Improved Gene Delivery and In Vivo Imaging

基因治疗目前受到缺乏非侵入性和临床上有用的手段来监测载体持久性和生物分布以及体内转基因表达的限制。目前项目的目标是测试策略，旨在改善腺病毒介导的基因治疗局部给药后基因表达的空间分布，使整个器官接受足够的病毒剂量，以尽可能少的病毒临床疗效（以限制毒性）。一个新的报告基因的基础上的钠碘同向转运体（NIS）的特点是它的能力，监测治疗基因的表达和对比与其他测量的催化活性正在开发中。有三个具体目标， 建议的研究。1.比较NIS报告基因表达与CD/HSV-1 TK治疗基因表达的分布。如果可以证明NIS活性是治疗性基因表达的替代标志物，NIS成像可以消除目前用于评估转基因表达的活检，并允许动态和全身监测转基因表达。2.评估新的载体制剂和注射条件，以优化腺病毒在前列腺内的放置，以获得最大的器官覆盖率。这项工作将为项目3中描述的临床试验提供科学依据。3.将前体药物标记的监测转基因表达的方法与NIS方法进行对比。这一目标 测试了旁观者效应可以测量并超过基因表达区域的假设。将使用结合的放射性标记的前药（[3 H]-更昔洛韦-MP、[14 C]-5-FdUMP）的放射自显影术测量旁观者效应，并与NIS转导后的碘定位进行比较。最后，测量将扩展到使用5-FU的[19 F]-MR化学位移成像和使用NIS活性的SPECT成像的123（I）摄取的非侵入性技术。拟议的研究需要大型动物模型。我们计划用大型杂种狗的前列腺。所描述的研究代表了“原理证明”，并将产生重要的新知识，这将有利于基因治疗和医学界。

Gene therapy is currently limited by the lack of a non-invasive and clinically useful means to monitor vector persistence and biodistribution and transgene expression in vivo. The goal of the current project is to test strategies devised to improve the spatial distribution of gene expression following the local administration of adenovirus-mediated gene therapy so that the entire organ receives a viral dose sufficient for clinical efficacy with as little virus as possible (to limit toxicity). A new reporter gene based on the sodium iodide symporter (NIS) will be characterized with regard to its ability to monitor therapeutic gene expression and contrasted with other measurements of catalytic activity under development. There are three specific aims of the proposed studies. 1. Compare the distribution of NIS reporter gene expression to that of CD/HSV-1 TK therapeutic gene expression. If it can be shown that NIS activity is a surrogate marker of therapeutic gene expression, NIS imaging could eliminate the biopsy currently used to assess transgene expression and allow for dynamic and whole body monitoring of transgene expression. 2. Evaluate new vector formulations and injections conditions designed to optimize placement of the adenovirus within the prostate to obtain maximal organ coverage. This work will provide the scientific basis for a clinical trial described in Project 3. 3. Contrast the prodrug-labeled approach of monitoring transgene expression to the NIS approach. This aim tests the hypothesis that the bystander effect can be measured and exceeds the area of gene expression. Bystander effect will be measured using autoradiography of bound radiolabeled prodrugs ([3H]-ganciclovir-MP, [14C]-5-FdUMP) and compared to iodide localization following transduction of NIS. Finally the measurements will be extended to non-invasive techniques using [19F]-MR chemical shift imaging of 5-FU and 123(I)uptake using SPECT imaging of NIS activity. A large animal model is needed for the proposed studies. We plan to use the prostates of large, mongrel dogs. The studies described represent "proof of principle" and will generate significant new knowledge that will benefit the gene therapy and medical communities.