High-content functional cancer drug testing on micro-cuboidal tumor dissections

微立方体肿瘤解剖的高内涵功能性癌症药物测试

基本信息

批准号：
10025143
负责人：
ALBERT FOLCH
金额：
$ 60.2万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-06 至 2023-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10025143
关键词：
Address Agonist Antibodies Antitumor Drug Screening Assays Autologous Behavior Bioinformatics Biopsy Breast Cancer Model Breast Cancer Patient CTLA4 gene Cell Death Cells Cellular immunotherapy Clinic Collaborations Collagen Cytotoxic agent Development Devices Diffuse Dissection Drug Combinations Drug Delivery Systems Drug Exposure Ensure Enzyme-Linked Immunosorbent Assay Evaluation Excision Extracellular Matrix Fluorescence Future Gel Glioma Goals Growth Human Hydrogels Immune Immune checkpoint inhibitor Immune system Immunohistochemistry Immunomodulators Immunotherapy In Situ Individual Innate Immune System Interleukin-10 Label Lasers Liquid substance Mammary Neoplasms Methodology Microdissection Microfluidic Microchips Microfluidics Modeling Mouse Mammary Tumor Virus Mus Optics Organoids Patients Pharmaceutical Preparations Pharmacotherapy Phase Polymethyl Methacrylate Preparation Production Reproducibility Retrieval Sampling Shapes Slice Stains T cell therapy T-Cell Activation T-Lymphocyte Testing Tissue Microarray Tissue Viability Tissues Tumor Tissue Tumor-Derived Tumor-infiltrating immune cells Xenograft Model base cancer immunotherapy cancer therapy cell killing cell type cost cytokine design drug development drug response prediction genetic analysis human tissue macrophage malignant breast neoplasm micrography next generation particle personalized cancer therapy preservation prevent programmed cell death ligand 1 programmed cell death protein 1 prototype response screening tumor tumor microenvironment wasting

项目摘要

ABSTRACT The goal of this project is to perform high-content analysis of drug and immunotherapy responses on hundreds of intact, live cultured fragments isolated from a single live tumor biopsy. In recent years, patient-derived tumor “organoids” have shown great promise to predict drug responses for personalized cancer treatment. Immunotherapy, including cellular immunotherapy, represents the next generation of cancer therapy, and many of the relevant drugs act on the local tumor microenvironment (TME). There is a pressing need for functional testing platforms that use human, intact and live tumor tissue to better predict traditional and immunotherapy responses. Such platforms should also retain as much of the native TME as possible. Present high-throughput testing platforms that have some of these features, e.g. based on patient-derived tumor organoids, require a growth step that alters the TME. On the other hand, the micro-dissection of tumor tissue into “spheroids” that contain the TME intact has shown promising responses to immunomodulators on native immune cells. We propose a microfluidic platform that enables drug treatment, exogenous T cell therapy, and high-content analysis using hundreds to thousands of similarly sized, precision-sliced cuboidal micro-tissues (CµTs) produced from a single tumor sample. Here we propose a combination of two methodologies to demonstrate the feasibility of our approach: 1) precision slicing methodology that will produce large numbers of cuboidal micro-tissues (CµTs) from a single tumor biopsy; and 2) microfluidic trapping of the CµTs in a multi-well platform, allowing for drug application to each individual CµT or groups of CµTs. We will be able to obtain several hundred patient-derived CµTs from each tumor resection. The size of the CµTs (initially 400 µm×400 µm×400 µm) will be reproducible and chosen to optimize viability and retention of the TME. As the CµTs are cultured, their cuboidal shape will relax into a more rounded one. We will study the viability of the CµTs and their TME composition as a function of size in various culture conditions, including collagen gels. We will focus on breast cancer immunotherapy using a syngeneic mouse breast tumor model. For this Aim, we will deliver various concentrations and combinations of immunomodulatory drugs, including antibody-based drugs, to breast tumor CµTs in the microfluidic device, and examine the effects on the resident immune system. We will assess cytokine production and use high-content immunohistochemistry and bioinformatics analysis to assess immune cell engagement with different cell types as well as cell death. We will apply the platform to deliver immune checkpoint inhibitors (CTLA4, PD-L1, PD-1) and other immunomodulators (such as IL-10) and examine the effect on the immune state, cell death, and the behavior of resident T cells (activation and localization). In the R33 phase we plan on applying our microfluidic platform to CµTs obtained from breast tumor patients (ongoing collaboration with Dr. V.K. Gadi, Fred Hutch).

抽象的该项目的目的是对数百种对药物和免疫疗法反应进行高含量分析完整的，从单个活肿瘤活检中分离出的活培养片段。近年来，患者衍生的肿瘤 “器官”表现出了巨大的希望，可以预测个性化癌症治疗的药物反应。免疫疗法，包括细胞免疫疗法，代表下一代癌症治疗，许多有关局部肿瘤微环境（TME）的相关药物法。迫切需要功能使用人类，完整和活肿瘤组织更好地预测传统和免疫疗法的测试平台回答。此类平台还应保留尽可能多的本机TME。目前的高通量具有其中一些功能的测试平台，例如基于患者来源的肿瘤器官，需要改变TME的增长步骤。另一方面，肿瘤组织的微截断成“球体” 包含完整的TME已显示出对天然免疫细胞上免疫调节剂的承诺反应。我们提案一个微流体平台，可实现药物治疗，外源T细胞疗法和高素质分析使用数百至数千个类似尺寸的精密切割的立方体微问题（CµTS）分析由单个肿瘤样品产生。在这里，我们提出了两种方法的组合来证明我们方法的可行性：1）精度切片方法，将从单个肿瘤中产生大量的立方体微问题（CµT）活检； 2）在多孔平台中CµTs的微流体捕获，允许在每个平台上使用药物单个CµT或CµT组。我们将能够从每个肿瘤切除。 CµTS的大小（最初为400 µm x 400 µm x 400 µm）将被重现并选择为优化TME的生存能力和保留。随着CµT的培养，它们的立方体形状会放松成更多四舍五入。我们将研究CµTS及其TME组成的生存能力随着各种大小的函数培养条件，包括胶原蛋白凝胶。我们将使用同步小鼠乳腺肿瘤模型专注于乳腺癌免疫疗法。为此，我们将提供免疫调节药物的各种浓度和组合，包括基于抗体药物，用于微流体装置中的乳腺肿瘤CµT，并检查对居民免疫系统的影响。我们将评估细胞因子的生产，并使用高内核免疫组织化学和生物信息学分析用不同的细胞类型和细胞死亡评估免疫细胞的接合。我们将将平台应用于输送免疫粘点抑制剂（CTLA4，PD-L1，PD-1）和其他免疫调节剂（例如IL-10）和检查对免疫状态，细胞死亡和居民T细胞行为的影响（激活和本土化）。在R33阶段，我们计划将微流体平台应用于从乳腺肿瘤获得的CµT 患者（与弗雷德·哈奇（Fred Hutch）的V.K. Gadi博士进行持续合作）。