Contextual Glioblastoma Screening For Efficacious Radiation Sensitizers

描述(申请人提供)：尽管经过几十年的研究，多形性胶质母细胞瘤(GBM)患者的预后仍然非常差。目前对新诊断的胶质母细胞瘤的治疗标准是在可行的范围内进行手术切除，然后辅以放射治疗和替莫唑胺化疗。GBM原位肿瘤细胞被认为是抗辐射的，这被经典地认为是细胞的固有属性。然而，最近 研究指出，胶质母细胞瘤干细胞(GSCs)和肿瘤微环境(TME)这两种非经典机制参与了GBM的辐射抵抗。虽然GSC使用明确的分子机制来导致辐射抗性，但这些机制在体内显著增强，表明TME具有强大的影响。鉴于GBM中的非经典辐射抵抗是由TME调制的，因此放射增敏剂的测试不能在细胞培养中进行。相反，需要新的测试平台，既要提供适当的生物学背景，考虑到TME，又要允许快速药物测试。这种检测由于基底膜中肿瘤间的异质性而进一步复杂化。四种具有不同预后的主要分子GBM亚型的鉴定促使人们担心，如果特定的放射增敏剂对一种亚型有效，而对其他亚型无效，这种异质性可能会混淆药物检测。在这里，我们建议实施一种新的方法来筛选背景的基底膜对放射增敏剂的反应，使用人体基底膜手术标本的器官类型培养来评估分子和细胞对原位辐射的反应。基于我们的临床前研究和当前GBM临床试验的知识，我们建议评估转化生长因子抑制作为增加GBM放射敏感性的一种手段来验证该测试平台。拟议的实验基于这样一种假设，即通过抑制转化生长因子β，通过减少对辐射诱导的双链DNA断裂(DSB)的识别和修复，增强了对辐射的反应。我们预测，抑制转化生长因子信号将阻止观察到的辐射诱导的器官型培养中GSCs患病率的增加，这是通过功能克隆形成试验和肿瘤启动潜能来衡量的。重要的是，我们将评估转化生长因子抑制剂作为放射增敏剂在代表上述所有分子亚型的人GBM标本中的相对有效性。我们假设，这种方法在体外环境下保留了TME和GSC对GBM放射生物学的贡献，将允许通过以下方式进行有效的药物筛选 结合细胞和功能读数的药物疗效，以及通过检查不同分子亚型的基底膜的药物效应。重要的是，我们设想这种方法将成为发现放射增敏剂的范例，可以应用于其他脑肿瘤。

DESCRIPTION (provided by applicant): The prognosis for patients with glioblastoma multiforme (GBM) remains extremely poor despite decades of research. The current standard of care for newly diagnosed glioblastoma is surgical resection to the extent feasible, followed by adjuvant radiotherapy and temozolomide chemotherapy. GBM tumor cells in situ are considered to be radioresistant, which is classically thought to be a cell-intrinsic property. However, recent studies point to the contribution of two non-classical mechanisms that contribute to radiation resistance in GBM: glioblastoma stem cells (GSCs) and the tumor microenvironment (TME). While GSCs employ defined molecular mechanisms that lead to radioresistance, these mechanisms are dramatically potentiated in vivo, suggesting a strong TME influence. Given that non-classical radioresistance in GBM is modulated by the TME, it follows that testing of radiosensitizing agents cannot be performed in cell culture. Instead, novel testing platforms are required that both provide appropriate biological context that takes into account the TME, as well as allow for rapid drug testing. Such testing is further complicated by intertumoral heterogeneity in GBM. The identification of four major molecular GBM subtypes that have different prognoses motivates concerns that such heterogeneity may confound drug testing if specific radiosensitizing agents are efficacious in one subtype but not others. Here we propose to implement a novel approach to screen contextual GBM response to radiosensitizers using organotypic culture of human GBM operative specimens to evaluate the molecular and cellular response to radiation in situ. Based on our preclinical studies and the knowledge of current GBM clinical trials, we propose to evaluate TGF¿ inhibition as a means to increase GBM radiosensitivity to validate this testing platform. The proposed experiments are based on the hypothesis that response to radiation is enhanced by inhibition of TGF¿ in the form of decreased recognition and repair of radiation-induced double-stranded DNA breaks (DSBs). We predict that inhibition of TGF¿ signaling will prevent the observed radiation-induced increase in the prevalence of GSCs in organotypic cultures, as measured by functional clonogenic assays and tumor initiation potential. Importantly, we will evaluate the relative efficacy of TGF¿ inhibitors as radiosensitizers in human GBM specimens representing all molecular subtypes previously described. We posit that this approach, which preserves TME and GSC contributions to GBM radiobiology in an ex vivo setting, will allow for efficient drug screening by incorporating both cellular and functional readouts for drug efficacy, as well as by examining drug effects in distinct molecular subtypes of GBM. Importantly, we envision this approach becoming a paradigm for discovery of radiosensitizing agents that can be applied to other brain tumors.