Oct 5, 2020 7:00 AM

A Common Plant Virus Is an Unlikely Ally in the War on Cancer

Researchers have seen promising results by injecting dog and mouse tumors with the cowpea mosaic virus. Now they’re aiming for a human trial.

Jack Hoopes spends a lot of time with dying dogs. A veterinary radiation specialist at Dartmouth College, Hoopes has spent his decades-long career treating canine cancers with the latest experimental therapies as a pathway for developing human treatments. Recently, many of Hoopes’ furry patients have come to him with a relatively common oral cancer that will almost certainly kill them within a few months if left untreated. Even if the cancer goes into remission after radiation treatment, there’s a very high chance it will soon reemerge.

For Hoopes, it’s a grim prognosis that’s all too familiar. But these pups are in luck. They’re patients in an experimental study exploring the efficacy of a new cancer treatment derived from a common plant virus. After receiving the viral therapy, several of the dogs had their tumors disappear entirely and lived into old age without recurring cancer. Given that around 85 percent of dogs with oral cancer will develop a new tumor within a year of radiation therapy, the results were striking. The treatment, Hoopes felt, had the potential to be a breakthrough that could save lives, both human and canine. “If a treatment works in dog cancer, it has a very good chance of working, at some level, in human patients,” says Hoopes.

The new cancer therapy is based on the cowpea mosaic virus, or CPMV, a pathogen that takes its name from the mottled pattern it creates on the leaves of infected cowpea plants, which are perhaps best known as the source of black-eyed peas. The virus doesn’t replicate in mammals like it does in plants, but as the researchers behind the therapy discovered, it still triggers an immune response that could be the key to more effective treatments for a wide variety of cancers.

The idea is to use the virus to overcome one of the gnarliest problems in oncology: A doctor’s best ally, their patient’s own immune system, doesn’t always recognize a cancerous cell when it sees one. It’s not the body’s fault; cancer cells have properties that trick the immune system into thinking nothing is wrong. Oncologists have puzzled over this for nearly a century, and it's only in the past decade that researchers have really started to get a grip on cancer’s immunosuppressive properties. Immunotherapy, which has emerged as one of the most promising types of cancer treatment, is all about developing techniques to help the body’s immune system recognize cancerous cells so it can fight back. It’s the medical equivalent of putting a big flashing neon sign on the tumor that reads “ATTACK HERE.” And that’s where the cowpea mosaic virus could help.

To treat his canine patients, Hoopes typically injects 200 micrograms of virus-like particles—about three times the dose of a typical flu vaccine—directly into their tumors. These particles are not live cowpea mosaic viruses; rather, they’re viruses that have had their genetic material removed or have been inactivated so they can’t replicate. Each pup receives four doses of the viral particles over two weeks while also taking standard radiation therapy. The dog’s immune system recognizes the pathogens as foreign bodies and goes into attack mode. When the body goes after the particles, it takes the cancerous cells down with them.

While other viruses could theoretically be used as immune system bait, CPMV has proven far more effective at triggering a response than any other pathogens the researchers have tried so far. They’re still not sure what makes this particular virus so uniquely effective, but the important thing is that it works. “It’s worked better than radiation by itself, which is a huge positive for us,” says Hoopes. “The immune system is more powerful than we thought.”

Viruses are microscopic zombies that are a natural analog of artificial nanoparticles. Not only are they small enough to invade cancerous cells—most are only a few dozen nanometers long—they can also be genetically reprogrammed to do specific tasks. Even better, cancer-fighting viruses are relatively cheap to make because they’re self-replicating, and they don’t require outside interventions once they’re injected into a tumor.

“Viruses are easy to work with and can be altered to obtain more favorable properties,” says Jan Carette, an immunologist at Stanford University and an expert on viral therapeutics, who was not involved with the CPMV research. “They’re highly flexible and manipulatable molecular machines that can be used in therapy, either through direct cancer-cell-killing properties or by stimulating anti-tumor immune responses.”

One approach to cancer immunotherapy uses genetically modified oncolytic viruses that invade tumor cells and start replicating until the cells explode. This releases a bunch of cancerous gunk into the body that signals to the immune system that something isn’t right. In response, it goes into hyperdrive trying to flush out the cancerous material.

“The real promise of the oncolytic virus is that it takes the virus’s natural ability to stimulate an immune response and transfers that to cancer cells by tricking the immune system into think that the tumor cell is actually a virally infected cell,” says Howard Kaufman, an expert on oncolytic viruses and immunotherapy at Harvard Medical School who led the Phase III clinical trial for the only oncolytic virus to be approved for treatments in the US.

There are concerns about uncontrolled viral replication in the patient and unintended transmission of the viral infection to other people, but mostly researchers have struggled to direct these viral suicide bombers to the tumor once they’re in the body. “The delivery issue is probably the biggest challenge for oncolytic viruses,” says Kaufman.

“None of the oncolytic viruses have shown a very good effect in clinical trials,” says Steve Fiering, an immunologist at Dartmouth College and one of the leaders of the research team working on plant viruses. To date, only three oncolytic virotherapies have been approved as cancer treatments worldwide, and only the one Kaufman helped develop is approved in the US. Two of these therapies are used for treating melanoma, and the other is for treating head and neck cancer. As detailed in a paper published earlier this year in Frontiers in Oncology, there are clinical trials underway around the world studying oncolytic viruses for treatment of liver, lung, pancreatic, ovarian, breast, and prostate cancers, but so far, the paper’s authors wrote, their efficacy “remains largely unknown.”

Like chemotherapy and many other cancer treatments, oncolytic virotherapies are typically delivered into the body with an intravenous injection and must search out the cancerous cells. The alternative is to do a local intervention such as surgery. But cancer is often not confined to a single region—this is known as metastasis—or surgically accessing the tumor may be difficult. And if you only treat the cancer in one spot, it could come back with a vengeance. By taking the systemic route and delivering the virus by IV, there’s a greater chance the virus will find any wayward cancer cells and kill them along with the main tumor. The trade-off is that fewer viruses will find their way to the main tumor and the immune response will be weaker.

“If you have a metastatic disease, oncologists always use systemic treatments,” says Fiering. “I think that’s fine, but it’s missing one of the fundamental ideas of immunology, which is that the response you get in one part of the body can distribute throughout the body.”

A familiar example of this is flu shots, which are delivered in your arm but trigger an immune response that protects against a respiratory infection. Fiering started to wonder if a similar approach might be taken with cancer. His idea was that if doctors injected something into a tumor that would cause the body’s immune system to start attacking it, the heightened immune response wouldn’t be limited to just the area around the tumor. The immune system’s T-cells—its frontline soldiers—would also track down any cancer cells that might be lurking elsewhere in the body.

It was an elegant idea, but Fiering had a hard time finding the right stuff to inject into a tumor that would alert the immune system to the attack target. At first, he focused on single-celled parasites and bacteria, but those didn’t elicit the sort of strong immune response that the body would need to take on a tumor. Mammalian viruses didn’t work much better. It was only after attending a talk about plant viruses in medicine by Nicole Steinmetz, a nanoengineer at the University of California, San Diego, that Fiering saw a way forward. Steinmetz and other researchers had shown that plant viruses have useful properties as vaccine-delivery platforms and adjuvants, an ingredient in a vaccine that increases the body’s immune response. It got Fiering thinking: Maybe he could harness this same effect to fight cancer, too.

For more than two decades, Steinmetz has been studying ways to modify plant viruses to do useful things like delivering cancer therapies and vaccines in animals, and treating diseases in plants. “I like to joke that we use dirt and sunlight to produce nanotechnology,” says Steinmetz. “But that’s essentially what we do. We grow plants, infect them, and then harvest the virus. The plant is our bioreactor.”

While he listened to Steinmetz present her work on plant viruses, it dawned on Fiering that those same pathogens might be useful in his work on cancer immunotherapies. After Steinmetz’s talk, he pitched her on a collaboration. It wasn’t something she'd tried before, but she was willing to give it a shot. “We had been developing virus-like particles as cancer therapies and vaccines, so the proposal made sense,” says Steinmetz. “We just never thought about injecting that material directly into the tumor.”

For Steinmetz, the question was which virus to use. There are just over 1,000 known species of plant viruses, but as Fiering and Steinmetz figured out, not all of them are equally good at stimulating the body’s immune system. Because plant viruses aren’t really a threat to humans, the body’s immune system typically doesn’t treat them like one.

In 2015, Steinmetz sent Fiering some cowpea mosaic viruses to test on mice in his lab. It’s one of the best characterized plant viruses; Steinmetz describes it as the “go-to virus” in her medical research. The viral particles are symmetrical, which makes it easy to precisely add molecules to the outside of each one, and they are easy to produce in plants in large quantities.

It seemed like as good a starting point as any, and when the team tested it on tumors in lab mice, it proved to be incredibly effective. As detailed in a paper published later that year in Nature Nanotechnology, the research team discovered that the cowpea mosaic virus was highly effective in treating melanoma, breast, ovarian, and colon tumor models in mice. (Tumor models are growths that are caused by the injection or implantation of cancerous cells into healthy mice.) They found that in all tumor models tested, the plant viral therapy reduced the rate of tumor growth. Depending on the tumor model, growth was slowed by an average of 50 to 100 percent over a two-week period. In some models, it caused the tumor to disappear completely.

“We were very lucky that we started with the cowpea mosaic virus or we wouldn’t necessarily have made this discovery,” says Steinmetz. “It sounds almost too good to be true.”

Moreover, the virus created an immune memory in most mice, which meant that once a tumor was gone, the cancer was highly unlikely to come back. The researchers tested this by re-injecting tumor cells in mice after the original tumor had completely regressed or been surgically removed. If the tumor cells didn’t replicate the second time around, it indicated that the mouse’s immune system “remembered” them and would begin attacking the cells to prevent a resurgence.

This is particularly important for breast cancer, which has a high rate of recurrence after the original tumor is treated. “We showed that it's effective in a variety of tumors and could eliminate many of those tumors,” says Fiering. “Like most therapies, it’s more effective against some tumors than others.” In these experiments, the team found that it was particularly effective against the melanoma and ovarian cancer models, and less effective against the breast cancer models. But Fiering cautions that the results don’t mean that the therapy is less effective against breast cancer in general; rather, the results are highly dependent on the immunosuppressive characteristics of the particular cancer model they’re tested on.

In 2017, Fiering and Steinmetz teamed up with Hoopes to begin testing their plant viral therapy in the dogs that he was treating. Dogs make a better model for studying cancer in humans than mice do. First, the dogs that Hoopes was treating had developed their cancers naturally, just like humans do; mice, by comparison, are injected with cancer cells by researchers. Dogs are also more genetically diverse, whereas lab mice are effectively clones of one another. Dogs also are prone to a wide variety of cancers, and their tumors are similar in size and cell count to those found in humans. “The dog is an incredible model, because they share the environment with us,” says Hoopes. “It’s the best model outside of human patients.”

Humans typically receive a combination of different cancer therapies, and the same is true of Hoopes’ canine patients. He treats them with a mix of radiation and an injection of cowpea mosaic virus particles in the tumor. That makes it challenging to separate out the effects of each treatment, but Fiering calls the results so far “striking.” The researchers have only published results on about six dogs with oral cancer, but Fiering says more than 20 dogs have been given the viral therapy at this point and that the team is beginning to try the therapy on other canine cancers. Of the six dogs with oral cancer—which has an 85 to 90 percent recurrence rate within a year of radiation treatment and a similarly high mortality rate—none had their cancer return after being treated with a combination of radiation and CMPV injections.

“A variety of the dog patients live much longer than would be predicted based on the prognosis of the cancer type with no signs of recurrence,” says Fiering. In other words, the combination of radiation and viral injections was more effective than radiation alone. But this doesn’t mean the CMPV injections are a cancer wonder drug. Fiering strongly emphasized that the virus injections are most effective when used in combination with other cancer therapies. And like any cancer treatment, it’s not going to work for everyone or kill every kind of tumor.

“I think this is a really original and creative concept with a lot of potential, especially because the nanoparticles can be further modified to better target cancer cells,” says Carette. But he also noted that this isn’t the first time that a promising immune system stimulant has been enlisted in the fight against cancer. About a decade ago, an experimental therapy that used modified bacteria to stimulate the body’s immune system in a similar way to the CMPV particles looked really promising in preclinical experiments, but it didn’t perform as well in clinical studies.

“Although I am enthusiastic about the potential, several challenges remain,” Carette says. One of the biggest unknowns is why CPMV appears to be so good at triggering a cancer-fighting immune response. He says if plant virus therapies are going to make it out of the lab and into the clinic, “a better understanding of how the nanoparticles stimulate the immune response would be useful.”

The CPMV team is working on it. In a paper published in September, Steinmetz and a team of researchers found that the cowpea mosaic virus was more effective as a cancer vaccine in mice than five other viruses—both plant and animal—with similar sizes and shapes. “Why this virus is so much more potent than other viruses is a huge focus of our research,” says Steinmetz. “Originally, we thought it could be the size and shape, but similar particles don’t induce these effects.” She and her team have also tested cowpea mosaic viruses with their genetic material removed and found that unmodified cowpea mosaic viruses appear to work best. She says the body’s immune system seems to recognize specific molecules in the virus. Now the team is working to figure out why this affinity exists.

The fact that the plant viruses appear to stimulate a sustained immune response could be a significant advantage compared to virotherapies that directly attack the tumor cells, says Kaufman. “An oncolytic virus is so effective at getting an immune response that the body can often clear the virus pretty quickly, and then you lose the cancer treatment effects,” he says. “That’s where this work is potentially very interesting, because plant viruses in general might be a little less immunogenic than mammalian viruses and they could offer a real advantage in the therapeutic setting. But that hypothesis awaits clinical confirmation.”

This summer, Steinmetz, Fiering, and Shaochen Chen, another nanoengineer at the University of California, San Diego, received a $2.9 million grant from the National Institutes of Health to develop a bioprinted implant to treat ovarian cancer with plant viruses. Most women diagnosed with ovarian cancer undergo surgery to remove the tumor, but the risk of recurrence of the cancer is high. Steinmetz says the idea is to insert the implant near the tumor during the surgery so that it can release plant viruses at regular intervals to ensure the cancer doesn’t return or crop up elsewhere. The five-year grant will fund research on an implant that can be bioprinted—a method of 3D printing using living cells instead of inorganic materials—and will then study the most effective schedules for releasing viruses into the body. Steinmetz says the team is still in the early phases of designing the implants, and they will be tested on mice—an important step toward convincing the FDA that the implant is safe enough for a clinical trial in people.

Earlier this year, Fiering and Steinmetz cofounded a company called Mosaic IE to facilitate a human trial using injected cowpea mosaic viruses. Before they can run that trial, they’ll have to do a large-scale toxicology study, which will involve systematically giving mice larger and larger doses of the virus to determine its toxicity. They’ll also have to demonstrate that they can reliably produce the virus at scale, which is critical to ensuring human safety. “Everything has to be very, very carefully documented and tested before you can begin clinical trials,” says Fiering. Still, the team is optimistic that an initial clinical trial could begin in just a few years.

While their viral therapy is certainly not the cure for cancer, it has the potential to improve survival rates for a variety of cancers when it's used together with other therapies. “Ultimately, the best use of these is going to be in combination with other drugs,” says Kaufman. Steinmetz and Fiering both agree, but that’s not a knock against the potential of plant virus therapies. In fact, if it works well with other therapies, that would be an advantage.

“Cancer is a statistics game,” says Fiering. “The best-case scenario would be that in a relatively broad range of cancers, it would improve the outcome for a significant proportion of people.” And in the fight against cancer, we need all the help we can get.