Elucidating the critical role of Wee1 in GIST

项目摘要 胃肠道间质瘤(GIST)治疗的管理已经发生了革命性的变化 KIT和PDGFRA的激活突变及酪氨酸激酶抑制剂(TKIs)的临床应用 疾病。然而，对所有已批准的TKI的耐药性是转移性肿瘤的一个危及生命的临床障碍。 GIST患者，他们的多克隆肿瘤由KIT/PDGFRA的各种耐药突变组成。 为难治性GIST制定有效的治疗策略需要确定新的靶点。vbl.使用 Kinome图谱和功能丧失分析，我们发现GIST对Wee1活性有很好的依赖性。 Wee1长期以来一直被认为是一个关键的G2/M细胞周期检查点把关人，控制基因组完整性和 控制扩散。然而，最近的研究表明，Wee1直接或间接地与其他 细胞功能，包括修复停滞的复制叉子和染色质重塑。转录组 对TKI敏感的GIST细胞株的分析强调致癌试剂盒/PDGFRA在DNA损伤反应中的作用 (DDR)和复制应激。我们的体外和体内研究表明阿达伐司替布(Wee1)具有显著的疗效 在TKI敏感试剂盒和PDGFRA突变型GIST中与avapritinib(KIT/PDGFRA抑制剂)联合使用 在患者来源的异种移植(PDX)模型中。此外，我们在将军澳支线的初步数据-耐火材料 环境展示了Wee1抑制和拓扑异构酶II抑制剂之间的治疗协同作用， 在一组对TKI耐药的GIST细胞系中，低纳摩尔浓度的阿霉素，提示Wee1 抑制使难治性GIST对阿霉素增敏，提供了KIT和PDGFRA以外的可行靶点。我们 假设致癌试剂盒/PDGFRA激活DDR蛋白以抑制复制应激和DNA损伤 并在GIST中保持基因组稳定，从而对DNA损伤剂产生抗药性。我们的初步调查结果 提供了一个强有力的前提，即Wee1是这一过程的关键调节因子，它的抑制作用在 胃肠道间质瘤的治疗。Wee1和Kit/PDGFRA协同作用机制的研究 GIST对DNA损伤和复制应激的反应是理解这一关键的细胞保护的保证 GIST的机制，这可能代表了一种新的治疗脆弱性。归根结底，这项提议旨在 测试Wee1抑制将是对GIST患者现有治疗的有价值的补充的假设 对TKI敏感或抵抗的疾病。我们相信，这项提案中概述的研究将提供新的见解。 研究Wee1在维持基因组完整性以及支持GIST临床试验的临床前数据方面的作用。

Project Summary Management of gastrointestinal stromal tumor (GIST) treatment has been revolutionized by the identification of activating mutations in KIT and PDGFRA and clinical application of tyrosine kinase inhibitors (TKIs) in advanced disease. However, resistance to all approved lines of TKIs is a life-threatening clinical obstacle in metastatic GIST patients, who have polyclonal tumors comprised of a variety of resistance mutations in KIT/PDGFRA. Development of effective treatment strategies for refractory GIST requires identification of novel targets. Using kinome profiling and loss-of function assays, we found that GIST are exquisitely dependent on Wee1 activity. Wee1 has long been known as a critical G2/M cell cycle checkpoint gatekeeper controlling genomic integrity and regulating proliferation. However, recent studies have implicated Wee1, both directly and indirectly, in other cellular functions, including restoration of stalled replication forks and chromatin remodeling. Transcriptome profiling of TKI-sensitive GIST cell lines highlighted a role for oncogenic KIT/PDGFRA in DNA damage response (DDR) and replication stress. Our in vitro and in vivo studies revealed significant efficacy of adavosertib (Wee1 inhibitor) in combination with avapritinib (KIT/PDGFRA inhibitor) in TKI-sensitive KIT and PDGFRA-mutant GIST lines and in a patient-derived xenograft (PDX) model. Furthermore, our preliminary data in the TKI-refractory setting demonstrated therapeutic synergies between Wee1 inhibition and the topoisomerase II inhibitor, doxorubicin, at low nanomolar concentrations in a panel of TKI-resistant GIST cell lines, suggesting Wee1 inhibition sensitizes refractory GIST to doxorubicin, providing viable targets beyond KIT and PDGFRA. We hypothesize that oncogenic KIT/PDGFRA activates DDR proteins to inhibit replicative stress and DNA damage and maintain genomic stability in GIST leading to resistance to DNA damaging agents. Our preliminary findings provide a strong premise that Wee1 is a critical mediator of this process and that its inhibition has potential in the treatment of GIST tumors. Studies interrogating mechanisms of Wee1 and KIT/PDGFRA cooperativity in response to DNA damage and replication stress in GIST are warranted to understand this critical cell-protective mechanism in GIST, which may represent a novel therapeutic vulnerability. Ultimately, this proposal seeks to test the hypothesis that Wee1 inhibition will be a valuable addition to current treatments for GIST patients with TKI-sensitive or -resistant disease. We believe that the studies outlined in this proposal will provide novel insights into the role of Wee1 in maintaining genomic integrity as well as preclinical data to support clinical trials in GIST.