In vivo reporters of gene expression

描述（由申请人提供）：器官移植变得越来越普遍，由于积极的免疫抑制药物治疗，过去20年来一年移植物存活率急剧上升。例如，肾移植现在在大多数移植中心的一年成功率超过90%。然而，在同一时期，长期存活率并没有显著提高。亚临床炎症（SCI）是移植恶化的主要原因，为了提高器官移植的长期成功率，在移植组织中测量SCI至关重要。SCI目前通过活检的侵入性测量进行阿塞塞，但通过炎症过程中激活的基因成像可以更有效地评估SCI。我们已经开发了一个基因表达报告系统，可用于成像基因表达在深部组织。报告基因是从所选启动子表达的适体，并且当它们结合其靶放射性配体时可视化。适体表现为细胞内受体，紧密结合其配体，从而提供启动子已被激活的细胞的图像。目前的经验丰富的团队，其中包括细胞和分子生物学，合成有机化学和放射化学的专业知识，将开发一种适体报告系统，通过实现以下特定目标来成像移植组织的局部炎症：1）制备稳定表达新霉素适体的哺乳动物细胞，作为调节基因（Lcn 2）或组成型表达基因（肌动蛋白）的报告基因，2）开发合成和放射性标记适体配体的化学程序，和3）在体内成像肌动蛋白和Lcn2基因表达。这里提出的研究对监测组织移植的健康状况具有广泛的意义，从而能够做出更明智的药物决策。这种监测预计将增加已通过纳入以下因素而改变的移植物的存活率： 适体SCI报告系统。相同的报告平台可以应用于其他成像需求，例如干细胞运输或其基因表达谱，以监测其分化阶段。

DESCRIPTION (provided by applicant): Organ transplants are becoming increasingly common and the one year graft survival rates have risen sharply over the past 20 years due to aggressive immunosuppressive drug treatments. For example, renal transplants now have one year success rates of over 90% in most transplant centers. However, the long-term survival rate has not increased significantly over this same period. Subclinical inflammation (SCI) is a major contributor to transplant deterioration and to increase long-term success rates of organ transplants it is vital to have a measure of SCI in the transplant tissue. SCI is currently assesse by the invasive measure of a biopsy but could be more effectively evaluated by imaging of gene(s) that are activated during inflammation. We have developed a gene expression reporter system that can be used for imaging gene expression in deep tissue. The reporters are aptamers that are expressed from a chosen promoter and are visualized when they bind their target radioligand. The aptamers behave as intracellular receptors, binding their ligands tightly and thereby providing an image of the cells in which the promoter has been activated. The current veteran team, which includes expertise in cell and molecular biology, synthetic organic chemistry and radiochemistry, will develop an aptamer reporter system for imaging local inflammation of transplanted tissues by achieving the following specific aims: 1) Prepare mammalian cells that stably express neomycin aptamers as reporters of either a regulated (Lcn2) or constitutively expressed (actin) gene, 2) Develop the chemical procedures to synthesize and radiolabel aptamer ligands, and 3) Image actin and Lcn2 gene expression in vivo. The studies proposed here have broad implications for monitoring the health of tissue transplants to enable more informed decisions regarding medications. Such monitoring is anticipated to increase the survival of transplants that have been modified by the inclusion of the aptamer SCI reporter system prior to their transplantation. The same reporter platform may be applied to other imaging needs such as stem cell trafficking or their gene expression profile to monitor their differentiation stage.