A Novel Anti-Cancer Therapy: CRISPR/Cas9 Gene Editing

Abstract

Cancer becomes one of the main causes of human deaths in the world due to the high incidence and mortality rate and produces serious economic burdens. With more and more attention is paid on cancer, its therapies are getting more of a concern. Previous research has shown that the occurrence, progression, and treatment prognosis of malignant tumors are closely related to genetic and gene mutation. CRISPR/Cas9 has emerged as a powerful method for making changes to the genome, which has extensively been applied in various cell lines. Establishing the cell and animal models by CRISPR/Cas9 laid the foundation for the clinical trials which possibly treated the tumor. CRISPR-Cas9-mediated genome editing technology brings a great promise for inhibiting migration, invasion, and even treatment of tumor. However, the potential off-target effect limits its clinical application, and the effective ethical review is necessary. The article reviews the molecular mechanisms of CRISPR/Cas9 and discusses the research and the limitation related to cancer clinical trials.

Keywords: CRISPR/Cas9; anti-cancer therapy; ethics; gene editing technology; off-target effect.

FIGURE 1

CRISPR/Cas9 system and DSB repair. (A) Mechanism of the CRISPR-Cas9 system. A short guide sgRNA associates with the Cas9 endonuclease to form the Cas9-sgRNA complex. Cas9 is a targeted DNA by PAM under the sgRNA. (B) cNHEJ : cNHEJ was the predominant pathway for repairing DSBs and was used for re-ligating broken DNA ends. Deletions or insertion mutations lead to gene frame shift mutations or premature generation of stop codes. (C) HDR: HDR was used a homologous DNA template to guide DSB repair. The DNA donor templates of HDR were used to insert or replace specific sequences into the genome. (D) MMEJ: MMEJ-mediated repair was capable of generating precise deletions between two short micro-homologous sequences (5–25 base pairs) at target loci.

FIGURE 2

Methods of CRISPR/Cas9 technology to construct animal models. (A) Editing gene in embryos. Collecting the zygote that was injected with Cas9 mRNA, sgRNA, and DNA. After the zygote develops into an embryo, it would be transferred into the animals to produce generation. (B) Editing gene in haploid ESCs and SSCs: haploid embryonic stem cells (ESCs) are pluripotent cells generated from oocytes. CRISPR/Cas9-mediated gene editing in ESCs and autologous transplant to pseudo-pregnant animals to produce generation. Spermatogonia stem cells (SSCs) would be transfected by RNP and autologous transplant to seminiferous tubule to produce the generation.