In vivo metal-free cycloaddition chemistry driven pretargeted cancer radiotherapy

描述（由申请人提供）：拟议研究的目标是开发用于癌症放射免疫治疗（RIT）的新型无金属"点击化学"预靶向方法。假设新的体内环加成化学将极大地促进预靶向抗体作为癌症治疗剂的使用，克服其相对缓慢的体内药代动力学性质并产生高肿瘤至正常组织定位。单克隆抗体（mAb）CC49将用作靶向剂。CC49结合-TAG 72粘蛋白抗原，该抗原在包括结肠癌在内的许多癌症上过表达。CC49 mAb将用反式环辛烯（TCO）衍生化，并用于在注射治疗性探针之前以2步预靶向方法靶向肿瘤。双（吡啶基）四嗪（四嗪）共轭金属将用？-发射极212 Bi或其母源212 Pb或其母源212 Bi。发射体177Lu，并在注射亲肿瘤抗体后数小时至数天注射。在体内，在肿瘤靶向抗体上的TCO和放射性标记的螯合剂-四嗪的四嗪部分之间将发生快速和高度特异性的环加成反应，产生共价缀合物。未反应的放射性标记螯合剂-四嗪将从体内迅速清除，导致肿瘤至正常组织的放射性定位较高。拟议研究的目的如下。目标1，预靶向化学开发，包括第二代环辛烯、螯合剂-四嗪缀合物和清除剂的合成和表征。目标2，放射化学和共轭研究，将集中在螯合剂-四嗪共轭物的放射化学标记效率和放射化学稳定性，将与212 Bi，212 Pb和177 Lu放射性标记。TCO-CC49和标记的四嗪螯合剂的生物活性将用表达TAG 72的细胞系进行评价。目标3，新的预靶向成分的评价，将优化对肿瘤的剂量，同时减少对正常组织的剂量，包括新开发的四嗪探针的血液动力学和生物分布。目标4，治疗研究，将采取最佳组合？然后呢？发射体预靶向分子，并检查它们在结肠癌异种移植小鼠模型中的治疗功效。该项目的重要性包括，将体内环加成反应提高10倍，使用pretargeted？RIT和针对性的五倍改善？是的。最后的里程碑将是证明预靶向治疗研究将在小鼠结肠癌模型中产生统计学显著的生存期延长。这种新的体内环加成化学在癌症放疗的预靶向方法中的应用是高度创新的。烯烃与四嗪在体内的环加成反应由于其前所未有的反应性而首次在体内临床相关条件下有效地使用，并且首次具有将这种类型的有机化学扩展到人类的潜力。该项目将是首次在体内应用具有预定靶向的TCO-四嗪环加成化学。粒子放射治疗

DESCRIPTION (provided by applicant): The goal of the proposed research is to develop novel metal-free "click chemistry" pretargeted approaches for cancer radioimmunotherapy (RIT). It is hypothesized that novel in vivo cycloaddition chemistry will greatly facilitate the use of pretargeted antibodies as cancer treatment agents, overcoming their relatively slow in vivo pharmacokinetic properties and yielding high tumor to normal tissue localization. The monoclonal antibody (mAb) CC49 will be used as the targeting agent. CC49 binds the-TAG72 mucin antigen that is over expressed on a number of cancers including colon carcinomas. The CC49 mAb will be derivatized with trans-cyclooctene (TCO) and used to target the tumor prior to injection of the therapeutic probe in a 2-step pretargeting approach. A bis(pyridinyl)tetrazine (tetrazine) conjugated metal will be radiolabeled with the ?-emitter 212Bi or its parent 212Pb or the ?-emitter 177Lu and injected hours to a few days post injection of the tumor-avid antibody. In vivo, a rapid and highly specific cycloaddition reaction will occur between the TCO on the tumor- targeted antibody and the tetrazine moiety of the radiolabeled chelator-tetrazine yielding a covalent conjugate. Unreacted radiolabeled chelator-tetrazine will be rapidly cleared from the body resulting in a high tumor to normal tissue localization of radioactivity. The aims of the proposed research are as follows. Aim1, pretargeting chemistry development, which encompassed second generation cyclooctene, chelator-tetrazine conjugate and clearing agent synthesis and characterization. Aim 2, radiochemistry and conjugation studies, will focus on the radiochemical labeling efficiencies and radiochemical stabilities of the chelator-tetrazine conjugates that will be radiolabled with 212Bi, 212Pb and 177Lu. The bioacitivties of TCO-CC49 and the labeled tetrazine-chelators will be evaluated with TAG72 expressing cell lines. Aim 3, evaluation of new pretargeting components, will optimize the dose to the tumor while reducing the dose to normal tissues, including blood kinetics and biodistribution of newly developed tetrazine probes. Aim4, therapy studies, will take the best combination of ? and ? emitter pretargeting molecules and examine their therapeutic efficacy in a colon cancer xenograft mouse model. Milestones for the project include, improving in vivo cycloaddition reaction by 10 fold, targeting a greater that five-fold improvement in dose to tumor versus normal tissue using pretargeted ? RIT and a five-fold improvement in targeted ? RIT. The final milestone will be to demonstrate that pretargeted therapy studies will yield statistically significantly extension in survival in a mouse colon cancer model. This application of novel in vivo cycloaddition chemistry in a pretargeted approach for cancer radiotherapy is highly innovative. The cycloaddition reaction of olefins with tetrazines in vivo is the first of its kind to be used effectively at clincally relevant conditions in vivo due to its unprecedented reactivity, and for the first time has the potential to extend this type of organic chemistry in to man. This project would be the first of it kind in vivo application of TCO-tetrazine cycloaddition chemistry with pretargeted ?-particle radiotherapy.