Targeting AVIL in Glioblastoma

多形性胶质母细胞瘤（GBM）是脑内最常见和最具侵袭性的恶性原发性脑肿瘤， 人类尽管多模式积极治疗，GBM是致命的-生存超过3年被认为是 长期的显然需要更多的治疗靶点和治疗选择。在初步研究中，我们 在GBM中发现了一种新的癌基因AVIL。AVIL基因在所有胶质母细胞瘤细胞系中过表达， 包括胶质母细胞瘤干细胞和临床病例，但在非癌星形胶质细胞、神经胶质细胞和神经胶质细胞中几乎检测不到。 干细胞和脑组织。沉默AVIL导致GBM细胞培养物中的完全根除，但 对星形胶质细胞控制细胞几乎没有影响。在动物模型中，沉默AVIL在体内显著抑制 小鼠异种移植物。相反，在细胞培养系统中过表达AVIL促进肿瘤发生， 成纤维细胞逃避接触抑制，并转化永生化星形胶质细胞。在患者队列中， 胶质瘤中AVIL的表达水平与预后相关。最重要的是，GBM细胞 对AVIL的小分子抑制剂敏感。这些证据支持AVIL是阿基里斯的前提 GBM的脚跟，瞄准可能是GBM的有效方法。在本申请中，我们提出 AVIL作为抗GBM新靶点的系统研究。目标1：确定靶向疗效 体外AVIL并测试AVIL表达作为敏感性的生物标志物。我们已经展示了AVIL在以下方面的关键作用： 一些GBM和GSC细胞系。在这里，我们将研究其在其他胶质母细胞瘤细胞系，GSC， GBM患者来源的异种移植物培养物的短期外植体和与 CDK4/MDM 2。我们将测试tet诱导的shRNAs靶向AVIL和小分子化合物的功效， 我们通过小分子筛选鉴定出来的。此外，我们将确定AVIL表达是否作为 对AVIL抑制敏感性的生物标志物。目的2：确定靶向AVIL在动物GBM中的功效 模型我们将在异种移植模型中使用shRNA和小分子抑制剂来测试其功效和 体内靶向AVIL的安全性。此外，我们将使用Avil敲除小鼠模型，通过将它们与 在神经胶质瘤小鼠模型中，以测试Avil表达是否是小鼠中神经胶质瘤肿瘤发生所必需的。目标3： 研究AVIL抑制的下游靶点和机制，并确定潜在的药效学 生物标志物。我们有初步的数据表明AVIL调节LIN 28 B，FOXM 1可能介导这种作用 在U87细胞中我们将研究LIN 28 B和FOXM 1是否是AVIL的功能关键靶点， 它们是否是评价AVIL靶向治疗疗效的潜在药效学生物标志物 疗法目的4：探讨AVIL靶向的耐药机制。目标是预测 在临床环境中出现之前，对基于AVIL的治疗提出挑战。拟议的研究将有一个 对理解和治疗GBM有重要影响。这些发现将为瞄准AVIL铺平道路， 一种新的致癌基因，并导致对致命疾病的新治疗方法的发展。

Glioblastoma multiforme (GBM), is the most common and most aggressive malignant primary brain tumor in humans. Despite multimodal aggressive therapy, GBM is fatal—with survival over 3 years being considered long-term. More therapeutic targets and treatment options are clearly needed. In our preliminary study, we identified a novel oncogene, AVIL in GBMs. AVIL gene is overexpressed in all the glioblastoma cell lines, including glioblastoma stem cells, and clinical cases, but is hardly detectable in non-cancer astrocytes, neural stem cells, and brain tissues. Silencing AVIL resulted in complete eradication in GBM cell cultures, but had little effect on the astrocyte control cells. In animal models, silencing AVIL dramatically inhibited in vivo xenografts in mice. Conversely, overexpressing AVIL in cell culture systems promoted tumorigenesis, enabled fibroblasts to escape contact inhibition, and transformed immortalized astrocytes. In patient cohorts, higher expression levels of AVIL in gliomas correlated with worse prognosis. Most importantly, GBM cells are more susceptible to small molecule inhibitors of AVIL. These evidences support the premise that AVIL is an Achilles heel of GBMs, targeting which may be an effective approach for GBMs. In this application, we propose the systematic investigation of AVIL as a novel target against GBMs. Aim 1: Determine the efficacy of targeting AVIL in vitro and test AVIL expression as a biomarker of sensitivity. We have shown the critical role of AVIL in some GBM and GSC cell lines. Here, we will investigate its role in additional glioblastoma lines, GSCs, and short-term explants of GBM patient derived xenograft cultures and potential synergistic effect with CDK4/MDM2. We will test the efficacy of tet-inducible shRNAs targeting AVIL, and small molecule compounds we identified through small molecule screen. In addition, we will determine whether AVIL expression serves as a biomarker for sensitivity to AVIL inhibition. Aim 2: Determine the efficacy of targeting AVIL in animal GBM models. We will use shRNA and small-molecule inhibitors in xenograft models to test both the efficacy and safety of targeting AVIL in vivo. In addition, we will use Avil knockout mouse model by crossing them with glioma mouse models to test whether Avil expression is necessary for glioma tumorigenesis in mouse. Aim 3: Investigate the downstream targets and mechanisms of AVIL inhibition and identify potential pharmacodynamic biomarkers. We have preliminary data suggesting AVIL regulates LIN28B, and FOXM1 may mediate the effect in U87 cells. We will investigate whether LIN28B and FOXM1 are functionally critical targets of AVIL, and whether they are potential pharmacodynamic biomarkers to evaluate treatment efficacy of AVIL-targeting therapy. Aim 4: Investigate potential resistance mechanisms of targeting AVIL. The goal is to anticipate challenges to AVIL-based treatment before they arise in the clinical setting. The proposed study will have a significant impact on the understanding and treatment of GBMs. The findings will pave ways to target AVIL as a novel oncogene, and lead to the development of novel therapeutic approaches for the deadly disease.