Nanoparticle delivery of Cas9 ribonucleoprotein and donor DNA in vivo induces homology-directed DNA repair

Lee, Kunwoo; Conboy, Michael J.; Park, Hyo Min; Jiang, Fuguo; Kim, Hyun Jin; Dewitt, Mark; Mackley, Vanessa A; Chang, Kevin; Rao, Anirudh; Skinner, Colin M.; Shobha, Tamanna; Mehdipour, Melod; Liu, Hui; Huang, Weijie; Lan, Freeman; Bray, Nicolas; Li, Song; Corn, Jacob E.; Kataoka, Kazunori; Doudna, Jennifer A.; Conboy, Irina M.; Murthy, Niren

doi:10.1038/s41551-017-0137-2

Cited by 585 publications

(487 citation statements)

References 45 publications

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“…Recently, AuNPs were functionalized with arginate that could bind oligo-glutamic acid tagged protein. [166,167] The arginine AuNPs can self-assemble with glutamate residues/NLS modified Cas9 RNP to form nanoassemblies through the charge interaction. In addition, the engineered proteins Cre recombinase and granzyme A could reserve bioactivity after cytosolic delivery.…”

Section: Metal-based Nanoparticlesmentioning

confidence: 99%

“…In addition, the engineered proteins Cre recombinase and granzyme A could reserve bioactivity after cytosolic delivery. [167] Both kinds of AuNPs could successfully deliver Cas9 RNP into the cytoplasm and induce gene editing in vitro or in vivo. To improve cellular uptaken efficiency and avoid endosomal sequestration, Rotello and co-workers designed a serious AuNP-stabilized capsule for effective protein delivery.…”

Section: Metal-based Nanoparticlesmentioning

confidence: 99%

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Rational Design of Nanocarriers for Intracellular Protein Delivery

et al. 2019

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Protein/antibody therapeutics have exhibited the advantages of high specificity and activity even at an extremely low concentration compared to small molecule drugs. However, they are accompanied by unfavorable physicochemical properties such as fragile tertiary structure, large molecular size, and poor penetration of the membrane, and thus the clinical use of protein drugs is hindered by inefficient delivery of proteins into the host cells. To overcome the challenges associated with protein therapeutics and enhance their biopharmaceutical applications, various protein‐loaded nanocarriers with desired functions, such as lipid nanocapsules, polymeric nanoparticles, inorganic nanoparticles, and peptides, are developed. In this review, the different strategies for intracellular delivery of proteins are comprehensively summarized. Their designed routes, mechanisms of action, and potential therapeutics in live cells or in vivo are discussed in detail. Furthermore, the perspective on the new generation of delivery systems toward the emerging area of protein‐based therapeutics is presented as well.

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Section: Metal-based Nanoparticlesmentioning

confidence: 99%

Section: Metal-based Nanoparticlesmentioning

confidence: 99%

Rational Design of Nanocarriers for Intracellular Protein Delivery

et al. 2019

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“…For example, DNA-conjugated gold NPs complexed with cationic polymers to deliver donor DNA and Cas9 RNP have been used to treat Duchenne muscular dystrophy (DMD). After intramuscular injection alongside cardiotoxin, these NPs corrected the DMD-causing mutation at a frequency of 5.4% with reduced muscle fibrosis in a DMD mouse model [30]. Gao et al used the Cas9 system for the treatment of autosomal dominant hearing loss using cationic lipids.…”

Section: Local Delivery In Vivomentioning

confidence: 99%

Debugging the genetic code: Non-viral in vivo delivery of therapeutic genome editing technologies

Piotrowski-Daspit

Glazer

Saltzman

2018

Current Opinion in Biomedical Engineering

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Efforts to precisely correct genomic mutations that underlie hereditary diseases for therapeutic benefit have advanced alongside the emergence and improvement of genome engineering technologies. These methods can be divided into two main classes: active nucleasebased platforms including the popular CRISPR/Cas9 system and oligo/polynucleotide strategies including triplex-forming oligonucleotides (TFOs), such as peptide nucleic acids (PNAs). These technologies have been successful in cell culture and in animals, but important challenges remain before these tools can be translated into the clinic; they must be effectively delivered to and taken up by specific cell types of interest, achieve correction levels in target cells that significantly ameliorate the disease phenotype, and demonstrate minimal off-target and toxicity effects. Here we review and compare the current strategies and non-viral delivery methods, mainly lipid and polymeric vehicles, proposed for genome editing of inherited disorders with a focus on in vivo delivery and efficacy. While the path to a safe and effective medical treatment may be arduous, the future outlook of therapeutic genome editing remains promising as long as precise technologies can be combined with efficient delivery.

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“…Although these results are somewhat encouraging, the cationic lipid delivery platforms such as those used in this study are known to be toxic, limiting their therapeutic potential 59 . Another recent study has reported the use of gold nanoparticles to co-localize Cas9 RNP, donor DNA and endosome-disrupting peptides to perform genetic correction in DMD model mice 60 . This strategy produced 3% correction of the disease-causing mutation and improved muscle function in the treated mice, though the approach may require improvement since delivery was limited to the local area of injection.…”

Section: Engineered Protein and Rna Delivery Platforms For In Vivo Gementioning

confidence: 99%

The Promise and Challenge of In Vivo Delivery for Genome Therapeutics

Wilson

Gilbert

2017

ACS Chem. Biol.

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CRISPR-based genome editing technologies are poised to enable countless new therapies to prevent, treat or cure diseases with a genetic basis. However, the safe and effective delivery of genome editing enzymes represents a substantial challenge that must be tackled to enable the next generation of genetic therapies. In this perspective we summarize recent progress in developing enzymatic tools to combat genetic disease and examine current efforts to deliver these enzymes to the cells in need of correction. Viral vectors already in use for traditional gene therapy are being applied to enable in vivo CRISPR-based therapeutics, as are emerging technologies such as nanoparticle-based delivery of CRISPR components and direct delivery of pre-assembled RNA-protein complexes. Success in these areas will allow CRISPR-based genome editing therapeutics to reach their full potential.

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Nanoparticle delivery of Cas9 ribonucleoprotein and donor DNA in vivo induces homology-directed DNA repair

Cited by 585 publications

References 45 publications

Rational Design of Nanocarriers for Intracellular Protein Delivery

Rational Design of Nanocarriers for Intracellular Protein Delivery

Debugging the genetic code: Non-viral in vivo delivery of therapeutic genome editing technologies

The Promise and Challenge of In Vivo Delivery for Genome Therapeutics

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