Novel Reagents for Rapid and Stable Thiol-Based Bioconjugations

用于快速、稳定的硫醇基生物共轭的新型试剂

• 批准号: 10640245
• 负责人: Brian Matthew Zeglis
• 金额: $ 32.37万
• 依托单位: HUNTER COLLEGE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-09 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10640245
• 关键词: Address; Antibodies; Biodistribution; Breast Cancer Model; Cancer Model; Chelating Agents; Chemistry; Clinic; Colorectal Cancer; Deferoxamine; Development; Dimethyl sulfone; Dissociation; ERBB2 gene; Exhibits; Extravasation; Generations; Goals; Health; Image; Immunoconjugates; Immunoglobulins; Isotopes; Kinetics; Label; Laboratories; Libraries; Ligation; Maleimides; Medicine; Methods; Modeling; Modification; Neuroblastoma; Patient-Focused Outcomes; Patients; Pentetic Acid; Performance; Physiological; Positron-Emission Tomography; Prosthesis; Protocols documentation; Radiation Dose Unit; Radioactive; Radioimmunoconjugate; Radioimmunotherapy; Radioisotopes; Radiolabeled; Reaction; Reagent; Reproducibility; Secure; Site; Solubility; Sulfhydryl Compounds; Technology; Testing; Therapeutic; Tissues; Trastuzumab; Treatment Protocols; Variant; Work; analog; clinical care; clinical efficacy; comparative; contrast imaging; design; experimental study; immunoreactivity; improved; in vivo; in vivo evaluation; member; mouse model; novel; nuclear imaging; pre-clinical; rational design; thioether; tool; uptake

Project Summary/Abstract For over three decades, the ligation between maleimide-bearing probes and thiols has been a cornerstone of the synthesis of site-specifically labeled antibodies. Yet despite its popularity, this approach to bioconjugation has serious limitations. Maleimide-based conjugates display limited stability in vivo because their succinimidyl thioether linkage can undergo a retro-Michael reaction that leads to the dissociation of the cargo or its exchange with circulating biomolecules. In the context of nuclear imaging and radioimmunotherapy, this retro- Michael reaction can lead to the release of the radioactive payload and the in vivo radiolabeling of endogeneous biomolecules, resulting in higher radiation doses to healthy tissues, reduced imaging contrast, and lower therapeutic ratios. In order to circumvent this obstacle, we have developed a modular, stable, and easily accessible phenyloxadiazolyl methyl sulfone reagent — `PODS' — as a platform for thiol-based bioconjugations. We have demonstrated that PODS can be used to reproducibly create homogenous, well- defined, highly immunoreactive, and highly stable radioimmunoconjugates with far superior in vivo performance compared to analogous probes synthesized via traditional, maleimide-based approaches. This proposal is centered upon the expansion and optimization of this technology. We will seek to build upon our preliminary work by optimizing the rapidity and selectivity of the PODS-thiol ligation and evaluating the in vivo performance of PODS-based radioimmunoconjugates labeled with 89Zr, 177Lu, 131I, and 225Ac. Specific Aim 1 (SA1) will be focused on the design, synthesis, and characterization of a library of second- generation PODS reagents, with the overall goal of identifying a single construct with the most favorable combination of stability, solubility, and reactivity. Specific Aim 2 (SA2) will be centered on the evaluation of the in vivo performance of radioimmunoconjugates synthesized using PODS. The most promising second- generation PODS reagent from SA1 will be used to synthesize 89Zr- and 177Lu-labeled radioimmunoconjugates, and their in vivo performance will be assessed in mouse models of cancer and compared to that of radiolabeled antibodies created using maleimide-based bioconjugations. Specific Aim 3 (SA3) will focus on the development of PODS-based prosthetic groups for the site-specific radiolabeling of antibodies with 131I and 225Ac. The in vivo performance of 131I- and 225Ac-labeled antibodies synthesized using these prosthetic groups will be evaluated in murine models of cancer and compared to analogues synthesized using current `gold- standard' strategies. We believe that this proposal could have a significant near-term impact on clinical care by developing and validating tools for the synthesis of highly stable radioimmunoconjugates with excellent in vivo performance, thereby improving imaging protocols, treatment regimens, and patient outcomes. Furthermore, we contend that this work could have a paradigm-shifting influence on bioconjugation chemistry, fundamentally changing the way biomolecular medicines are synthesized in the laboratory and clinic.

项目概要/摘要 三十多年来，马来酰亚胺探针和硫醇之间的连接一直是 然而，尽管这种生物共轭方法很受欢迎，但合成了位点特异性标记的抗体。 基于马来酰亚胺的缀合物在体内表现出有限的稳定性，因为它们的琥珀酰亚胺基。 硫醚键可以发生逆迈克尔反应，导致货物或其分子解离 在核成像和放射免疫治疗的背景下，这种逆转录酶与循环生物分子进行交换。 迈克尔反应可以导致放射性有效负载的释放和体内放射性标记 内源性生物分子，导致健康组织受到更高的辐射剂量，降低成像对比度， 为了克服这个障碍，我们开发了一种模块化的、稳定的和较低的治疗率。 易于获得的苯基恶二唑基甲基砜试剂 - “PODS” - 作为硫醇基的平台 我们已经证明 PODS 可用于可重复地产生均质的、良好的。 明确的、高度免疫反应性和高度稳定的放射免疫缀合物，在体内具有远远优越的性能 与通过传统的基于马来酰亚胺的方法合成的类似探针相比的性能。 该提案的重点是我们将寻求构建该技术的扩展和优化。 通过优化 PODS-硫醇连接的快速性和选择性并评估我们的初步工作 用 89Zr、177Lu、131I 和 225Ac 标记的基于 PODS 的放射免疫缀合物的体内性能。 具体目标 1 (SA1) 将重点关注第二个库的设计、合成和表征 一代 PODS 试剂，总体目标是识别具有最有利的单一构建体 稳定性、溶解度和反应性的组合将集中于评估 使用 PODS 合成的放射免疫缀合物的体内性能 最有前途的第二。 SA1 生成的 PODS 试剂将用于合成 89Zr 和 177Lu 标记的放射免疫缀合物， 他们的体内表现将在小鼠癌症模型中进行评估，并与 使用基于马来酰亚胺的生物共轭创建的放射性标记抗体将重点关注特定目标 3 (SA3)。 开发基于 PODS 的假体基团，用于用 131I 和 225Ac。使用这些辅基合成的 131I 和 225Ac 标记抗体的体内性能 将在小鼠癌症模型中进行评估，并与使用当前“金- 我们相信该提案可能会对临床护理产生重大的近期影响。 开发和验证用于合成具有优异体内活性的高度稳定的放射免疫缀合物的工具 性能，从而改善成像方案、治疗方案和患者结果。 我们认为这项工作可能会对生物共轭化学产生范式转变的影响，从根本上来说 改变生物分子药物在实验室和临床的合成方式。