Nanoparticle Modified Human Fat Derived Mesenchymal Stem Cells for Brain Cancer (Change of Organization Application)

纳米颗粒修饰的人类脂肪源性间充质干细胞治疗脑癌（组织申请变更）

• 批准号: 9551197
• 负责人: Jordan Green
• 金额: $ 36.46万
• 依托单位: MAYO CLINIC JACKSONVILLE
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-19 至 2020-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9551197
• 关键词: Adipose tissue; Adult; Affect; Age; American; Animals; Antineoplastic Agents; Artificial nanoparticles; Benchmarking; Blood - brain barrier anatomy; Bone Marrow; Brain; Brain Neoplasms; Bypass; Cell Culture Techniques; Cell Survival; Cells; Characteristics; Clinical Trials; Destinations; Devices; Disease; Drug Carriers; Effectiveness; Engineering; Family; Fatty acid glycerol esters; Formulation; Freeze Drying; Future; Gene Delivery; Genes; Genetic Engineering; Glioblastoma; Glioma; Goals; Grant; Home environment; Human; Human Engineering; In Vitro; Incidence; Insertional Mutagenesis; Invaded; Investigation; Lead; Malignant Neoplasms; Malignant neoplasm of brain; Malignant neoplasm of lung; Medical; Mesenchymal Stem Cells; Methods; Microfluidics; Modeling; Modification; Morbidity - disease rate; Mus; Muscle; Oncogenic; Operative Surgical Procedures; Organizational Change; Patients; Phenotype; Primary Brain Neoplasms; Property; Protein Engineering; Proteins; Protocols documentation; Radiation; Radiation therapy; Research; Rodent; Safety; Stem cells; Surface; Survival Rate; Techniques; Technology; Testing; Therapeutic; Time; Tissues; Transfection; Translating; Tropism; Tumor Burden; Tumor Initiators; Virus; Xenograft Model; biodegradable polymer; bone morphogenic protein; brain parenchyma; cancer cell; cell motility; cell type; chemoradiation; chemotherapy; clinical application; clinically relevant; effective therapy; immunogenicity; in vivo; ineffective therapies; interest; malignant breast neoplasm; minimally invasive; mortality; mouse model; nanobiotechnology; nanoparticle; neoplastic cell; new technology; non-viral gene delivery; novel therapeutics; personalized medicine; public health relevance; statistics; temozolomide; therapeutic protein; therapy resistant; tumor; viral gene delivery

 DESCRIPTION (provided by applicant): Glioblastoma (GBM) is the most common primary brain tumor in adults, and accounts for 20% of all primary brain tumors. GBM has a median survival rate of only 14.6 months despite current best treatment practices including surgery and chemoradiation. A significant reason for this morbidity and mortality is the ability of GBM to invade normal brain parenchyma, making localized treatment ineffective. There is increasing evidence of a small subset of cells, brain tumor initiating cells (BTICs) that are responsible for the disease's treatment resistance. In order for treatment to be effective, these invading cells need to be targeted. One promising approach involves the use of mesenchymal stem cells (MSCs), which have been found to migrate preferentially to and home in on cancer cells. Moreover, MSCs can be engineered to synthesize and release anti-tumor proteins, like bone morphogenic protein 4 (BMP4), which affects BTICs. MSCs can be obtained from bone marrow (BM- MSC) and adipose tissue (AMSCs). BM-MSCs are difficult to obtain, have limited ex vivo proliferation capacity, and decrease in effectiveness with donor age. Unlike BM-MSCs, AMSCs are more abundant in supply, easier to obtain from fat tissue, express higher levels of surface markers implicated in cell migration, and have been shown to resist oncogenic transformation. AMSCs may therefore be a better option. The viral gene delivery method, though commonly used to modify AMSCs, is associated with insertional mutagenesis and immunogenicity, and, therefore, has potentially limited translational ability for use in human patients. Biodegradable, polymeric nanoparticles enable effective non-viral gene delivery to multiple cell types, including human AMSCs (hAMSCs), while avoiding the problems typical of viruses. In this grant, we propose a novel technology to combine Freshly-extracted Adipose Tissue (F.A.T.) and nanoparticles to non-virally engineer the primary hAMSCs contained within F.A.T without prior culture to secrete anti-cancer proteins while maintaining the cells' ability to migrate toward tumo cells. Our overall hypothesis is that nanoparticle-modified hAMSCs obtained from F.A.T. retain their tumor suppressive characteristics in a clinically relevant in vivo human GBM model. To test this hypothesis, we will pursue the following specific aims: (1) To effectively deliver exogenous genes of interest to Freshly-extracted Adipose Tissue (F.A.T.) from patients via lyophilized biodegradable nanoparticles. (2) To determine if nanoparticle-modified BMP4-secreting hAMSCs retain an anti-glioma effect in vitro. (3) To determine the safety and efficacy of nanoparticle-modified BMP4-secreting hAMSC treatment in combination with targeted radiation therapy on human GBM in an in vivo murine model. Aim 1 involves investigation and optimization of a unique technology of combining nanoparticles with F.A.T. from our patients. For aims 2 and 3, using nanoparticles already tested in commercial hAMSCs, we will now investigate the modification of primary hAMSCs that have been isolated and cultured prior to adding the nanoparticles. The techniques to be used in vitro and in vivo in this proposal have been developed and further characterized by our teams. In vitro studies will be conducted using new advancements in the fields of microfluidics and nanobiotechnology. In vivo studies will employ a mammalian xenograft model that engrafts human GSC-derived GBM, which bests recapitulates human GBM. Further, in the in vivo studies, animal subjects will be treated with radiation using Small Animal Radiation Research Platform (SARRP), thus recreating traditional conformal beam radiotherapy for humans on the scale of a mouse. The results of this study will determine whether nanoparticle-modified hAMSCs can provide a treatment that is safe and effective for not only patients with GBM, but many types of primary and metastatic brain cancers. For future clinical application, the nanoparticles could be administered either to hAMSCs obtained from patient fat after culturing for a few days or then given IV as a treatment or to F.A.T. with the resulting engineered hAMSCs re- administered during surgery. This may lead to clinical trials, with a revolutionary new way of treating patients with brain cancer and facilitating personalized medicine.

 描述（由申请人提供）：胶质母细胞瘤（GBM）是成人中最常见的原发性脑肿瘤，占所有原发性脑肿瘤的20%。尽管目前采用包括手术和放化疗在内的最佳治疗方法，GBM 的中位生存率仅为 14.6 个月。这种发病率和死亡率的一个重要原因是 GBM 能够侵入正常脑实质，导致局部治疗无效。越来越多的证据表明，一小部分细胞，即脑肿瘤起始细胞（BTIC），与该疾病的治疗耐药性有关。为了使治疗有效，需要针对这些入侵细胞。一种有前景的方法涉及使用间充质干细胞（MSC），人们发现这种细胞会优先迁移并定位于癌细胞。此外，MSC 可以被改造为合成和释放抗肿瘤蛋白，例如影响 BTIC 的骨形态发生蛋白 4 (BMP4)。 MSC 可以从骨髓 (BM-MSC) 和脂肪组织 (AMSC) 中获得。 BM-MSC 很难获得，离体增殖能力有限，并且随着供者年龄的增长，有效性降低。与 BM-MSC 不同，AMSC 供应更丰富，更容易从脂肪组织中获得，表达更高水平的与细胞迁移有关的表面标志物，并且已被证明能够抵抗致癌转化。因此，AMSC 可能是更好的选择。病毒基因递送方法虽然常用于修饰 AMSC，但与插入突变和免疫原性相关，因此在人类患者中使用的翻译能力可能有限。可生物降解的聚合物纳米粒子能够有效地将非病毒基因传递到多种细胞类型，包括人类 AMSC (hAMSC)，同时避免病毒的典型问题。在这笔资助中，我们提出了一种新技术，将新鲜提取的脂肪组织 (F.A.T.) 和纳米粒子结合起来，以非病毒方式改造 F.A.T 中包含的原代 hAMSC，无需事先培养即可分泌抗癌蛋白，同时保持细胞向肿瘤细胞迁移的能力。我们的总体假设是纳米颗粒修饰的 hAMSCs 从 F.A.T. 获得。在临床相关的体内人类 GBM 模型中保留其肿瘤抑制特性。为了检验这一假设，我们将追求以下具体目标：（1）通过冻干的可生物降解纳米粒子，有效地将感兴趣的外源基因传递到患者新鲜提取的脂肪组织（F.A.T.）。 (2) 确定纳米颗粒修饰的分泌BMP4的hAMSCs是否在体外保留抗神经胶质瘤作用。 (3) 在体内小鼠模型中确定纳米颗粒修饰的 BMP4 分泌 hAMSC 治疗联合靶向放射治疗对人 GBM 的安全性和有效性。目标 1 涉及研究和优化将纳米颗粒与 F.A.T 相结合的独特技术。来自我们的患者。对于目标 2 和 3，使用已经在商业 hAMSC 中测试过的纳米颗粒，我们现在将研究在添加纳米颗粒之前已分离和培养的原代 hAMSC 的修饰。我们的团队已经开发并进一步表征了本提案中要使用的体外和体内技术。将利用微流体和纳米生物技术领域的新进展进行体外研究。体内研究将采用哺乳动物异种移植模型，该模型植入人类 GSC 衍生的 GBM，这最好地再现了人类 GBM。此外，在体内研究中，动物受试者将使用小动物放射研究平台（SARRP）接受放射治疗，从而在小鼠的规模上为人类重现传统的适形束放射治疗。这项研究的结果将确定纳米颗粒修饰的 hAMSC 是否可以为 GBM 患者以及多种类型的原发性和转移性脑癌提供安全有效的治疗。对于未来的临床应用，纳米颗粒可以施用到从患者脂肪中培养几天后获得的 hAMSC，然后进行静脉注射作为治疗，也可以施用到 F.A.T。手术期间重新施用由此产生的工程化 hAMSC。这可能会导致临床试验，以革命性的新方法治疗脑癌患者并促进个性化医疗。