Advanced tools for HCMI model genetic perturbation and metastasis characterization

用于 HCMI 模型遗传扰动和转移表征的高级工具

基本信息

批准号：
10229465
负责人：
John Doench
金额：
$ 78.99万
依托单位：
BROAD INSTITUTE, INC.
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：
2020-08-05 至 2023-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10229465
关键词：
3-Dimensional Address Bar Codes Benchmarking Brain CRISPR screen CRISPR/Cas technology Cancer Model Cancer cell line Cell model Cells Clinical Clinical Data Clustered Regularly Interspaced Short Palindromic Repeats Communities Complex Computational Science Consumption Data Development Dropout Ensure Enzymes Epigenetic Process Future Gene Expression Profiling Generations Genetic Genetic Models Genetic Screening Growth Guide RNA Human Immunodeficient Mouse Injections Kidney Knock-out Libraries Liver Lung Malignant Neoplasms Maps Measures Mediating Methods Modeling Molecular Monitor Neoplasm Metastasis Organ Organoids Outcome Phenotype Physiological Population Proliferating Protocols documentation Reagent Research Resources System Systems Biology Time Translational Research Ultrasonography anticancer research base bone cell growth cost experimental study functional genomics gene discovery genome editing genome-wide high throughput screening in vivo innovation model development multidisciplinary neoplastic cell next generation patient response precision medicine precision oncology scale up screening single-cell RNA sequencing targeted treatment technology development tissue culture tool tumor tumor microenvironment vector

项目摘要

ABSTRACT The Human Cancer Models Initiative (HCMI) is creating next generation cancer models that will drive the future of cancer precision medicine research. Historical cancer cell lines have been selected for their rapid proliferation on tissue culture plastic, which has made them amenable to high throughput screening such as genome-wide CRISPR/Cas9 knock-out screens. However, the historical lines have large gaps in their representation of the diversity of human cancer, and they may lack physiological relevance given their optimization for rapid proliferation. Next generation HCMI models address these concerns, but will require the development of new methods to make them useful. Specifically, standard approaches to genome editing (involving first creating Cas9- stably expressing lines and then introducing guide RNAs) will not work for slowly proliferating cells often growing in 3D. We will therefore develop all-in-one genome editing vector systems that will make it possible to bring the power of genome editing to HCMI models. In addition, standard viability read-outs of such “drop-out” screens involve the growth of cells over many population doublings. But for slowly proliferating HCMI models, alternative readouts will be required for efficient screening. We will therefore develop short-term single cell RNA sequencing (scRNAseq) methods that will serve as surrogate read-outs for long-term viability. Given the clinical annotation associated with HCMI models, there is also enormous opportunity to expand the use of these models beyond viability measures to more complex, physiologically relevant phenotypes such as organ-specific metastatic potential. We will therefore develop methods that make it possible to determine the metastatic potential for next generation cancer models, and we will create a public resource of the metastasis map (MetMap) for at least 50 HCMI models. All data and protocols will be made publicly available without restriction, all reagents will be made available via Addgene, and all modified models made available to ATCC for distribution. Importantly, throughout the project, all cell models will be rigorously monitored for evidence of genetic and epigenetic drift. At the conclusion of the proposed project, we expect to have generated a set of tools and data that will help propel the future of cancer precision medicine based on next generation cancer models.

抽象的人类癌症模型计划 (HCMI) 正在创建下一代癌症模型，推动未来发展癌症精准医学研究的历史癌细胞系因其快速增殖而被选择。在组织培养塑料上，这使得它们适合高通量筛选，例如全基因组筛选然而，CRISPR/Cas9 敲除筛选的历史线在其代表性方面存在很大差距。人类癌症的多样性，并且考虑到它们对快速的优化，它们可能缺乏生理相关性下一代 HCMI 模型可以解决这些问题，但需要开发新的模型。具体来说，是基因组编辑的标准方法（包括首先创建 Cas9-）。稳定表达细胞系，然后引入向导RNA）对于经常生长的缓慢增殖细胞不起作用因此，我们将开发一体化基因组编辑载体系统，使之成为可能。 HCMI 模型的基因组编辑的力量此外，这种“退出”屏幕的标准活力读数。涉及多次群体倍增的细胞生长，但对于缓慢增殖的 HCMI 模型，可以采用替代方法。因此，我们将开发短期单细胞 RNA 测序。 (scRNAseq) 方法将作为长期可行性的替代读数。与 HCMI 模型相关，还有巨大的机会将这些模型的使用扩展到其他领域对更复杂、生理相关表型（例如器官特异性转移）的活力测量因此，我们将开发能够确定下一步转移潜力的方法。生成癌症模型，我们将创建至少 50 个转移图谱（MetMap）的公共资源 HCMI 模型将不受限制地公开，所有试剂均将公开。重要的是，所有修改后的模型均可通过 Addgene 提供给 ATCC 进行分发。在该项目中，所有细胞模型都将受到严格监测，以获取遗传和表观遗传漂移的证据。拟议项目结束后，我们预计将生成一套工具和数据，以帮助推动基于下一代癌症模型的癌症精准医学的未来。