RNA delivered using a retooled retrovirus

Could a new way of delivering disease-treating RNA have been hiding in our genomes all along? Michael Segel and Feng Zhang at the Broad Institute of MIT and Harvard think so. Segel, a postdoc in Zhang’s lab, has repurposed remnants of viral DNA found in our genes to selectively deliver CRISPR-Cas gene-editing machinery into target cells. The researchers call the new method selective endogenous encapsidation for cellular delivery (SEND) and suggest it could be a new piece of the RNA delivery toolkit (Science 2021, DOI: 10.1126/science.abg6155).

It is unsurprising that Zhang’s lab is interested in RNA delivery. The CRISPR pioneer has a long interest in trying to apply gene editing for therapeutic use, but he says delivery of RNA required to do this is a significant hurdle. Currently, researchers repurpose other viruses like lentiviruses or adenoviruses, or they use lipid nanoparticles to carry the sequence of interest into a cell. Zhang wants more selective delivery options that can target specific cell types. To do that, the researchers looked inside our own DNA.

It turns out that around 8% of the human genome is sequences that originally came from other places, like viruses. In 2018, a different team of researchers working on a human gene with viral origins realized that the gene encoded a protein that formed virus-like particles that packaged its own RNA (Cell, DOI: 10.1016/j.cell.2017.12.024). If such sequences in the human genome can assemble into viral-like compartments, Zhang’s team wondered if they could find one that would work to deliver gene editing machinery. They searched through the human genome for proteins that might form the protective packages called capsids and found nearly 50 potential options, but after a few experiments, one of those proteins stood out. This protein, PEG10, looked like it would have all the pieces to make a virus-like particle, but Segel says they really became interested when they realized the protein was one of the most-efficiently secreted by mammalian cells, suggesting the protein might already be being used for RNA transport.

Segel and colleagues used the PEG10 protein to engineer a new CRISPR delivery vehicle by adding regulatory parts of the PEG10 gene to either end of CRISPR RNA so that PEG10 would recognize the RNA as cargo and package it along with the other PEG10 RNA. The team then introduced into cells DNA-encoded instructions to make the delivery nanoparticles. The completed nanoparticles also carry additional proteins, called fusogens, on the outside of the particle that help the newly made parcels enter target cell types and let the CRISPR machinery do its work.

So far, the researchers have only edited genes in human and mouse cells in the lab. Frank Sainsbury, who engineers viral capsids for applications like drug delivery at Griffith University, says the work is a significant advance but there’s a lot of characterization and optimization work still to do before SEND is used therapeutically.

Because the main protein involved, PEG10, is already found in humans, Segel says he “hopes and expects” that the proteins won’t trigger the immune system. But Sainsbury says an immune response is still a risk and if so, that could trigger an autoimmune response that is “potentially much worse” than reacting to a protein not typically found in the body. Even if this is the case, Sainsbury suggests the tool might still benefit labs working on RNA. “I think things like this can be super useful if they’re made easy to use,” he says.

For now, Zhang and Segel agree there’s still a lot of work to do, including solving the immunogenicity question. Although the team has patented SEND, it is not yet part of one of Zhang’s biotech companies or a new spin-out. “We’re focused on continuing to understand it more.” Zhang says. “There’s still a lot to do. And it’s still early days for this technology.”