Tools like CRISPR that cut DNA to alter its sequence are tantalizingly approaching the clinic as a treatment for some genetic diseases. But away from the limelight, researchers are increasingly excited about an alternative that leaves a DNA sequence unchanged. These molecular tools target the epigenome, the chemical tags that adorn DNA and surrounding proteins that govern a gene’s expression and how it ultimately behaves.
A number of studies in mice in recent years suggest that epigenome editing is a potentially safer and more flexible way to turn genes on or off than DNA editing. In an example described at a gene therapy meeting in Washington, DC last month, an Italian team reduced the expression of a gene in mice to lower the animals’ cholesterol levels for months. Other groups are exploring epigenome editing to treat everything from cancer to pain to Huntington’s disease, a fatal brain disease.
Unlike DNA editing, where changes are permanent and may include unintended results, epigenomic changes are less likely to cause harmful off-target effects and can be undone. They can also be more subtle, increasing or decreasing the activity of a gene slightly, instead of blasting it full force or erasing it altogether. “What’s exciting is that there are so many different things you can do with technology,” says Charles Gersbach, longtime researcher on epigenome editing at Duke University.
Adding or removing chemical tags on DNA and the histone proteins around which it wraps (see illustration, p. 1035) can deaden a gene or expose its DNA base sequence to other proteins that activate. Some cancer drugs remove or add these chemical tags, but as disease fighters they have had limited success. One problem is that the drugs are not focused, they act on many genes at the same time, not just those related to cancer, which means they have toxic side effects.
But epigenome editing can be made precise by harnessing the same enzymes cells use to turn their genes on and off. Researchers link key components of these proteins to a gene-modifying protein, such as a “dead” version of CRISPR’s Cas9 protein, capable of reaching a specific point in the genome but unable to cut DNA. Their effects can vary: one editor might remove tags from histones to activate a gene, while another might add methyl groups to DNA to repress it.
Two decades ago, biotech company Sangamo Therapeutics designed an epigenome editor using this method that produced a gene called VEGF, which helps promote blood vessel growth, in hopes of restoring blood flow in people with diabetic neuropathy. The company injected editor-encoding DNA into the leg muscles of about 70 patients in a clinical trial, but the treatment didn’t work very well. “We couldn’t deliver it efficiently,” says Fyodor Urnov, a former Sangamo scientist now at the University of California (UC), Berkeley, Innovative Genomics Institute.
So the company turned to an adeno-associated virus (AAV), a harmless virus that has long been used in gene therapy to efficiently deliver DNA to cells. The cell’s protein-making machinery, it was thought, would use the DNA it codes for an epigenome editor to provide a constant supply. This strategy looks more promising: Over the past 3 years, Sangamo has reported that in mice it can reduce brain levels of tau, a protein involved in Alzheimer’s, as well as levels of the protein that causes Huntington’s disease.
Other teams working with mice are using the AAV delivery approach to raise abnormally low levels of a protein to treat an inherited form of obesity, as well as Dravet syndrome, a severe form of epilepsy. Last year, a group used epigenome editing to turn off a gene involved in pain perception for months, a potential alternative to opioid drugs. Another team recently activated a gene with an epigenome editor provided by a virus other than AAV. They injected it into young rats exposed to alcohol; alcohol dampened the activity of a gene, which in turn left the animals anxious and inclined to drink. The publisher of the epigenome has awakened the gene and relieved symptoms, the team reported in May Science advances.
The AAVs tested by many groups are expensive, and these DNA carriers, along with the foreign proteins they encode, can trigger an immune response. Another drawback is that the DNA loop that codes for the epigenome editor is gradually lost in the cells as they divide.
Last month at the American Society of Gene and Cell Therapy annual meeting in Washington, DC, gene editing experts offered an alternative to avoiding the downsides of AAVs. A fundamental step for the group, led by Angelo Lombardo at the San Raffaele Telethon Institute of Gene Therapy, came in 2016, when he, Luigi Naldini, and others reported in Cell that adding a cocktail of three different epigenome editors to cells in a petri dish repressed gene expression and that this lasted as the cells divided.
This meant that instead of relying on AAVs to carry DNA for their epigenome editors – and force endless expression – they could use lipid nanoparticles, a kind of bubble of fat, to carry its blueprint as messenger RNA (mRNA ). In this way, cells produce the protein for only a short time, which is less likely to trigger an immune response or make changes to the epigenome in unexpected places. Such nanoparticles are widely considered safe, especially after being injected into hundreds of millions of people over the past 2 years to provide mRNA for COVID-19 vaccines.
It took several more years for the Italian team to turn their laboratory study into a success on an animal. At the genomics meeting, postdoc Martino Cappelluti from the Lombardo lab detailed how the team injected mRNA-carrying fat particles into mice that encode for epigenome editors designed to silence a live gene. PCSK9, which affects cholesterol levels. The strategy worked, with an injection that suppressed blood levels of the PCSK9 protein by 50% and reduced low-density lipoprotein, or “bad” cholesterol, for at least 180 days.
“I see this as tremendous progress,” says Urnov, who hopes the lipid nanoparticle approach will soon be extended to other disease genes. “The key thing here is that you don’t need to have a continuous expression of the epigenome editor,” says Jonathan Weissman of the Whitehead Institute. Weissman co-directed the work reported last year at Cell on improved CRISPR-based epigenome editors that make long-lasting changes.
The researchers say epigenome editing could be particularly useful for checking more than one gene, which is more difficult to do safely with DNA editing. It could cure diseases like Dravet syndrome where a person produces some of the necessary proteins but not enough, because like a brightness regulator, the strategy can modulate gene expression without turning it on or off completely. Several new companies are hoping to commercialize treatments using epigenome editors. (Gersbach and Urnov founded one, Tune Therapeutics; Lombardo, Naldini, and Weissman are among the founders of another, Chroma Medicine.)
Despite the enthusiasm, the researchers warn that it will take some time for epigenome editing to have a broad impact. Publishers don’t always work as advertised on some genes, says David Segal, an epigenetics researcher at UC Davis. This may be partly because, as epigenetics researcher John Stamatoyannopoulos of the University of Washington, Seattle worries, researchers don’t understand exactly what editors do once they infiltrate cells. “It’s a black box,” he says.
However, Stamatoyannopoulos agrees that epigenome editing has “enormous promise”. Now, researchers need to fine-tune their epigenome editors, test them on other disease genes and tissues, and test them in larger animals for safety before moving on to people.