HARNESSING IN VIVO CLICK CHEMISTRY FOR 211AT-BASED PRETARGETED RADIOIMMUNOTHERAPY

项目摘要/摘要 在过去的15年里，使用放射核素标记的抗体的放射免疫治疗(RIT) 在治疗癌症方面显示出了巨大的希望。然而，单抗的多天循环时间 抗体意味着它们必须标记有a辐射的放射性核素，其物理半衰期为数天： 225Ac(T1/2~9.9d)。这种不可避免的长期生物和物理半衰期的组合可能会产生高 健康组织的辐射剂量率，这一并发症阻碍了225Ac标记的临床增殖 放射免疫结合物。规避这一问题的一种方法是进行预靶向放射免疫治疗。 (PRIT)。PRIT的预测是将免疫球蛋白和放射性核素去偶联，在注射前注射前者 后者，并使它们能够在体内的目标位置重组。用FAST标记抗体- 在体内达到最佳生物分布后移动的小分子放射性配基限制了循环。 组装的放射免疫结合物在血液中的时间，并使半衰期放射性核素的使用成为可能 它们通常与全长IGG不兼容。因此，这种方法可以产生高活跃度 在靶组织中的浓度仅为健康器官辐射剂量的一小部分， 直接标记的放射免疫结合物。 这份F31提案的重点是创建一种新的PRIT方法，该方法利用了短暂的、 放射性卤素Asatine-211(211At；T1/2~7.2h)的体内生物正交连接 四嗪(TZ)和反式环辛烯(TCO)。为此，我们将采用基于概念验证的模型系统 针对A33抗原的huA33抗体以及小鼠结直肠癌模型的研究。具体目标1 (SA1)将集中于合成、表征和体内评估TZ放射性配位体的文库 ~(131)I(T1/2~8.0d)标记，是~(211)At的廉价替代物。在具体目标2(SA2)中，我们将创建 211At标记的在SA1中发现的四个最有希望的放射性配基的变体。然后我们将使用这些211 At- 标记四嗪和huA33-TCO在异种皮下移植体内的预靶向生物分布研究 结直肠癌模型的建立。这些生物分布数据将被用来评估体内的性能 用每个~(211)At-Tz进行预靶向并启用剂量计算，旨在阐明 每种策略都以肿瘤组织和健康器官为目标。最后，具体目标3(SA3)将以验证为中心 211At-Prit皮下移植、原位移植和患者来源异种移植的治疗效果 人结直肠癌模型的建立。将对两个最有希望的项目进行纵向PRIT研究 211At-TZ放射性配基，以确定适合临床翻译的单个构建物。就在附近 从长远来看，这项研究可能会为患者创造一种更安全、更有效的治疗方式 患有结直肠癌。从长远来看，这项工作可能有助于开创一个基于211At的时代 放射性药物在多种恶性肿瘤患者的内放射治疗中起着至关重要的作用。

PROJECT SUMMARY/ABSTRACT Over the last fifteen years, radioimmunotherapy (RIT) using antibodies labeled with a-emitting radionuclides has shown significant promise for the treatment of cancer. Yet the the multi-day circulation time of monoclonal antibodies means that they must be labeled with an a-emitting radionuclide with a multi-day physical half-life: 225Ac (t1/2 ~ 9.9 d). This unavoidable combination of long biological and physical half-lives can create high radiation dose rates to healthy tissues, a complication that has hampered the clinical proliferation of 225Ac-labeled radioimmunoconjugates. One approach to circumventing this issue lies in pretargeted radioimmunotherapy (PRIT). PRIT is predicated on decoupling the immunoglobulin and radionuclide, injecting the former prior to the latter, and empowering them to recombine at target sites within the body. Labeling the antibody with a fast- moving, small molecule radioligand after it has reached an optimal biodistribution in vivo limits the circulation time of the assembled radioimmunoconjugate in the blood and enables the use of radionuclides with half-lives that are normally incompatible with full-length IgGs. As a result, this approach can produce high activity concentrations in target tissues at only a fraction of the radiation dose to healthy organs produced by traditional, directly-labeled radioimmunoconjugates. This F31 proposal is focused upon creating a novel approach to PRIT that leverages the short-lived, a- emitting radiohalogen astatine-211 (211At; t1/2 ~ 7.2 h) and the in vivo bioorthogonal click ligation between tetrazine (Tz) and trans-cylooctene (TCO). To this end, we will employ a proof-of-concept model system based on the A33 antigen-targeting huA33 antibody as well as murine models of colorectal carcinoma. Specific Aim 1 (SA1) will be focused on the synthesis, characterization, and in vivo evaluation of a library of Tz radioligands labeled with iodine-131 (t1/2 ~ 8.0 d), an inexpensive surrogate for 211At. In Specific Aim 2 (SA2), we will create 211At-labeled variants of the four most promising radioligands identified in SA1. We will then use these 211At- labeled tetrazines and huA33-TCO to perform pretargeted biodistribution studies in a subcutaneous xenograft model of colorectal carcinoma. These biodistribution data will be used to assess the performance of in vivo pretargeting with each 211At-Tz and enable dosimetry calculations designed to elucidate the radiation dose of each strategy to tumor tissue and healthy organs. Finally, Specific Aim 3 (SA3) will be centered on validating the therapeutic efficacy of this approach to 211At-PRIT in subcutaneous, orthotopic, and patient-derived xenograft models of human colorectal carcinoma. Longitudinal PRIT studies will performed with the two most promising 211At-Tz radioligands from SA2 in order to identify a single construct suitable for clinical translation. In the near term, this investigation could lead to the creation of a safer and more effective treatment modality for patients with colorectal cancer. And in the long term, this work could help usher in an era in which 211At-based radiopharmaceuticals play a vital role in the endoradiotherapy of patients with a wide variety of malignancies.