Researchers develop new tool for targeted cell monitoring

Thanks to new RNA vaccines, we humans have been able to protect ourselves incredibly quickly against new viruses like SARS-CoV-2, the virus that causes COVID-19. These vaccines insert a short-lived piece of genetic material into the body’s cells, which then read its code and produce a specific protein – in this case, telltale “spikes” that stud the outside of the coronavirus – preparing the immune system to fight off the virus. future invaders.

The technique is effective and shows promise for all sorts of therapies, says Eerik Kaseniit, a Ph.D. student in bioengineering at Stanford. At the moment, however, these types of RNA therapies cannot focus on specific cells. Once injected into the body, they indiscriminately manufacture the encoded protein in every cell they enter. If you want to use them to treat a single type of cell, like those inside a cancerous tumor, you’ll need something more specific.

Kaseniit and his adviser, Assistant Professor of Chemical Engineering Xiaojing Gao, may have found a way to make this possible. They’ve created a new tool called an RNA “sensor” – a lab-made strand of RNA that only reveals its contents when it enters particular tissues in the body. The method is so precise that it can target both cell types and cell states, activating only when its target cell creates a certain RNA, Gao says. The couple published their findings on October 5 in the journal Natural biotechnology.

“For the first time, you can directly make only cells of interest produce a protein under very specific circumstances,” Gao adds. “That kind of precision just wasn’t possible before.” The protein produced could be an antigen – a foreign substance that triggers an immune response – as in the case of vaccines, an enzyme that restores function to a broken cell, a fluorescent protein that can be used to track specific cells in a research study, or a protein that triggers cell death to eliminate pathogenic or otherwise unwanted cells, among other possibilities.

Take advantage of the immune system

The pair’s new system, dubbed RADAR, is essentially made up of two sections: a “sensor” region that locks onto specific RNAs in the body, and a “payload” region that a cell will read and convert into protein. The two sections are separated by a stop codon, a portion of RNA sequence that makes part of RADAR’s genetic code inaccessible.

If RADAR’s sensor region successfully locks onto its target, the stop codon will disappear, making the remaining region – its “payload” – suddenly readable. In theory, this payload could contain instructions to make any protein, in any type of cell, at any time.

The process takes place through an existing set of enzymes called ADAR (adenosine deaminases acting on RNA) – a byproduct of an ongoing viral arms race that has raged in the human body for millennia, Gao says. .

Some viruses, like SARS-CoV-2, influenza, and norovirus, are just a protein shell with RNA tucked inside. In the process of replication, these viruses create very long stretches of double-stranded RNA. Since viruses can have devastating effects on the body, our immune system has gradually learned to see these double-stranded RNAs as a threat and will shut them down quickly.

“It’s a kind of danger signal – if a cell sees double-stranded RNA, it panics immediately,” says Kaseniit.

In a strange twist of evolution, however, our own bodies also make double-stranded RNA. As viruses have attacked us for millennia, burrowing into our cells and messing with our genetic machinery, some of their genes have been absorbed and incorporated into our DNA. (It’s no coincidence: it’s happened so many times in the past that today the human genome is almost 8% virus.)

To solve this problem, ADAR evolved as a sort of “test” system – a way for the body to tell whether a piece of double-stranded RNA is friend or foe. If it finds one created by our own genome, ADAR modifies it slightly to make it less threatening, causing holes or spaces to open between the two strands, such as removing a few stitches in the middle of a seam of tissue. The immune system, which has bigger fish to fry, quickly ignores this irregular-looking RNA and continues to fight the real enemy.

RADAR takes advantage of this mechanism. When its “sensor” module locks onto a specific target molecule (another piece of RNA), ADAR sees the resulting double-stranded pair as a friendly, harmless variety and faithfully modifies it so that the immune system ignores it. In the process, it erases the tiny molecular “stop” sign the researchers built in the middle of the RNA strand. Once removed, the payload section of RADAR is visible to the cell and the code it contains is transformed into protein.

Potential for new programmable therapies

Currently, Kaseniit, Gao and their collaborators are still testing RADAR in various settings, but the results look promising. Together with co-authors, associate professor of chemical engineering Elizabeth Sattely and postdocs Diego Wengier and Will Cody, they even tried it in plants, which don’t naturally have ADAR systems, but after adding ADAR enzymes in the mix, they were able to achieve the same results. In the future, they say, the flexibility and precision of RADAR could offer a valuable tool for both research and medicine, allowing scientists to focus on specific cells in the laboratory or deliver therapies in the body.

“It’s the hope and dream of RNA as a platform, because you can just code whatever protein you want on a piece of RNA and the cells will make it. Now with these control elements , we can specify which target cell it will be activated in. It’s very powerful,” says Kaseniit.