma. These cells are known for their radiation resistant properties. We have also observed that HIF1a, whose mRNA translation is promoted by YB1, is likewise found in the PVN, where we hypothesize that it promotes maintenance of the stem cell-like phenotype, which also confers radiation resistance. HIF1a is normally degraded under normoxic conditions, however our preliminary studies indicate that it is stabilized in a reactive oxygen species (ROS) and NADPH oxidase (Nox4)-dependent manner. Here, we propose to use mouse Shh medulloblastoma models to test the hypothesis that pharmacologically targeting YB1 in vivo can induce tumor cell death in the perivascular niche, and that in vitro and in vivo treatment of mouse medulloblastoma with inhibitors of ROS or NADPH oxidase will destabilize HIF1a and similarly increase radiation response. Finally, we propose a translational aim wherein we will utilize a novel microfluidic drug delivery platform to test the combined effects of YB1 and ROS inhibition on cell viability in freshly resected human medulloblastoma tumor slices, along with analysis of HIF1a, YB1, and Nox4 localization in primary and recurrent medulloblastoma specimens from a newly established biorepository at Children’s Hospital of Atlanta. Our studies have the potential to validate YB1 activity and HIF1a stabilization as novel therapeutic targets in medulloblastoma, and to set a precedent for screening patient samples to predict response to drugs that could be given in the event of relapse, or to improve radiation response in the primary tumor.