One Dose of This Gene Editor Could Defeat a Host of Genetic Diseases Suffered by Millions

On the surface, cystic fibrosis and Tay-Sachs disease have nothing in common. Although both are inherited genetic disorders, one causes thick mucus buildup in the lungs, making it progressively harder to breathe; the other gradually leads to a buildup of fatty molecules that kills brain cells.

But under the hood, the diseases share a common villain: Nonsense mutations.

Like a molecular “stop sign,” these mutations instruct cells to abandon making certain proteins, resulting in truncated versions that don’t work and lead to disease. Gene-editing tools can correct mutated genes by targeting them one by one for each disease. While this approach can save lives, it takes time and a lot of resources to develop.

Why not aim for the common villain?

This month, a team led by David Liu at Harvard University developed a gene-editing tool to correct nonsense mutations. Called PERT, the tool tackles the mutations head on, allowing cells to ignore the mutations and continue producing proteins to their full length and potential.

In cultured human cells and mice with nonsense mutations, a single dose of PERT rescued protein production and eased disease symptoms. Because PERT is inserted into the genome, the treatment is, in theory, one-and-done.

The gene therapy could be a boon for rare diseases. More than 7,000 inherited diseases affect hundreds of millions of people around the world. Nonsense mutations are involved in roughly 30 million. Few have any treatment or cure.

“The most effective strategy for addressing this unmet need would be to develop therapies that are effective against multiple rare disorders,” wrote Kim Keeling at the University of Alabama, who was not involved in the study.

A more universal gene editor fits the bill. If proven safe and effective in humans, PERT sets the stage for more affordable gene therapies, faster development times, and more importantly—hope for people with rare diseases that have been largely sidelined in the past.

Hacking the Protein Machine

Proteins are the workhorses of our bodies. They’re the foundational building blocks of tissues and organs and are in charge of intricate biological functions, from regulating immune responses to digesting food.

The blueprints for proteins are embedded in our DNA as three-letter codons. Each codon represents a single amino acid, the basic molecules that make up proteins. These codons are transcribed into molecular transports called mRNA, which shuttle the information to the cell’s protein-making factory for assembly.

The factory reads the codons one by one and instructs a team of molecular chauffeurs called tRNA to grab the correct amino acid and bring it to the assembly line. In this way, the factory translates the body’s genetic code into a ribbon-like protein chain.

Nonsense mutations bring the process to a screeching halt. The factory needs instructions on when a protein chain is complete so it can be released for further processing. These instructions are called “stop codons” and are made of several unique genetic letter combinations.

Some genetic diseases have a single genetic mutation that turns a protein-making codon into a stop codon—basically, pulling an emergency switch to shut down production. Rather than making the entire functional protein, the cell destroys mRNA shuttles and truncates the resulting proteins. These proteins are less stable or struggle to perform their roles.

Previous studies found a workaround: Nonsense suppression.

Engineered suppressor tRNA molecules can skip over nonsense mutations. Like molecular smugglers, these synthetic RNA molecules sneak amino acids to spots where the protein should have been terminated. This trick rewires the code and lets the protein-making factory skip the stop command and keep making the rest of the protein.

The strategy has already had successes. In one study, synthetic suppressor tRNA molecules delivered by a virus were shown safe and effective in mice with a nonsense mutation, and beneficial effects from a single treatment lasted for more than half a year. Another suppressor tRNA molecule wrapped up in a fatty bubble for delivery—a commonly used system in gene therapies—restored production of a protein in mice with cystic fibrosis, allowing them to better breathe.

Both methods have downsides though. Viral carriers, even when stripped of their disease-causing traits, can still stir up immune responses. And although using fatty bubbles to deliver therapies is relatively safer, they require multiple doses in chronic genetic diseases.

Precision Editor

Liu and colleagues brainstormed a one-and-done therapy that directly inserts instructions for suppressor tRNA molecules into cells or an animal’s genetic code.

After screening thousands of tRNA variants, they found a highly active candidate as a starting point. Using prime editing, a type of small and precise gene editor, they altered natural versions of tRNA into suppressor versions that recognized a specific mutated stop codon.

Rather than terminating the building project, the engineered tRNA shuttled an amino acid into place to override the mutation and finish constructing the full-length protein.

The team tested the new tool, called PERT, in several human cell types in petri dishes. The cells harbored nonsense mutations for different genetic diseases, including cystic fibrosis and Tay-Sachs disease. A single dose increased working proteins by 20 to 70 percent regardless of the disease.

The therapy also worked in mice with a nonsense mutation causing a severe disease called Hurler syndrome in humans. Here, the body struggles to make a protein that degrades a type of sugar molecule, which builds up and causes cellular mayhem. Seven weeks after a single treatment, the mice had 8 percent more of the protein—enough to decrease harmful sugars and alleviate symptoms.

Making Sense Out of Nonsense

PERT’s strength is in its versatility. In a screen of over 14,000 mutated stop codons, the gene editor bypassed mutations roughly 70 percent of the time.

But while the results are promising, altering nonsense mutations can be fickle.

Inserting an amino acid into a growing protein chain can impact its function and stability. Proteins largely depend on their 3D structures to interact with other biological molecules, and a single change in amino acid makeup could alter the overall architecture.

It’s therefore unlikely “that the engineered tRNA will recover optimal function for all proteins” that have nonsense mutations, wrote Keeling. The current study focused on one stop codon: UGA. Several others exist and are now targets for other suppressor tRNA molecules.

Using prime editing, the molecules can linger in the body to continually produce the engineered versions without need for repeated jabs. From here, scientists must conduct long-term animal studies to test the edited tRNA’s stability and side effects.

There’s also the dosage problem. An optimal amount for liver tissue may be too large or ineffective for the heart or lungs. Eventually the team envisions a library of PERT tools, tailored to each organ and frozen in a fridge for use on command. With their work, the team has brought the therapeutic use of suppressor tRNA molecules “a step closer,” wrote Keeling.