WIRED, March 20, 2012
There’s a moment in the history of medicine that’s so cinematic it’s a wonder no one has put it in a Hollywood film. The scene is a London laboratory. The year is 1928. Alexander Fleming, a Scottish microbiologist, is back from a vacation and is cleaning up his work space. He notices that a speck of mold has invaded one of his cultures of Staphylococcus bacteria. It isn’t just spreading through the culture, though. It’s killing the bacteria surrounding it.
Fleming rescued the culture and carefully isolated the mold. He ran a series of experiments confirming that it was producing a Staphylococcus-killing molecule.
And Fleming then discovered that the mold could kill many other species of infectious bacteria as well. “I had a clue that here was something good, but I could not possibly know how good it was,” he later said.
No one at the time could have known how good penicillin was. In 1928, even a minor wound was a potential death sentence, because doctors were mostly helpless to stop bacterial infections. Through his investigations into that peculiar mold, Fleming became the first scientist to discover an antibiotic—an innovation that would eventually win him the Nobel Prize. Penicillin saved countless lives, killing off pathogens from staph to syphilis while causing few side effects. Fleming’s work also led other scientists to seek out and identify more antibiotics, which collectively changed the rules of medicine. Doctors could prescribe drugs that effectively wiped out most bacteria, without even knowing what kind of bacteria was making their patients ill.
Of course, even if bacterial infections were totally eliminated, we would still get sick. Viruses—which cause their own panoply of diseases from the common cold and the flu to AIDS and Ebola—are profoundly different from bacteria, and so they don’t present the same targets for a drug to hit. Penicillin interferes with the growth of bacterial cell walls, for example, but viruses don’t have cell walls, because they aren’t even cells—they’re just genes packed into “shells” made of protein. Other antibiotics, such as streptomycin, attack bacterial ribosomes, the protein-making factories inside the pathogens. A virus doesn’t have ribosomes; it hijacks the ribosomes inside its host cell to make the proteins it needs.
We do currently have “antiviral” drugs, but they’re a pale shadow of their bacteria-fighting counterparts. People infected with HIV, for example, can avoid developing AIDS by taking a cocktail of antiviral drugs. But if they stop taking them, the virus will rebound to its former level in a matter of weeks. Patients have to keep taking the drugs for the rest of their lives to prevent the virus from wiping out their immune system.
Viruses mutate much faster than bacteria, and so our current antivirals have a limited shelf life. And they all have a narrow scope of attack. You might treat your flu with Tamiflu, but it won’t cure you of dengue fever or Japanese encephalitis. Scientists have to develop antivirals one disease at a time—a labor that can take many years. As a result, we still have no antivirals for many of the world’s nastiest viruses, like Ebola and Nipah virus. We can expect more viruses to leap from animals to our own species in the future, and when they do, there’s a good chance we’ll be powerless to stop them from spreading.
Virologists, in other words, are still waiting for their Penicillin Moment. But they might not have to wait forever. Buoyed by advances in molecular biology, a handful of researchers in labs around the US and Canada are homing in on strategies that could eliminate not just individual viruses but any virus, wiping out viral infections with the same wide-spectrum efficiency that penicillin and Cipro bring to the fight against bacteria. If these scientists succeed, future generations may struggle to imagine a time when we were at the mercy of viruses, just as we struggle to imagine a time before antibiotics.
Three teams in particular are zeroing in on new antiviral strategies, with each group taking a slightly different approach to the problem. But at root they are all targeting our own physiology, the aspects of our cell biology that allow viruses to take hold and reproduce. If even one of these approaches pans out, we might be able to eradicate any type of virus we want. Someday we might even be faced with a question that today sounds absurd: Are there viruses that need protecting?
At five a.m. one day last fall, in San Francisco’s South of Market district, Vishwanath Lingappa was making rabies soup. At his lab station, he injected a syringe full of rabies virus proteins into a warm flask loaded with other proteins, lipids, building blocks of DNA, and various other molecules from ground-up cells. It cooked for hours on Lingappa’s bench, and occasionally he withdrew a few drops to analyze its chemistry. By spinning the fluid in a centrifuge, he could isolate small clumps of proteins that flew toward the edge as the bigger ones stayed close to the center.
To his mix, Lingappa had added a particular protein he wanted to study. He suspected that the rabies virus used this protein in the infected cell to assemble the capsid, or external shell, of replicated viruses. He had tagged the target protein with radioactive atoms, allowing him to follow it as it interacted with other elements in the soup.
At around 10 in the morning, Lingappa took pictures of the mixture. By lunchtime, seven hours into his workday, the images were developed and ready to show off to his staff. In the conference room, a table was strewn with take-out sandwiches, and an abandoned bowl of oatmeal sat on a credenza. As Lingappa held up the films to the light, his colleagues crowded behind him to make out black streaks across the images.
As predicted, the tagged protein had joined with other proteins, creating the microscopic machines that in a real infection would assemble the rabies virus shell. Why would this matter? Because a drug developed by Lingappa’s firm, Prosetta Antiviral, has been shown to interfere with this protein, blocking it from functioning in these shell-making machines. If his gamble pays off, this is the pathway by which an antiviral drug will stop cells from replicating the rabies virus. At moments like these, Lingappa’s voice swells from its normal, mellow pedagogical tone to a near-talk-radio thunder; he clenches his fist and paraphrases one of his favorite lines from the philosopher John Dewey. “The givens of experience are not given,” he says. “They are taken! With great difficulty!”
Lingappa came relatively late to his obsession with antivirals. He trained as a cell biologist in the late 1970s in the laboratory of Günter Blobel, a Rockefeller University cell biologist who went on to win the Nobel Prize in 1999. Blobel studied how cells work by grinding them up and running experiments on their loose contents. This type of cellular soup, known as a cell-free system, can simulate the inner workings of a cell, including the assembly of new genes and proteins. By adjusting its composition—leaving out a single enzyme, for example—scientists can figure out how a cell’s molecules work together to keep it alive. Under Blobel’s tutelage, Lingappa became a cellular chef de cuisine in his own right. For example, he ran experiments to figure out how newly made proteins were ferried through a cell to the place where they were needed. After earning his PhD, Lingappa headed west to UC San Francisco to continue his research.
He might have experimented his way to a quiet retirement had it not been for his younger sister Jaisri, who was treating AIDS patients at the UCSF Medical Center. She spent a summer at Rockefeller many years beforehand, at Vishwanath’s urging, and now saw that cell-free systems might shed some light on viruses. At the time, the prevailing dogma was that once a host cell made new virus genes, the capsid could self-assemble around them. But Jaisri was skeptical. She suspected that viruses needed help from host enzymes to mold the shell into its proper shape. By experimenting in a cell-free system, she reasoned, she might be able to identify those host enzymes that the virus depended upon—and figure out how to block them. She asked her brother whether there was any chance her idea could work. “I haven’t a clue,” Vishwanath replied. “Let’s try it.”
HIV was still dauntingly mysterious at the time, so the Lingappas began their experiments on hepatitis B, a relatively simple virus that scientists already knew a great deal about. They figured out how to get cell-free systems to generate hepatitis B capsids. Next they tinkered with the soup’s recipe, taking out various enzymes and observing whether there was any change to the shells it produced. Before long they had found that a number of enzymes were essential to making the capsids. When these enzymes were present, the cell-free system produced perfect shells. Without them the system could manage only stunted, half-formed shells. As the Lingappas had predicted, the capsids could not put themselves together.
They and their colleagues went on to run the same experiments on HIV, and again they found that the viruses needed lots of help. Host enzymes had to join together to form complicated biological machines with the right shape—the right set of pockets, grooves, and clefts—to grab parts of viruses and push them into their proper place to build the shell. For each capsid-making machine, the Lingappas reasoned, there should be a molecule they could lodge in some key pocket, making it useless for hauling capsid proteins into place. The machine would thereby be immobilized, and the infected cell could no longer build viruses.
By 2003, Vishwanath had so much faith in his idea that he retired early from UCSF to launch Prosetta. (Jaisri helped him start the company, but she has remained in academic life, now at the University of Washington.) The first thing researchers at Prosetta had to do was search for promising candidates—molecules of just the right shape to lodge into the capsid-making machinery. They screened 80,000 compounds by testing each of them in a cell-free system. Most of the compounds couldn’t stop capsids from forming, but a few dozen did. Instead of focusing on one, Lingappa decided to pursue almost all of them at once on the premise that a victory against any one virus would help Prosetta extend its strategy to all of them.
“We distributed our eggs in every basket conceivable,” he says. It was a gutsy strategy, and so far it’s paying off. Preliminary studies—in both cell cultures and on animals—are showing that Prosetta’s approach can stop rabies, Ebola, influenza, and a number of other viruses. If, as Lingappa suspects, all viruses need help from their host cells to assemble, he may have found a strategy that can work against every virus that could ever make us sick.
Up until now, antiviral drugs have tended to work by interfering with viruses themselves. Consider the case of Tamiflu, our best drug against influenza after infection has already occurred. Tamiflu binds to neuraminidase, a protein on the surface of flu viruses. When new viruses form, they use neuraminidase to pry open a passageway out of their host cell. Tamiflu disables the protein, trapping the flu viruses so that they can’t spread to other cells. The shape of the molecule in Tamiflu is exquisitely well matched to the neuraminidase protein on flu viruses, and as a result it can’t bind to proteins on the surface of other viruses.
The Tamiflu strategy—targeting the components of individual viruses—can yield effective drugs, but it also has some serious drawbacks. The biggest of these is resistance. Many viruses, such as the flu and HIV, mutate at a blinding pace: a million times faster than our own genome does. Every now and then, one of those mutations will alter the target of an antiviral drug. The drug will have a harder time latching onto the mutant virus, which will then be free to reproduce and flourish unchecked. In 2007, for instance, a mutant form of flu virus emerged that could resist Tamiflu. It had evolved a differently shaped neuraminidase that Tamiflu couldn’t grab. So even as governments around the world stockpiled 200 million doses of the drug to prepare for the next great flu pandemic, Tamiflu-resistant strains spread across the world. “It’s a game-breaker,” says Vincent Racaniello, a Columbia University virologist and the author of the textbook Principles of Virology. “Anything you make, the viruses will become resistant to.” (So far in Lingappa’s experiments, influenza viruses have failed to evolve resistance to the Prosetta drug. The reason: It’s easy for viruses to evolve a new shape that allows them to shake off Tamiflu, but it’s difficult—potentially impossible—for them to evolve a way to assemble themselves if their host’s proteins won’t cooperate.)
Prosetta’s approach is so intriguing because it alters our own cellular machinery instead of attacking the virus directly. Roughly speaking, this is the chief insight that animates all of the new strategies for antiviral drugs: They focus on the host instead of the virus. A second approach along these lines would boost our own immunological response to viruses, while a third strategy would take an even more radical step: rewiring our cells to commit suicide when they get infected.
The Replication Process
Enter Each type of virus has its own unique way of hijacking cells. In the case of influenza, the virus shell contains proteins called hemagglutinin, which bind to the surface of healthy cells and trigger them to open up.
Replicate Once inside, the virus sheds its shell, and its genes worm their way into the cell’s nucleus, hijacking the host to copy the viral genes instead of its own. In most cases, as virus genes are copied, they take the form of double-stranded RNA, or dsRNA.
Neutralize cell defense Certain proteins in the cell will sense the dsRNA and trigger the production of antiviral molecules called interferons. But some viruses (including some influenza strains) have evolved counterattacks against the interferon response.
Mature Viruses coordinate dozens of host proteins to become molecular machines, which then assemble the new virus particles. Once each particle matures, it binds to the inner surface of the cell and opens a passageway out. It is now ready to infect a new cell.
The Strategies
A. Trigger Suicide
An artificial molecule called Draco attaches itself to dsRNA, triggering the cell to commit suicide. Because viruses have never been exposed to Draco before, their counterattacks are useless. The infected cells die before the viruses can mature.
B. Amp Up Defenses
Many viruses are programmed to prevent a cell from making its own interferons. By injecting us with synthetic interferons, doctors could trigger an antiviral response—no matter what virus is attacking us.
C. Inhibit Maturation
Prosetta’s molecules bind to the proteins that viruses need to create their shells, stopping the maturation process. And because these proteins come together only to make viruses, the drugs should be nontoxic to patients.
That second approach is being spearheaded by Eleanor Fish of the University of Toronto. She and other researchers worldwide are developing drugs that might replace or supplement interferons, our own catchall viral response. Essentially the idea is to accelerate the body’s own virus-killing powers. Our cells can sense a viral invasion because of a quirk in the way most viruses replicate: Using the host cell’s machinery, they copy their own genes by making a peculiar molecule called double-stranded RNA. So our cells are equipped with proteins whose sole job is to detect double-stranded RNA. When they do, they immediately relay a signal throughout the cell that an intruder has invaded.
The cell then produces a second type of protein, called interferons, which launch a massive counterattack. They in turn trigger the production of more than 300 other kinds of proteins, each with its own role to play in killing viruses. Some of the proteins slice up the virus’s genes and wreck its proteins. Others tell the cell to become rigid, making it harder for new viruses to escape. The infected cell also sends interferons to surrounding cells, creating a firebreak that stops the spread of the infection. “The very first response to a virus—whether it’s a respiratory infection, whether it’s a virus that gets into your gut, whether it gets into a skin tear—our very first response is an interferon response,” Fish says. But our natural defenses are far from perfect: Viruses often replicate fast enough to keep ahead of the interferon patrol, and certain kinds of viruses, such as influenza and SARS, make proteins whose sole job is to shut down interferons. Since the 1990s, some natural interferons have been approved for use against certain viruses (hepatitis C, for example), but their track record has been disappointing: They work in some patients and not others, they can cause toxic side effects, and they are expensive, delicate drugs.
So Fish and other groups of scientists are trying to build a drug that would do the interferons’ job better. By tacking on polyethylene glycol—a cluster of hydrogen, carbon, and oxygen atoms—they’ve created synthetic interferons that last days instead of hours and will wipe out hepatitis C viruses completely in up to 81 percent of treated patients, depending on the strain. During Toronto’s SARS outbreak, Fish tested synthetic interferons on a small pool of patients and found that their lungs healed significantly faster than those of control patients, allowing them to get off supplemental oxygen sooner. Like Lingappa, Fish has huge ambitions for these synthetics. If one of them succeeds, it could become a single drug to fight not just one virus or a few, but nearly every virus.
The third and arguably the most radical approach to broad-spectrum antivirals was conceived in the shower, of all places. It was 12 years ago, and Todd Rider was struck with a blinding insight about viruses—specifically, an idea for how to stop any virus in the world. By the time the shower was over, he knew exactly what to do.
Rider is a biological engineer. At MIT’s Lincoln Laboratory in the late 1990s, he built a technology called Canary, a sensor for dangerous airborne pathogens like anthrax and smallpox. This ingenious box houses white blood cells, each of which is engineered to detect a particular kind of bacteria or virus. Canary is now installed in several government buildings in the Washington, DC, area. But for all the success of his invention, the experience left him profoundly dissatisfied. “I realized that if we detected bacteria, it was fine,” Rider says. Doctors could prescribe antibiotics, after all. “But if we detected viruses, there really wasn’t anything out there.” Rider had read everything he could about traditional antivirals, and he had sensed an opportunity to create something quite different. “I wanted a treatment that was broad spectrum, that was effective against a wide range of viruses, and that would be difficult for viruses to evolve resistance to,” he says.
That something—the thing Rider conceived of in the shower that day—was an artificial protein. To make it, he would need to marry parts of two natural proteins. One string would detect double-stranded RNA, the telltale sign of most invading viruses. The other would lead the infected cell to kill itself. Rider wanted to make a poison pill for cells: a protein that, when it grabbed onto the double-stranded RNA of a virus, would trigger instant cellular suicide. That may sound like a dangerous kind of therapy, but our bodies already rely on it to naturally fight both infections and cancer.
Rider dubbed his theoretical molecule Draco, for Double-stranded RNA activated caspase oligomerizer. On paper, at least, it looked great. Viruses that quickly evolve resistance to targeted antivirals would have no way of evading this weapon: Doing so “would need a whole new set of genes,” Rider says.
The idea had come in a flash, but testing it proceeded slowly. Rider started in 2000, when manufacturing microbiological materials was still a struggle. (Today you can custom-order genes online and get them a few days later in a FedEx envelope.) Rider also had to stretch a lean budget, cobbled together from grants from the National Institutes of Health and the Department of Defense. In online databases, he found sequences for two genes that performed the functions he wanted to combine—detect double-stranded RNA and trigger apoptosis, or cell death—and then he joined the two into a single sequence. He gave his new protein an increasingly challenging series of tests. In one early experiment, he found that the protein could get inside cells and stay there for as long as 11 days. Next he injected rhinoviruses (which cause the common cold) into both ordinary human lung cells and lung cells carrying Draco. In the unprotected batches of cells, the rhinovirus multiplied, spreading from cell to cell and wiping out the entire population. When Rider added the virus to the Draco-protected cells, the infected cells promptly destroyed themselves, right on cue; the rest of the cells were left unharmed. Rider also found that he could stop the spread of the virus even if he injected Draco as long as three days after infection.
To see how Draco would fare in a living body, he began to study it in mice. To deliver the drug to the right place in the mice’s bodies, Rider added a kind of molecular “address label” to the Draco protein such that the cells in a particular organ would be targeted. The mice seemed unharmed by the initial introduction. So Rider exposed the mice to flu viruses. Without Draco, four out of five mice died. But if he spritzed Draco into their nostrils just before or after the infection, they all lived.
Having published the results in the journal PLoS One last July, Rider is now preparing to test Draco against hemorrhagic fever and other viruses in mice and hopes to license the technology to a company that can take it to human trials. Rider sees his potential drug as a true broad-spectrum antiviral, but the “address label” can tailor it to go directly to particular organs. So if you have the flu, you would get lung-directed Draco; if you have a brain infection, it would go to your head.
Many virologists remain doubtful that any of these approaches will succeed in creating a wonder drug. Just consider the arduous path still faced by the rabies drug that Prosetta researchers have invented. They’re now collaborating with the rabies division at the Centers for Disease Control and Prevention to investigate it further. To do so, they’ll have to start running experiments on neurons, the cells that rabies actually infects. They’ll have to work out the proper dosage of the drug. They’ll have to start testing it in animals. Potential failure lurks at every step. Sometimes the immune system will attack antiviral drugs as if they were an invader. Drugs that work in mice are sometimes useless in humans.
Even success might bring unanticipated problems. That’s certainly the lesson from the story of broad-spectrum antibiotics. If you swallow a Cipro pill for a salmonella infection, it will wipe out not just the salmonella but many other beneficial bacteria in your gut. Once the salmonella is gone, it may take weeks, months, or even years for the microbial ecosystem to return to something resembling its former state. This disruption can, ironically, allow other pathogens to sneak in and establish themselves. We also depend on bacteria in our bodies to guide our immune systems on the proper path of development. A number of studies suggest that children who get a lot of antibiotics are more at risk of developing immune disorders such as allergies and asthma. (See “Your Own Personal Ecosystem,” an atlas of the human microbiome, in issue 19.10.)
Our bodies are rife not just with bacteria but with viruses too. Even when we’re perfectly healthy, we have trillions of viruses inside of us. Scientists are only beginning to survey this viral ecology, but some suspect that it may actually be essential to our health. Many animals depend on viruses. Aphids, for example, need a virus that makes a toxin that prevents wasps from laying eggs inside their bodies. Scientists have found that infecting mice with lymphotrophic viruses protects them from developing diabetes. Other viruses attack cancer cells.
We may have such beneficial viruses inside our own bodies as well, waiting to be discovered. These viruses may not even infect our own cells but could instead be inside the bacteria that colonize us. Some species might keep the populations of their microbial hosts in check, like predators thinning a herd. Some viruses merge with bacteria rather than killing them, providing their hosts with useful genes for feeding or fighting off competitors. All of these microbe-infecting viruses may ultimately help us stay healthy.
It’s conceivable that a broad-spectrum antiviral could devastate this complex, poorly understood biological jungle. As beneficial viruses disappeared, we might pay the price, developing diseases that the viruses used to keep at bay. Even Lingappa concedes that virus-killing could potentially go too far. “I don’t think we want to kill all viruses,” he says. “You only know about a virus when it does something bad. We’ve evolved with them. There’s probably some virus out there doing something good.”
There’s a book floating around the desks at Prosetta these days: The Mold in Dr. Florey’s Coat. It’s about the discovery of penicillin, but the book’s author, Eric Lax, has dredged up parts of the story that are often forgotten. In particular, few people realize that Fleming abandoned research on penicillin not long after he discovered it. For more than a decade, while Fleming worked on vaccines and other projects, the potential of the drug went untapped. Untold lives were lost to bacterial infections during that time. Fleming’s work was all but forgotten until 1938, when the Oxford pathologist Howard Florey stumbled across his papers and decided to take up where Fleming had left off.
No one wanted to fund Florey’s work at first, so he had to scrape by until he was finally ready to treat his first patient in 1941. The patient, who had been pricked by a rose thorn and developed an infection, died anyway. It took another two years for Florey to build a case powerful enough to persuade the pharmaceutical giant Eli Lilly to start making penicillin on an industrial scale. Florey, along with his collaborator Ernst Chain, shared the Nobel Prize with Fleming. But history tends to forget the 15 years of inaction and struggle that separated the iconic discovery of penicillin from its mass production.
Lingappa wants everyone at Prosetta to read this book. He considers it a necessary education for anyone who would dare to make the penicillin of viruses. “Everyone remembers Fleming,” he says. “Fleming tried and failed, and it took Florey to come a decade later and ask, whatever happened to that stuff Fleming was working on?”
If the true history of penicillin can be any guide for scientists working on antivirals, it teaches them to hunker down and get ready to work for years—to prepare for disappointment, setbacks, and obscurity on the way to finding a drug that works not just in a dish of cells or a mouse but in a human being. The scientists who are working on antivirals today may not be the ones who find that drug. But if they do, future generations may someday tell the story of Vishwanath Lingappa’s rabies or of Todd Rider’s fateful shower.
Copyright 2012 WIRED Magazine. Reprinted with permission.