In vivo reporters of gene expression

描述(申请人提供)：器官移植正变得越来越普遍，在过去的20年里，由于积极的免疫抑制药物治疗，移植物一年存活率急剧上升。例如，现在大多数移植中心的肾移植一年成功率超过90%。然而，在同一时期，长期存活率并没有显著增加。亚临床炎症(SCI)是导致移植恶化的主要因素，为了提高器官移植的长期成功率，对移植组织中SCI进行测量是至关重要的。目前脊髓损伤的评估方法是侵入性的活检，但更有效的评估方法是对炎症过程中被激活的基因(S)进行成像。我们开发了一种基因表达报告系统，可以用于对深层组织中的基因表达进行成像。记者是从选定的启动子表达的适体，当它们与目标放射性配基结合时，可以看到它们。适配子充当细胞内的受体，紧密地结合它们的配体，从而提供启动子被激活的细胞的图像。目前的资深团队包括细胞和分子生物学、合成有机化学和放射化学方面的专业知识，他们将开发一种适体报告系统，用于对移植组织的局部炎症进行成像，以实现以下特定目标：1)制备稳定表达新霉素适配子的哺乳动物细胞，作为受调控(Lcn2)或成分表达(Actin)基因的报告者，2)开发合成和放射标记适体配体的化学程序，以及3)图像肌动蛋白和Lcn2基因在体内的表达。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，从而能够更明智地做出有关药物的决定。这种监测预计将提高因纳入 适体脊髓损伤报告系统移植前。同样的报告平台可以应用于其他成像需求，如干细胞贩运或其基因表达谱，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.