Progress of delivery methods for CRISPR-Cas9

ABSTRACT

Introduction

Gene therapy is becoming increasingly common in clinical practice, giving hope for the correction of a wide range of human diseases and defects. The CRISPR/Cas9 system, consisting of the Cas9 nuclease and single-guide RNA (sgRNA), has revolutionized the field of gene editing. However, efficiently delivering the CRISPR-Cas9 to the target organ or cell remains a significant challenge. In recent years, with rapid advances in nanoscience, materials science, and medicine, researchers have developed various technologies that can deliver CRISPR-Cas9 in different forms for in vitro and in vivo gene editing. Here, we review the development of the CRISPR-Cas9 and describe the delivery forms and the vectors that have emerged in CRISPR-Cas9 delivery, summarizing the key barriers and the promising strategies that vectors currently face in delivering the CRISPR-Cas9.

Areas covered

With the rapid development of CRISPR-Cas9, delivery methods are becoming increasingly important in the in vivo delivery of CRISPR-Cas9.

Expert opinion

CRISPR-Cas9 is becoming increasingly common in clinical trials. However, the complex nuclease and protease environment is a tremendous challenge for in vivo clinical applications. Therefore, the development of delivery methods is highly likely to take the application of CRISPR-Cas9 technology to another level.

KEYWORDS:

• CRISPR
• Cas9
• gene editing
• drug delivery
• review

Article highlights

• CRISPR-Cas9 technology offers a powerful capability to manipulate genomic sequences and regulate gene expression, making it ideal for treating various human diseases and defects.

• The CRISPR-Cas9 system can be delivered in three formats, and the delivery technologies for the CRISPR-Cas9 system can be divided into three main categories: physical delivery, viral vectors, and non-viral vectors.

• Physical methods include microinjection, electroporation, hydrodynamic tail vein injection (HTVI), ultrasonic microbubbles, and laser. Physical methods for in vivo delivery of CRISPR-Cas9 have many limitations and are more often used in laboratory studies and in vitro gene editing.

• The main advantage of delivering the CRISPR-Cas9 system via viral vectors is the high efficiency in gene editing. However, viral vectors have potentially oncogenic properties, and immunogenicity can lead to insertional mutations. It is also difficult to achieve precisely targeted delivery with viral vectors.

• Non-viral vectors have high CRISPR-Cas9 encapsulation capacity, low immunogenicity, and are easy to assemble. Although they generally have less efficiency in gene editing than viral vectors, they are the most likely mode of delivery for future generations of in vivo CRISPR-Cas9 delivery.

Declaration of interest

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

Reviewer disclosures

Peer reviewers on this manuscript have no relevant financial or other relationships to disclose.

Additional information

Funding

The authors were supported by the National Key Research and Development Program of China (2020YFA0908200), National Natural Science Foundation of China General Program (81871472); Research Fellow (Grant No. 328933), Solution for Health Profile (336355), and InFLAMES Flagship (337531) grants from Academy of Finland; Finland China Food and Health International Pilot Project funded by the Finnish Ministry of Education and Culture; and Guangci Professorship Program of Ruijin Hospital Shanghai Jiao Tong University School of Medicine.