Development of an MRI gene and application to probing the tumor microenvironment

描述（由申请人提供）：癌症和免疫系统之间的关系在很大程度上涉及疾病的结果。不幸的是，几乎没有方法可以直接监测肿瘤周围的免疫微环境。本项目的目标是开发一种新的基于磁共振成像（MRI）的报告基因，并将其应用于癌症免疫学领域。这将通过（1）开发基于HaloTag蛋白的报告基因来实现，该报告基因可以在MR图像上以对比度报告基因表达，以及（2）在（3）最终使用它来监测黑色素瘤肿瘤附近白细胞介素-10（IL-10）的存在之前，证明其对简单模型生物体的适用性。米德实验室最近的工作已经产生了一套Gd（III）造影剂，其与HaloTag蛋白紧密结合，并在结合时显著增加MR信号。结合这些技术创造了构建基于MRI的报告基因系统的潜力，从而基因的表达在MR图像上产生亮点。这种技术将有无数的用途，特别是在发育生物学、癌症和免疫学领域。这些领域中的每一个都将大大受益于MRI提供的无与伦比的深部组织分辨率。MR报告基因系统将在非洲爪蟾（一种发育生物学中常用的模式生物）中进行测试，方法是将HaloTag置于血管系统中活性启动子的控制下。为了原位检测IL-10，将使用基于细胞的生物传感器，该生物传感器包括模块化细胞外感觉结构，以在与IL-10结合时引起特异性基因表达。在所提出的系统中，IL-10的结合将导致外源性细胞外HaloTag的表达。暴露于HaloTag靶向造影剂将导致MR对比度增强，特别是在IL-10浓度升高的位置。这种技术有望推广到各种各样的细胞因子，最终允许在局部水平上详细描述肿瘤-免疫系统相互作用。

DESCRIPTION (provided by applicant): The relationship between cancer and the immune system is heavily implicated in the outcome of the disease. Unfortunately few methods exist to monitor the immune microenvironment immediately around the tumor. The goal of this project is to develop a novel magnetic resonance imaging (MRI) based reporter gene and apply it to the field of cancer immunology. This will be accomplished by (1) developing a reporter gene based on the HaloTag protein that can report gene expression with contrast on an MR image and (2) proving its applicability to a simple model organism before (3) finally using it to monitor the presence of Interleukin-10 (IL-10) in vicinity of melanoma tumors. Recent work in the Meade lab has produced a suite of Gd(III) contrast agents that bind tightly to the HaloTag protein and dramatically increase MR signal when bound. Combining these technologies creates the potential to construct an MRI based reporter gene system, whereby the expression of a gene produces a bright spot on an MR image. Such a technology would have innumerable uses, particularly in the fields of developmental biology, cancer, and immunology. Each of these fields would benefit greatly from the unparalleled deep tissue resolution offered by MRI. The MR reporter gene system will be tested in Xenopus laevis, a common model organism used in developmental biology, by placing HaloTag under the control of a promoter active in the vasculature. In order to detect IL-10 in situ a cell-based biosensor will be used that includes a modular extracellular sensory architecture to cause specific gene expression upon binding to IL-10. In the proposed system, the binding of IL-10 will result in expression of exogenous, extracellular HaloTag. Exposure to HaloTag targeted contrast agents will result in MR contrast enhancement specifically in locations of elevated IL-10 concentration. This technique is expected to be generalizable to a wide variety of cytokines, eventually allowing detailed descriptions of tumor-immune system interactions on a local level.