基于CRISPR/Cas工具的肿瘤基因线路构建及应用

宋斐; 刘宇辰; 蔡志明; 黄卫人

doi:10.12211/2096-8280.2021-046

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基于CRISPR/Cas工具的肿瘤基因线路构建及应用

特约评述 | 更新时间：2025-03-20

- 基于CRISPR/Cas工具的肿瘤基因线路构建及应用
- Construction of tumor gene circuits using CRISPR/Cas tool and their applications
- 合成生物学 2022年3卷第1期页码：53-65
- 作者机构：
  
  1.深圳大学第一附属医院，深圳市第二人民医院，泌尿外科，广东深圳 518035
  2.广东省泌尿生殖肿瘤系统生物学与合成生物学重点实验室，广东深圳 518035
- 作者简介：
  
  [ "宋斐（1979—），女，副研究员，硕士生导师。E-mail：lst121@outlook.com" ]
  [ "黄卫人（1980—），男，研究员，博士生导师。主要研究方向：（1）肿瘤基因组学，应用多组学手段鉴定肿瘤及微环境诊疗标志物，开发相关临床应用；（2）肿瘤类器官，利用体外培养系统还原肿瘤体内生长，药物筛选及耐药机制研究；（3）医学合成生物学，创新肿瘤治疗新方法。E-mail：pony8980@163.com" ]
- 基金信息：
  
  国家重点研发项目(2019YFA0906000);广东省自然科学基金(2021A1515012488)
- DOI：10.12211/2096-8280.2021-046
  中图分类号： Q812
- 收稿：2021-04-15，
  
  修回：2021-11-24，
  
  纸质出版：2022-02-28
- 稿件说明：
移动端阅览
宋斐，刘宇辰，蔡志明，黄卫人. 基于CRISPR/Cas工具的肿瘤基因线路构建及应用［J］. 合成生物学， 2022， 3（1）： 53-65

SONG Fei， LIU Yuchen， CAI Zhiming， HUANG Weiren. Construction of tumor gene circuits using CRISPR/Cas tool and their applications［J］. Synthetic Biology Journal， 2022， 3（1）： 53-65
宋斐，刘宇辰，蔡志明，黄卫人. 基于CRISPR/Cas工具的肿瘤基因线路构建及应用［J］. 合成生物学， 2022， 3（1）： 53-65 DOI： 10.12211/2096-8280.2021-046.

SONG Fei， LIU Yuchen， CAI Zhiming， HUANG Weiren. Construction of tumor gene circuits using CRISPR/Cas tool and their applications［J］. Synthetic Biology Journal， 2022， 3（1）： 53-65 DOI： 10.12211/2096-8280.2021-046.

摘要

以恶性肿瘤为代表的难治性疾病仍是困扰我国乃至全球的一个重大公共卫生问题。近年来，合成生物学的蓬勃发展，极大推动了疾病创新治疗策略，并显示出巨大潜力。人工构建的基因线路可特异性地识别、区分疾病细胞和正常细胞，并控制疾病细胞命运，为精准治疗提供了新途径。然而构建基因线路用于疾病治疗的挑战之一是缺乏有效、可编程、安全和序列特异性的基因编辑工具。CRISPR/Cas自从首次被用于哺乳动物基因编辑以来，已被广泛应用于研究、工业和医学领域。除Cas9诱导的indel突变外，CRISPR/Cas能够进行DNA/RNA的碱基编辑，为基因线路设计提供了高效的工具，极大提高了每个线路元件效率。因此，基于CRISPR技术的智能化基因线路的应用，可有效保证癌症等疾病治疗的安全性、高效性和特异性。本文介绍了CRISPR/Cas技术应用于医学合成生物学基因线路设计的研究进展和潜在挑战。评估了使用CRISPR系统元件的合成基因线路在肿瘤治疗中的效果，尤其利用人工开关诱导的Cas9干预肿瘤细胞治疗的安全性和有效性；介绍了CRISPR/Cas9介导的信号传感器设计，其可同时识别多个蛋白质信号，实现了多基因同步调控，以及新一代体系小、特异性强、基因调控效率高的CRISPR/Cas12a系统及其遗传改造。基于无启动子表达CRISPReader元件的精简基因线路设计，其高效启动的特点极大提升了智能化基因线路在未来精准癌症治疗中的应用潜力。

Abstract

Refractory diseases such as malignant tumors are still a major challenge for public health in the world. In recent years

the fast development of synthetic biology has greatly promoted novel treatment strategies for diseases with great potentials for effective treatment. The artificially constructed gene circuits can specifically identify and distinguish diseased cells from normal cells

and control their fate to provide a new way for precise treatment. However

the ultimate challenge in constructing gene circuits is the lack of effective

programmable

safe

and sequence-specific gene editing tools. The clustered regularly interspaced short palindromic repeat (CRISPR) system and CRISPR-associated RNA-guided endonuclease Cas9-targeted (CRISPR-associated protein 9) genome editing tool have recently been applied in engineering gene circuits for its unique properties: manipulability

high efficiency and programmability. In addition to indel mutations induced by Cas9

CRISPR/Cas can perform DNA/RNA base editing

providing efficient tools for gene circuit design

and greatly improve efficiency for designing circuit elements. The traditional single-targeted treatment cannot effectively distinguish tumor cells from normal cells

and gene therapy has poor anti-tumor effects

which severely limits its application. Currently

the design of gene circuits using tumor-specific targets based on CRISPR/Cas systems provides a new strategy for precision cancer therapy. Scientists have developed a series of efficient and targeted transcription factor components based on CRISPR technology to maximize the performance of gene circuits. These novel designs extend the toolbox for gene editing

and enable the construction of intelligent gene circuits such as logic gates

signal conductors

analogue computing circuits

counters and memory devices. Therefore

the application of intelligent gene circuits based on CRISPR technology can effectively ensure safety

efficiency and specificity for cancer treatment. This article introduces the updated progress

prospects and potential challenges of CRISPR/Cas technology for the design and construction of gene circuits in biomedical field. Firstly

the efficacy of synthetic gene circuits using CRISPR system components in tumor treatment is evaluated

especially the safety and effectiveness of using Cas9 induced by artificial switch to intervene tumor growth. Secondly

the design of CRISPR/Cas9-mediated signal conductors is introduced

which can recognize multiple protein signals simultaneously and realize the simultaneous regulation of multiple genes. In addition

the CRISPR/Cas12a system with smaller size

strong specificity and high gene regulation efficiency is a new generation alternative. Finally

the simplified gene circuit design based on the promoter-free CRISPReader expression elements is addressed

and its high-efficiency startup feature will greatly enhance the potential application of intelligent gene circuits in precise cancer treatments in the future.

关键词

Keywords

references

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CRISPR-Cas9系统在肿瘤生物学中的应用及前景