ENZYME - ACTIVATED PROTON MRI CONTRAST AGENTS

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 基因治疗已经成为治疗前列腺癌的一种潜在的有前途的策略。然而，由于难以评估转基因的成功，尤其是评估转基因表达的位置、大小和持久性，广泛的实施受到了阻碍。报告基因和相关分子应该可以评估基因的表达。编码大肠杆菌β-半乳糖苷酶的LacZ基因已被公认为最常用的报告系统。此外，前列腺特异性膜抗原(PSMA)作为最成熟、最具限制性的前列腺癌细胞表面抗原，已被确定为前列腺癌的理想抗原靶点。 许多出色的研究致力于开发非治疗性报告基因和成像方案，以非侵入性地在体内揭示基因表达和癌症本身，如核成像(PET、SPECT)、荧光、生物发光和新的近红外报告分子。磁共振成像技术在小动物的发育过程中和临床实践中开辟了以非常高的分辨率进行成像的领域。最近，新一代的MRI造影剂，即所谓的“智能”造影剂，为特定疾病状态的生物标志物开发MRI测试，并帮助早期检测和诊断，在基因治疗领域有着巨大的希望。 我们建议开发一种新型的“智能”的基于Gd3+的MRI造影剂，用于体内检测β-Gal或PSMA的活性。这一新概念的基于Gd3+的智能磁共振造影剂由三部分组成：(A)信号增强基团，如Gd-DOTA或Gd-PCTA；(B)Fe3+螯合基团；(C)β-D-半乳糖或β-谷氨酸。在前列腺癌细胞中被LacZ转基因或PSMA切割后，释放的、激活的苷元Fe3+-配体将自发地在酶活性位置捕获内源性Fe3+，形成高度稳定的络合物。值得注意的是，这种络合物将表现出Gd3+络合物的受限运动，增强了弛豫性能。此外，基于Fe3+的顺磁性络合剂将产生很强的弛豫。这一过程将提供基于酶β-半乳糖或PSMA刺激的局部积累的对比。Gd3+和Fe3+造影剂联合用于体内检测β-Gal或PSMA活性是前列腺癌基因活性体内监测或体内成像的一种新方法。 我们计划合成一系列专为检测前列腺癌细胞培养中的β-半乳糖或PSMA活性而设计的新型“SMART”磁共振造影剂，探索将最有希望的类似物应用于体内生长的小鼠和大鼠细胞的可行性，并与血管扩张和坏死进行比较，以分析新型“SMART”磁共振造影剂对肿瘤大小和生长的响应的相关性。 最终目的是开发一种用于体内评估β-半乳糖或PSMA活性的“智能”MRI造影剂的新概念，从而通过细胞外MRI方法评估PSMA，从而建立一个新的平台，用于前列腺癌基因治疗中LacZ转基因表达的体内监测或前列腺癌本身的体内成像。基本上，这个新的概念结合了所有可能达到最高松弛的方法，三个功能部分的组合是基于癌症治疗的临床应用策略。我们相信，将这项技术转化为前列腺癌的临床研究，并有望在体内监测前列腺癌的基因治疗和体内成像，将提高疗效，同时减少副作用，从而改善前列腺癌患者的生活质量，该技术将对探索其他疾病的研究人员具有广泛的实用价值。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Gene therapy has emerged as a potentially promising strategy for treatment of prostate cancer. However, widespread implementation is hindered by difficulties in assessing the success of transfection: in particular, assessing the location, magnitude and persistence of transgene expression. Reporter genes and associated molecules should allow assessment of gene expression. lacZ gene encoding E. coli beta-galactosidase (beta-gal) has already been recognized as the most commonly used reporter system. Moreover, prostate-specific membrane antigen (PSMA), as the most well-established, highly restricted prostate cancer cell surface antigen, has been identified as an ideal antigenic target in prostate cancer. Much elegant research has been directed at developing non-therapeutic reporter genes and imaging protocols to non-invasively in vivo reveal gene expression and cancer itself, such as nuclear imaging (PET, SPECT), fluorescence, bioluminescence and new near infrared reporter molecules. MRI techniques have opened the realm of imaging at very high resolutions in small animals during development and in clinical practice. Recently, a new emerging generation of MRI contrast agents, so-called "smart" contrast agents of developing MRI tests specific for biomarkers indicative of particular disease states and aiding in the early detection and diagnosis, hold great promise in the gene therapy arena. We propose to develop a novel class of "smart" Gd3+-based MRI contrast agents for in vivo detection of beta-gal or PSMA activity. This new concept of the "smart" Gd3+-based MRI contrast agents is composed of three moieties: (A) a signal enhancement group, such as Gd-DOTA or Gd-PCTA; (B) an Fe3+ chelating group; (C) beta-D-galactose or beta-glutamate. Following cleavage by lacZ transgene or PSMA in prostate cancer cells, the released, activated aglycone Fe3+-ligand will spontaneously trap endogenous Fe3+ at the site of enzyme activity forming a highly stable complex. Significantly, this complex will exhibit restricted motion of the Gd3+ chelates enhancing relaxivity. In addition, a paramagnetic chelator based on the Fe3+ will generate strong relaxation. This process will provide contrast based on enzyme beta-gal or PSMA stimulated local accumulation. The combination of Gd3+ and Fe3+ contrast agents for in vivo detection of beta-gal or PSMA activity is fundamentally new approach for in vivo monitoring gene activity or in vivo imaging of prostate cancer. We plan to synthesize series of novel "smart" MRI contrast agnets well-designed for detecting beta-gal or PSMA activity in prostate cancer cell culture, explore the feasibility of applying the most promising analogies to cells grown in vivo in mice and rats, and compare with vascular extent and necrosis to analyse the correlation of novel "smart' MRI contrast agents response to tumor size and growth. The ultimate objective is to develop a new concept of "smart" MRI contrast agents for in vivo assessment of beta-gal or PSMA activity, thereby, to establish a novel platform for in vivo monitoring of lacZ transgene expression in prostate cancer gene therapy or in vivo imaging of the prostate cancer itself by assessing PSMA through an extracellular MRI approach. Basically, this novel concept combines all the approaches of reaching the highest possible relaxivities, and the combination of three functional moieties is based on the clinically applied strategies on cancer therapy. We believe the technique translation to clinical investigations with the high promise of in vivo monitoring gene therapy and in vivo imaging of prostate cancer will improve efficacy while reducing side effects and hence improving quality of life for patients with prostate cancer, and the technique will be of widespread utility to investigators probing other diseases.