Multi-modal129 Xe Probes for Tumor Detection

描述(由申请人提供)： 该项目的长期目标是开发多模式(磁共振、荧光)129Xe生物传感器，用于癌症早期检测和可能的治疗干预。与其他有机分子相比，隐形分子在其内部与氙气的结合更紧密(Kd约为100微米)，使这些笼子对氙气成像实验具有吸引力。氙气对其分子环境很敏感，隐形原子附近的微小扰动会导致129Xe核磁共振化学位移的巨大变化。此外，氙气在生物上是相容的，很容易在体内扩散，并且可以被超极化，将核磁共振信号增强10,000倍。这些特性构成了一种灵敏的成像试剂的基础，该试剂能够同时监测体内的多个分析物，每个分析物都会导致独特的129Xe化学位移。这种“多路传输”能力为肿瘤发生的分子成像提供了无与伦比的机会，同时识别多个癌症标记物将加快诊断和改善治疗。有两个特别具有挑战性但也很重要的目标是及早发现胰腺癌和脑癌，因为这些肿瘤是美国癌症死亡的主要原因，而且人们在分子水平上对它们的调控知之甚少。在该提案的R21部分，目标#1是合成具有增强的生物传感能力的新型密码材料，包括对129Xe的更高亲和力、更长的T1和新的化学位移。~(129)Xe~*的结合将通过核磁共振研究来表征。第二个目标是将目前对氙气亲和力最高的笼子--隐语-A与各种荧光团一起功能化，用于双峰检测，以及靶向药物，如环多巴胺、叶酸、葡萄糖和生长抑素。在AIM#1中开发的增强型密码也将被功能化。目的3是为了证明组织培养中的特异性递送。为了获得最佳的细胞递送和特异性，将以最小的毒性合成隐语。在R33阶段，目标4是使用针对特定目标优化的氙气递送剂，同时检测癌细胞系中的多个标记物。数据收集程序将在体内应用进行测试：其中包括优化129Xe核磁共振成像参数，传输密码和超极化氙气的协议，以及简化数据分析。到这一阶段，一大批密码武器将在手中，它们的毒性将在小鼠身上进行测试。最后，第五个目标是演示使用MRI对小鼠胰腺和/或脑肿瘤的早期检测。生物传感器的可交付性、肿瘤选择性、标志物特异性和129Xe信号强度将在小鼠肿瘤模型中进行评估。129Xe磁共振成像结果将使用荧光标记的载体进行验证，随后在体内和体外对组织进行成像：这样的“地面真实情况”将有助于评估这项新技术的诊断准确性。这项提案将专注于开发129Xe试剂，用于体内检测与胰腺癌和脑癌相关的多个生物标志物。

DESCRIPTION (provided by applicant): The long-term objective of this project is to develop multi-modal (MRI, fluorescence) 129Xe biosensors for early cancer detection, and possible therapeutic intervention. Cryptophanes bind xenon more tightly within their interior (Kd equals approximately 100 mu m) than other organic molecules, making these cages attractive for xenon imaging experiments. Xenon gas is sensitive to its molecular environment, and small perturbations near the cryptophane produce large changes in the 129Xe NMR chemical shift. Moreover, xenon is biologically compatible, diffuses readily in vivo, and can be "hyperpolarized" to enhance the NMR signal 10,000-fold. These properties form the basis for a sensitive imaging reagent capable of simultaneously monitoring multiple analytes in vivo, each causing a unique 129Xe chemical shift. This "multiplexing" capability offers unparalleled opportunities for molecular imaging of tumorigenesis, where simultaneous identification of multiple cancer markers would speed diagnosis, and improve treatment. Two particularly challenging, but important goals are the early detection of pancreatic and brain cancers, as these neoplasms are a leading cause of cancer death in the U.S., and their regulation is poorly understood at the molecular level. In the R21 portion of this proposal, Aim #1 is to synthesize novel cryptophanes with enhanced biosensing capabilities, including higher affinity for 129Xe, longer T1, and new chemical shifts. 129Xe*cryptophane binding will be characterized by NMR studies. Aim #2 is to functionalize cryptophane-A, currently the cage with highest affinity for xenon, with a variety of fluorophores for bimodal detection, as well as targeting agents such as cyclopamine, folate, glucose, and somatostatin. Enhanced cryptophanes developed in Aim #1 will also be functionalized. Aim #3 is to demonstrate specific delivery in tissue culture. Cryptophanes will be synthesized for optimal cell delivery and specificity with minimal toxicity. In the R33 phase, Aim #4 is to detect multiple markers simultaneously in cancer cell lines, using xenon delivery agents optimized for specific targets. Data collection procedures will be tested for in vivo applications: These include optimizing 129Xe NMR imaging parameters, protocols for delivering cryptophanes and hyperpolarized xenon, and streamlining data analysis. By this stage, an "arsenal" of cryptophanes will be in hand, and their toxicity will be tested in mice. Finally, Aim #5 is to demonstrate the early detection of pancreatic and/or brain tumors in mice using MRI. Biosensor deliverability, tumor selectivity, marker specificity, and 129Xe signal intensity will be evaluated in mouse tumor models. 129Xe MRI results will be validated using fluorescently labeled carriers and subsequently imaging tissue in vivo and ex vivo: such "ground truthing" will help to evaluate the diagnostic accuracy of this new technology. This proposal will focus on developing 129Xe reagents for the in vivo detection of multiple biomarkers associated with pancreatic and brain cancers.