Advance Could Also Work Against Other Viruses
In research published online today in Science, a team of scientists describe a new therapeutic strategy to target a hidden Achilles’ heel shared by all known types of Ebola virus. Two antibodies developed with this strategy blocked the invasion of human cells by all five ebolaviruses, and one of them protected mice exposed to lethal doses of Ebola Zaire and Sudan, the two most dangerous. The team included scientists from Albert Einstein College of Medicine, U.S. Army Medical Research Institute of Infectious Diseases (USAMRIID), Integrated Biotherapeutics, Vanderbilt University Medical Center, and The Scripps Research Institute.
Ebola viruses cause a highly fatal disease for which no approved vaccines or treatments are available. About two dozen Ebola outbreaks have been documented since 1976, when infections first occurred in villages along the Ebola River in Africa. The largest outbreak in history—the 2014-2015 Western Africa epidemic—caused more than 11,000 deaths and infected approximately 29,000 people.
Monoclonal antibodies, which bind to and neutralize specific pathogens and toxins, have emerged as the most promising treatments for Ebola patients. A critical problem, however, is that most antibody therapies target only one specific ebolavirus. For example, the most promising experimental therapy—ZMappTM, a cocktail of three monoclonal antibodies—is specific for Ebola virus Zaire, and doesn’t work against the other two viruses (Sudan and Bundibugyo), which have both caused major outbreaks. The broad-spectrum antibodies developed by the research team represent an important advance against one of the world’s most dangerous pathogens.
Exploiting Ebola’s Achilles’ Heel
In 2011, a team that included co-senior authors Kartik Chandran, Ph.D. professor of microbiology & immunology at Einstein, and John M. Dye, Ph.D., chief of viral immunology at USAMRIID, discovered that all filoviruses (the family to which ebolaviruses and the more distantly related Marburg virus belong) have an Achilles’ heel: To infect and multiply in human cells, they must all bind to a host-cell protein called Niemann-Pick C1 (NPC1).
But capitalizing on that knowledge required a completely new approach to targeting viruses: exploiting the fact that Ebola and many other viruses must enter host cell compartments called lysosomes. Once safely inside the lysosomes, the viruses transform and expose key portions of their exterior that the research team successfully targeted using monoclonal antibodies.
To gain entry to cells, filoviruses bind to the host cell’s outer membrane via glycoproteins (proteins to which carbohydrate chains are attached) that bristle from the virus’s surface. (See illustration.) A portion of the cell membrane then surrounds the virus and pinches off, eventually developing into a lysosome—a membrane-bound, intracellular compartment filled with enzymes to digest foreign and cellular components.
Filoviruses then use the host cells’ resources to break out of their lysosomal “prisons” so they can enter the host cell’s cytoplasm to multiply. Enzymes in the lysosome slice a “cap” from the virus’s glycoproteins, unveiling a site that binds to the NPC1 embedded in the lysosome membrane. NPC1, which normally helps transport cholesterol within the cell, offers Ebola virus its only means of escaping the lysosome and multiplying. By fitting its protein “key” into the NPC1 “lock,” the virus fuses itself to the lysosome membrane. (See illustration close-up.) Now the virus can propel its RNA from the lysosome and into the cell’s cytoplasm, where it can finally replicate itself.
Penetrating an Invisibility Cloak
The research team realized that monoclonal antibodies could potentially thwart all filovirus infections by neutralizing the viral protein that binds to NPC1, or by neutralizing NPC1 itself. There was just one problem: Reflecting Ebola’s ingenuity, both targets reside only in lysosomes deep within cells—making them invisible to the immune system and shielded from attack by conventional antibodies.
Dr. Chandran, Dr. Dye and co-senior author Jonathan R. Lai, Ph.D., associate professor of biochemistry at Einstein and an expert in engineering antibodies, devised a clever “Trojan Horse” strategy for overcoming the virus’s invisibility cloak: Just as the citizens of Troy unwittingly pulled a wooden horse filled with Greek soldiers into their walled city, they tricked the viruses into carrying the means of their own destruction along with them into host cells.
To do so, the research team synthesized two types of “bispecific” antibodies, each consisting of two monoclonal antibodies combined into one molecule. One bispecific antibody was devised to neutralize the viral protein that binds to NPC1, the other to target NPC1. Both had one monoclonal antibody in common: antibody FVM09, which binds to the surface glycoproteins of all ebolaviruses while the virus is outside cells, allowing the bispecific antibodies to hitch a ride with the virus into the lysosome. FVM09 was developed by co-senior author M. Javad Aman, Ph.D. at Integrated Biotherapeutics.
Once in the lysosome, the bispecific antibodies are released from the viral surface when enzymes in the lysosome slice off the glycoprotein caps—allowing the business ends of the bispecific antibodies to swing into action.
One bispecific antibody combined FVM09 with antibody MR72, which was isolated from a human survivor of Marburg virus infection by co-senior author James E. Crowe Jr., M.D., director of the Vanderbilt Vaccine Center. MR72 targets the NPC1-binding viral protein that is unveiled by all filoviruses in lysosomes. The second bispecific antibody links FVM09 to antibody mAb-548, developed at Einstein, which zeroes in on NPC1. With one bispecific antibody targeting the “lock” (NPC1) and the other targeting the “key” (the virus’s NPC1-binding protein), both had the potential for preventing Ebola virus from interacting with NPC1 and escaping from the lysosome into the cytoplasm.
Putting Antibodies to the Test
The researchers then tested their bispecific antibodies against ebolaviruses in the lab. They initially used a harmless virus (vesicular stomatitis virus) that had been genetically engineered to display glycoproteins from all five ebolaviruses on its surface. The researchers incubated the bispecific antibodies with the Ebola-like viruses and then added the mixtures to human cells in tissue culture. Both bispecific antibodies successfully neutralized all five viruses. Work in the high-containment facilities at USAMRIID confirmed that these antibodies also blocked infection by the actual Zaire, Sudan, and Bundibugyo ebolaviruses.
Next came studies at USAMRIID to test whether the two bispecific antibodies could protect mice infected with the two most dangerous ebolaviruses, Zaire and Sudan. Researchers, led by Dr. Dye, administered the bispecific antibodies two days after mice were exposed to a lethal dose of virus.
The bispecific antibody that targeted the viral binding protein provided good protection to mice exposed to both viruses. As expected, the bispecific antibody that targeted NPC1 did not protect mice. It was designed to bind specifically to human NPC1, which differs slightly in structure from the NPC1 protein found in mice.
As a next step, both bispecific antibodies will need to be tested in nonhuman primates, the current gold standard for anti-Ebola therapeutics.
Scientists have identified a new “multicomponent” virus — one containing different segments of genetic material in separate particles — that can infect animals, according to research published today in the journal Cell Host & Microbe.
This new pathogen, called Guaico Culex virus (GCXV), was isolated from several species of mosquitoes in Central and South America. GCXV does not appear to infect mammals, according to first author Jason Ladner, Ph.D., of the U.S. Army Medical Research Institute of Infectious Diseases (USAMRIID). However, the team also isolated a related virus — called Jingmen tick virus, or JMTV — from a nonhuman primate. Further analysis demonstrates that both GCXV and JMTV belong to a highly diverse and newly discovered group of viruses called the Jingmenvirus group.
Taken together, the research suggests that the host range of this virus group is quite diverse–and highlights the potential relevance of these viruses to animal and human health.
“Animal viruses typically have all genome segments packaged together into a single viral particle, so only one of those particles is needed to infect a host cell,” Ladner explained. “But in a multicomponent virus, the genome is divided into multiple pieces, with each one packaged separately into a viral particle. At least one particle of each type is required for cell infection.”
Several plant pathogens have this type of organization, but the study published today is the first to describe a multicomponent virus that infects animals.
Working with collaborators including the University of Texas Medical Branch and the New York State Department of Health, the USAMRIID team extracted and sequenced virus from mosquitoes collected around the world. The newly discovered virus is named for the Guaico region of Trinidad, where the mosquitoes that contained it were first found.
In collaboration with a group at the University of Wisconsin-Madison, the USAMRIID investigators also found the first evidence of a Jingmenvirus in the blood of a nonhuman primate, in this case a red colobus monkey living in Kibale National Park, Uganda. The animal showed no signs of disease when the sample was taken, so it is not known whether the virus had a pathogenic effect.
Jingmenviruses were first described in 2014 and are related to flaviviruses — a large family of viruses that includes human pathogens such as yellow fever, West Nile and Japanese encephalitis viruses.
“One area we are focused on is the identification and characterization of novel viruses,” said the paper’s senior author Gustavo Palacios, Ph.D., who directs USAMRIID’s Center for Genome Sciences. “This study allowed us to utilize all our tools–and even though this virus does not appear to affect mammals, we are continuing to refine those tools so we can be better prepared for the next outbreak of disease that could have an impact on human health.”
While it is difficult to predict, experts believe that the infectious viruses most likely to emerge next in humans are those already affecting other mammals, particularly nonhuman primates.
See NPR.org for a more detailed explanation . . .
The United States Army Medical Research Institute of Infectious Diseases (USAMRIID; pronounced: you-SAM-rid) is the U.S Army’s main institution and facility for defensive research into countermeasures against biological warfare.
It is located on Fort Detrick, Maryland and is a subordinate lab of the U.S. Army Medical Research and Materiel Command (USAMRMC), headquartered on the same installation.
USAMRIID is the only U.S. Department of Defense (DoD) laboratory equipped to study highly hazardous viruses at Biosafety Level 4 within positive pressure personnel suits.
USAMRIID employs both military and civilian scientists as well as highly specialized support personnel, in all about 800 people. In the 1950s and ’60s, USAMRIID and its predecessor unit pioneered unique, state-of-the-art biocontainment facilities which it continues to maintain and upgrade. Investigators at its facilities frequently collaborate with the Centers for Disease Control and Prevention, the World Health Organization, and major biomedical and academic centers worldwide.
USAMRIID was the first bio-facility of its type to research the Ames strain of anthrax, determined through genetic analysis to be the bacterium used in the 2001 anthrax attacks.