In vivo imaging of encapsulated stem cells in mouse models of tumor resection

描述(由申请人提供)：恶性多形性胶质母细胞瘤(GBM)的标准治疗方法包括最大限度地手术切除肿瘤，然后进行放射治疗和辅助化疗。然而，GBM的复发率和相关患者的死亡率几乎为100%。尽管肿瘤切除在临床GBM治疗中起着关键作用，但大多数临床前研究都集中在治疗颅内完整的实性GBM肿瘤上，而不是模仿手术切除的临床情景。因此，在总结临床疾病特征的小鼠模型中实施肿瘤切除对于开发临床可移植的GBM治疗方法至关重要。在拟议的研究中，我们将首先根据3种不同的表型(侵袭性、半侵袭性和结节型)，使用患者来源的CD133+GBM来创建和表征不同的GBM切除模型。虽然原发肿瘤切除术已显示出临床上的好处，但系统地输送化疗药物或直接注射病毒和 将卡莫司汀(BCNU)晶片放置在肿瘤切除腔中提供的额外好处非常有限。基于我们最近的发现，包裹间充质干细胞(MSC)对于防止干细胞移植后在肿瘤切除腔中的快速“洗出”是必要的，我们将把经工程设计的人MSC包裹到合成的细胞外基质(SECM)中，并评估它们在不同表型的GBM小鼠切除模型中的保留、存活和肿瘤归巢。我们的初步研究表明，一种新型的抗肿瘤药物，包括分泌版本的表皮生长因子受体靶向纳米体融合肿瘤坏死因子凋亡诱导配体(eNB-TRAIL)，在TRAIL耐药和敏感的GBM中诱导GBM细胞杀伤，包裹的MSC-eNB-TRAIL将在不同的GBM切除模型中进行测试。为了确保我们的方法的安全性，我们最终将改造人类MSC-eNB-TRAIL来表达HSV-胸苷激酶(TK)，这是一种可激活的细胞自杀基因，将使我们能够选择性地在GBM治疗后根除MSC。将基因工程荧光和生物发光成像(BLI)标记整合到MSC和GBM中，将使我们能够通过体内BLI、活体显微镜(IVM)、磁共振成像(MRI)和正电子发射断层扫描(PET)来跟踪GBM细胞的侵袭、MSC的命运、治疗蛋白的药代动力学和它们的有效性，从而对所提出的方法进行微调。一旦得到验证，我们将启动一项临床研究，在脑肿瘤手术时，将切除主要的肿瘤肿块，并引入包裹SECMs的MSC，以靶向大脑中广泛残留的肿瘤细胞和微侵袭肿瘤沉积。这将对挽救许多脑癌患者的生命产生重大影响。

DESCRIPTION (provided by applicant): The standard treatment for malignant glioblastoma multiforme (GBM) includes maximal surgical tumor resection followed by radiation therapy and adjuvant chemotherapy. However, recurrence rates of GBM and the associated patient mortality are nearly 100%. Despite the key role of tumor resection in clinical GBM therapy, most of the pre-clinical studies focus on treating solid intact intracranial GBM tumors without mimicking the clinical scenario of surgical resection. Therefore, implementation of tumor resection in mouse models that recapitulate the clinical disease features are critical in developing clinically translatable therapies for GBM. In the proposed studies, we will first create and characterize different GBM resection models using patient derived CD133+ GBMs based on 3 different phenotypes (invasive, semi-invasive and nodular). While resection of primary tumor has shown clinical benefit, systemically delivered chemotherapeutic agents or direct injection of viruses and placement of carmustine (BCNU) wafers in tumor resection cavities has provided very limited additional benefit. Based on our recent findings that encapsulation of mesenchymal stem cells (MSC) is necessary to prevent rapid "wash- out" of stem cells post-transplantation in the tumor resection cavity, we will encapsulate human MSC engineered to express in vitro and in vivo imaging markers into synthetic extracellular matrices (sECMs) and evaluate them for their retention, survival and tumor homing in mouse resection models of GBM with different phenotypes. Based on our preliminary studies which indicate that a novel anti-tumor agent consisting of a secretable version of epidermal growth factor receptor targeted nanobody fused to tumor necrosis factor apoptosis inducing ligand (Enb-TRAIL), induces GBM cell killing in both TRAIL resistant and sensitive GBMs, encapsulated MSC-Enb- TRAIL will be tested in different GBM models of resection. To ensure the safety of our approach, we will ultimately engineer human MSC-Enb-TRAIL to express HSV-thymidine kinase (TK), an activatable cellular suicide gene that will allow us to selectively eradicate MSC post-GBM treatment. The incorporation of genetically engineered fluorescent and bioluminescent imaging (BLI) markers into MSC and GBMs will allow us to follow GBM cell invasion, fate of MSC and pharmacokinetics of therapeutic proteins and their efficacy by in vivo BLI, intravital microscopy (IVM), magnetic resonance imaging (MRI) and positron emission tomography (PET) and thus to fine tune the proposed approaches. Once validated, we will initiate a clinical study in which at the time of brain tumor surgery, the main tumor mass will be removed and sECMs encapsulated MSC will be introduced to target a broad spectrum of remaining tumor cells and micro-invasive tumor deposits in the brain. This will have a major impact in saving the lives of many brain cancer patients.