Virotherapy for Neuroblastoma Stem Cells

神经母细胞瘤干细胞的病毒疗法

• 批准号: 7664471
• 负责人: TIMOTHY P CRIPE
• 金额: $ 20.25万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7664471
• 关键词: Animals; Biology; Brain Neoplasms; Calcium Channel; Cancer Relapse; Cell Culture Techniques; Cell Line; Cell Separation; Cell division; Cells; Characteristics; Child; Cues; Cytolysis; Cytotoxic Chemotherapy; Data; Development; Disease; Disease remission; Dose; Doxorubicin; Drug resistance; Exhibits; Flow Cytometry; Gene Expression; Gene Mutation; Genes; Genotype; Growth; Herpesviridae; Human; Injection of therapeutic agent; Lytic; Malignant Neoplasms; Measures; Modeling; Molecular Profiling; Mus; Natural regeneration; Neuroblastoma; Oncolytic; Oncolytic viruses; Pathogenesis; Patients; Pharmaceutical Preparations; Phenotype; Population; Primary Neoplasm; Property; Proteins; Radiosurgery; Recurrent disease; Relapse; Resistance; Resolution; Sampling; Serum-Free Culture Media; Side; Simplexvirus; Solid Neoplasm; Sorting - Cell Movement; Stem cells; Testing; Therapeutic; Tumorigenicity; Virus; Virus Diseases; Xenograft Model; Xenograft procedure; base; cancer cell; cancer stem cell; chemotherapy; conventional therapy; cytotoxic; experience; high risk; improved; killings; mutant; neoplastic cell; nerve stem cell; nestin protein; notch protein; promoter; public health relevance; relating to nervous system; resistance mechanism; stem; stem cell biology; stemness; theories; therapeutic target; tool; tumor; tumor growth; tumor xenograft; tumorigenic; validation studies

DESCRIPTION (provided by applicant): Neuroblastoma is the most common extracranial solid tumor in children. Disease remission can frequently be achieved in patients with high-risk neuroblastoma using surgery, radiation, and chemotherapy. Unfortunately, relapse is common, even many years later, and the majority of patients ultimately die of disease. The emerging stem cell theory of cancer suggests that cancer relapse may be caused by a rare tumorigenic progenitor cell that is resistant to conventional therapy. If true for neuroblastoma, improved cure rates will likely only be achieved by identification and therapeutic targeting of the neuroblastoma stem cell. Our overarching hypotheses are that (1) neuroblastomas arise from a cancer stem cell that can be isolated, purified, and characterized and (2) these cells can be specifically included along with bulk cancer cells for destruction using transcriptionally regulated oncolytic herpes viruses. Using previously described properties of neural stem cells, we identified and isolated cell subpopulations in human neuroblastoma cell cultures suggestive of a neuroblastoma stem cell. Cells express the neural stem cell marker CD133, exhibit a calcium channel-dependent side population, form spheres in serum-free media (that exhibit properties shared with bona fide neural stem cell-derived neurospheres), show evidence of asymmetric cell division, and are relatively resistant to the cytotoxic chemotherapy drug doxorubicin. Some of these cell populations are enriched for the neuroblastoma stem cell, demonstrated by their increased efficiency in forming tumors in mice at low cell inoculums. We propose to further refine the identification and isolation of the neuroblastoma stem cell from both cell lines and primary tumor samples using a combination of these characteristics (aim 1). Rather than rely exclusively on marker expression, this function-based, multi-parameter selection is unique, and should allow us to more precisely define the neuroblastoma cancer stem cell. We have also shown that sphere-forming cells express nestin, similar to bulk cells, and these cells are susceptible to virus-induced cytolysis by an oncolytic virus containing a critical virus gene under control of the nestin promoter. We propose to determine whether this virus exhibits a more pronounced antitumor effect in neuroblastoma xenografts, and if such an effect is due to improved targeting of the neuroblastoma stem cells (aim 2). The first aim should advance the field of cancer stem cell biology by pioneering a multi-parameter selection schema, and should advance the field of neuroblastoma biology by more precisely defining the stem cell phenotype and genotype. The second aim should provide justification for further development of the nestin-targeted virus as a therapeutic for neuroblastoma. It will also help open a new avenue for potential therapeutic strategies that are cytotoxic for the bulk of tumor cells but also include destruction of neuroblastoma stem cells. PUBLIC HEALTH RELEVANCE: Aside from brain tumors, neuroblastoma is the most common solid tumor in children. We have partly isolated human neuroblastoma tumor initiating cells (cancer stem cells) that may be responsible for relapsed disease. Here we will further purify and characterize these cells from cell lines and primary human neuroblastomas. We will also determine if a new type of therapy in development, oncolytic herpes viruses, can be directed to kill neuroblastoma stem cells and give better treatment results in mouse tumor models. The results should not only deepen our understanding of the pathogenesis of neuroblastoma but will also be a cornerstone for the development of herpes viruses and other new experimental therapeutics that target neuroblastoma stem cells.

描述（由申请人提供）：神经母细胞瘤是儿童最常见的颅外实体瘤。通过手术、放疗和化疗，高危神经母细胞瘤患者通常可以达到疾病缓解。不幸的是，复发是常见的，甚至许多年后，大多数患者最终死于疾病。新兴的癌症干细胞理论表明，癌症复发可能是由一种罕见的致瘤祖细胞引起的，这种祖细胞对常规治疗有抵抗力。如果对神经母细胞瘤来说是真的，那么只有通过识别和治疗神经母细胞瘤干细胞才有可能提高治愈率。我们的首要假设是：（1）神经母细胞瘤产生于一种可以分离、纯化和表征的癌症干细胞，（2）这些细胞可以与大量癌细胞一起沿着被转录调节的溶瘤疱疹病毒破坏。利用先前描述的神经干细胞的特性，我们在人神经母细胞瘤细胞培养物中鉴定并分离出提示神经母细胞瘤干细胞的细胞亚群。细胞表达神经干细胞标记物CD 133，表现出钙通道依赖性侧群，在无血清培养基中形成球体（表现出与真正的神经干细胞衍生的神经球共有的特性），显示出不对称细胞分裂的证据，并且对细胞毒性化疗药物阿霉素具有相对抗性。这些细胞群中的一些富集成神经细胞瘤干细胞，这通过它们在低细胞接种下在小鼠中形成肿瘤的效率增加来证明。我们建议使用这些特征的组合进一步完善从细胞系和原发性肿瘤样品中鉴定和分离神经母细胞瘤干细胞（目的1）。这种基于功能的多参数选择是独特的，而不是完全依赖于标记物表达，应该使我们能够更精确地定义神经母细胞瘤癌症干细胞。我们还表明，球形成细胞表达巢蛋白，类似于散装细胞，这些细胞容易受到病毒诱导的细胞溶解的溶瘤病毒含有一个关键的病毒基因控制下的巢蛋白启动子。我们建议确定这种病毒是否在神经母细胞瘤异种移植物中表现出更明显的抗肿瘤作用，以及这种作用是否是由于神经母细胞瘤干细胞的靶向性提高（目的2）。第一个目标应该通过开创多参数选择模式来推进癌症干细胞生物学领域，并且应该通过更精确地定义干细胞表型和基因型来推进神经母细胞瘤生物学领域。第二个目标应该为进一步开发巢蛋白靶向病毒作为神经母细胞瘤的治疗方法提供理由。它还将有助于为潜在的治疗策略开辟一条新途径，这些治疗策略对大部分肿瘤细胞具有细胞毒性，但也包括破坏神经母细胞瘤干细胞。公共卫生相关性：除了脑肿瘤，神经母细胞瘤是儿童最常见的实体瘤。我们已经部分分离出了可能导致复发性疾病的人类神经母细胞瘤肿瘤起始细胞（癌症干细胞）。在这里，我们将进一步从细胞系和原代人神经母细胞瘤中纯化和鉴定这些细胞。我们还将确定一种正在开发的新型疗法，溶瘤疱疹病毒，是否可以直接杀死神经母细胞瘤干细胞，并在小鼠肿瘤模型中获得更好的治疗效果。这些结果不仅加深了我们对神经母细胞瘤发病机制的理解，而且也将成为开发疱疹病毒和其他靶向神经母细胞瘤干细胞的新实验疗法的基石。

项目成果

Neuroblastomas vary widely in their sensitivities to herpes simplex virotherapy unrelated to virus receptors and susceptibility.

• DOI: 10.1038/gt.2015.105
• 发表时间: 2016-02
• 期刊: Gene therapy
• 影响因子: 5.1
• 作者: Wang PY;Swain HM;Kunkler AL;Chen CY;Hutzen BJ;Arnold MA;Streby KA;Collins MH;Dipasquale B;Stanek JR;Conner J;van Kuppevelt TH;Glorioso JC;Grandi P;Cripe TP
• 通讯作者: Cripe TP