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肿瘤类器官在药物筛选和个性化用药中的研究进展*
Progress in the Research and Application of Tumor Organoids in Drug Screening and Personalized Drug Treatment
({{custom_author.role_en}}), {{javascript:window.custom_author_en_index++;}}恶性肿瘤是影响人类生命健康的重大疾病之一,药物治疗是常见的治疗手段。近年来,“精准治疗”已经成为肿瘤治疗的趋势。要实现对恶性肿瘤有效、精准的药物治疗,药物筛选模型至关重要。肿瘤类器官是近年来新兴的一种三维细胞模型,具有经长期传代还保留亲本肿瘤的特征和异质性、培养成功率高、周期短和能够高通量筛选药物等优点,已被用于药物筛选、预测患者对治疗的反应以及为个性化用药提供指导等。重点介绍了肿瘤类器官在药物筛选及个性化用药中的研究进展和面临的挑战。
Malignant tumor is one of the major diseases that threaten human life, and drug therapy is a common method. Currently, precision treatment has become a trend of tumor therapy. To achieve effective and precise drug therapy for malignant tumors, the drug screening models are very important points. Tumor organoids are three-dimensional cell models which have emerged in recent years. They have the advantages of retaining the characteristics and heterogeneity of parental tumors during long-term culture, with high success rate of culture and short culture cycle, and they can be used for high-throughput drug screening. They have been used for drug screening, predicting patients’ response of drug therapy, and providing guidance for personalized drug treatment. The progress of tumor organoids in drug screening and personalized drug treatment, and possible challenges were introduced.
肿瘤类器官 / 药物筛选 / 个性化用药 / 精准治疗 / 肿瘤 {{custom_keyword}} /
Tumor organoids / Drug screening / Personalized drug treatment / Precision treatment / Tumor {{custom_keyword}} /
表1 应用于抗体偶联与固相表面的分析技术Table 1 Application of surface analysis techniques for immobilized antibody |
| 技术方法 | 输入 | 输出 | 信息 | 应用 |
|---|---|---|---|---|
| XPS[60,61] | 单色X射线 | 光电子 | 元素和化学成分 | 定量载体表面抗体的密度 |
| SE[62,63] | 椭圆偏振光 | 光的相位和强度的变化 | 厚度、折射率、表面粗糙度 | 模式化分析、推断抗体形态 |
| DPI[64,65] | 激光 | 衰逝波的变化 | 质量、膜厚度、折射率、密度 | 通过质量和膜厚度推断抗体形态 |
| SPR[66,67] | 多角度单色激光 | 反射光和吸收光的变化 | 折射率、膜厚度 | 通过抗体和抗原的光吸收特性推断其形态 |
| NR[68,69] | 中子束 | 中子束反射后角度和波长的变化 | 折射率、膜厚度、表面粗糙度 | 模式化分析、推断抗体形态 |
| AFM[70,71,72,73] | 反馈驱动悬臂式纳米尖端 | 抗体在载体表面的高度、表面张力 | 表面粗糙度、相位信息、图像 | 精确定位高于载体表面14nm的抗体 |
| QCM[74,75] | 微量天平的共振频率 | 频率和振幅的变化 | 质量吸收系数、生物亲和性 | 通过压电石英晶体对抗体的吸收性和质量推断抗体形态 |
| ToF-SIMS[76,77] | 电离金属簇,“一级离子” | 电离片段,“二级离子” | 元素半定量、化学成分、分子质量 | 分辨F(ab')2和Fc片段氨基酸的种类及数量 |
| [1] |
There has been a resurgence of interest in the use of phenotypic screens in drug discovery as an alternative to target-focused approaches. Given that oncology is currently the most active therapeutic area, and also one in which target-focused approaches have been particularly prominent in the past two decades, we investigated the contribution of phenotypic assays to oncology drug discovery by analysing the origins of all new small-molecule cancer drugs approved by the US Food and Drug Administration (FDA) over the past 15 years and those currently in clinical development. Although the majority of these drugs originated from target-based discovery, we identified a significant number whose discovery depended on phenotypic screening approaches. We postulate that the contribution of phenotypic screening to cancer drug discovery has been hampered by a reliance on 'classical' nonspecific drug effects such as cytotoxicity and mitotic arrest, exacerbated by a paucity of mechanistically defined cellular models for therapeutically translatable cancer phenotypes. However, technical and biological advances that enable such mechanistically informed phenotypic models have the potential to empower phenotypic drug discovery in oncology.
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高彤, 丁冠守, 苏萍萍, 等. 药物筛选模型的研究进展. 海峡药学, 2021, 33(7): 1-5.
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Through the introduction of a steadily growing variety of preclinical test models drug development and biomarker research has advanced. Next to classical used 2D cell line cultures, tissue-slice cultures, 3D organoid cell cultures, genetically engineered mouse models, cell line derived mouse models and patient derived xenografts may be selected for a specific question. All models harbor advantages and disadvantages. This review focuses on the available preclinical test models, novel developments such as humanized mice and discusses for which question a particular model should be employed.Copyright © 2017 Elsevier Ltd. All rights reserved.
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Human liver cancer research currently lacks in vitro models that can faithfully recapitulate the pathophysiology of the original tumor. We recently described a novel, near-physiological organoid culture system, wherein primary human healthy liver cells form long-term expanding organoids that retain liver tissue function and genetic stability. Here we extend this culture system to the propagation of primary liver cancer (PLC) organoids from three of the most common PLC subtypes: hepatocellular carcinoma (HCC), cholangiocarcinoma (CC) and combined HCC/CC (CHC) tumors. PLC-derived organoid cultures preserve the histological architecture, gene expression and genomic landscape of the original tumor, allowing for discrimination between different tumor tissues and subtypes, even after long-term expansion in culture in the same medium conditions. Xenograft studies demonstrate that the tumorogenic potential, histological features and metastatic properties of PLC-derived organoids are preserved in vivo. PLC-derived organoids are amenable for biomarker identification and drug-screening testing and led to the identification of the ERK inhibitor SCH772984 as a potential therapeutic agent for primary liver cancer. We thus demonstrate the wide-ranging biomedical utilities of PLC-derived organoid models in furthering the understanding of liver cancer biology and in developing personalized-medicine approaches for the disease.
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冯紫伊, 梁珊珊, 于炜婷, 等. 患者来源肿瘤类器官的培养与研究及应用. 中国组织工程研究, 2021, 25(25): 4082-4088.
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高坚钧, 秦伟, 王浩, 等. 类器官技术在肿瘤研究中的应用与展望. 中国组织工程研究, 2019, 23(7): 1136-1141.
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李甜瑞, 赵瑞波, 张权, 等. 类器官及其应用的研究进展. 生物化学与生物物理进展, 2019, 46(8): 737-750.
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张健. 基于转录组数据挖掘的肿瘤异质性与肿瘤免疫微环境研究. 北京: 军事科学院, 2019.
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Colorectal cancer (CRC) liver metastasis is highly unfavorable for patient outcome and is a leading cause of cancer-related death. Pre-clinical research of CRC liver metastasis predominately utilizes CRC cell lines grown in tissue culture. Here, we demonstrate that CRC liver metastases organoids derived from human specimens recapitulate some aspects of human disease.Human CRC liver metastases pathological specimens were obtained following patient consent. Tumor disaggregates were plated and organoids were allowed to expand. CRC markers were identified by immunofluorescence. Stem cell genes were analysed by QPCR and flow cytometry. Response to drug therapy was quantified using time-lapse imaging and MATLAB analysis.Organoids showed global expression of the epithelial marker, EpCAM and the adenocarcinoma marker, CEA CAM1. Flow cytometry analysis demonstrated that organoids express the stem cell surface markers CD24 and CD44. Finally, we demonstrated that CRC liver metastases organoids acquire chemotherapy resistance and can be utilized as surrogates for drug testing.These data demonstrate that CRC liver metastases organoids recapitulate some aspects of human disease and may provide an invaluable resource for investigating novel drug therapies, chemotherapy resistance and mechanism of metastasis.Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.
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In Rspondin-based 3D cultures, Lgr5 stem cells from multiple organs form ever-expanding epithelial organoids that retain their tissue identity. We report the establishment of tumor organoid cultures from 20 consecutive colorectal carcinoma (CRC) patients. For most, organoids were also generated from adjacent normal tissue. Organoids closely recapitulate several properties of the original tumor. The spectrum of genetic changes within the "living biobank" agrees well with previous large-scale mutational analyses of CRC. Gene expression analysis indicates that the major CRC molecular subtypes are represented. Tumor organoids are amenable to high-throughput drug screens allowing detection of gene-drug associations. As an example, a single organoid culture was exquisitely sensitive to Wnt secretion (porcupine) inhibitors and carried a mutation in the negative Wnt feedback regulator RNF43, rather than in APC. Organoid technology may fill the gap between cancer genetics and patient trials, complement cell-line- and xenograft-based drug studies, and allow personalized therapy design. PAPERCLIP.Copyright © 2015 Elsevier Inc. All rights reserved.
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The KHP PCaBB was established in 2013 and recruits donors from the Urology or Oncology Departments at Guy's Hospital in London (UK). Prostate cancer patients may be approached to give their consent for biobanking at any point in their treatment pathway, which allows residual material from their earlier diagnosis to be transferred and used by the Biobank. Currently, patients are specifically asked to donate samples of blood and surplus prostate tissue as well as permitting access to their clinical and pathological data that continues to be added throughout the course of their disease. Between 2013 and 2015, 549 prostate cancer patients gave their consent to the biobank and, the tissue repository collected 489 blood samples, 120 frozen prostate tissue samples and 1064 formalin fixed paraffin embedded diagnostic blocks.Prostate cancer has become a chronic disease in a large proportion of men, with many men receiving multiple subsequent treatments, and their treatment trajectory often spanning over decades. Therefore, this resource aims to provide an ideal research platform to explore potential variations in treatment response as well as disease markers in the different risk categories for prostate cancer.A recent audit of the KHP PCaBB revealed that between 2013 and 2015, 1796 patients were diagnosed with prostate cancer at King's Health Partners (KHP), out of which 549 (30.6%) gave their consent to KHP PCaBB. Comparisons between demographic and clinical characteristics of patients who had consented compared to the total patient population revealed that the KHP PCaBB is demographically representative of the total prostate cancer patient population seen in Guy's and St Thomas' NHS Foundation Trust (GSTT). We observed no differences in distribution of ethnicity (p = 0.507) and socioeconomic status (p = 0.097). Some differences were observed in clinical characteristics, specifically with treatment type-which differed significantly between the patients who had given consent and total patient population.The KHP PCaBB has thereby amassed a rich data and tissue repository that is largely reflective of both the demographic and clinical diversity within the total prostate cancer patient population seen at KHP, making it an ideal platform for prostate cancer research.
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颜畅, 胡艺冰, 赵晖, 等. CEA-TCB双特异性抗体cibisatamab增强T细胞对胃癌类器官的免疫杀伤作用. 华中科技大学学报(医学版), 2020, 49(5): 511-516.
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T cells are key players in cancer immunotherapy, but strategies to expand tumor-reactive cells and study their interactions with tumor cells at the level of an individual patient are limited. Here we describe the generation and functional assessment of tumor-reactive T cells based on cocultures of tumor organoids and autologous peripheral blood lymphocytes. The procedure consists of an initial coculture of 2 weeks, in which tumor-reactive T cells are first expanded in the presence of (IFNγ-stimulated) autologous tumor cells. Subsequently, T cells are evaluated for their capacity to carry out effector functions (IFNγ secretion and degranulation) after recognition of tumor cells, and their capacity to kill tumor organoids. This strategy is unique in its use of peripheral blood as a source of tumor-reactive T cells in an antigen-agnostic manner. In 2 weeks, tumor-reactive CD8 T-cell populations can be obtained from ~33-50% of samples from patients with non-small-cell lung cancer (NSCLC) and microsatellite-instable colorectal cancer (CRC). This enables the establishment of ex vivo test systems for T-cell-based immunotherapy at the level of the individual patient.
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Patient-derived organoids (PDOs) have recently emerged as robust preclinical models; however, their potential to predict clinical outcomes in patients has remained unclear. We report on a living biobank of PDOs from metastatic, heavily pretreated colorectal and gastroesophageal cancer patients recruited in phase 1/2 clinical trials. Phenotypic and genotypic profiling of PDOs showed a high degree of similarity to the original patient tumors. Molecular profiling of tumor organoids was matched to drug-screening results, suggesting that PDOs could complement existing approaches in defining cancer vulnerabilities and improving treatment responses. We compared responses to anticancer agents ex vivo in organoids and PDO-based orthotopic mouse tumor xenograft models with the responses of the patients in clinical trials. Our data suggest that PDOs can recapitulate patient responses in the clinic and could be implemented in personalized medicine programs.Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.
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Effective treatment of advanced metastatic disease remains the primary challenge in the management of inoperable pancreatic cancer. Current therapies such as oxaliplatin (OxPt)-based chemotherapy regimens (FOLFIRINOX) provide modest short-term survival improvements, yet with significant toxicity. Photodynamic therapy (PDT), a light-activated cancer therapy, demonstrated clinical promise for pancreatic cancer treatment and enhances conventional chemotherapies with non-overlapping toxicities. This study investigates the capacity of neoadjuvant PDT using a clinically-approved photosensitizer, benzoporphyrin derivative (BPD, verteporfin), to enhance OxPt efficacy in metastatic pancreatic cancer. Treatment effects were evaluated in organotypic three-dimensional (3D) cultures, clinically representative models that bridge the gap between conventional cell cultures and models. The temporally-spaced, multiparametric analyses demonstrated a superior efficacy for combined PDT+OxPt compared to each monotherapy alone, which was recapitulated on different organotypic pancreatic cancer cultures. The therapeutic benefit of neoadjuvant PDT to OxPt chemotherapy materialized in a time-dependent manner, reducing residual viable tissue and tumor viability in a manner not achievable with OxPt or PDT alone. These findings emphasize the need for intelligent combination therapies and relevant models to evaluate the temporal kinetics of interactions between mechanistically-distinct treatments and highlight the promise of PDT as a neoadjuvant treatment for disseminated pancreatic cancer.
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Cancer immunotherapies have shown substantial clinical activity for a subset of patients with epithelial cancers. Still, technological platforms to study cancer T-cell interactions for individual patients and understand determinants of responsiveness are presently lacking. Here, we establish and validate a platform to induce and analyze tumor-specific T cell responses to epithelial cancers in a personalized manner. We demonstrate that co-cultures of autologous tumor organoids and peripheral blood lymphocytes can be used to enrich tumor-reactive T cells from peripheral blood of patients with mismatch repair-deficient colorectal cancer and non-small-cell lung cancer. Furthermore, we demonstrate that these T cells can be used to assess the efficiency of killing of matched tumor organoids. This platform provides an unbiased strategy for the isolation of tumor-reactive T cells and provides a means by which to assess the sensitivity of tumor cells to T cell-mediated attack at the level of the individual patient.Copyright © 2018 Elsevier Inc. All rights reserved.
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罗升昌, 王颖, 王士斌, 等. 基于微流控技术构建3D肿瘤模型用于药物筛选. 科学通报, 2021, 66(34): 4395-4410.
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图1 抗体的结构及固定化抗体的空间取向图2 EDC/NHS活化羧基表面的反应机理图3 光反应性探针辅助的亚硼酸连接法用于抗体固定图4 通过Diels-Alder反应制备Herceptin 单抗偶联纳米粒图5 末端含有硫酯的EYFP及连接有EYFP的脂质体的制备图6 位点特异性定向共价偶联生物素化IgG的策略图7 紫外线辅助下的核苷酸结合位点的抗体固定图8 紫外线辅助下基于金属鳌合的抗体固定
表1 应用于抗体偶联与固相表面的分析技术/
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