Neural organoid models of the immunological microenvironment of glioblastoma for drug discovery applications

项目摘要/摘要 胶质母细胞瘤(GBM)是成人中最常见的原发脑肿瘤，其发病率极低。 存活率和基本不变的护理标准。虽然面临着许多挑战， 开发更好的GBM治疗方法，主要挑战之一是免疫抑制 常见于GBM肿瘤内的环境。这种免疫抑制的性质导致了一种 不适合建立对GBM细胞的免疫反应的肿瘤，呈现出 免疫疗法无效。为了解决这个问题，合适的模型可以审问 基底膜癌细胞与小胶质细胞及其外周来源的复杂相互作用 巨噬细胞对于靶标识别、新疗法的筛选和 行动研究的模式。肿瘤相关的小胶质细胞和巨噬细胞尤其令人感兴趣。 由于它们在形成基底膜肿瘤的免疫环境中起主要作用。人类 有机体技术非常适合于模拟人类体内复杂的多细胞相互作用 像组织一样的环境。茎制药公司的水凝胶激活的神经有机体可以进行整合 可复制的96孔板中的非神经细胞群体，如小胶质细胞和巨噬细胞 适用于筛选申请的格式。因此，这项提案中的工作将发展和 通过掺入小胶质细胞验证人体外胶质母细胞瘤器官模型， 巨噬细胞和病人来源的神经器官中的GBM细胞。具体目标将1) 鉴定含有GBM的有机体，评估GBM的存活、侵袭和增殖； 并鉴定细胞类型对GBM的特异性转录反应，并与亲本进行比较 肿瘤和公开可用的数据集。2)表现出免疫抑制活性 神经器体内的小胶质细胞和巨噬细胞对浸润性基底膜细胞的反应。 多重细胞因子面板，共刺激和检查点分子表达，以及直接 外周血单个核细胞来源的T细胞的免疫抑制试验将用于 评估小胶质细胞和巨噬细胞的免疫抑制作用。最后，用三个小治疗 已知的调节巨噬细胞激活的分子将在类有机物中进行评估，以 证明有能力调节小胶质细胞和巨噬细胞对GBM细胞的反应。 成功完成这些特定的目标将导致一个强大的体外有机物模型 询问GBM侵袭和随后的小胶质细胞和巨噬细胞的新功能 免疫抑制。这将为制药合作伙伴提供研究以下疗法的能力 之前由于这种免疫抑制环境而失败，并测试了新的治疗方法 接近了。第二阶段研究将扩大可用的患者来源样本的数量 更好地捕捉基底膜肿瘤的多样性和异质性，探索与性别相关的差异，以及 在GBM模型中评估CART和联合治疗的有效性 目标是为这种毁灭性的疾病患者带来更好的治疗选择。

Project Summary/Abstract Glioblastoma (GBM) is the most prevalent primary brain tumor in adults with extremely poor survival rates and largely unchanged standard of care. While there are many challenges to developing better GBM treatments, one of the major challenges is the immune-suppressive environment commonly found within GBM tumors. This immune-suppressive nature results in a tumor that is not suitable for mounting an immune response to GBM cells, rendering emerging immunotherapies ineffective. To address this issue, suitable models that can interrogate the complex interactions between GBM cancer cells and microglia and peripherally derived macrophages would be invaluable for target identification, screening of novel therapeutics and for mode of action studies. Tumor-associated microglia and macrophages are of particular interest due to their primary role in shaping the immunological environment of GBM tumors. Human organoid technology is well-suited for modeling complex, multicellular interactions in a human tissue-like environment. Stem Pharm’s hydrogel-enabled neural organoids allow for incorporation of non-neural populations such as microglia and macrophages in a reproducible, 96-well plate format amenable to screening applications. Therefore, work in this proposal will develop and validate a human in vitro glioblastoma organoid model through incorporation of microglia, macrophages, and patient derived GBM cells in our neural organoids. Specific aims will 1) characterize organoids incorporating GBM, evaluate GBM survival, invasion, and proliferation; and characterize cell-type specific transcriptional responses to GBM and compare them to parent tumors and publicly available data sets. 2) demonstrate immunosuppressive activation of microglia and macrophages within the neural organoid in response to infiltrating GBM cells. Multiplex cytokine panels, co-stimulatory and checkpoint molecule expression, and a direct immunosuppression assay with peripheral blood mononuclear cell-derived T-cells will be used to evaluate microglia and macrophage immunosuppression. Finally, treatment with three small molecules known to modulate macrophage activation will be assessed within the organoids to demonstrate the ability to regulate the microglial and macrophage response to GBM cells. Successful completion of these specific aims will result in a robust in vitro organoid model with novel capabilities to interrogate GBM invasion and subsequent microglia and macrophage immunosuppression. This will provide pharma partners with the ability to study therapies that previously failed due to this immunosuppressive environment, and test new therapeutic approaches. Phase II studies will expand the number of available patient-derived samples to better capture the diversity and heterogeneity of GBM tumors, explore sex-linked differences, and evaluate the effectiveness of CAR-Ts and combination therapies within the GBM model with the goal of bringing better treatment options to patients for this devastating condition.