Genetically engineered rat gliomas: PDGF-driven tumor initiation and progression in <i>tv-a</i> transgenic rats recreate key features of human brain cancer

Previously rodent preclinical research in gliomas frequently involved implantation of cell lines such as C6 and 9L into the rat brain. More recently, mouse models have taken over, the genetic manipulability of the mouse allowing the creation of genetically accurate models outweighed the disadvantage of its smaller brain size that limited time allowed for tumor progression. Here we illustrate a method that allows glioma formation in the rat using the replication competent avian-like sarcoma (RCAS) virus / tumor virus receptor-A (tv-a) transgenic system of post-natal cell type-specific gene transfer. The RCAS/tv-a model has emerged as a particularly versatile and accurate modeling technology by enabling spatial, temporal, and cell type-specific control of individual gene transformations and providing de novo formed glial tumors with distinct molecular subtypes mirroring human GBM. Nestin promoter-driven tv-a (Ntv-a) transgenic Sprague-Dawley rat founder lines were created and RCAS PDGFA and p53 shRNA constructs were used to initiate intracranial brain tumor formation. Tumor formation and progression were confirmed and visualized by magnetic resonance imaging (MRI) and spectroscopy. The tumors were analyzed using histopathological and immunofluorescent techniques. All experimental animals developed large, heterogeneous brain tumors that closely resembled human GBM. Median survival was 92 days from tumor initiation and 62 days from the first point of tumor visualization on MRI. Each tumor-bearing animal showed time dependent evidence of malignant progression to high-grade glioma by MRI and neurological examination. Post-mortem tumor analysis demonstrated the presence of several key characteristics of human GBM, including high levels of tumor cell proliferation, pseudopalisading necrosis, microvascular proliferation, invasion of tumor cells into surrounding tissues, peri-tumoral reactive astrogliosis, lymphocyte infiltration, presence of numerous tumor-associated microglia- and bone marrow-derived macrophages, and the formation of stem-like cell niches within the tumor. This transgenic rat model may enable detailed interspecies comparisons of fundamental cancer pathways and clinically relevant experimental imaging procedures and interventions that are limited by the smaller size of the mouse brain.此前，胶质瘤的啮齿类动物临床前研究通常将C6、9L等细胞系植入大鼠脑部。近年来，小鼠模型逐渐成为主流：小鼠具备更高的遗传可操作性，可构建遗传准确性更优的模型，这一优势弥补了其脑部尺寸更小、限制肿瘤增殖观察时长的缺陷。本研究展示了一种可在大鼠体内诱导胶质瘤形成的方法，该方法基于复制型禽肉瘤病毒（RCAS）/肿瘤病毒受体-A（tv-a）转基因系统，可实现出生后细胞类型特异性基因转移。RCAS/tv-a模型可实现对单个基因转化的空间、时序及细胞类型特异性调控，并能从头生成与人类胶质母细胞瘤（GBM）分子亚型高度匹配的神经胶质肿瘤，因此已成为通用性强且准确性高的建模技术。本研究构建了巢蛋白启动子驱动的tv-a（Ntv-a）转基因斯普拉格-道利大鼠创始品系，并使用RCAS PDGFA与p53短发夹RNA（shRNA）构建载体启动颅内脑瘤形成。通过磁共振成像（MRI）与磁共振波谱分析，研究人员对肿瘤的形成与进展进行了验证与可视化观测。采用组织病理学与免疫荧光技术对肿瘤进行分析后发现，所有实验动物均形成了与人类胶质母细胞瘤（GBM）高度相似的大型异质性脑瘤。从肿瘤启动至中位生存期为92天，从首次通过MRI观测到肿瘤至中位生存期为62天。每只荷瘤动物均通过MRI与神经学检查，呈现出恶性进展为高级别胶质瘤的时间依赖性特征。死后肿瘤分析显示，该模型具备人类胶质母细胞瘤（GBM）的多项关键特征：肿瘤细胞增殖水平高、假栅栏状坏死、微血管增生、肿瘤细胞侵袭周围组织、瘤周反应性星形胶质增生、淋巴细胞浸润、大量肿瘤相关小胶质细胞与骨髓源性巨噬细胞存在，以及肿瘤内干细胞样细胞龛的形成。该转基因大鼠模型可实现对核心癌症通路的跨物种比较研究，以及受限于小鼠脑部尺寸的临床相关实验成像操作与干预手段的相关研究。