基因工程菌的构建、性能研究及其在疾病诊断和治疗中的应用

随着肠道微生物组与宿主关系研究的深入和基因工程的迅速发展,基因工程菌（genetically engineered bacteria,GEB）在医学领域的应用成为研究热点。GEB是指经基因工程改造而具有高效表达外源蛋白质或分子化合物能力的细菌,相比传统药物具有诸多优势。GEB的构建过程包括底盘的选择、功能基因的获取、基因转移和重组这几个基本步骤,包裹技术能够提高其存活率和定殖能力,合成基因回路的应用可使其智能化。功能稳定性、有效性和安全性是评价GEB的一般指标,也是性能优化过程中需要重点关注的方面。GEB在炎症性疾病、肿瘤、代谢性疾病、感染性疾病和神经系统疾病等疾病中已有广泛应用,但实现临床转化还有很多问题亟待解决。本文介绍了GEB的构建和性能研究方法,总结了近年来其在疾病诊断和治疗中的应用,指出了现存问题并提出了展望。

Genetically engineered bacteria: construction, performance evaluation, and applications in disease diagnosis and treatment

With the in-depth study of the relationship between intestinal microbiome and host and the rapid development of genetic engineering, the application of genetically engineered bacteria (GEB) in the medical field has become a research hotspot. GEB refer to the bacteria that have been genetically engineered to efficiently express exogenous proteins or compounds to achieve specific goals. Compared with traditional drugs, GEB have a variety of advantages. The construction process of GEB includes the selection of chassis, the acquisition of functional genes, and gene transfer and recombination. At present, the bacteria serving as GEB chassis can be classified into two categories:generally regarded as safe (GRAS) strains and commensal strains. The application of multi-omics facilitates the selection of chassis. Functional genes can be obtained by PCR, DNA synthesis, CRISPR-Cas9 or Red/ET recombination system according to their sizes. Heat shock and electroporation are widely used for plasmid transfer in bacteria. Homologous recombination can directly integrate the target gene into the host''s chromosome. The encapsulation technology can improve GEB''s survival rate and colonization ability, and synthetic gene circuits can make GEB intelligent. Functional stability, effectiveness, and safety are the general indicators for evaluating GEB. The instability of plasmid is an inherent defect of the GEB constructed by plasmid-mediated gene transfer, while the GEB constructed by gene integration have strong stability. In addition, to achieve long-term stable expression of functional genes, researchers need to evaluate and alleviate the impact of load. The efficacy and safety of GEB need to be evaluated in vitro, in animal models and clinical trials. There are some methods that have successfully achieved optimization of the above indicators. GEB have been widely used in the diagnosis and treatment of inflammatory diseases, tumors, metabolic diseases, infectious diseases, neurological diseases, and other diseases, playing a unique role. However, there are still problems regarding construction methods, performance evaluation and optimization, and large-scale production, which limit the clinical application. In this paper, we introduced the methods for construction and performance evaluation of GEB, summarized the application in disease diagnosis and treatment in recent years, pointed out the existing problems, and prospected the development of this field in the future.