ULTRASOUND-TARGETED MICROBUBBLE DESTRUCTION IN COMBINATION WITH THE PIGGYBAC

结合 PIGGYBAC 进行超声波定向微泡破坏

• 批准号: 8360328
• 负责人: Johann Urschitz
• 金额: $ 7.58万
• 依托单位: UNIVERSITY OF HAWAII AT MANOA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8360328
• 关键词: Adenocarcinoma Cell; Affect; Biogenesis; Breast Adenocarcinoma; Cabbage - dietary; Cell Membrane Permeability; Cells; Centers of Research Excellence; Clinical; Contrast Media; Country; DNA Transposable Elements; DNA delivery; Developed Countries; Early Diagnosis; Enzymes; Funding; Ganciclovir; Gene Delivery; Gene-Directed Enzyme Prodrug Therapy; Genes; Genome; Grant; HSV-Tk Gene; Human; Immune response; In Vitro; Incidence; Institutes; Mammalian Cell; Mediating; Microbubbles; Mobile Genetic Elements; Moths; Mus; National Center for Research Resources; Organ; Phosphorylation; Principal Investigator; Prodrugs; Research; Research Infrastructure; Resources; Simplexvirus; Source; System; Techniques; Thymidine Kinase; Time; Tissues; Transgenes; Ultrasonography; United States National Institutes of Health; Viral; Virus; Woman; Xenograft procedure; base; cancer cell; cancer diagnosis; cancer therapy; cost; cytotoxic; cytotoxicity; extracellular; gene delivery system; gene therapy; immunogenicity; in vivo; interest; killings; macromolecule; malignant breast neoplasm; mortality; neoplastic cell; non-viral gene therapy; novel; nucleotide analog; plasmid DNA; research study; suicide gene; tool; transgene expression

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Breast cancer is the most common form of cancer diagnosed in women worldwide, affecting an estimated 10% of women. More than half of the cases are in industrialized countries and while the rate of mortality as a result of breast cancer has decreased due to early detection, the incidence of breast cancer has risen by 30% in these countries in the last quarter of a century. Developing novel techniques for treating breast cancer should therefore be of primary interest and gene therapy offers a promising approach in that regard. Suicide-gene or gene-directed enzyme prodrug therapy (GDEPT) is a strategy developed to selectively target cancer cells. Here, treatment of cancer is based on the transfer of non-mammalian genes into tumor cells. These transgenes encode enzymes that selectively convert non-toxic prodrugs to highly toxic metabolites. For example, expression of the herpes simplex virus thymidine kinase (HSVtk) gene in mammalian cells renders these cells susceptible to the cytotoxic effects of innocuous nucleotide analogues such as ganciclovir (GCV) after phosphorylation by the viral enzyme. Treatment with HSVtk/GCV not only kills HSVtk expressing cells but also neighboring cells that do not express the HSVtk gene, greatly enhancing the efficacy of HSVtk-mediated cytotoxicity. Viral systems are the most efficient delivery systems; however, problems with immunogenicity, the potential for mutagenicity and the difficulty and expense of producing large amounts of pure virus hamper their successful application in a clinical setting. In contrast, non-viral systems have several advantages: they are easy and inexpensive to produce, are able to incorporate large transgenes and do no illicit an immune response. However, to date many non-viral gene therapy strategies have been limited by the efficiency of this gene delivery systems and the lack of stable chromosomal integration. Transposable elements (transposons) are mobile genetic elements that represent a novel non-viral DNA delivery system that provides long-term expression of transgenes as they are able to efficiently and permanently integrate into the host genome. PiggyBac (pB), a transposable element originally isolated from the cabbage looper moth Trichoplusia ni showed efficient transposition and long-term transgene expression in vitro in a variety of cells and in vivo in mouse experiments. Recently, ultrasound-targeted microbubble destruction (UTMD) has been proposed as safe and efficient means for gene delivery. Microbubbles are commonly used as contrast agents in ultrasound imaging and it has also been shown that microbubble enhanced ultrasound can alter cell membrane permeability for a short time, allowing extracellular macromolecules such as plasmid DNA to instantaneously enter cells without cytotoxicity. Temporal and spatial administration of the ultrasound allows for controlled delivery of genes to specific tissues or organs at precise time points. In this application we propose to evaluate the feasibility of using UTMD in combination with the piggyBac transposon system as a tool to enhance gene directed enzyme prodrug therapy in human breast adenocarcinoma cell xenografts.

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 乳腺癌是全世界女性确诊的最常见的癌症形式，估计有10%的女性受到影响。一半以上的病例发生在工业化国家，虽然由于及早发现乳腺癌导致的死亡率有所下降，但在过去25年里，这些国家的乳腺癌发病率上升了30%。因此，开发治疗乳腺癌的新技术应该是人们的首要兴趣，而基因疗法在这方面提供了一种很有前途的方法。自杀基因或基因导向的酶前药治疗(GDEPT)是一种选择性靶向肿瘤细胞的策略。在这里，癌症的治疗是基于将非哺乳动物基因转移到肿瘤细胞中。这些转基因编码的酶选择性地将无毒的前体药物转化为剧毒的代谢物。例如，单纯疱疹病毒胸苷激酶(HSVtk)基因在哺乳动物细胞中的表达使这些细胞在被病毒酶磷酸化后容易受到无害的核苷酸类似物如更昔洛韦(GCV)的细胞毒性作用。HSVtk/GCV不仅可以杀死表达HSVtk基因的细胞，还可以杀死邻近不表达HSVtk基因的细胞，大大增强了HSVtk介导的细胞毒效应。病毒系统是最有效的递送系统；然而，免疫原性、潜在的诱变性以及生产大量纯病毒的困难和费用阻碍了它们在临床环境中的成功应用。相比之下，非病毒系统有几个优势：它们容易生产，成本低廉，能够整合大量转基因，并且不会产生非法的免疫反应。然而，到目前为止，许多非病毒基因治疗策略一直受到这种基因递送系统的效率和缺乏稳定的染色体整合的限制。转座元件(转座子)是一种可移动的遗传元件，它代表了一种新型的非病毒DNA递送系统，提供了转基因的长期表达，因为它们能够有效和永久地整合到宿主基因组中。PiggyBac(PB)是一种从卷心菜二化蛾中分离出来的转座元件，在体外多种细胞和小鼠体内实验中都显示出高效的转座和长期的转基因表达。近年来，超声靶向微泡破坏(UTMD)被认为是一种安全有效的基因传递手段。微泡是超声成像中常用的造影剂，也有研究表明，微泡增强超声可以在短时间内改变细胞膜的通透性，使质粒DNA等细胞外大分子瞬间进入细胞，而不会产生细胞毒性。超声波的时间和空间管理允许在精确的时间点将基因控制地传递到特定的组织或器官。在这项应用中，我们建议评估联合使用UTMD和PiggyBac转座子系统作为工具来加强对人乳腺腺癌细胞异种移植瘤的基因导向的酶前药治疗的可行性。