Magnetic‐Activated Nanosystem with Liver‐Specific CRISPR Nonviral Vector to Achieve Spatiotemporal Liver Genome Editing as Hepatitis B Therapeutics

• Published in: Advanced functional materials Vol. 33; no. 7
• Main Authors: Zhuo, Chenya, Kong, Huiming, Yi, Ke, Chi, Chun‐Wei, Zhang, Jiabing, Chen, Ran, Wang, Haixia, Wu, Caixia, Lao, Yeh‐Hsing, Tao, Yu, Li, Mingqiang
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc 01.02.2023
• Subjects: CRISPR; CRISPR/Cas9; Editing; gene editing; Genetic modification; Genomes; Hepatitis B; hepatitis B virus (HBV); Interferon; Liver; liver‐specific promoters; magnetic nanosystems; Materials science; Nanoparticles; Polyethyleneimine
• DOI: 10.1002/adfm.202210860
• ISSN: 1616-301X

Chronic hepatitis B infection remains incurable due to the stable presence of various forms of hepatitis B virus (HBV) genome, especially the HBV covalently closed circular DNA (cccDNA). The emergence of clustered regularly interspaced short palindromic repeat (CRISPR) technology provides a new opportunity to potentially cure the HBV infection. However, the efficiency and specificity remain unsatisfactory, especially for nonviral CRISPR/Cas9 delivery. To tackle these, a liver‐specific CRISPR/Cas9 magnetic nanosystem FMNPpAG333/sgXPP is constructed based on fluorinated polyethylenimine‐coated magnetic nanoparticles and liver‐specific ApoE.HCR.hAAT promoter‐driven Cas9‐T2A‐EGFP plasmid with dual sgRNAs. The elaborate system enables magnetic field‐induced spatially specific distribution and hepatocyte‐specific promoter‐driven liver‐specific gene editing. Moreover, this CRISPR/Cas9 magnetic nanosystem is designed to disrupt the two conserved sites in X opening reading frame and Pol opening reading frame of the HBV genome, thereby significantly inactivating the HBV genome without showing off‐target effects. Treatment with FMNPpAG333/sgXPP for 7 days reduces serum HBsAg levels by 76% with a total editing efficiency of ≈20% in the two conserved sites. Collectively, this study demonstrates spatiotemporal liver genome editing as well as the feasibility of applying a nonviral CRISPR/Cas9 vector for HBV treatment, which may open up a new application for CRISPR therapeutics. A liver‐specific CRISPR/Cas9 magnetic nanosystem, FMNPpAG333/sgXPP, can enhance liver accumulation under the magnetic activation and improve the transfection efficiency, while the liver‐specific promoter further confines the Cas9 expression in the hepatocyte, which offers an additional specificity at the cellular level. This nanosystem represents an in vivo spatiotemporal liver genome editing approach as a HBV CRISPR therapeutic.