The Tetrazine Ligation for Efficient 18F Labeling and Pretargeted Imaging/Radiotherapy of Cancer

描述(由申请人提供)：建议开发一种简单、快速和通用的策略，将放射性核素结合到生物分子中，应用于癌症成像和预靶向免疫治疗。该策略基于福克斯实验室开发的生物偶联反应：四嗪与反式环辛烯(TCO)之间的反应。四嗪-TCO配位反应的快速动力学(K2e250,000 L*摩尔/S-1)使低微摩尔浓度下的快速反应在几分钟内即可完成，且没有过量的任何一种反应物。这种快速的反应为蛋白质和抗体的修饰以及在体内进行反应提供了前所未有的机会。该提案涉及以下方面的合作努力 南加州大学分子成像中心(MIC)和福克斯研究小组(UD)。这项应用的一个目的是利用四嗪连接来开发有效的方法来构建具有高比活性的18F标记蛋白用于肿瘤成像。此外，我们还将开发一种新的预靶向成像和放射免疫治疗方法，这种方法将允许使用成像/治疗性同位素对单抗进行普遍和直接的标记，而不会对正常器官造成严重辐射。为了实现这些目标，我们将首先在四嗪-TCO连接的基础上开发具有快速动力学和体内稳定性的生物连接反应。作为四嗪-TCO的快速动力学 连接能够在几分钟内在低微摩尔浓度下快速反应，而不需要过量的任何一种反应物，我们将把这种反应转化为有效的方法，可以用于构建具有高比活性的18F标记蛋白。最后，我们还将建立一种基于四嗪-TCO连接和抗EphB4抗体的化学预靶向方法，用于肿瘤的预靶向成像/放射免疫治疗。综上所述，在这项应用中，我们正在开发高效的18F标记蛋白质的方法，以及一种可以用于乳腺癌的预靶向成像和放射免疫治疗的化学预靶向方法。这些方法的可行性将通过EphB4抗体、F(ab‘)2和Fab进行测试。这些新的成像和治疗方法的成功可以使乳腺癌诊断成为可能，使直接监测对治疗干预的反应成为可能，并可能显著提高乳腺癌的治疗效果。此外，这些新开发的方法可以在许多其他癌症类型中有重要的应用，从而对非常大量的癌症患者产生重大的临床影响

DESCRIPTION (provided by applicant): Proposed is the development of a simple, fast and universal strategy for incorporating radionuclides into biomolecules for applications for cancer imaging and pretargeted immunotherapy. The strategy is based on a bioconjugation reaction developed in the Fox laboratories: the reaction between tetrazines and trans-cyclooctenes (TCO). The fast kinetics of the 'tetrazine-TCO ligation' (k2 e 250,000 L*mol/s-1) enable fast reactivity at low micromolar concentrations within minutes and without an excess of either reactant. This fast reactivity provides unprecedented opportunities for protein and antibody modification and for performing reactions in vivo. This proposal involves collaborative effort from the Molecular Imaging Center (MIC) at University of Southern California (USC) and the Fox group (UD). An objective of this application is to use the tetrazine ligation to develop efficient methods for the construction of 18F labeled proteins with high specific activity for cancer imaging. Moreover, we will also develop a novel pretargeted imaging and radioimmunotherapy method that will allow for universal and straightforward tagging of monoclonal antibodies (mAbs) with an imaging/therapeutic isotope without severe radiation exposure towards normal organs. To achieve these objectives, we will first develop bioconjugation reactions with fast kinetics and robust in vivo stability based on the tetrazine-TCO ligation. As the fast kinetics of tetrazine-TCO ligation enable fast reactivity at low micromolar concentrations within minutes and without an excess of either reactant, we will transform this reaction into efficient methods that could be used for the construction of 18F labeled proteins with high specific activity. At last, we will als establish a chemical pretargeting approach that for pretargeted imaging/radioimmunotherapy of cancer based on the tetrazine-TCO ligation and anti-EphB4 antibody. In summary, in this application, we are developing efficient methods for proteins labeling with 18F, and a chemical pretargeting approach that could be used for pretargeted imaging and radioimmunotherapy of breast cancer. The feasibility of these approaches will be tested with EphB4 antibody, F(ab')2, and Fab. The success of these novel imaging and therapy approached could enable breast cancer diagnosis, make possible direct monitoring of responses to therapeutic interventions, and may significantly improve breast cancer treatment efficacy. Moreover, these newly developed approaches could have important applications in many other cancer types, and thus have a significant clinical impact on a very large number of cancer patients