Precision targeting of bladder cancer using CRISPR technology

利用 CRISPR 技术精确靶向膀胱癌

• 批准号: 10289763
• 负责人: MATS LJUNGMAN
• 金额: $ 18.23万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10289763
• 关键词: Automobile Driving; Benchmarking; Bioluminescence; CRISPR/Cas technology; Cancer Model; Cancer cell line; Cells; Chemistry; Chromosome Pairing; Chromosomes; Clinic; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Colon Carcinoma; Complex; DNA Double Strand Break; DNA Sequence Alteration; Data Set; Development; Drug Design; Drug Targeting; Elements; Encapsulated; Gene Dosage; Generations; Growth; Guide RNA; In Vitro; Malignant Neoplasms; Malignant neoplasm of urinary bladder; Monitor; Mus; Oncogenes; Oncogenic; Pathway interactions; Pharmaceutical Preparations; Precision therapeutics; Proteins; RNA; Reagent; Site; System; Technology; Testing; Therapeutic; Time; Tumor Biology; Tumor Burden; Tumor Suppressor Genes; Xenograft procedure; base; cancer cell; cancer type; carcinogenesis; design; endonuclease; genome sequencing; in vivo; lipid nanoparticle; mouse model; multidisciplinary; nanoparticle; nanoparticle delivery; neoplastic cell; novel; novel strategies; personalized approach; pre-clinical; protein complex; targeted cancer therapy; tumor; whole genome

ABSTRACT Chromosome rearrangements are common in cancers and typically occur early in carcinogenesis. They drive tumor development by altering the expression or function of oncogenes and tumor suppressor genes by copy number alterations, formation of oncogenic fusions or by alterations of DNA elements responsible for regulating cancer genes. Despite being a well-known feature of cancer, discovered over hundred years ago, this common cancer hallmark has not yet been therapeutically exploited. Now, for the first time, technological advancements in whole genome sequencing and the development of CRISPR technologies for precision targeting makes it possible to identify and target chromosome rearrangement junctions (CRJs) in tumor cells. In addition, recent breakthroughs in in vivo delivery of CRIPR reagents involving nanoparticles will open new avenues into clinical utilization. In this R21 application, we present a precision CRISPR approach that aims at targeting cancer-specific CRJs as an untapped Achilles heel common to all tumors. This CRISPR-based precision approach consists of pairs of CRJ-targeting guide RNAs (gRNAs) that bring together two parts of a dCas9-conjugated endonuclease, Fok1, leading to its activation and the induction of lethal DNA double strand breaks (DSBs) specifically in cancer cells. Regardless of the tumor-driving mechanism, the presence of CRJs will be exploited to selectively force these cancer cells to die. We have obtained proof-of-principle of the successful targeting of CRJs with this approach in both colon and bladder cancer cell lines. We have assembled a multi- disciplinary team with expertise in CRISPR technology and design, lipid nanoparticle delivery systems and bladder cancer mouse models. With this team in place and with strong evidence of efficient targeting of CRJs with Fok1-dCas9 in cancer cell lines, we are now primed to develop and test our approach in pre-clinical cancer models using recently developed breakthrough lipid nanoparticle delivery technologies. If successfully implemented, this paradigm-shifting precision- targeting cancer therapeutic platform could be transformative in designing uniform treatments for all cancer types irrespective of the biology of the tumor.

摘要 染色体重排在癌症中很常见，通常发生在癌发生的早期。 它们通过改变癌基因和肿瘤细胞的表达或功能来驱动肿瘤的发展。 抑制基因的拷贝数改变，形成致癌融合或改变 负责调节癌症基因的DNA元件。尽管这是一个众所周知的特点， 癌症，发现超过一百年前，这种常见的癌症标志还没有 治疗性剥削现在，第一次，整个基因组的技术进步 测序和CRISPR技术的发展使其成为精确靶向的技术， 可以识别和靶向肿瘤细胞中的染色体重排连接（CRJ）。在 此外，最近在涉及纳米颗粒的CRIPR试剂的体内递送方面的突破将 为临床应用开辟了新的途径。在这个R21应用中，我们提出了一种精确的CRISPR， 该方法旨在针对癌症特异性CRJ，将其作为癌症常见的未开发的阿喀琉斯之踵 所有肿瘤这种基于CRISPR的精确方法由成对的CRJ靶向指导RNA组成 （gRNA）将dCas 9缀合的内切核酸酶Fok 1的两个部分结合在一起，导致其 激活和诱导致死性DNA双链断裂（DSB），特别是在癌细胞中。 无论肿瘤驱动机制如何，CRJ的存在将被利用来选择性地 迫使癌细胞死亡我们已经获得了成功瞄准的原理证明， 在结肠癌和膀胱癌细胞系中使用这种方法的CRJ。我们已经建立了一个多- 拥有CRISPR技术和设计、脂质纳米粒递送等专业知识的学科团队 系统和膀胱癌小鼠模型。有了这个团队，有了强有力的证据， 在癌细胞系中用Fok 1-dCas 9有效靶向CRJ，我们现在准备开发 并使用最近开发的突破性脂质在临床前癌症模型中测试我们的方法 纳米粒子输送技术。如果成功实施，这种范式转换的精度- 靶向癌症治疗平台可能在设计统一的治疗方法方面具有变革性， 所有癌症类型，无论肿瘤的生物学。