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A team of researchers, including Bowen Li from the University of Toronto’s Leslie Dan Faculty of Pharmacy, has developed a new lipid nanoparticle with potential to deliver gene editing tools to cells in the lung – a promising step toward developing new, inhalable therapies for lung diseases such as cystic fibrosis.

“The discovery of this lipid nanoparticle is a significant step forward,” said Li, adding that it demonstrates the potential to deliver gene editing tools to the lung via inhalation. “This has been very difficult to achieve to date and it has huge potential for clinical translation.” 

A first author of , Li is currently an assistant professor at the Leslie Dan Faculty of Pharmacy and helped lead the research during a post-doctoral stint at the Massachusetts Institute of Technology (MIT).

Daniel Anderson, senior author and professor in MIT’s Department of Chemical Engineering and a member of MIT’s Koch Institute for Integrative Cancer Research and Institute for Medical Engineering and Science, said it was the first demonstration of highly efficient delivery of RNA to the lungs. “We are hopeful that it can be used to treat or repair a range of genetic diseases, including cystic fibrosis,” . 

The research team also designed an innovative high-throughput platform that enabled it to quickly develop and test hundreds of candidate lipid nanoparticles, significantly speeding up identification of the lead candidate. The novel platform has potential to be used to develop lipid nanoparticles for countless other applications – work that Li is continuing in his research program at the Leslie Dan Faculty of Pharmacy.

“Behind this work is our high-throughput platform, and we aim to repurpose this platform to identify nanoparticles for delivering various RNA therapies to different disease-affected organs,” says Li who completed his post-doctoral fellowship with Anderson and Robert Langer, Institute professor at MIT and senior author of the study.

In 2019, Anderson’s group developed nanoparticles that could deliver mRNA that produced a bioluminescent protein to lung cells. The current research takes the approach one step farther by delivering mRNA that produces a protein capable of editing the genome.

mRNA explored as novel approach to deliver gene editing tools

The CRISPR-Cas9 gene editing tool holds promise as a potential approach to treat diseases caused by genetic mutations because it can change specific genetic sequences to produce proteins that have a therapeutic effect. It is composed of RNA that targets a specific DNA sequence and the Cas9 enzyme, which “cuts” the genome to add or remove sections.

In the past, researchers delivered CRISPR-Cas9 to targeted cells using a modified virus. But the tool’s comparatively large size and the potential of the virus to stimulate an unwanted immune response have blunted its success.

However, new mRNA technology – similar to what is used in COVID-19 vaccines – has potential for gene editing. Since mRNA is involved in making proteins, it may be possible to deliver mRNA that allows the target cell to produce the Cas9 enzyme instead of delivering the enzyme itself. This advance could lessen the risk of an immune response and allow for repeated doses.

To be successful, this approach requires identifying a lipid nanoparticle that can surround the mRNA and help it cross the cell membrane, allowing the mRNA to be delivered to the right cell.

Innovative approach identifies candidate lipid nanoparticle

Delivering mRNA directly to lung cells has proven to be particularly challenging. A layer of mucus acts as an additional barrier for lipid nanoparticles and, while the lung contains several types of cells, only a few are relevant targets for gene editing to treat lung disease.

The research team designed several hundred lipid nanoparticles and used a novel, high-throughput platform to quickly test and find the best candidates that could deliver gene editing tools to lung cells.

They identified a previously undescribed lipid nanoparticle that was able to reach the target lung cells, cross the cell membrane and escape the endosome to deliver the mRNA where it could be translated into Cas9 and, ultimately, resulted in the genome being edited.

The candidate lipid nanoparticle was able to deliver mRNA to two specific types of lung cells – club cells and ciliary cells – that are implicated in cystic fibrosis and other lung diseases. The ability to deliver to multiple cells could also result in a longer-lasting therapeutic effect.

“Club cells live longer than ciliary cells and they can differentiate into the ciliary cells. Even if the ciliary cell dies, the edited club cells could differentiate into ciliary cells and maintain the efficacy,” Li explains.

Research will now focus on testing the lipid nanoparticle under disease conditions, such as in models of cystic fibrosis where the mucus is very thick, as well as on developing a formulation that can be inhaled. And Li plans to continue using the high-throughput platform in his current research program to develop and test lipid nanoparticles for a range of applications.

The research received support from Translate Bio, the National Institutes of Health, the Leslie Dan Faculty of Pharmacy startup fund, a post-doctoral fellowship, the American Cancer Society, and the Cystic Fibrosis Foundation.