An Efficient Antiviral Strategy for Targeting Hepatitis B Virus Genome Using Transcription Activator-Like Effector Nucleases

Abstract

The hepatitis B virus (HBV) is a DNA virus that can cause chronic hepatitis B (CHB) in humans. Current therapies for CHB infection are limited in efficacy and do not target the pre-existing viral genomic DNA, which are present in the nucleus as a covalently closed circular DNA (cccDNA) form. The transcription activator-like (TAL) effector nucleases (TALENs) are newly developed enzymes that can cleave sequence-specific DNA targets. Here, TALENs targeting the conserved regions of the viral genomic DNA among different HBV genotypes were constructed. The expression of TALENs in Huh7 cells transfected with monomeric linear full-length HBV DNA significantly reduced the viral production of HBeAg, HBsAg, HBcAg, and pgRNA, resulted in a decreased cccDNA level and misrepaired cccDNAs without apparent cytotoxic effects. The anti-HBV effect of TALENs was further demonstrated in a hydrodynamic injection-based mouse model. In addition, an enhanced antiviral effect with combinations of TALENs and interferon-α (IFN-α) treatment was observed and expression of TALENs restored HBV suppressed IFN-stimulated response element-directed transcription. Taken together, these data indicate that TALENs can specifically target and successfully inactivate the HBV genome and are potently synergistic with IFN-α, thus providing a potential therapeutic strategy for treating CHB infection.

Transcription Activator-Like Effector (TALE) Nucleases and Repressor TALEs for Antiviral Gene Therapy

“…Cleavage of the HBV DNA also reduced the number of circulating viral particles, indicating disruption of HBV replication. Later in 2013, these results were supported by Chen and colleagues [23], who also showed that TALENs designed to bind to the C and polymerase (P) open reading frames reduced viral antigen and pregenomic RNA expression in vitro and in vivo. They found that the antiviral effect was not limited to a single genotype, suggesting that a TALEN pair may be used to target multiple genotypes.…”

Genome editing and the next generation of antiviral therapy

“…Since this first study, numerous other groups have employed similar strategies to target HBV. The first follow-up studies showed antiviral activity in vitro by ZFNs delivered using adeno-associated virus (AAV) vectors (Weber et al 2014b), or by TALENs in vitro and in a mouse hydrodynamic injection model of HBV lacking cccDNA (Bloom et al 2013; Chen et al 2014). Importantly, the in vitro studies using TALENs showed for the first time that HBV cccDNA could be cleaved and disrupted directly.…”

New horizon for radical cure of chronic hepatitis B virus infection

“…In addition, several genome editing technologies have been developed to silence sequencespecific cle avage of cccDNA. These include zinc finger nucleases (ZFNs) [100,101] , transcription activatorlike effector nucleases (TALENs) [102,103] , and the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR associated system (Cas). These sequence-specific genome editing technologies could induce doublestranded breaks at certain DNA sites.…”