Nanoparticle-Based System Developed for Delivery of CRISPR/Cas9

CRISPRs (clustered regularly interspaced short palindromic repeats) are segments of prokaryotic DNA containing short repetitions of base sequences. Each repetition is followed by short segments of "spacer DNA" from previous exposures to a bacterial virus or plasmid. Since 2013, the CRISPR/Cas system has been used in research for gene editing (adding, disrupting, or changing the sequence of specific genes) and gene regulation. By delivering the Cas9 enzyme and appropriate guide RNAs (sgRNAs) into a cell, the organism's genome can be cut at any desired location. The conventional CRISPR/Cas9 system is composed of two parts: the Cas9 enzyme, which cleaves the DNA molecule and specific RNA guides that shepherd the Cas9 protein to the target gene on a DNA strand. Efficient genome editing with Cas9-sgRNA in vivo has required the use of viral delivery systems, which have limitations for clinical applications.

To bypass the need for a viral carrier, investigators at the Massachusetts Institute of Technology (Cambridge, USA) identified regions of sgRNA that could be modified while maintaining or enhancing genome-editing activity. They then developed an optimal set of chemical modifications for in vivo applications.

The investigators reported in the November 13, 2017, online edition of the journal Nature Biotechnology that by using lipid nanoparticle formulations of these enhanced sgRNAs and mRNA encoding Cas9, they were able to show that a single intravenous injection of this mixture into mice induced specific editing of the enzyme Pcsk9 (Proprotein convertase subtilisin/kexin type 9) in the livers of more than 80% of the treated animals. Serum Pcsk9 was reduced to undetectable levels, and cholesterol levels were significantly lowered by about 35 to 40% in these animals.

"What is really exciting here is that we have shown you can make a nanoparticle that can be used to permanently and specifically edit the DNA in the liver of an adult animal," said senior author Dr. Daniel Anderson, associate professor of chemical engineering at the Massachusetts Institute of Technology.

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