‘Gene therapy 2.0’ is changing medicine
Scientists are rushing to figure out how to use the gene-editing tool to stop devastating diseases. ANTONIO REGALADO reports.
DOCTORS SAY THE DISEASE is terminal, but they tell you less about living with it. About the girls who don’t see past your wheelchair, or how the phone stops ringing. It’s you and Mum counting the birthdays and figuring out what you can’t do this year. Dupree says he got by in high school, but in college depression gripped him. “I didn’t know how I could keep going,” he says.
The problem is that Dupree’s body doesn’t make dystrophin, a protein in muscle fibres that acts like a shock absorber. Without it, your biceps, calf muscles and diaphragm slowly turn to a fat-like substance. You end up on a ventilator, and then your heart stops.
Dystrophin is manufactured by a gene that is among the largest in the human genome. It consists of 79 components known as exons, each an instruction for one ingredient of the protein. Dupree’s problem, he says, is a “pseudo” exon – as though in the middle of this epic recipe someone added an instruction that read: “Stop the cooking.” There are thousands of ways a gene this size can go wrong, and Dupree’s mutation – a single letter of DNA that reads “G” instead of “T” – is unique, so far as scientists know.
Dupree, who majored in biochemistry and hopes to become a genetic counsellor, has sometimes imagined what life would be like if that small error were not there.
A year ago, in December 2015, he learnt how a technology called CRISPR might make that possible. A scientist named Eric Olson had requested some of Dupree’s blood a few months earlier, and Dupree had agreed. Soon he was rolling through the lab on his Tilite wheelchair so Olson, a biologist at the University of Texas Southwestern Medical Centre, could show him the results – and what some scientists now predict is the likeliest way to cure Duchenne.
Using CRISPR, which makes it possible to snip DNA open at a precisely chosen spot, a team at the hospital had modified Dupree’s cells in a dish, cutting through the extra exon. The editing process required only a single step and had taken three days. In an image taken with a microscope, Dupree’s cells were clouded with green puffs of perfect dystrophin.
“I try to be realistic with my expectations,” he says. “But that gave me a sense of, ‘Wow, this is here’.”
The potential to precisely and easily “edit” any genome using CRISPR is changing the way we think about nature. The CRISPR technique is often likened to a “search and replace” function for DNA. To laboratory scientists, it might better be compared to the discovery of fire. Every day they publish an average of eight scientific articles describing new uses of the technology – or merely reflecting on its exponentially expanding possibilities, such as designer babies engineered with desirable traits and mosquitoes with DNA programmed to make them go extinct.
Among these possibilities, the chance to end the pain and suffering of people like Dupree is CRISPR’S most compelling, if still distant, promise. In early-stage lab experiments, academic scientists are showing that gene editing offers new ways to attack cancer, to knock out HIV and hepatitis infections, even to reverse blindness and deafness. Companies aren’t far behind. Three startups in the Boston area have already raised a combined US$1 billion and partnered with some of the world’s biggest drug companies such as Bayer and Novartis. “None of us can anticipate where this technology will end up,” says Olson. “I’m operating under the premise that it will take us farther than we can imagine.”
Scientists know the gene errors responsible for about 5,000 inherited disorders and sequencing labs discover some 300 more each year. Some are one-in-a-billion syndromes. Duchenne is at the other extreme; it is one of the most common
‘ I AM MORE EXCITED ABOUT THE IMPLICATIONS SCIENTIFICALLY THAN ANY TREATMENT FOR ME’
inherited diseases, affecting 1 in 4,000 boys. Girls are affected rarely and to a lesser degree.
Gene editing could be a way to erase such diseases with a one-time, permanent alteration of a person’s DNA. It’s a step beyond conventional gene therapy – the 30-year-old idea of inserting entire replacement genes into a person’s cells, usually using a virus. That approach is impractical for some diseases. The gene for dystrophin, for instance, is too large to fit inside a virus, as CRISPR’S DNAsnipping proteins can. Sometimes a faulty gene that’s doing harm needs to be silenced, so adding a new one won’t help. CRISPR’S ability to delete and swap out genetic letters makes a huge new range of treatments possible.
Some doctors are now calling CRISPR “gene therapy 2.0”. To be sure, even gene therapy 1.0 has yet to fully arrive. After 30 years of research, scientists are still learning how to use viruses to move genetic instructions into a living person’s cells. Only two gene-replacement treatments for inherited disease have ever been approved, both in Europe.
But Olson says he is convinced CRISPR is the most plausible way to stop Duchenne. Early this year, he showed he could repair mutations in mice with muscular dystrophy after sending viruses stuffed with CRISPR ingredients into their veins. “A mouse is not a boy, but we think we know exactly what needs to be done,” Olson says. If it works, he adds, “this is a cure, not a treatment”.
Olson says the very first human test of a CRISPR therapy in a patient with Duchenne could begin in two years in what would be a small, exploratory clinical trial involving just a few boys. Working with Jerry Mendell of Nationwide Children’s Hospital in Ohio, a centre for gene-therapy studies, they expect to give the treatment to monkeys during the next 12 months, a prelude to human tests. The researchers will also be looking to see whether the CRISPR gene therapy has unexpected effects. Accidental edits are a particular concern.
Dupree, who is following events in the lab, says he’s not expecting much for himself. He knows the studies could take years and since his mutation is unique, he’d need a therapy tailored just for him. “I am more excited about the implications scientifically than any treatment for me,” he says. But his mother, Debbie Dupree, says chat boards and Facebook pages where parents gather are already alight with questions. “There is a lot of talk. People want to know when it will be available,” she says.
Duchenne patients and their families won’t be the only ones anxiously asking that question. Countless others facing deadly cancers or HIV, as well as sickle-cell anemia and numerous other genetic diseases, could soon be watching the fate of those Crispr-altered cells in Olson’s lab. Are they the beginning of a new era of medicine or merely one more promising research result that will never make it out of the lab? In particular, researchers
will need to solve the next challenge: safely and effectively editing DNA in cells throughout a human body, thus turning CRISPR from an invaluable lab tool into a medical cure.
CRISPR EVOLVED INSIDE BACTERIA, over billionyear time scales, as a form of immunity against viruses. Bacteria collect and store short snippets of DNA from viruses that have invaded them, spacing the segments out through their own genome in a pattern called “clustered regularly interspaced short palindromic repeats” – the term that gives CRISPR its acronym. When reinfected with one of these viruses, bacteria can create copies of these genetic snippets, which zip up letter for letter with the new virus’s DNA – signalling to a specialised cutting enzyme that it should attach itself and close, pincer-like, onto the viral genome and sever it.
By 2013, teams of scientists in Boston, Berkeley and Seoul separately showed this naturally occurring bacterial immune process could be simplified and repurposed to cut DNA in human cells. Though scientists had previously created gene-editing proteins, these were difficult to design and build compared with the solution bacteria had devised. “Instead of version 2 or version 3, it was version 3 trillion,” says Tom Barnes, chief scientist of the CRISPR startup Intellia Therapeutics in Cambridge, Massachusetts. “It went from no labs working on it to everyone working on it.”
Intellia is one of a trio of startups that have set up shop around Boston and raised about US$300 million each to create CRISPR treatments; the others are Editas Medicine and CRISPR Therapeutics. Barnes says CRISPR vastly simplifies gene editing because of the way the cutting works. Just as bacteria spot and slice the viral genetic material, CRISPR can zero in on specific stretches of human DNA. The only ingredients needed are an editing enzyme – one named Cas9 is used most often – and a short “guide”, or length of genetic letters, to tell it where to cut.
It seems simple, but using it to create human treatments is anything but. And there’s one hitch that’s often overlooked: “editing” is a bit of a misnomer. Scientists have mastered cutting into DNA, which gives them something akin to a “delete” key for genes, in addition to the “add” function offered by traditional gene therapy. But they can’t as easily rewrite genes letter for letter, an aspect of the technology still being developed.
For now, that mostly limits them to situations where deleting genes, or parts of them, is useful. Duchenne is one of those. Another is sickle-cell disease, a condition that in the United States affects mostly African-americans. Medical researchers have given it relatively little attention in the past, but there’s an obvious DNA cut that might solve it, meaning a potentially elegant cure. Now Mitchell Weiss, a haematologist who treats people with sickle-cell at St. Jude Children’s Research Hospital in Memphis, says every gene-editing company is calling him. “The interest right now is incredible,” he says. “Before, no one was interested. No one cared. But they need a proof of principle, and this is a good one.”
In addition to finding the kind of genetic problem to which CRISPR offers a solution, companies need a way to get the CRISPR instructions into the body. Most are counting on viruses for that job, but Intellia’s strategy is to package CRISPR into fatty blobs that liver cells suck up, just as if they were cholesterol. In August 2016, at the annual CRISPR meeting in Cold Spring Harbor, New York, researchers from the company showed that a single dose could alter the genomes of at least half the cells in a mouse’s liver. If Intellia can successfully edit liver cells in a person, that may let the company treat a slew of previously unassailable metabolic conditions like a form of hereditary amyloidosis, in which painful plaques build up in the body.
What’s obvious is that it will be easier to get CRISPR to work in some parts of the body than others. The easiest task is probably deleting genes in blood cells, since these cells can be removed from a patient and then put back. Already, a Chinese drug company has opened a study to create supercharged immune cells to battle cancer, and scientists at the University of Pennsylvania have announced similar plans with the financial backing of the billionaire internet entrepreneur Sean Parker.
If you’re looking for gene editing’s Everest, it’s probably rewriting DNA in the human brain – say, to treat Huntington’s disease. Editing muscle cells lies somewhere in the middle of the difficulty scale. Genetically, it’s a good candidate. Even with just a delete key, Olson says, up to 80% of muscular dystrophy cases could be treated. Initially, the editing treatment he’s working on will target a hot spot in the dystrophin gene – exon 51, in which Editas has also signalled an interest. Deleting that exon could treat about 13% of Duchenne cases.
The most significant unknown is whether it will be possible to edit enough muscle cells and make enough dystrophin in a human body. “I think this represents the most promising strategy,” Olson says. “But the thing that has to go right is that it has to be efficient.” Muscles, including the heart, glutes, and biceps, make up 40% of a person’s body mass – billions and billions of cells. So far, in his mice, Olson has succeeded in producing dystrophin in 5-25% of muscle fibres. It’s half calculation and half speculation, but he thinks that editing 15% of the muscle cells in a boy will be enough to slow, if not halt, muscular dystrophy.
When I last spoke to Olson, he was rushing to a phone meeting to drum up commercial support for his idea of starting a human test for a Duchenne treatment. He has been talking with several companies including Editas, probably the best-known of Boston’s trio of CRISPR startups. It has Bill Gates and Google as investors.
The company, founded by several of the inventors of CRISPR technology, also declared an early interest in Duchenne, licensing work done at Duke University. But its chief operating officer, Sandra Glucksmann, says it isn’t providing updates on the Duchenne program.
In fact, Editas has been lying low. CRISPR could potentially treat so many different diseases that the company has been reluctant to announce what its do-or-die project will be, and proving any CRISPR drug is effective could easily take a decade.
That puts Glucksmann in a tough position. On weekends she answers emails from desperate parents: “Could CRISPR cure my child?”. In theory the answer may be yes, but about a quarter of the time Glucksmann has never even heard of the illness before. The answer Editas has been giving to the parents of boys with muscular dystrophy has been particularly disappointing: “I am very sorry to hear about your son. Unfortunately, we are still in the very earliest stages of research.”
ONE THING THAT’S ALREADY APPARENT is that many inherited genetic diseases will require tailoring a CRISPR treatment to very specific mutations – those affecting small subsets of patients or even individual people. Take Dupree, who lives less than a mile from Olson in a Dallas suburb. His mutation is unique, and it’s not near exon 51, so he wouldn’t be helped by the first CRISPR treatment Olson is developing.
But there’s no question in Olson’s mind that Dupree’s mutation is correctable too, given the technique can potentially target any spot on the genome. Dupree now sees at least a glimmer of a chance that someone could make a CRISPR treatment just for him. “It’s only given once, and maybe it’s not that expensive,” he says. “It made me think about how it could be done, because I see things moving closer.”
Paediatrician-in-chief Ronald Cohn at Toronto’s Hospital for Sick Children, who is also a muscular dystrophy doctor, is certain that with CRISPR one-of-a-kind treatments are possible and even likely. Last December, he published a paper showing corrections of several rare mutations – again in cells in a lab dish, including some taken from a child with dwarfism and others from another boy with Duchenne. That boy, 14-year-old Gavriel Rosenfeld, is the son of close friends of Cohn’s in London. They run a charitable foundation that Cohn advises.
Cohn is a newcomer to CRISPR. A few years ago, he was studying hibernating squirrels. They don’t move for months, yet their muscles aren’t any worse for it. That is the sort of “we might just find something” approach favoured in basic-research labs. Now, with gene editing, he sees a direct path to curing someone he knows. Since correcting Gavriel’s cells, Cohn’s lab has also created a mouse model that shares his mutation. Like Dupree’s, the mutation is one of a kind, and within a few weeks Cohn’s lab will start treating the mice.
But then what? Cohn says he doesn’t know. How would you even test a drug designed for one person? Who would pay for it? He says he visited Health Canada, the country’s regulator, and was told to come back if he cured the mice. “This is going to require a significant rethinking,” he says. “The fact that you and I are having this conversation is the beginning of the paradigm shift.”
Cohn’s approach of correcting individual mutations has stirred hope among parents of boys with Duchenne. “This is a CURE!!!” one wrote on the web. His lab has used CRISPR to fix mutations in cells taken from several boys he knows, and a waiting list he keeps in a spreadsheet currently lists 53 children with muscular dystrophy. The parents of all of them want to know if their child could be helped by gene editing.
If a gene-therapy study like the one Olson plans is successful, and if CRISPR reaches enough muscle cells, there might be a strong argument that a oneoff treatment would work. After all, to aim at a new mutation all you would do is tweak the component of CRISPR that zeroes in on a specific DNA sequence. The price of manufacturing a single
‘ IT MAY TAKE A LOT LONGER THAN WE THINK, BUT SOONER OR LATER, GENE EDITING WILL CHANGE WHAT MEDICINE LOOKS LIKE.’
dose also might not be an obstacle. Two existing gene therapies approved in Europe cost US$1 million and US$665,000. Even if it cost twice that, a one-time gene fix with CRISPR would be cheaper than a lifetime of costly drugs, wheelchairs and dependency.
In holding out the hope of individual cures, Cohn admits he has created some new problems. He has invited parents to the lab, and little boys have tottered among the lab stools. But he and his students have decided to stop referring to “Gavriel’s cells” or “Jake’s cells” and use numerical code names instead. They still know who is who, but this gives them space to be impartial. “I know in the back of my head, but you want to stay unbiased,” says a graduate student in the lab, Tatianna Wong. “I can’t work on this case just because I feel bad for him. I have scientific questions to answer.”
SOME VETERANS OF GENE THERAPY roll their eyes when they hear what newcomers think CRISPR will do. I visited the vector development centre at St. Jude, touring a cramped L-shaped lab with Byoung Ryu, an expert in making viruses, who chops the air above his head and says, “people’s expectations are up here.” Ryu warns that basic, unresolved biological problems remain. One is whether editing will work often enough in cells such as those in the bone marrow, the type that need to be changed to correct sickle-cell disease. If too few cells end up edited, the treatments won’t be effective. “It’s a numbers game,” he says.
Ryu was the first employee at a Boston-area gene-therapy company, Bluebird Bio, whose stock price staggered down the chart after its first few patients didn’t all respond the same way. “I’m not negative on CRISPR, but there is a reality check,” Ryu says. “It’s not coming to people next year. It works in the petri dish every single time, but my perspective is that genome editing may happen in the future but not in the near term.”
CRISPR’S future as a treatment depends heavily on the skills of gene therapists like Ryu. They’ve been making progress, yet so far only two gene therapies – the kind that add an entire gene – have reached the market to address an inherited disorder. One, called Strimvelis, provides an outright cure for a fatal immune deficiency and was approved in 2016 in Europe. It took 15 years to test it on 18 children, and similar trials had failed. “What I learned about gene therapy is that the rabbit does not win the race,” says Weiss, who leads the St. Jude effort to apply gene editing to sickle-cell disease. “The tortoise wins the race.”
Side effects could also be an obstacle. CRISPR has the potential to cause accidental, unwanted edits that could not be erased if they ended up written into a person’s genome. Currently researchers rely on academic computer programs to predict such effects. But a program can’t predict everything. Two early tests of gene therapy in the 2000s accidentally caused leukemia in several children. No one had anticipated that consequence of changing the genome. Although Olson says he has not seen ill-effects in his mice, he allows that CRISPR can cause “inadvertent changes in DNA that are important for life”. Editing billions of individual cells in a person’s body, scientists acknowledge, will be the surest way to discover how CRISPR can go wrong.
It may take a lot longer than we think, but sooner or later, gene editing will change what medicine looks like. The biotechnology industry began in the 1970s when someone grafted insulin into E. coli, showing that a human protein could be manufactured outside the body. Now there’s a way to change DNA where it lies, inside your genes.
When he looked through a microscope at his own cells in Olson’s lab, Dupree tried to take the rational view: here was a solution for the next generation of boys. His mother, however, allowed herself to hope. “I was ecstatic. I remember thinking: this could be something that works,” Debbie says. Duchenne is a ticking clock. Parents can’t help making the calculations: this long for animal studies, this many years for the first human trial, that much more time until they know if it really works. Luckily, Dupree’s disease is the slow-moving kind. The doctors said he’d be gone by 19, but he’s still here. Maybe he’ll still be here in 10 years, says his mother, “so they can try it on him”.
Wheelchair bound Ben Dupree is cautiously hoping that the results being cooked up in the lab will one day help him.
A virtuoso gene editor: CRISPR- Cas9 (3d model) at work on a strand of DNA.
If CRISPR can reach enough of these wasted muscle cells a one- off treatment could work.