Research Roundup: Viral hepatitis, treatment for persistent Ebola, E. coli–based vaccines

Earlier this year, the World Health Organization (WHO) declared the outbreak of Zika in Latin America a Public Health Emergency of International Concern. The virus, spread by the Aedes aegypti mosquito, was once considered innocuous, causing a mild illness and lasting for only a handful of days. However, the virus has since been linked to devastating birth defects and neurological complications and has spread to more than 60 countries. The virus’s virulence took the research community by storm: although it was discovered in 1947, only 25 scientific papers were published on the topic prior to the current outbreak. As the scientific community learns more about the virus and the disease, the need for tools to prevent, detect, and treat it has become undeniable. On February 22, US President Barack Obama submitted a request to Congress for US$1.9 billion in emergency funding to combat the virus, a critical move, as the US government is unrivaled as the largest funder of global health research and development (R&D). However, 146 days later, Congress has yet to reach a deal, and the US agencies leading the fight against Zika are running out of money. In the meantime, these agencies have made impressive strides in advancing Zika R&D using limited funding and leveraging cross-sector partnerships. An unprecedented number of private-sector partners have also entered the hunt for Zika countermeasures, building out a robust pipeline in record time. Below, we highlight the most promising vaccine, drug, and diagnostic candidates as well as vector control approaches. Vaccines There are currently more than 35 Zika vaccine projects underway, and at least 3 have completed successful animal trials. Inovio Pharmaceuticals and GeneOne Life Sciences partnered to develop a DNA-based vaccine, using a unique delivery approach—electroporation—in which a device is applied to the injection site, sending out electrical pulses that enhance the vaccine’s ability to enter human cells. The vaccine successfully prompted an immune response, including the production of protective antibodies, in both mice and monkeys. In late June, the experimental vaccine became the first to be approved for human clinical trials by the US Food and Drug Administration (FDA). The phase 1 trial, which will enroll 40 healthy adults, is intended to evaluate the safety and tolerability; however, the team will also monitor its ability to induce an immune response. Meanwhile, two more vaccine candidates have proven effective in mice. The candidates include a DNA-based vaccine and one composed of an inactivated version of the Zika virus, the first supported by the US National Institutes of Health (NIH) and the second developed by the US Walter Reed Army Institute of Research. The latter will be tested in phase 1 clinical trials by the US National Institute of Allergy and Infectious Diseases and then transferred to French pharmaceutical company Sanofi for further clinical development. Sanofi is also working on a vaccine of its own; however, it won’t enter clinical trials until 2017. After several months of feasibility studies, British pharmaceutical company GlaxoSmithKline has announced that it will be partnering with the NIH to test its RNA-based vaccine in animals. To date, no DNA or RNA vaccines have been approved for use in humans, although they are often called the future of vaccines. Drugs Due to the dearth of knowledge about the virus and the difficulty of conducting clinical trials and using novel drugs in pregnant women—a key target population for Zika control—the WHO has stated, “It is unlikely at this stage that therapeutics will constitute a priority activity.” However, efforts are underway, both to discover novel treatments and to repurpose existing medicines. Australian biotech company Biotron announced recently that two compounds in their library have proven effective against Zika in test tubes. They will continue to conduct laboratory tests to better assess the safety and efficacy of the compounds in humans. Researchers at the US National Center for Advancing Translational Sciences are screening their compound library—with half a million entities— and have identified a novel drug, which is safe for use in humans and currently in clinical trials for cancer, that prevented the virus from killing progenitor cells, which play a critical role in the neurological system and fetal development. Researchers at the University of California, San Diego are embarking on an ambitious study to virtually screen more than 20 million compounds against the virus. The program—OpenZika—uses a research collaboration platform to harness the power of volunteers’ computers and smartphones. Participants download the software, and when their computer or phone is inactive, it runs calculations to predict how a compound will interact with the virus in the lab. Researchers are also looking at repurposing existing drugs as Zika treatments. Chloroquine—which both prevents and treats malaria—has successfully protected against Zika virus in vitro. A team at Katholieke Universiteit Leuven has identified an experimental treatment for hepatitis C that delayed the manifestation of Zika-related symptoms in mice. And scientists at the University of California, San Francisco reported that the antibiotic azithromycin—which has long been used in pregnant women—blocked the virus from multiplying in petri dishes. Diagnostics As of March, more than 30 diagnostics for the virus were under development, and the FDA has sanctioned the use of five tests—four that detect current infection and one to assess previous exposure—under its Emergency Use Authorization program. However, rapid, point-of-care diagnostics are desperately needed. Eighty percent of cases are asymptomatic, and initial symptoms—fever, rash, joint pain—are nonspecific. Thus, early and accurate diagnosis is critical to ensure appropriate treatment. A team of Harvard University and Massachusetts Institute of Technology researchers have designed a paper-based test that can diagnose cases within a few hours. The device is appropriate for low-resource settings: it’s cheap, it can be stored or shipped at room temperature, and the results are easy to interpret. The tool contains miniscule sensors that change color in the presence of a specific genetic sequence. To ensure accurate detection of the virus, the team developed sensors that can detect 24 different genetic sequences found in the Zika genome. The device has been tested using samples from infected monkeys and has proven to be effective at detecting even low quantities of the virus. Although the results are easy to read—a positive sample turns the strip purple—the team has also created a simple electronic reader that determines the quantity of the virus in the sample. Vector Control As vaccines, drugs, and diagnostics against the virus inch their way through the R&D pipeline, health officials are ramping up vector control efforts against the Aedes aegypti mosquito. New techniques are also underway, and the FDA is currently accepting public comment on a proposal to expand testing of genetically engineered mosquitoes. The mosquitoes mate with the local population, resulting in offspring that die before reaching adulthood and are consequently not able to spread diseases. A trial of the mosquitoes in Brazil led to an 82 percent decrease in the Aedes aegypti population in just nine months. Scientists are using innocuous bacteria and fungi to combat the spread of Zika. Male mosquitoes infected with the Wolbachia bacteria do not produce viable offspring, and tests across the United States have reduced the population of the closely related Aedes albopictus mosquito by an estimated 70 percent. Meanwhile, the Metarhizium brunneum fungus is used as a natural pesticide, killing mosquito larvae. Meanwhile, in North Hempstead, New York, bats have been enlisted in the fight against Zika. The city is building and hanging bat houses in parks to attract the animals, as they can eat more than 1,000 mosquitoes in an hour.

Just one case of yellow fever in an unvaccinated population is considered an outbreak, according to the World Health Organization (WHO). And now, an outbreak in Angola has spread to Kinshasa, a city of 12 million in the Democratic Republic of Congo. Health experts are calling it the worst yellow fever outbreak in Africa in decades. The outbreak in Angola has resulted in nearly 350 deaths, and unrelated outbreaks are ongoing in Uganda and Peru. Meanwhile, the WHO’s emergency stockpile of the yellow fever vaccine—the only available countermeasure against the virus—has been depleted twice this year, and is currently running low at only 6 million doses, raising concerns about global ability to respond to growing outbreaks.Background: The illness and impact of a historic virus Yellow fever is a mosquito-borne disease—spread by both the Aedes and Haemogogus species—found in both urban and rural settings. The disease is endemic in forty-seven countries across Africa and Latin America, and each year results in anywhere from 84,000 to 170,000 severe cases and 29,000 to 60,000 deaths. Many cases of yellow fever are asymptomatic, and when they manifest, initial symptoms are nonspecific: fever, fatigue, aches and pains, nausea, and vomiting. Fifteen percent of cases progress to a more severe illness  characterized by high fevers, liver and kidney damage, jaundice—thus the name yellow fever—and even hemorrhaging. Half of those who enter this toxic stage of infection die. While the yellow fever virus has plagued the world since the 17th Century, it was first isolated in 1900 by American pathologist Walter Reed, earning it the distinction of being the first human virus ever to be discovered. A safe and effective vaccine was first introduced in 1936; however, in the eight decades since, few new tools against the virus have been developed. Below, we break down the status of research and development (R&D) for vaccines, drugs, and diagnostics for yellow fever.Status of R&D and access to yellow fever countermeasures Vaccines Yellow Fever vaccine manufacturing facility Val de Reuil Sanofi Pasteur site, France (Photo: Sanofi Pasteur / Patrick Boulen)The yellow fever vaccine is effective, safe, and affordable. Just one dose provides lifetime protection from the virus for 99 percent of those vaccinated. However, the vaccine contains live virus, and in extremely rare cases, can result in illness. Since 2000, the price of the vaccine has risen 30 percent annually. However, this increase has not incentivized manufacturers to scale up production. There are only four manufacturers of the vaccine, and it’s notoriously difficult to produce. Consequently, the global demand for the vaccine surpasses availability by 42 percent. Yellow fever vaccine production relies on eggs from pathogen-free chickens flocks, which are hard to come by. The embryos are infected with the yellow fever virus, and after three to four days, the embryos are harvested under sterile conditions. Each egg can provide anywhere from 100 to 300 doses of the vaccine. The entire production process takes about one year. A vaccine candidate with an inactivated version of the virus completed successful phase 1 clinical trials in 2011. However, since GE Healthcare acquired the intellectual property for the vaccine in 2012, no additional clinical trials have been undertaken. Fortunately, last month PnuVax Incorporated entered into an agreement with GE Healthcare, under which PnuVax will advance the vaccine through clinical trials and will partner with GE Healthcare for manufacturing. In the meantime, the WHO is recommending that the vaccine be administered in fractional doses, using just one-fifth of the standard amount. Studies suggest that this approach still confers at least 12 months of protection from the virus, however, booster shots would be required to sustain immunity when a global shortage is no longer imminent. Additionally, fractional dosing has not been tested in children, raising concerns that this approach might not be sufficient for kids, whose immune responses, including to vaccines, are typically weaker. Drugs There are no specific therapies for yellow fever. Current treatment involves supportive care (i.e., addressing the symptoms). Animal studies suggest that the widely-used antiviral ribavirin and an active carboxamide drug reduce mortality in hamsters, and the molecule interferon-α, used in some cancer therapies, effectively treats the disease in monkeys. However, these treatments were only effective when administered within a few days of infection. Thus, even if they were to prove effective in humans, their application would be limited. The yellow fever virus incubates for three to four days in the body before symptoms appear, and as aforementioned, initial symptoms are nonspecific and easily confused with anything from malaria to dengue fever or leptospirosis to poisoning. Diagnostics As yellow fever is hard to distinguish from other illnesses based on initial symptoms alone, the development of rapid, point-of-care diagnostics is critically needed. Currently, cases can only be confirmed in laboratory settings. Diagnosis requires examining blood or serum for either the virus itself or antibodies against the virus. However, researchers at the Massachusetts Institutes of Technology are currently developing a paper-based diagnostic that simultaneously tests for Ebola, dengue fever, and yellow fever. The test is designed for use in low-resource settings, requiring no electricity or specialized lab equipment and the results are easy to interpret. Testing requires a small amount of blood, and colored nanoparticles appear—red for Ebola, orange for yellow fever, and green for dengue—based on the proteins in the blood. While this new tool is showing promise, it could be some years before testing is completed and regulatory approval is secured. With a limited pipeline of countermeasures against yellow fever, the global community is scrambling to ramp up vaccine production, expand vaccination coverage, and scale up vector control efforts. However, R&D for rapid, point-of care diagnostics; lifesaving treatments; and a safe, affordable, easy-to-produce vaccine is critical to combat this virus after a centuries-long battle.