Probing INTS11 as a novel target in neuroblastoma

项目总结 在美国，每年有近千名儿童被诊断出患有神经母细胞瘤(NBL)， 到目前为止，这种癌症是婴儿中最常见的类型。可悲的是，高风险的NBL，其中约70% 所有NBL病例的五年存活率均为50%。高危NBL的特点是有 MYCN基因的扩增和第一染色体1p36(D1p36)区域的缺失 包含多个肿瘤抑制物。但这些特征很难作为药理作用加以利用 MYCN的抑制已被证明是难以捉摸的，并在 基因组损失几乎是不可能的。与MYCN在增加全球市场份额方面的既定角色一致 转录，具有扩增的MYCN表达的高危NBL独特地依赖于细胞 转录和RNA处理设备。在这些机制中，对MYCN功能至关重要的是 整合子复合体，负责共转录剪接所必需的USnRNA生物合成 促进RNA聚合酶II(RNAPII)的周转和可塑性。积分器是一个14个成员的复合体 与RNAPII相关，是调节转录输出的基础。我的实验室已经 对积分器的分子机制进行了十多年的研究(19-34)，我们发现 整合子亚单位11(INTS11)是该复合体的关键酶组分。INTS11利用其 切割剪接体USnRNA 3‘端的RNA内切酶活性是Pre-Pre-DNA所必需的 信使核糖核酸剪接此外，我们最近发现，INTS11也是切割新生蛋白质所必需的- 编码RNA以促进RNAPII循环。这两种功能都是支持高转录的关键 MYCN生产过剩时观察到的情况。除了MYCN扩增的肿瘤所需的外，大多数 将整合子提名为NBL潜在的药物靶点的吸引人的特征是INTS11基因是 位于1p36内，在NBLS中经常被删除。为了支持这一观察结果，来自DEPMAP的数据 NBL细胞系对INTS11基因的部分缺失高度敏感，表明 对这一基因的强烈依赖。总之，这些观察结果支持了INTS11的假设 代表了NBL中一个新的和未探索的目标(图1)，我们下面的具体目标旨在 直接测试这一点。特异性目的1.测定MYCN扩增的NBL细胞和D1p36 NBL细胞的敏感性 到INTS11下调监管。特定目的2.测试INTS11的候选抑制剂对小鼠的差别毒性 高危NBL细胞系。

PROJECT SUMMARY Nearly a thousand children are diagnosed each year with Neuroblastoma (NBL) in the United States, and this cancer is, by far, the most prevalent type in infants. Sadly, high-risk NBL, which comprises ~70% of all NBL cases, has a five-year survival rate of <50%. High-risk NBL is characterized by having amplification of the MYCN gene and deletion of the 1p36 (D1p36) region of chromosome one, which houses multiple tumor suppressors. But these signatures are difficult to exploit as pharmacologic inhibition of MYCN has proven elusive, and re-activating tumor suppressor gene expression after genomic loss is virtually impossible. Consistent with MYCN’s established role in increasing global transcription, high-risk NBLs with amplified MYCN expression are uniquely dependent on the cellular transcription and RNA processing apparatus. Among this machinery vital to MYCN function is the Integrator Complex, which is responsible for UsnRNA biosynthesis essential for co-transcriptional splicing and promoting RNA polymerase II (RNAPII) turnover and plasticity. Integrator is a 14-membered complex associated with RNAPII and is fundamental in regulating transcriptional output. My laboratory has been investigating the molecular mechanism of Integrator for over ten years(19-34), and we have found that Integrator subunit 11 (INTS11) is the critical enzymatic component of the complex. INTS11 utilizes its RNA endonuclease activity to cleave the 3'-end of spliceosomal UsnRNA and is thus required for pre- mRNA splicing. Moreover, we recently found that INTS11 is also necessary to cleave nascent protein- coding RNA to promote RNAPII recycling. Both functions are critical to support high transcription observed when MYCN is overproduced. Beyond being required in MYCN-amplified tumors, the most attractive feature nominating Integrator as a potential druggable target in NBL is that the INTS11 gene is located within 1p36 and is frequently deleted in NBLs. In support of this observation, data from DEPMAP reveals that NBL cell lines are highly sensitive to partial depletion of INTS11, indicating a unique and robust dependency on this gene. Altogether, these observations support a hypothesis that INTS11 represents a novel and unexplored target in NBL (Fig. 1), and our Specific Aims below are designed to test this directly. Specific Aim 1. Determine the sensitivity of MYCN-amplified and D1p36 NBL cell lines to INTS11 downregulation. Specific Aim 2. Test candidate inhibitors of INTS11 for differential toxicity in high-risk NBL cell lines.