Virus World

A new monoclonal antibody cocktail neutralizes a dangerous virus circulating West Africa: Lassa Virus.  Monoclonal antibodies are among our greatest assets in treating and preventing virus-induced disease. While the spotlight has focused squarely on Covid-19 monoclonal antibodies throughout the pandemic, antibody candidates for other severe pathogens have also made strides forward. Here we describe a new antibody cocktail that neutralizes a lesser known but nonetheless dangerous virus circulating West Africa: Lassa Virus.

What is Lassa Virus?

Lassa Virus is a pathogen responsible for Lassa hemorrhagic fever, which afflicts between 300,000 and 500,000 people per year, commonly in West African countries, including Nigeria, Liberia, Sierra Leone, Guinea, and Ghana. Lassa fever has a mortality rate of roughly one percent. Pregnant women are at the greatest risk of death, with up to a 90% fatality rate. The symptomatic cases present issues such as fever, headaches, vomiting, and muscle pain. Spread typically occurs via contact with infected mice excrement or urine, though direct contact spread from person to person is also common. Unfortunately, a vaccine is not yet available for the virus, and antivirals are weak at best. Lassa Virus, like SARS-CoV-2, is an RNA virus that constantly mutates. A therapy or prophylactic to fill the void of Lassa Virus drugs is needed.

Lassa Virus Monoclonal Antibody Therapy

Fortunately, a new candidate may be just around the corner. A group of researchers from the La Jolla Insitute for Immunology in California led by Dr. Erica Ollmann Saphire have identified a cocktail of three antibodies that bind and neutralize the Lassa Virus. This same group discovered a broadly accepted cocktail of monoclonal antibodies for Ebolavirus, which we have previously described. This cocktail arrived just in time as the current to treat the current epidemic caused by the Sudan strain of Ebolavirus in Nigeria.  The new Lassa Virus therapy, denoted Arevirumab-3, is comprised of three distinct antibodies, each one binding distinct regions of the Lassa Virus glycoprotein. The Lassa Virus spike protein comes in a set of three identical subunits embedded into the membrane. The virus is synthesized as a single long polypeptide and is cleaved into three parts, a leader sequence, GP1 which encodes the receptor-binding function, and GP2 embedded into the membrane (Figure 1).

8.9F inhibits virion-cell attachment by blocking the GPC/α-DG interaction

The first antibody, 8.9F, binds the top of the spike trimer, stretching across all three portions of the glycoprotein. 8.9F is an unusual antibody as the single antibody binds all three faces of the trimer. The antibody also binds the three cleavage sites of the GP1 subunits. It binds the portion of the glycoprotein that attaches to the host cell, directly inhibiting virion-cell attachment by mimicking the cell alpha-dystroglycan receptor. The 8.9F antibody also specifically binds a glycan, N119, which is required for neutralization activity. This glycan is required for alpha-dystroglycan receptor-binding, so 8.9F’s recognition is surprising.

A native N89 glycan plays a central role in 12.1F/LASV GPC recognition

The second antibody, 12.1F, binds to a separate site on GP1, directly interacting with N-glycans for neutralization activity. The antibody binds more membrane-proximal and has direct interaction with six crucial amino acid residues and critical glycans N89 and N109. To infect a cell, the Lassa Virus must bind the cell membrane and be engulfed by a cellular endosome. Then the protein undergoes a structural change to bind the LAMP-1 protein in the endosome interior. Binding to LAMP-1 is essential for fusion of viral and cell membranes and entry into the cytoplasm where replication occurs. 37.2D neutralizes LASV by locking the GPC trimer in an inactive configuration. The third antibody, 37.2D, binds two adjacent subunits of GP2. The antibody stretches across subunits, locking the trimer in place by binding conserved peptides and conserved glycans N390 and N395. 37.2D is special in that it attacks GP2 at a unique angle, stabilizing the pre-fusion trimer, preventing fusion activation and, therefore, infection. These three mechanisms work together to yield a highly effective antibody cocktail.  One critical question for monoclonal antibody treatment, is can the virus escape via mutation in the binding sites? Dr. Ollmann Saphire's team shows that such is the case for treatment with a single 8.9 F antibody. However, no resistance mutations arose when using the cocktail of all three antibodies, nor was the virus capable of escaping. The Arevirumab-3 antibody may provide a long-awaited answer to the lack of treatment and prevention of Lassa infections. This is particularly important for cases involving pregnant women and their fetuses. It will be important to reduce the costs of this antibody cocktail as much as possible so that it is available when needed in West Africa. Current technologies allow monoclonal antibodies to be produced at $200 and $250 per gram.

The next step in the development of the control of Lassa Fever is the development of a vaccine effective against all Lassa Virus variants. This work may serve as a guide for the creation of such a vaccine.

The vaccine provided protection from Ebola virus, Sudan virus, Marburg virus and Lassa virus. Scientists funded by the National Institutes of Health have developed an investigational vaccine that protected cynomolgus macaques against four types of hemorrhagic fever viruses endemic to overlapping regions in Africa. The University of Texas Medical Branch in Galveston and Profectus BioSciences of New York are developing and testing the candidate quadrivalent VesiculoVax vaccine, with support from NIH’s National Institute of Allergy and Infectious Diseases (NIAID) and Redeemer’s University in Nigeria.

The newly published study in the Journal of Clinical Investigation describes how the vaccine was created using a live-attenuated (weakened) vesicular stomatitis virus to deliver proteins that elicit protective immune responses. The proteins are from Ebola virus (Kikwit strain), Sudan virus (Boniface strain, which also causes Ebola virus disease), Marburg virus (Angola strain) and Lassa virus (Josiah strain). There are no licensed vaccines to provide protection from any of those viruses — all of which can cause severe disease and death — although the European Medicines Agency has recommended licensing a VSV-Ebola vaccine. Importantly, the monkeys infected in the study were exposed to different strains of Sudan virus (Gulu) and Lassa virus (0043/LV/14) than those in the candidate vaccine to help the researchers determine whether the vaccine would be cross-protective. Lassa 0043/LV/14 is circulating in an outbreak in Nigeria that began in 2018. Previous studies indicate that the investigational Ebola virus (Kikwit) vaccine will protect against other strains of Ebola virus.

The scientists inoculated 20 macaques with a primary and booster dose of quadrivalent VesiculoVax. The animals had five blood draws to check for an immune response, including on the day of initial vaccination and on days 10 and 28, then on day 56 when they received a booster inoculation, and again on day 66. On day 84 scientists infected the macaques with the four different hemorrhagic fever viruses and monitored them to day 112. Twelve additional macaques in the study who were infected with the four viruses but not vaccinated all became sick, but none of the vaccinated animals did. Only one of the 20 vaccinated animals had any of the four hemorrhagic fever viruses detectible (Lassa) following the study. The scientists state that the addition of the Lassa virus component to their multivalent vaccine is an exciting research advance as they already had developed an investigational trivalent vaccine that provided protection against Ebola, Sudan and Marburg viruses. The researchers now plan further vaccine tests against other strains of Lassa virus, and they want to further evaluate whether a single-dose quadrivalent vaccine appears safe and effective.

Published in the Journal of Clinical Investigation (October 22, 2019):

https://doi.org/10.1172/JCI131958

Filoviruses such as Ebola virus (EBOV) cause outbreaks of viral hemorrhagic fevers for which no FDA-approved vaccines or drugs are available. The 2014–2016 EBOV outbreak in West Africa infected approximately 30,000 people, killing more than 11,000 and affecting thousands more in areas still suffering from the effects of civil wars. Sierra Leone and Liberia reported EBOV cases in every county demonstrating the efficient spread of this highly contagious virus in the well-connected societies of West Africa.

In communities, canines are often in contact with people while scavenging for food, which may include sickly bush animals or, as reported from the outbreak, EBOV infected human bodies and excrement. Therefore, dogs may serve as sentinel animals for seroprevalence studies of emerging infectious viruses. 

The authors  used a multiplex Luminex-based microsphere immunoassay (MIA) to detect dog IgG binding to recombinant filovirus antigens or LASV glycoprotein (GP) in serum from dogs that were old enough to be present during the EBOV outbreak. We identified 47 (73%) of 64 dog serum samples as potentially exposed to filoviruses and up to 100% of the dogs from some communities were found to have elevated levels of EBOV antigen-binding IgG titers.

These data support the feasibility of canines as EBOV sentinels and provides evidence that seroprevalence studies in dogs can be conducted using suitable assays even under challenging field conditions. 

Study published July 22  on PLOS Neglected Tropical Diseases:

https://doi.org/10.1371/journal.pntd.0007614

Long before COVID-19, scientists had been working to identify animal viruses that could potentially jump to people. These efforts have led to a Web-based platform called SpillOver, which ranks the risk that various viruses will make the leap. Developers hope the new tool will help public health experts and policymakers avoid future outbreaks. Jonna Mazet, an epidemiologist and disease ecologist at the University of California, Davis, has led this work for more than a decade. It began with the USAID PREDICT project, which sought to go beyond well-tracked influenza viruses and identify other emerging pathogens that pose a risk to humans. Thousands of scientists scoured more than 30 countries to locate and identify animal viruses, discovering many new ones in the process. But not every virus is equally threatening. So Mazet and her colleagues decided to create a framework to interpret their findings. “We wanted to move beyond scientific stamp collecting [simply finding viruses] to actual risk evaluation and reduction,” she says. The team was surprised to find very little existing research on categorizing threats from viruses that are currently found only in animals but are in viral families that can likely cause disease in people.

So the researchers started from scratch, identifying 31 factors pertaining to animal viruses (such as how they are transmitted), to their hosts (such as how many and varied they are), and to the environment (human population density, frequency of interaction with hosts, and more). These are summed up in a risk score out of 155; the higher the score, the more likelihood of spillover. Cornell University virologist Colin Parrish, who was not involved in the study, says the factors examined were important in previous spillovers. But he notes that other viruses' crossover risk may be heightened by unforeseeable factors that crop up later. “It's a bit like the stock market,” he says. The new study, published in the Proceedings of the National Academy of Sciences USA, ranks 887 animal-borne viruses. Twelve known human pathogens scored at the top—with the virus that causes COVID-19 in second place, just under the rat-carried Lassa virus. (Influenza would have topped the list if included, Mazet says, but flu variants are already tracked elsewhere.) Parrish notes that the list also omits insect-borne viruses and those from domesticated animals. “This is a work in progress,” he says. “I'm sure it will be iterated into a more powerful tool as more information and data become available. SpillOver is publicly editable, and scientists around the world are already contributing their own findings. Mazet hopes it catches the attention of public health practitioners and leaders, too. With targeted action, Mazet says, “we can ensure that we don't have these spillovers at all. Or if we do, we're ready for them—because we're watching.”

See also research published in P.N.A.S. (April 13, 2021):

https://doi.org/10.1073/pnas.2002324118

A new study, published in the August 8, 2019, issue of Cell by a team of researchers led by Instructor Kathryn Hastie, Ph.D., and Professor Erica Ollmann Saphire, Ph.D., at La Jolla Institute for Immunology (LJI),  identified and then reverse engineered the molecular properties shared by antibodies that are particularly efficient at inactivating or “neutralizing” the virus. The team’s findings also revealed that most neutralizing antibodies bind to the same spot on the surface of Lassa virus, providing a map for rational vaccine design.As this year's Lassa fever outbreak in Nigeria is finally ebbing, the total tally came to more than 600 infected people, one-quarter of them dead. Thousands more die each year, uncounted in rural villages throughout West Africa. With an annual wave of infections and new viral strains emerging, it has never been more important to understand the characteristics of a broadly protective immune response in order to develop effective treatments, or better yet, a vaccine.

“The beauty of structural biology is that it gives you the ability to dissect the molecular details at high resolution to explain precisely how something works,” says structural immunologist Ollmann Saphire. “Once you do, you have a blueprint to engineer potent immunotherapeutics or a vaccine that elicits the desired immune response.”

Identified 50 years ago and named for the town in Nigeria where the first known cases cropped up, Lassa virus is endemic in West Africa where it infects hundreds of thousands of people every year. For the majority of infected people, symptoms are mild and the infection mostly goes undiagnosed. But in 20 percent of patients, the disease causes a more serious illness including neurological symptoms and hemorrhage, which can result in multi-organ failure and death.

For the current study, Hastie compared the structure of three different neutralizing antibodies of varying potency—high, moderate and low—bound to the glycoprotein. The side-by-side comparison highlighted specific amino acid residues that drive high potency and enabled the researchers to precision-engineer mediocre antibodies to turn them into highly effective ones.

“Not only were we able to increase the antibody’s potency, which means you can deliver much less antibody, we were also able to make it pan-Lassa. It can hit every Lassa virus lineage characterized so far,” says Hastie.  But few naturally infected people generate neutralizing antibodies and current vaccine efforts focus on eliciting T cell immunity. “Historically, researchers have found that development of antibodies is not a good correlate of protection in natural Lassa infections,” says Hastie. “It is actually very difficult to induce neutralizing antibodies.” 

The study was published in Cell on August 8, 2019:

https://doi.org/10.1016/j.cell.2019.07.020