[Corrected, 1/6/16, 8:05pm. See below.] Keith Joung, a scientist at Massachusetts General Hospital in Boston, and his colleagues say they have come up with a way to make the gene editing system known as CRISPR-Cas9 far more precise when it enters cells to snip DNA.
But as Joung, who is also a founder of Cambridge, MA-based gene editing company Editas Medicine, was quick to note, there’s much more work to do to prove the technique makes CRISPR-Cas9 precise enough—and safe enough—to speed up the already competitive race to test CRISPR-based therapeutics in humans.
“I’m a physician scientist myself, and I’m quite bullish on the prospects for CRISPR-Cas9 therapeutics,” said Joung, a pathologist who runs a lab devoted to DNA engineering. The lab’s paper published today in Nature. “But it’s important to do the best possible job we can to define the risks.”
Cas9 is an enzyme that, when used as a gene editing tool, acts as molecular scissors to snip genes, forcing cells to make repairs that either delete or replace the genes. When it cuts, it also comes into contact with the DNA in multiple places along the backbone of the double helix structure to stabilize itself. Imagine an octopus sawing through a log. One arm holds the saw, while the other seven grip the log in other spots for support.
Joung and colleagues have altered four chemical fingerprints on Cas9 so it can still cut genes but doesn’t bind to the DNA backbone in as many places.
His team’s hypothesis: Weaken the hold, and perhaps the CRISPR-Cas9 complex—which is Cas9 attached to a short RNA “guide” that matches the scissors with the sequence of DNA it’s looking for—would be forced to compensate by making sure it had a perfect match with the DNA.
[This sentence has been changed. It previously gave an incorrect full meaning of HF.] “The new version, called SpCas9-HF1, for “high fidelity,” seems to work when tested in human cells in test tubes, as far as Joung and his lab mates can tell. (The Sp refers to the bacterium S. pyogenes, the source of the Cas9 version that is being used by labs around the world.)
They used a homegrown error detection system called GUIDE-Seq, then confirmed the results with deep targeted sequencing, which hones in on one nucleotide, or letter, of a gene at a time.
Off target effects have been a major concern in the field of gene editing, particularly for scientists who want to treat human disease and regulators who oversee experiments. A previous generation of gene therapy was practically brought to a halt by treatments gone awry. In France and the U.K. last decade, experimental treatments to cure X-linked severe combined immune deficiency disorder (the “bubble boy disease”) triggered leukemia in at least five children, and in 1999, teenager Jesse Gelsinger died from an immune reaction while being treated for a less severe form of a genetic liver disease in a trial at the University of Pennsylvania.
Anything similar with gene editing, whether with CRISPR-Cas9 or other systems, could be devastating to the field, especially as the public begins to grapple with the profound potential of these new tools.
Joung and colleagues are one of several teams who have been developing off-target detection technology. (I wrote about three methods, including Joung’s, and the larger problem of off-target CRISPR cuts, last year.)
Joung was careful to acknowledge, however, that GUIDE-Seq and other current detection systems are probably too blunt to catch all possible wayward cuts. He noted that current systems have error rates of 1 in every 1,000 or 10,000 alleles, or gene variants.
If a therapeutic requires putting millions of copies of CRISPR-Cas9 into millions of human cells, that error rate isn’t good enough, Joung said. “For me, the challenge is now to develop methods more sensitive than what we have.”
For diseases that require treatment of fewer cells, or when treating cells with a lower risk of triggering cancer, such exquisite precision might not be necessary. Indeed, there is an ongoing debate over how much off-target cutting is tolerable. Those less conservative argue that human cells are repairing their own DNA constantly, so misdirected cuts from CRISPR might not add much to that manageable burden.
The only gene editing system to be tested in people is called zinc finger nucleases, owned by Sangamo Biosciences (NASDAQ: SGMO), which has advanced a therapy that aims to cure HIV into Phase 2. No CRISPR-based treatment has yet reached human clinical trials. The first could come from Editas, which is aiming to start a trial for a rare genetic blindness in 2017.
Even though Joung is a founder, it’s unclear if Editas would move forward with its blindness program, or any program, with Joung’s “high fidelity” Cas9. As regulatory filings from Editas noted, the company does not necessarily gain license to the inventions of its scientific founders. On Monday, the company filed initial paperwork for an IPO and noted that it has no rights to another CRISPR development, a new enzyme called Cpf1 described by Editas founder Feng Zhang of the Broad Institute.