The Code Breaker, page 34
However, because there are still huge risks that may be unknown, she feels that CRISPR should be used only when it is medically necessary and there are no good alternatives. “That means we have no reason to be doing it yet,” she says. “That’s why I had a problem with He Jiankui’s use of CRISPR to attempt to achieve immunity to HIV. There were other ways of doing that. It wasn’t medically necessary.”
One moral issue that continues to loom large for her is inequality, especially if the wealthy are able to buy genetic enhancements for their children. “We could create a gene gap that would get wider with each new generation,” she says. “If you think we face inequalities now, imagine what it would be like if society became genetically tiered along economic lines and we transcribed our financial inequality into our genetic code.”
By limiting gene edits to those that are truly “medically necessary,” she says, we can make it less likely that parents could seek to “enhance” their children, which she feels is morally and socially wrong. The line between medical treatment and enhancement can be blurry, she acknowledges, but it is not totally meaningless. We know the difference between correcting a very harmful gene variant and adding some genetic trait that is not medically necessary. “As long as we are correcting genetic mutations by restoring the ‘normal’ version of the gene—not inventing some wholly new enhancement not seen in the average human genome—we’re likely to be on the safe side.”
She is confident that the good that can come from CRISPR will eventually outweigh the dangers. “Science doesn’t move backwards, and we can’t unlearn this knowledge, so we need to find a prudent path forward,” she says, reprising the phrase in the title of the report she wrote after her 2015 Napa Valley meeting. “We’ve never seen anything like this before. We now have the power to control our genetic future, which is awesome and terrifying. So we must move forward cautiously and with respect for the power we’ve gained.”
PART EIGHT Dispatches from the Front
Here’s to the crazy ones. The misfits. The rebels. The troublemakers. The round pegs in the square holes. The ones who see things differently. They’re not fond of rules. And they have no respect for the status quo. You can quote them, disagree with them, glorify or vilify them. About the only thing you can’t do is ignore them. Because they change things. They push the human race forward. And while some may see them as the crazy ones, we see genius. Because the people who are crazy enough to think they can change the world are the ones who do.
—Steve Jobs, Apple’s “Think Different” ad, 1997
Samuel Sternberg
CHAPTER 44 Quebec
Jumping genes
While attending the 2019 CRISPR Conference in Quebec, I am struck by the realization that biology has become the new tech. The meeting has the same vibe as those of the Homebrew Computer Club and the West Coast Computer Faire in the late 1970s, except that the young innovators are buzzing about genetic code rather than computer code. The atmosphere is charged with the catalytic combination of competition and cooperation reminiscent of when Bill Gates and Steve Jobs frequented the early personal computer shows, except this time the rock stars are Jennifer Doudna and Feng Zhang.
The biotech nerds, I realize, are no longer the outsiders. The CRISPR revolution and coronavirus crisis have turned them into the cool kids on the edge, just as happened to the awkward pioneers who once populated the cyber-frontier. As I wandered around reporting dispatches from the front lines of their revolution, I noticed that even as they pursue their new discoveries they feel tugged, sooner than the digital techies did, to engage in a moral reckoning about the new age they are creating.
* * *
The buzz in Quebec is about a fascinating breakthrough that reignited the tension between Doudna’s realm and that of Zhang. It involves dueling discoveries of an efficient way to add new sequences into DNA. Instead of making a cut in the double-stranded DNA, the newly discovered CRISPR system would insert a new chunk of DNA by harnessing transposons, known as “jumping genes,” which are big segments of DNA that can hop from one place to another on chromosomes.
Sam Sternberg, the whip-smart biochemist who studied under Doudna and then was recruited to open his own lab at Columbia, has just published in Nature his first major paper as an assistant professor. It describes a CRISPR-guided system that inserts a tailored jumping gene into a desired DNA location. But to Sternberg’s surprise, Zhang was able to get a similar paper of his published online in Science a few days earlier.1
Sternberg seems deflated when he arrives in Quebec, and his friends, including Doudna, are angry. He had submitted his paper to Nature on March 15, and word of his discovery began to spread after one of his graduate students gave a talk about it. “Feng then quietly raced to get his paper published first,” Martin Jinek tells me at the conference. To Doudna, it was typical of Zhang: “His network of people will tell him about a paper and he will rush ahead.”2
She and Eric Lander had both conceded to me, when recalling the 2012 race, that rushing a paper into print when you sense competition is fair play. Nevertheless, Zhang’s publication on transposons causes resentment. He had submitted his paper to Science on May 4, seven weeks after Sternberg had submitted his, but Zhang’s was published online on June 6 and Sternberg’s did not appear until June 12.
I find it hard to share the Doudna camp’s outrage about Zhang. The two papers both involve harnessing jumping genes, but they differ in important ways and each makes a distinctive contribution to the progress of CRISPR. I happened to be visiting Zhang at his Broad Institute lab the day after his paper went online, which was ten days before the Quebec conference, and he described to me the research he had done on transposons. His paper was not a rush job. It had been in the works for a long time. But when he heard footsteps, he pushed Science to get it reviewed and online expeditiously—just as Doudna had done with the seminal 2012 paper she coauthored with Charpentier when she heard the footsteps of Virginijus Šikšnys and others.3
* * *
On the first day of the Quebec conference, Sternberg’s friends, including Doudna, both celebrate and commiserate with him in the hotel lobby bar over Romeo’s gin, a fragrant Canadian product. His personality is so naturally ebullient that he seems to get over his annoyance by the time he does his presentation the next day, following one given by Zhang. After all, his discovery is an important triumph and step in his career, one not diminished by Zhang’s complementary finding. So Sternberg is gracious in his talk. “We heard from Feng earlier today about how CRISPR-Cas12 can mobilize transposable elements,” he says. “What I am going to tell you about is a recently published work on type-one systems that work in similar but also different ways to mobilize these bacterial transposons.” He makes sure to heap credit on the PhD student in his Columbia lab, Sanne Klompe, who carried out the main experiments.
“Is there any field that is more cutthroat and competitive than biological research?” one of the participants asks me after Zhang and Sternberg give their dueling talks. Well, yes, I think, almost every field can be, from business to journalism. What distinguishes biological research is the collaboration that is woven in. The camaraderie of being rival warriors in a common quest suffuses the Quebec conference. The desire to win prizes and patents tends to create competition, which spurs the pace of discoveries. But equally motivating, I think, is the passion to uncover what Leonardo da Vinci called the “infinite wonders of nature,” especially when it comes to something so breathtakingly beautiful as the inner workings of a living cell. “The jumping gene discoveries show just how fun biology is,” Doudna says.
Seared bison
When the first day of presentations is over, Doudna and Sternberg go to a casual restaurant in Old Quebec City, but I accept an invitation from Feng Zhang to join him and a small group of his friends for dinner. Not only do I want to hear his perspective, but I also want to check out the inventive new restaurant he has chosen, Chez Boulay, which features crispy seal meatloaf, huge raw scallops, Arctic char, seared bison, and cabbage blood sausage. Our group of a dozen diners includes Kira Makarova of the U.S. National Center for Biotechnology Information, who was a coauthor of Zhang’s jumping-gene paper; the CRISPR pioneer Erik Sontheimer, who was Luciano Marraffini’s mentor but has stayed above the personal rivalries of the CRISPR world; and April Pawluk, who had been a postdoc in Doudna’s lab and was now an editor at Cell, a peer-reviewed journal that competes with Science and Nature. There is a symbiotic relationship between top researchers, who want to make sure their papers get speedy and favorable treatment, and smart journal editors such as Pawluk, who want to publish the most important new discoveries.
Sontheimer orders the wine, which comes from Quebec and is unexpectedly good, and we drink a toast to transposons. When the talk turns from science to the ethical issues hovering over CRISPR, most of the diners agree that, when it’s safe and practical, genetic editing—even making inheritable edits in the human germline—ought to be used if necessary to fix bad single-gene mutations, such as Huntington’s disease and sickle-cell anemia. But they recoil at the idea of using gene editing for human enhancements, such as trying to give our kids more muscle mass or height or perhaps someday higher IQ and cognitive skills.
The problem is that the distinction is difficult to define and even more difficult to enforce. “There’s a blurry line between fixing abnormalities and making enhancements,” Zhang says. So I ask him, “What is wrong with making enhancements?” He pauses for a long time. “I just don’t like it,” he says. “It’s messing with nature. And from a longer term population perspective, you may be reducing diversity.” He took the famous Harvard course on moral justice taught by the philosopher Michael Sandel, and he has clearly wrestled with these issues in a profound way. But like the rest of us, he hasn’t found easy answers.
A looming ethical issue, everyone at the table agrees, is that gene editing could exacerbate, and even encode, inequality in society. “Should rich people be allowed to buy the best genes they can afford?” Sontheimer asks. It is true, of course, that all of society’s benefits, including medical ones, are unequally distributed, but creating a marketplace for inheritable genetic enhancements would kick that issue into an entirely new orbit. “Look at what parents are willing to do to get kids in college,” Zhang says. “Some people will surely pay for genetic enhancement. In a world in which there are people who don’t get access to eyeglasses, it’s hard to imagine how we will find a way to have equal access to gene enhancements. Imagine what that will do to our species.”
Gavin Knott showing how to edit
CHAPTER 45 I Learn to Edit
Gavin Knott
Now that I had become immersed in the world of CRISPR pioneers, I decided that I should, in my own small way, be initiated into the club. I should learn how to edit DNA using CRISPR.
So I arrange to spend a few days in Doudna’s open-space lab amid the dozens of workspaces, cluttered with centrifuges and pipettes and Petri dishes, where her students and postdocs perform their experiments. I want to replicate the major advances I’ve recounted: using CRISPR-Cas9 to edit DNA in a test tube, like Doudna and Charpentier described in June 2012, and then using it to make an edit in a human cell, as Zhang, Church, Doudna, and others described in January 2013.
For the first, I am helped by Gavin Knott, a young postdoc from western Australia with a trim beard and easygoing manner. As a graduate student, he decided he wanted to find CRISPR-associated enzymes that attack RNA rather than DNA, and he wrote a letter to Doudna proposing that he come to her lab to do that. Doudna’s team was already on the case, working with an enzyme known as Cas13. “She had her finger on the pulse much more than I did,” Knott says. But she invited him to be a postdoc in her lab anyway. Among other duties, he became part of the group working on the Safe Genes project for DARPA.1
When we go into the secure part of the Doudna Lab where experiments are done, I put on my lab coat and goggles, spray my gloved hands to sterilize them, and instantly feel like a pro. Knott takes me to one of the hoods, a tabletop workspace that is partially enclosed by plastic sides and specially ventilated. Just before we begin work, Doudna buzzes through, wearing a white lab coat over jeans and a black Innovative Genetics Institute T-shirt. She briefly checks on the experiments being done by each of her students (and me), before heading off to an all-day strategy retreat with the institute’s top researchers.
The experiment that Knott walks me through involves a snippet of DNA that contains a gene that can make bacteria resistant to the antibiotic ampicillin. This is not a good thing, especially if you’re a person who’s been infected by such bacteria. So Knott concocts for me some Cas9 with a guide RNA that is designed to eliminate the gene. The lab had brewed all of this from scratch. “The Cas9 we need is encoded on a piece of DNA, and anyone who can grow bacteria in a lab can produce large quantities of it,” he assures me. My look probably conveys that I’m not sure this is part of my skill set. “Don’t worry,” he says. “If you don’t want to make it all from scratch, you can just buy the Cas9 from companies like IDT on the web. You can even buy the guide RNAs. If you want to edit genes, it’s easy to order the components online.”
(Later, I go online to see. The IDT website advertises “all of the reagents needed for successful genome editing,” with kits designed for delivery into human cells beginning at $95. Over at a site called GeneCopoeia, a Cas9 protein with a nuclear location signal starts at $85.)2
Some of the vials that Knott prepared are lined up in an old-fashioned chill box, one that uses ice to keep liquids cool. “This chill box has a significant history,” he says, turning it around. On the back is etched the name “Martin.” It had been Jinek’s before he left to start his own lab at the University of Zurich. “I inherited it,” Knott says proudly. I feel part of a historic chain. The experiments we are about to do mirror Jinek’s from 2012: taking a piece of DNA and incubating it with the Cas9 and guide RNA to cut it in the desired location. It’s sweet to be using his chill box.
Knott walks me through a variety of steps, using pipettes to combine the ingredients and then incubating it for ten minutes. We add a dye to help us visualize the results, and then we are able to create an image of what we had done by using a process called electrophoresis, which puts an electric field through a gel to separate DNA molecules of different sizes. The resulting printout shows bands at different locations along the gel, indicating if and how they were cut by Cas9. “Textbook success!” Knott exclaims as he takes the image off the printer. “Look at the differences in these bands.”
On the way out of the lab, I run into Jamie Cate, Doudna’s husband, by the elevator, and I show him my printouts. He points to blurry bars at the bottom of two of the columns and asks, “What are those?” I actually know the answer (thanks to Knott’s tutorial). “It’s the RNA,” I say. Later that day, Cate sends out a tweet attached to a picture of Knott and me working at the lab bench, saying, “And Walter Isaacson passed my pop quiz!” For just a moment, until I realize that Knott did all of the real work, I feel like a true gene editor.
Jennifer Hamilton
The next challenge is to edit a gene in a human cell. In other words, I want to take the step that the labs of Zhang and Church and Doudna accomplished at the end of 2012.
For that I team up with another postdoc in Doudna’s lab, Jennifer Hamilton, a Seattle native who earned her doctorate in microbiology at Mount Sinai Medical Center in New York City. With her big glasses and even bigger smile, Hamilton radiates enthusiasm for harnessing viruses to deliver gene-editing tools into human cells. When Doudna came to give a talk to the Women in Science group at Mount Sinai in 2016, Hamilton served as her student escort. “I felt instantly a connection with her,” Hamilton recalls.
Doudna was then beginning to build the Innovative Genomics Institute at Berkeley, which would bring together researchers from around the Bay Area. Part of its mission was to find ways to deliver CRISPR editing tools into human cells for medical treatments. So she recruited Hamilton. “I had skills in engineering viruses, and I wanted to apply them to figuring out delivery methods for getting CRISPR into humans,” Hamilton says.3 It was a specialty that would prove valuable when the lab took on the coronavirus pandemic and needed to find ways to deliver CRISPR-based treatments into human cells.
When we begin our attempt to edit DNA in a human cell, Hamilton stresses that it is more challenging than doing it in a test tube. The strands of DNA that I had edited the day before with Knott contained only 2.1 kilobases (2,100 pairs of DNA base letters) versus the 6.4 million kilobases in the cell we plan to use, which was derived from a human kidney cell. “The challenge with human gene editing,” she tells me, “is to get your editing tools past the cell’s outer plasma membrane and past its nuclear membrane to get to where the DNA is, and then you also have to get your tools to find the location in the genome.”
Hamilton’s explanation of our planned procedure seems to support, albeit inadvertently, Zhang’s argument that it is not a simple step to move from editing DNA in a tube to editing it in a human cell. However, the fact that I was about to do it could be used, I guess, to make the opposite argument.
Our plan, Hamilton says, is to make a double-strand break at a targeted place in the DNA of the human cell. In addition, we will supply a template so that a new gene will be inserted. The human cell we start with has been engineered to have a gene that creates a fluorescent protein that glows blue. In one of our procedures, we will use CRISPR-Cas9 to cut the gene and thus deactivate it. This means that the cell should no longer glow. In another sample, we will supply a template that the cell will then incorporate, changing three base pairs of the cell’s DNA in order to make the fluorescent protein change from blue to green.
