Foundations of Pretargeted Radioimmunotherapy

预定位放射免疫治疗的基础

基本信息

批准号：
7446643
负责人：
Karl Dane Wittrup
金额：
$ 25.01万
依托单位：
MASSACHUSETTS INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-08-01 至 2010-05-11
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7446643
关键词：
Adenocarcinoma Cell Affinity Animals Anti-CEA Antibody Antibodies Antibody Affinity Antigen Targeting Antigens Binding Biodistribution Biological Biological Models Biotin Bismuth Bispecific Antibodies Bolus Infusion Bone Marrow Carcinoembryonic Antigen Cell Line Cells Chelating Agents Colon Adenocarcinoma Colorectal Cancer Cultured Cells Diffusion Dissociation Dose Dose-Limiting Drug Kinetics Engineering Ensure Exhibits Experimental Models Exposure to Faculty Fluorescence Microscopy Foundations Gallium Generations Half-Life Human Immunoglobulin Fragments Immunoglobulin G Ionizing radiation Ions Isotopes Kidney Kinetics Label Libraries Link Metabolism Micrometastasis Modeling Nude Mice Penetration Performance Process Protein Engineering Proteins Radiation Radiation Dosage Radiation Oncology Radioimmunoconjugate Radioimmunotherapy Radioisotopes Radiometry Rate Reagent Research Personnel Saccharomyces cerevisiae Set protein Solid Solid Neoplasm Streptavidin Study models Surface System Testing Therapeutic Index Time Tissues Toxic effect Tumor Cell Line Week Xenograft Model Xenograft procedure Yeasts Yttrium anti-CEA scFv antibody engineering base cell killing directed evolution immunogenicity in vitro Model in vivo mathematical model multidisciplinary neoplastic cell size sound stable isotope tool trafficking tumor tumor xenograft uptake

项目摘要

DESCRIPTION (provided by applicant): Delivery of cell-killing doses of ionizing radiation to tumors is the objective of antibody-directed radioimmunotherapy (RIT). In practice, however, collateral damage to healthy bone marrow and kidneys limits the maximum delivered radiation dose. Pretargeted RIT (PRIT) aims to overcome this limitation by separating the pharmacokinetics of tumor targeting and radionuclide delivery. First-generation PRIT utilizes the streptavidin/biotin interaction for radionuclide capture, and exhibits promise in animal tumor xenograft models. However, problems with streptavidin immunogenicity, kidney localization, and endogenous biotin intrinsically limit this approach. The perspective of this proposal is that the principle of PRIT is sound, but that to reach its full potential the protein targeting agents must be optimized, and the pharmacokinetics of tumor penetration must be subjected to rigorous engineering analysis. This project brings together faculty from Biological Engineering and Radiation Oncology to collaboratively develop the essential reagents and dosing strategies to enable PRIT to be maximally effective. Protein engineering by directed evolution will be applied to: 1) construct human antibody fragments that capture radiometal chelates (DOTA with yttrium, gallium, or bismuth) effectively irreversibly; and 2) to develop anti-CEA antibody fragments that bind persistently through cycles of endocytic trafficking through acidic compartments. The processes of diffusion, binding, and metabolism of antibody fragments in micrometastases will be characterized by quantitative fluorescence microscopy of single LS174T human colon adenocarcinoma tumor cells and spheroid cultures, and analyzed within a mathematical modeling framework to determine the limiting kinetic processes and predict necessary concentrations and times for saturation binding. These predictions will be tested with quantitative biodistribution studies in LS174T xenografts in nude mice. Taken together, these studies will establish a firm foundation from which to optimize PRIT. In this second and final allowed revision of the proposal, we have eliminated radiation dosimetry and focused on endocytic trafficking of antibody fragments as a critical process limiting the permeation and retention of bispecific antibodies, based on direct evidence for rapid antibody uptake by LS174T cells. This hypothesis informs the planned efforts in protein engineering, cell culture, and in vivo biodistribution studies.

描述（由申请人提供）：向肿瘤递送细胞杀伤剂量的电离辐射是抗体导向的放射免疫疗法（RIT）的目标。然而，在实践中，对健康骨髓和肾脏的附带损害限制了最大放射剂量。预靶向 RIT (PRIT) 旨在通过分离肿瘤靶向和放射性核素递送的药代动力学来克服这一限制。第一代 PRIT 利用链霉亲和素/生物素相互作用来捕获放射性核素，并在动物肿瘤异种移植模型中展现出前景。然而，链霉亲和素免疫原性、肾脏定位和内源性生物素的问题本质上限制了这种方法。该提案的观点是，PRIT 的原理是合理的，但为了充分发挥其潜力，必须优化蛋白质靶向剂，并且必须对肿瘤渗透的药代动力学进行严格的工程分析。该项目汇集了生物工程和放射肿瘤学的教师，共同开发必要的试剂和剂量策略，以使 PRIT 发挥最大效力。定向进化的蛋白质工程将应用于：1）构建有效不可逆地捕获放射性金属螯合物（钇、镓或铋的 DOTA）的人类抗体片段； 2) 开发抗 CEA 抗体片段，该片段在通过酸性区室的内吞运输循环中持续结合。将通过单个 LS174T 人结肠腺癌细胞和球体培养物的定量荧光显微镜来表征微转移中抗体片段的扩散、结合和代谢过程，并在数学模型框架内进行分析，以确定限制动力学过程并预测必要的浓度和饱和结合时间。这些预测将通过裸鼠 LS174T 异种移植物的定量生物分布研究进行检验。总而言之，这些研究将为优化 PRIT 奠定坚实的基础。在第二次也是最后一次允许的提案修订中，我们取消了辐射剂量测定，并根据 LS174T 细胞快速摄取抗体的直接证据，将抗体片段的内吞运输作为限制双特异性抗体渗透和保留的关键过程。这一假设为蛋白质工程、细胞培养和体内生物分布研究的计划工作提供了信息。