CRISPR/Cas9 cleavage of viral DNA efficiently suppresses hepatitis B virus

Ramanan, Vyas; Shlomai, Amir; Cox, David Benjamin Turitz; Schwartz, Robert E.; Michailidis, Eleftherios; Bhatta, Ankit; Scott, David; Zhang, Feng; Rice, Charles M.; Bhatia, Sangeeta N.

doi:10.1038/srep10833

Cited by 270 publications

(213 citation statements)

References 29 publications

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“…We found that Cas9 (0.56 mg/kg) and sg21(0.25 mg/ kg) significantly decreased all measurements of HBV viral loads, including liver and serum HBsAg levels, liver and serum HBV e antigen (HBeAg) levels, and liver HBV RNA and DNA levels, when compared with sgGFP ( Figure 1F). The efficacy of Cas9+sg21 LLNs in reducing HBV viral loads is comparable to a previous study using hydrodynamic injection of a plasmid encoding Cas9 and sg21 [11]. sgB5 identified by our screening had a significant effect in 4 out of the 6 measurements ( Figure 1F).…”

Section: Dear Editorsupporting

confidence: 70%

“…Several previous studies have explored the potential of the CRISPR/Cas9 system to target HBV cccDNA in mice by hydrodynamic injection of plasmid DNA [7][8][9][10][11]; however, therapeutically relevant in vivo delivery methods are still lacking [12]. To systematically identify potential sgRNAs targeting HBV DNA, we used the CRISPR DESIGN tool (http:// crispr.mit.edu) to design potential sgRNAs.…”

Section: Dear Editormentioning

confidence: 99%

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A non-viral CRISPR/Cas9 delivery system for therapeutically targeting HBV DNA and pcsk9 in vivo

et al. 2017

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Section: Dear Editorsupporting

confidence: 70%

Section: Dear Editormentioning

confidence: 99%

A non-viral CRISPR/Cas9 delivery system for therapeutically targeting HBV DNA and pcsk9 in vivo

et al. 2017

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“…The dCas9 has been used for transcriptional activation and repression [134][135][136][137], for localizing fluorescent protein labels [138], and for recruiting histone modifying enzymes [139,140]. Other applications of Cas9 include building gene circuits [141][142][143], creating new anti-microbials [144], antivirals [145][146][147], and large-scale gain-and loss-of-function screening [148][149][150][151] (Table 1).…”

Section: Genome Editing Applicationsmentioning

confidence: 99%

Diverse evolutionary roots and mechanistic variations of the CRISPR-Cas systems

Mohanraju

Makarova

Zetsche

et al. 2016

Science

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Background: Prokaryotes have evolved multiple systems to combat invaders such as viruses and plasmids. Examples of such defence systems include receptor masking, restriction-modification (R-M systems), DNA interference (Argonaute), bacteriophage exclusion (BREX or PGL) and abortive infection, all of which act in an innate, non-specific manner. In addition, prokaryotes have evolved adaptive, heritable immune systems, i.e. clustered regularly interspaced palindromic repeats (CRISPR) and the CRISPR-associated proteins (CRISPR-Cas). Adaptive immunity is conferred by the integration of DNA sequences from an invading element into the CRISPR array (adaptation), which is transcribed into long pre-CRISPR (pre-cr) RNAs and processed into short crRNAs (expression), which guide Cas proteins to specifically degrade the cognate DNA on subsequent exposures (interference). Advances:A plethora of distinct CRISPR-Cas systems are represented in genomes of most archaea and almost half of the bacteria. The latest CRISPR-Cas classification scheme delineates two classes that are each subdivided into three types. Integration of biochemistry and molecular genetics has contributed significantly to revealing many of the unique features of the variant CRISPR-Cas types. Additionally, structural analysis and single molecule studies have further advanced our understanding of the molecular basis of CRISPR-Cas functionality. Recent progress includes relevant steps in the adaptation stage, when fragments of foreign DNA are processed and incorporated as new spacers into the CRISPR array. In addition, three novel CRISPR-Cas types (IV, V, and VI) have been identified, and in particular, the type V interference complexes have been experimentally characterized. Moreover, the ability to easily program sequence-specific DNA targeting and cleavage by CRISPRCas components, as demonstrated for Cas9 and Cpf1, allows for the application of CRISPR-Cas components as highly effective tools for genetic engineering and gene regulation in a wide range of eukaryotes and prokaryotes.The pressing issue of off-target cleavage by the Cas9 nuclease is being actively addressed using structure-guided engineering.Outlook: Although our understanding of the CRISPR-Cas system has increased tremendously over the past few years, much remains to be done. About the evolution of CRISPR-Cas systems, the continuing discovery of novel CRISPR-Cas variants will provide direct tests of the recently proposed modular scenario. The recent discovery and characterization of new CRISPR-Cas types with many unique features implies that our current knowledge has relatively limited power for predicting the functional details of distantly related variants. Hence, newly discovered CRISPR-Cas systems need to be thoroughly dissected with the aforementioned multi-disciplinary approaches to gain insight in their biological role, to unravel their molecular mechanism, and to harness their potential for biotechnology. As to the biology, key outstanding questions include the ecological roles of microbial ...

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“…Researchers have shown that DNA plasmids encoding Cas9 and sgRNA can be delivered to the livers of mice infected with hepatitis B virus (HBV) via a hydrodynamic tail vein injection. The resulting Cas9:sgRNA complexes cleave HBV viral DNA, resulting in a gradual decrease in HBV expression in the animals (Lin et al, 2014a;Zhen et al, 2015;Ramanan et al, 2015). This technique has also been used to generate mouse liver cancer models.…”

Section: Delivery Of Genome-editing and Epigenome-editing Agentsmentioning

confidence: 99%

CRISPR-Based Technologies for the Manipulation of Eukaryotic Genomes

Komor¹,

Badran²,

Liu³

2017

Cell

270

214

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The CRISPR-Cas9 RNA-guided DNA endonuclease has contributed to an explosion of advances in the life sciences that have grown from the ability to edit genomes within living cells. In this review we summarize CRISPR-based technologies that enable mammalian genome editing and their various applications. We describe recent developments that extend the generality, DNA specificity, product selectivity, and fundamental capabilities of natural CRISPR systems, and some of the remarkable advancements in basic research, biotechnology, and therapeutics development that these developments have facilitated.

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CRISPR/Cas9 cleavage of viral DNA efficiently suppresses hepatitis B virus

Cited by 270 publications

References 29 publications

A non-viral CRISPR/Cas9 delivery system for therapeutically targeting HBV DNA and pcsk9 in vivo

A non-viral CRISPR/Cas9 delivery system for therapeutically targeting HBV DNA and pcsk9 in vivo

Diverse evolutionary roots and mechanistic variations of the CRISPR-Cas systems

CRISPR-Based Technologies for the Manipulation of Eukaryotic Genomes

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