针对PTEN突变或缺失恶性脑胶质瘤的多层次靶向药物筛选与研究

Malignant gliomas are the most common type of brain tumors, and are among the most lethal type of human cancers. Due to the lack of efficacious therapy, even patients receiving maximized treatments survive only about one year as average. Especially, up to 36% of gliomas carry mutation(s) or loss of PTEN, an essential tumor suppress gene, causing a significant reduce of average survival and worse prognosis. However, currently no effective therapeutics are available, nor a systematic way to find drug for this type of gliomas. To find the Achilles' heel of PTEN mutated gliomas, we employed a systematic solution with multi-level screening based on the hypotheses that PTEN deficient cells’ addition on hyper activation of PI3K-mTOR signaling, the synthetic lethality caused by the loss of PTEN, and the importance of brain penetration capability of drug. Upon screening a group of PI3K-mTOR targeted inhibitors, we identified a PI3K inhibitor ZSTK474 and a dual PI3K/mTOR inhibitor NVP-BEZ235 with potent anti-proliferation activity and desirable brain penetration. Furthermore, they demonstrated in vivo efficacy in an orthotropic xenograft glioma model. Next, we plan to use isogenic PTEN positive and deficient glioma cell lines for genome-wide synthetic lethal screening and high-throughput small molecule (targeted agents library) screening. Further in vivo efficacy study and mechanism study will be carried on based on the candidate drugs/drug targets we identified from above screens. Our study will provide preclinical supports for personalized and precision therapy for PTEN mutant malignant glioma patients, and facilitate the understanding and treatment seeking of other types of tumors with PTEN mutation.

恶性脑胶质瘤是人类最致命的癌症之一，患者平均生存期仅为一年。尤其是高达36%的脑胶质瘤患者的抑癌基因PTEN发生了缺失、突变，导致其生存期显著缩短，预后更差。但目前缺乏针对这类肿瘤的治疗药物以及系统性寻找药物的方法。为找到PTEN突变脑胶质瘤的阿基琉斯之踵，我们通过肿瘤细胞对PI3K-mTOR异常激活的依赖性，PTEN缺失带来的潜在合成致死效应以及药物穿透血脑屏障的能力进行多层次系统性的药物筛选。我们前期已经对多种PI3K-mTOR靶向药物进行筛选并发现能穿透血脑屏障并有效抑制脑胶质瘤生长的小分子药物。下一步我们将用PTEN正常与缺失的胶质瘤细胞进行功能基因组合成致死筛选, 靶向药物库筛选结合血脑屏障穿透性筛选，并对候选药物、药靶进行后续的药效与机制的研究。本课题将为精准治疗PTEN缺失恶性脑胶质瘤提供临床前支持，并对研究、治疗PTEN缺失、突变的其他类型肿瘤提供基础。

脑胶质瘤是最为普遍的一种原发性脑肿瘤，其中半数以上为脑胶质母细胞瘤等高等级脑胶质瘤，该类型肿瘤是人类最为致命的癌症之一。30%-60%的脑胶质母细胞瘤患者的抑癌基因PTEN（磷酸酯酶与张力蛋白同源物）发生突变或缺失。这类病人预后差，且目前缺乏有效治疗药物。PI3K/mTOR抑制剂可抑制因PTEN突变导致的PI3K-AKT通路激活，但由于血脑屏障的存在以及肿瘤异质性和脑胶质母细胞瘤的多通路异常激活而疗效有限。本项目从三个方向去研发针对PTEN突变脑胶质瘤的有效治疗方案：根据合成致死原理鉴定与PTEN突变产生致死效应的药物或药物靶点；筛选能与PI3K/mTOR抑制剂产生协同抑制脑胶质瘤效应的靶向药物；筛选能有效穿透血脑屏障入脑的PI3K/mTOR靶向药物。项目完成情况如下：1）通过靶向药物文库筛选鉴定得到抗凋亡蛋白MCL1的选择性抑制剂UMI-77能在体外与体内原位胶质瘤小鼠中有效抑制PTEN缺失脑胶质母细胞瘤生长；阐明了MCL1与PTEN之间的合成致死机制： PTEN缺失导致MCL1上调，而抑制PI3K能降低MCL1表达并上调多种凋亡蛋白，同时UMI-77也能抑制PI3K通路，联合治疗令肿瘤细胞重新回到凋亡命运；发现了UMI-77与目前临床脑胶质瘤一线化疗药物替莫唑胺的协同抗胶质瘤效应。2）建立了一种能与PI3K抑制剂产生抗胶质瘤协同效应的高通量筛选方法；鉴定了MTH1抑制剂TH588 与PI3K抑制剂的抗胶质瘤协同作用并对其协同机制进行了探究。3）筛选多种PI3K/mTOR抑制剂并鉴定得到ZSTK474不是血脑屏障药物转运蛋白ABCB1和ABCG2的底物，因而能更有效的穿越血脑屏障和滞留脑内，同时评估了其单药治疗原位自发脑胶质瘤的疗效。根据申请书设立的计划和目标，项目完成情况良好。在该项目基金的资助下，成果已发表在Mol Cancer Ther（2020）和Cancer Cell Int（2020），并获得国家发明专利授权2项，受邀在多个会议或论坛做学术报告，成功培养博士1名和硕士3名。