Endosomal Escape and Delivery of CRISPR/Cas9 Genome Editing Machinery Enabled by Nanoscale Zeolitic Imidazolate Framework.

CRISPR/Cas9 is a combined protein (Cas9) and an engineered single guide RNA (sgRNA) genome editing platform that offers revolutionary solutions to genetic diseases. It has, however, a double delivery problem owning to the large protein size and the highly charged RNA component. In this work, we report the first example of CRISPR/Cas9 encapsulated by nanoscale zeolitic imidazole frameworks (ZIFs) with a loading efficiency of 17% and enhanced endosomal escape promoted by the protonated imidazole moieties. The gene editing potential of CRISPR/Cas9 encapsulated by ZIF-8 (CC-ZIFs) is further verified by knocking down the gene expression of green fluorescent protein by 37% over 4 days. The nanoscale CC-ZIFs are biocompatible and easily scaled-up offering excellent loading capacity and controlled codelivery of intact Cas9 protein and sgRNA.

[1]  Mark E. Davis,et al.  Imidazole groups on a linear, cyclodextrin-containing polycation produce enhanced gene delivery via multiple processes. , 2006, Journal of controlled release : official journal of the Controlled Release Society.

[2]  Hao Zhu,et al.  Non-Viral CRISPR/Cas Gene Editing In Vitro and In Vivo Enabled by Synthetic Nanoparticle Co-Delivery of Cas9 mRNA and sgRNA. , 2017, Angewandte Chemie.

[3]  Kang Liang,et al.  Metal-Organic Frameworks at the Biointerface: Synthetic Strategies and Applications. , 2017, Accounts of chemical research.

[4]  J. Doudna,et al.  A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity , 2012, Science.

[5]  Chao Wang,et al.  Self-assembled DNA nanoclews for the efficient delivery of CRISPR-Cas9 for genome editing. , 2015, Angewandte Chemie.

[6]  S. Bell,et al.  Biomimetic mineralization of metal-organic frameworks as protective coatings for biomacromolecules , 2015, Nature Communications.

[7]  Yi-Wei Lee,et al.  In Vivo Delivery of CRISPR/Cas9 for Therapeutic Gene Editing: Progress and Challenges. , 2017, Bioconjugate chemistry.

[8]  Alexander Deiters,et al.  Optical Control of CRISPR/Cas9 Gene Editing. , 2015, Journal of the American Chemical Society.

[9]  M. Carreon,et al.  Highly permeable zeolite imidazolate framework-8 membranes for CO2/CH4 separation. , 2010, Journal of the American Chemical Society.

[10]  Michael O'Keeffe,et al.  Synthesis, structure, and carbon dioxide capture properties of zeolitic imidazolate frameworks. , 2010, Accounts of chemical research.

[11]  Nisheeth Agarwal,et al.  Gene silencing by CRISPR interference in mycobacteria , 2015, Nature Communications.

[12]  Yi-Wei Lee,et al.  Direct Cytosolic Delivery of CRISPR/Cas9-Ribonucleoprotein for Efficient Gene Editing. , 2017, ACS nano.

[13]  M. O'keeffe,et al.  Design and synthesis of an exceptionally stable and highly porous metal-organic framework , 1999, Nature.

[14]  J. Keith Joung,et al.  Efficient Delivery of Genome-Editing Proteins In Vitro and In Vivo , 2014, Nature Biotechnology.

[15]  Keiichiro Suzuki,et al.  Use of the CRISPR/Cas9 system as an intracellular defense against HIV-1 infection in human cells , 2015, Nature Communications.

[16]  Gérard Férey,et al.  Metal-organic frameworks in biomedicine. , 2012, Chemical reviews.

[17]  David R. Liu,et al.  Discovery and characterization of a peptide that enhances endosomal escape of delivered proteins in vitro and in vivo. , 2015, Journal of the American Chemical Society.

[18]  F. Shieh,et al.  Imparting functionality to biocatalysts via embedding enzymes into nanoporous materials by a de novo approach: size-selective sheltering of catalase in metal-organic framework microcrystals. , 2015, Journal of the American Chemical Society.

[19]  S. Dowdy.  Overcoming cellular barriers for RNA therapeutics , 2017, Nature Biotechnology.

[20]  Gang Bao,et al.  CRISPR/Cas9-Based Genome Editing for Disease Modeling and Therapy: Challenges and Opportunities for Nonviral Delivery. , 2017, Chemical reviews.

[21]  Ping Yu,et al.  Mitochondria Targeted Nanoscale Zeolitic Imidazole Framework-90 for ATP Imaging in Live Cells. , 2017, Journal of the American Chemical Society.

[22]  A. Chakraborty,et al.  Zeolitic Imidazole Framework (ZIF) Nanospheres for Easy Encapsulation and Controlled Release of an Anticancer Drug Doxorubicin under Different External Stimuli: A Way toward Smart Drug Delivery System. , 2015, Molecular pharmaceutics.

[23]  Christopher A. Trickett,et al.  Principles of Designing Extra-Large Pore Openings and Cages in Zeolitic Imidazolate Frameworks. , 2017, Journal of the American Chemical Society.

[24]  James E. DiCarlo,et al.  RNA-Guided Human Genome Engineering via Cas9 , 2013, Science.

[25]  Le Cong,et al.  Multiplex Genome Engineering Using CRISPR/Cas Systems , 2013, Science.