Functional Inhibition of Deubiquitylase HAUSP to Disrupt Glioblastoma Stem Cells

 描述（由申请人提供）：胶质母细胞瘤（GBM）是一种预后极差的高致死性脑肿瘤。GBM的治疗由于治疗抗性而仍然是姑息性的。GBM显示出显著的细胞异质性，其中肿瘤起始细胞（BTIC）或GBM干细胞（GSC）群体处于分化层级的顶点。我们先前证明GSC促进治疗抗性、肿瘤血管生成和癌症侵袭，表明GSC的治疗靶向可以有效地改善GBM治疗。GSC的干细胞样表型由包括SOX 2的一组核心转录因子维持。SOX 2的蛋白质稳定性受到翻译后修饰的严格调控。我们的初步研究表明，SOX 2在胶质瘤细胞中受到泛素化（Ub）和去泛素化（Dub）的调节。SOX 2被E3泛素连接酶Huwe1泛素化，然后在非干细胞胶质瘤细胞中靶向降解。在GSC中，SOX 2蛋白通过去泛素化酶HAUSP（疱疹病毒相关泛素特异性蛋白酶，也称为USP 7）介导的去泛素化而稳定。相对于神经祖细胞，HAUSP优先在GSC中表达。通过shRNA靶向HAUSP减少了SOX 2蛋白，促进了细胞分化，破坏了GSC维持并有效抑制了GSC肿瘤生长。小分子抑制剂对HAUSP的功能性抑制也抑制了原位GBM异种移植物中的肿瘤生长，并使GSC对辐射或替莫唑胺（TMZ）敏感。基于这些初步数据，我们假设HAUSP介导的去泛素化拮抗Huwe 1介导的泛素化，以稳定GSC维持中的SOX 2，提出了破坏GSC和抑制GBM肿瘤生长的治疗靶向策略。我们提出以下具体目标：1。确定HAUSP抑制对抑制GBM肿瘤生长的治疗影响。2.定义HAUSP介导的GSC维持的分子机制。3.评价HAUSP抑制和当前GBM疗法的协同作用。本提案的目的是在临床前研究中评价HAUSP抑制的治疗潜力。我们将确定靶向HAUSP是否可以作为改善GBM治疗的有效治疗策略。这些研究将为开发新的抗GSC治疗药物以有效提高GBM患者的生存率奠定坚实的基础。

 DESCRIPTION (provided by applicant): Glioblastomas (GBMs) are highly lethal brain tumors with extremely poor prognosis. The treatment of GBMs remains palliative due to therapeutic resistance. GBM displays striking cellular heterogeneity with a population of tumor-initiating cell (BTICs) or GBM stem cells (GSCs) at the apex of differentiation hierarchy. We previously demonstrated that GSCs promote therapeutic resistance, tumor angiogenesis and cancer invasion, indicating that therapeutic targeting of GSCs may effectively improve GBM treatment. The stem cell-like phenotype of GSCs is maintained by a set of core transcription factors including SOX2. The protein stability of SOX2 is tightly regulated by post-translational modifications. Our preliminary study indicated that SOX2 is regulated by both ubiquitylation (Ub) and deubiquitylation (Dub) in glioma cells. SOX2 is ubiquitylated by the E3 ubiquitin ligase, Huwe1, and then targeted for degradation in non-stem glioma cells. In GSCs, SOX2 protein is stabilized through deubiquitylation mediated by the deubiquitylase, HAUSP (Herpesvirus-Associated Ubiquitin-Specific Protease, also known as USP7). HAUSP is preferentially expressed in GSCs relative to neural progenitors. Targeting HAUSP by shRNA reduced SOX2 protein, promoted cell differentiation, disrupted GSC maintenance and potently inhibited GSC tumor growth. Functional inhibition of HAUSP by a small molecule inhibitor also inhibited tumor growth in orthotopic GBM xenografts and sensitized GSCs to radiation or temozolomide (TMZ). Based on these preliminary data, we hypothesize that HAUSP-mediated deubiquitylation antagonizes Huwe1-mediated ubiquitylation to stabilize SOX2 in the maintenance of GSCs, presenting a therapeutic target strategy to disrupts GSCs and suppress GBM tumor growth. We propose the following specific aims: 1. Determine the therapeutic impact of HAUSP inhibition on suppressing GBM tumor growth. 2. Define the molecular mechanisms underlying HAUSP-mediated GSC maintenance. 3. Evaluate synergy of HAUSP inhibition and current GBM therapies. The goal of this proposal is to evaluate the therapeutic potential of HAUSP inhibition in a preclinical study. We will determine whether targeting HAUSP can serve as an effective therapeutic strategy to improve GBM treatment. The proposed studies will lay a solid foundation for the development of novel anti-GSC therapeutics to effectively improve the survival of GBM patients.