The Code Breaker, page 14
One especially informed reaction, a mix of excitement and envy, came from Erik Sontheimer. He had been among the first to predict that CRISPR would become a gene-editing tool. When Jinek and Chylinski finished their presentation, he raised his hand to ask a question: How could the single-guide technology be used for gene editing in eukaryotic cells, meaning ones that had a nucleus? More specifically, would it work in human cells? They suggested it could be adapted, just as many previous molecular technologies had been. After the discussion, Sontheimer, a gentle and old-school type of scientist, turned to Doudna, who was sitting two rows behind him, and mouthed the words “Let’s talk.” During the next break, they ducked out to meet in a hallway.
“I felt comfortable talking to her because, even though we were going to try to do similar things, I knew she was trustworthy,” Sontheimer says. “I told her that I was trying to make CRISPR work in yeast. She said she wanted to keep talking, because adapting CRISPR for eukaryotic cells was going to happen fast.”
* * *
That evening, Doudna took a walk into downtown Berkeley to eat at a sushi restaurant with three of the researchers who had been, and would continue to be, both colleagues and competitors: Erik Sontheimer and the two men whose paper had just been overshadowed by hers, Rodolphe Barrangou and Virginijus Šikšnys. Rather than being upset that they had been scooped, Barrangou said he realized that they had been bested fairly. In fact, as they were walking down the hill to the restaurant, he asked Doudna whether he and Šikšnys might do well to withdraw their paper that was still pending publication. She smiled. “No, Rodolphe, your paper will be fine,” she said. “Don’t withdraw it. It makes its own contribution, just like we all try to do.”
At the dinner, the four shared where each of their labs might go from there. “It was all very warm, despite the potential for awkwardness,” Sontheimer says. “Just a very exciting dinner at a very exciting time when we were all just recognizing how important this was going to be.”
The Doudna-Charpentier paper, published online on June 28, 2012, galvanized an entire new field of biotechnology: making CRISPR work in the editing of human genes. “We all knew we were going to be in a big race to do this in human cells,” says Sontheimer. “It was an idea whose time had come, and it was going to be a sprint to get there first.”
PART THREE Gene Editing
How beauteous mankind is!
O brave new world,
That has such people in’t!
—William Shakespeare, The Tempest
CHAPTER 20 A Human Tool
Gene therapies
The road to engineering human genes began in 1972 when Professor Paul Berg of Stanford discovered a way to take a bit of the DNA of a virus found in monkeys and splice it to the DNA of a totally different virus. Presto! He had manufactured what he dubbed “recombinant DNA.” Herbert Boyer and Stanley Cohen discovered ways to make these artificial genes more efficiently and then clone millions of copies of them. Thus the science of genetic engineering—and the business of biotechnology—was launched.
It took another fifteen years before scientists began to deliver engineered DNA into the cells of humans. The goal was similar to creating a drug. There was no attempt to change the DNA of the patient; it was not gene editing. Instead, gene therapy involved delivering into the patient’s cells some DNA that had been engineered to counteract the faulty gene that caused the disease.
The first trial came in 1990 on a four-year-old girl with a genetic mutation that crippled her immune system and left her at risk for infection. Doctors found a way to get functioning copies of the missing gene into the T cells of her blood system. The T cells were removed from her body, given the missing gene, and then reintroduced into her body. This led to a dramatic improvement of her immune system and allowed her to live a healthy life.
The field of gene therapy initially showed modest success, but soon there were setbacks. In 1999, a clinical trial in Philadelphia came to a halt when a young man died due to a massive immune response caused by the virus transporting the therapeutic gene. In the early 2000s, a gene therapy procedure for an immune-deficiency disease inadvertently triggered a cancer-causing gene that led to five patients developing leukemia. Tragedies such as these froze for at least a decade most of the clinical trials, but incremental improvements in gene therapies would lay the groundwork for the more ambitious field of gene editing.
Gene editing
Instead of treating genetic problems through gene therapy, some medical researchers began looking for ways to fix the problems at their source. The goal was to edit the flawed sequences of DNA in the relevant cells of the patient. Thus was born the endeavor called gene editing.
Harvard professor Jack Szostak, Doudna’s thesis advisor, discovered in the 1980s one of the keys to editing a gene: causing a break in both strands of the DNA double helix, known as a double-strand break. When this happens, neither strand can serve as a template to repair the other. So the genome repairs itself in one of two ways. The first is called “nonhomologous end-joining.” (“Homologous” comes from the Greek word for “matching.”) In such cases, the DNA is repaired by simply stitching two ends together without trying to find a matching sequence. This can be a sloppy process resulting in unwanted inserts and deletions of genetic material. A more precise process, “homology-directed repair,” occurs when the cut DNA finds a suitable replacement template nearby. The cell will usually copy and insert the available homologous sequence where the double-strand breaks occurred.
The invention of gene editing required two steps. First, researchers had to find the right enzyme that could cut a double-strand break in DNA. Then they had to find a guide that would navigate the enzyme to the precise target in the cell’s DNA where they wanted to make the cut.
The enzymes that can cut DNA or RNA are called “nucleases.” In order to build a system for gene editing, researchers needed a nuclease that could be instructed to cut any sequence that the researchers chose to target. By 2000, they had found a tool to do this. The FokI enzyme, which is found in some soil and pond bacteria, has two domains: one that serves as scissors that can cut DNA and another that serves as a guide telling it where to go. These domains can be separated, and the first can be reprogrammed to go anywhere the researchers want.1
Researchers were able to devise proteins that could serve as a guide to get the cutting domain to a targeted DNA sequence. One system, zinc-finger nucleases (ZFNs), came from fusing the cutting domain with a protein that has little fingers shaped by the presence of a zinc ion, which allow it to grasp on to a specified DNA sequence. A similar but even more reliable method, known as TALENs (transcription activator–like effector nucleases), came from fusing the cutting domain with a protein that could guide it to longer DNA sequences.
Just when TALENs were being perfected, CRISPR came along. It was somewhat similar: it had a cutting enzyme, which was Cas9, and a guide that led the enzyme to cut a targeted spot on a DNA strand. But in the CRISPR system, the guide was not a protein but a snippet of RNA. This had a big advantage. With ZFNs and TALENs, you had to construct a new protein guide every time you wanted to target a different genetic sequence to cut; it was difficult and time consuming. But with CRISPR you merely had to fiddle with the genetic sequence of the RNA guide. A good student could do it quickly in a lab.
There was one question, which was either a big one or a trivial one, depending on your perspective and your side in the patent wars that would later erupt. The CRISPR systems worked in bacteria and archaea, which are single-cell organisms that have no nucleus. But that left the question: Would they work in cells that do have a nucleus, especially multicell organisms such as plants, animals, you, and me?
As a result, the Doudna-Charpentier paper in June 2012 set off a furious sprint in many labs around the world, including Doudna’s, to prove that CRISPR-Cas9 could work in human cells. That triumph was accomplished in five places in about six months. This rather quick success could be taken as evidence, as Doudna and her colleagues would later argue, that making CRISPR-Cas9 work in human cells was an easy and obvious step that was not a separate invention. Or it could be used to argue, as Doudna’s competitors have, that it was a major inventive step that came after a fiercely competitive race.
On that question would hang patents and prizes.
CHAPTER 21 The Race
Competition drives discovery. Doudna calls it “the fire that stokes the engine,” and it certainly stoked hers. Ever since she was a child, she was not embarrassed to appear ambitious, but she knew how to balance this by being collegial and forthright. She had learned about the importance of competition from reading The Double Helix, which describes how the perceived footsteps of Linus Pauling were a catalyst for James Watson and Francis Crick. “Healthy rivalries,” she later wrote, “have fueled many of humankind’s greatest discoveries.”1
Scientists are mainly motivated by the joy that comes from understanding nature, but most will admit that they are also driven by the rewards, both psychic and substantive, of being the first to make a discovery: papers published, patents granted, prizes won, and peers impressed. Like any human (is it an evolutionary trait?), they want credit for their accomplishments, payoff for their labor, acclaim from the public, and prize ribbons placed around their necks. That’s why they work late into the night, hire publicists and patent attorneys, and even invite writers (like me) into their labs.
Competition gets a bad rap.2 It’s blamed for discouraging collaboration, constricting the sharing of data, and encouraging people to keep intellectual property proprietary rather than allowing it to be free and open for common use. But the benefits of competition are great. If it hastens the discovery of a way to fix muscular dystrophy, prevent AIDS, or detect cancer, fewer people will die early deaths. To take an example relevant to these days, the Japanese bacteriologist Kitasato Shibasaburō and his Swiss rival Alexandre Yersin both rushed to Hong Kong in 1894 to investigate the pneumonic plague epidemic and, working with different methods, discovered the responsible bacteria within days of each other.
There was one competition in Doudna’s life that stands out for becoming heated and then bitter: the race in 2012 to show how CRISPR could edit the genes of humans. It may not be up there with Charles Darwin and Alfred Russel Wallace converging on the idea of evolution or Newton and Leibniz disputing who first figured out calculus. But it is our contemporary counterpart to the race between Pauling and the team of Watson and Crick to discover the structure of DNA.
Doudna entered this competition handicapped by not having a team of collaborators who were experts in working with human cells. Her lab did not specialize in such experiments; its researchers were mainly biochemists comfortable working with molecules in test tubes. So Doudna ended up struggling to keep pace in what turned out to be a six-month frenzied competition.
There were many labs around the world that engaged in this race, but the primary drama—emotionally and personally as well as scientifically—involved three players. All were competitive in their own way, but they were very different in how comfortable they were with their competitiveness:
Feng Zhang of the Broad Institute of MIT and Harvard. Although as competitive as any star researcher, he was blessed with a cheery sweetness that made him uncomfortable displaying that trait. With deep values imbued by his mother, he had a natural humility that often masked his equally natural ambition. It was as if he had dual cores, one competitive and one beatific, that coexisted quite comfortably. He had a warm smile that rarely left his face except in those moments when the talk turned to competition—or the importance of Doudna’s achievements—at which point his lips would continue to smile, but his eyes no longer joined in. He tended to be shy of the limelight, but he was pushed by his mentor Eric Lander, the brilliant and sparky mathematician-turned-scientist who directed the Broad Institute, to compete for credit as well as for discoveries.
George Church of Harvard, Doudna’s longtime friend, who considered himself, at least for a while, to be Zhang’s mentor and academic advisor. Both on the surface and as deep as my eye can discern, he was the least competitive of them all. A Santa-bearded vegan who wants to use genetic engineering to bring back the woolly mammoth, he was driven by a playful and earnest curiosity.
And finally there was Doudna, who was not only competitive but also comfortable with her competitiveness. It was one of the reasons a certain coolness developed between her and Charpentier, who expressed some amusement and a bit of disdain for Doudna’s drive for credit. “She is sometimes stressed about credit, which made her seem insecure or not fully grateful for her success,” Charpentier says. “I am French and not as worked up, so I was always telling her, ‘Surf on the good wave.’ ” But when pressed, Charpentier admits that the competitiveness that Doudna exhibits is the force that drives most scientific pioneers, and thus science itself. “If it were not for competitive people like Jennifer, our world would not be as good,” she says. “Because what drives people to do good things is recognition.”3
CHAPTER 22 Feng Zhang
Des Moines
When I first approached Feng Zhang to ask if I could spend some time with him, I was nervous. I had told him that I was doing a book focused on Jennifer Doudna, his rival, and I thought he would be put off, perhaps would even push back.
Instead, when I visited him at his lab at the Broad Institute near MIT, with its high windows offering views of the Charles River and the spires of Harvard, he was exceedingly gracious, as he was at our subsequent conversations, lunches, and dinners. I could not tell whether his geniality was genuine or arose from an assessment that it would lead to his being portrayed better in my book. But the more time I spent with him, the more I became convinced that it was the former.
* * *
Zhang’s journey, which is worthy of a book of its own, is one of those classic immigration tales that has made America great. He was born in 1981 in Shijiazhuang, an industrial city of 4.3 million people southwest of Beijing. His mother taught computer science, his father was a university administrator. The streets of the city were festooned with China’s customary banners of exhortations, most notably those touting the patriotic duty to study science. Zhang was sold. “I grew up playing with robot kits and fascinated by anything to do with science,” he recalls.1
In 1991, when Zhang was ten, his mother came to the United States as a visiting scholar at the University of Dubuque, a gem nestled in an architecturally rich Iowa city along the Mississippi River. One day she visited a local school, where she marveled at the computer lab and the lack of emphasis on rote memorization. Like any loving parent, she imagined it through the eyes of her child. “She thought I would enjoy being in such a lab and school, so she decided to stay and bring me over,” Zhang recalls. She got a job at a paper company in Des Moines and with her H-1B visa was able to bring her son to America the next year.
His father soon followed, but he never learned English well, so Zhang’s mother became the driving force in the family. She was the one who pioneered the path to America, got a job, made friends at work, and volunteered to set up computers at local charities. Because of her, and because of the hospitality gene ingrained in heartland towns, the family always had invitations to neighbors’ houses for Thanksgiving and other holidays.
“My mother always told me to keep my head down and not be arrogant,” Zhang says. She bestowed upon him the gift of easygoing humility, which he wore lightly. But she also instilled in him an ambition to be innovative and never passive. “She pushed me to make things, even on a computer, rather than play with things that other people had made.” Years later, as I was writing this book, Zhang’s mother had moved in part time with him and his wife in Boston to help take care of their two young kids. As he talks about her while picking at a hamburger in a Cambridge seafood restaurant, Zhang lowers his head and pauses for a moment. “I’m sure going to miss her when she’s gone,” he says in a very soft voice.
At first Zhang seemed likely to follow the path of so many super-smart kids in the 1990s and become a computer geek. When he got his first computer (a PC, not a Mac) at age twelve, he learned to take it apart and use the components to build other computers. He also became a wizard at using open-source Linux operating system software. So his mother sent him to computer camp and, just to make sure he was wired for success, debate camp as well. It was the type of enhancement that privileged parents can do even without gene editing.
Instead of pursuing computer science, however, Zhang became a forerunner of what will, I think, soon be common among aspiring geeks: his interests shifted from digital tech to biotech. Computer code was something his parents and their generation did. He became more interested in genetic code.
Zhang’s path to biology began with his Des Moines middle school’s Gifted and Talented Program, which included a Saturday enrichment class in molecular biology.2 “Until then, I didn’t know much about biology and didn’t find it interesting, because in seventh grade all they did was give you a tray with a frog and tell you to dissect it and identify the heart,” he recalls. “It was all memorization and not very challenging.” In the Saturday enrichment class, the focus was on DNA and how RNA carried out its instructions, with an emphasis on the role played in this process by enzymes, those protein molecules that act as catalysts to spark actions in a cell. “My teacher loved enzymes,” Zhang says. “He told me that whenever you face a tough question in biology, just say ‘Enzymes.’ It’s the correct answer to most questions in biology.”
