Scientists at the Jackson Laboratory (JAX) have developed an antibody-based therapy against influenza A virus (IAV), which they suggest could change the way the world fights flu, one of the deadliest infectious diseases. The team, headed by JAX immunologist Silke Paust, PhD, demonstrated that a cocktail of non-neutralizing antibodies targeting distinct, overlapping epitopes within influenza A virus’s conserved Matrix Protein 2 ectodomain (M2e) protected mice—including those with weakened immune systems—from nearly every strain of influenza tested, including avian and swine variants that pose pandemic threats.
Unlike current FDA-approved flu treatments, which target viral enzymes and can quickly become useless as the virus mutates, the new antibody therapy did not allow viral escape, even after a month of repeated exposure in animals. The team says this difference could prove crucial in future outbreaks, when survival may depend on how quickly and effectively doctors can deploy treatments, and vaccine development will take about six months.
The new developments challenge a long-held belief that for antibodies to be useful as a therapy against viruses, they must be neutralizing antibodies that bind directly to viruses and block them from infecting cells. Rather than prevent infection, the non-neutralizing antibodies developed by Paust and colleagues tag infected lung cells and recruit the body’s immune system to clear the infection. This new approach could reshape how scientists design treatments for other viruses.
Broad protection with extended therapeutic window
“This is the first time we’ve seen such broad protection coupled with an extended therapeutic window against flu in a living system,” said Paust, who is senior author of the researchers’ published paper in Science Advances. “Even when we gave the therapy days after infection, most of the treated mice survived … The majority of antibodies our bodies make are non-neutralizing, but medicine has largely ignored them. We show they can be lifesaving. Even with lethal strains like H5 and H7 avian flu, this therapy saved lives long after infection had taken hold.”
Paust told GEN, “Our work may help broaden acceptance of non-neutralizing antibodies as viable therapeutics. It also highlights how much we still need to learn about their mechanisms of action in vivo, mechanisms that may well be tissue-specific, depending on the immune cell compositions of the tissue. I hope these findings will encourage renewed investment in basic science to address these fundamental questions, as well as greater interest from the pharmaceutical industry in developing non-neutralizing antibodies as a new class of therapeutics.”
The team’s published paper in Science Advances is titled “Non-neutralizing antibodies to influenza A matrix-protein-2-ectodomain are broadly effective therapeutics and resistant to viral escape mutations.”
The authors cite figures indicating that every year, influenza virus causes about one billion infections, including 3 to 5 million cases of severe illness and the death of about 500,000 people worldwide. In humans, infections with influenza A viruses outnumber influenza B virus infections and are more likely to lead to more severe disease. “Influenza A viruses remain a global health threat, yet no universal antibody therapy exists,” the scientists reported. IAV also has “substantial pandemic potential,” they pointed out, due to antigenic shift and drift in two regions of the virus’s immunodominant epitopes, hemagglutinin (HA) and neuraminidase.
The need for broadly effective, off-the-shelf IAV treatments
“These antigenic changes thwart the development of monoclonal antibody (mAb)–based therapeutics …” resulting in the need for seasonal vaccination, the team added. “Broadly effective, escape mutant-resistant off-the-shelf therapeutics are likely our best option for reducing lethality during future IAV pandemics and may provide us with the necessary window to develop and disseminate a pandemic subtype–specific vaccine … Off-the-shelf IAV treatments must be universally effective and, thus, should target highly conserved viral epitopes that are resistant to mutations leading to viral escape.”
Rather than focus on neutralizing mAbs, Paust and colleagues developed three non-neutralizing IgG2a monoclonal antibodies (mAbs) targeting the small, highly conserved M2e region of the influenza A virus Matrix Protein 2. This part of the virus is essential for its life cycle and remains nearly unchanged across infected cells in all flu strains, including human, avian, and swine variants.
Tests in mice infected with a range of human and zoonotic IAV strains, including highly pathogenic variants, showed that the team’s newly developed M2e-mAb antibody combination did not lead to viral resistance even after repeated exposure, and sequencing confirmed no mutations in the virus’s M2 region after 24 days of treatment. “Serial passaging in triple M2e-mAb–treated immunocompetent and immunodeficient hosts failed to generate viral escape mutants,” the investigators reported.
While the team tested the efficacy of the three antibodies individually, success came from combining them, as this approach reduces the virus’s chances of escaping three different antibodies. “The virus didn’t mutate away even when using individual antibodies,” Paust said. “But in a flu season with millions of people taking this therapy, I would be much more confident that we can prevent escape from the therapy if we use the cocktail.”
Antibody cocktail effective at low doses
Paust and her team found that the antibodies were effective at low doses, both before and after influenza infection. “Combined at low dose, this ‘triple M2e-mAb’ confers robust prophylactic and therapeutic protection in mice challenged with diverse human and zoonotic IAV strains, including highly pathogenic variants,” the team stated. The cocktail significantly reduced disease severity and viral load in lungs, and improved survival rates in both healthy and immunocompromised mice. Mechanistic studies further established that “… protection from IAV lethality requires M2e-mAb–triggered FcγRI and FcγRIII and/or FcγRIV-mediated effector functions.”
Paust commented to GEN, “Science advances not only through successes but also through the many failures of those who came before us. It became clear to us that neutralizing antibodies, and perhaps single antibodies more generally, were not sufficient to prevent influenza infection in animal models or in humans, nor to block the emergence of viral escape mutants resistant to therapy. More recent studies have suggested that neutralization may not be the primary mechanism by which influenza antibodies function in vivo. Even antibodies that neutralize in vitro, when administered systemically, must act in infected lung tissue rather than at the point of viral entry. In those settings, Fc effector functions appear to be the critical driver of therapeutic activity.”
When testing the new treatment against H7N9, a type of bird flu that can be deadly to both animals and people, the team found that just one dose of the combined antibody therapy reduced the amount of virus in the lungs, even when it was given four days after infection. The reduced viral loads correlated with better survival rates. All mice survived when treated with the antibody cocktail on the first three days after infection, while 70% and 60% survived on days four and five, respectively.
Antibody cocktail superior to FDA-approved treatments
“Disease severity, as determined by overall weight loss, was also significantly ameliorated when the M2e-mAb triple cocktail therapy was administered as late as day 4 after the H7N9 IAV challenge,” the authors wrote. “Until our study, therapeutic efficacy had not been evaluated for published M2e-specific mAbs administered later than 2 days postinfection … These data establish our triple M2e-mAb therapy as robustly effective against one of the most lethal IAV serotypes and superior to FDA-approved treatments.”
Paust added, “We can use very low doses, which is also promising because potential therapies could be cheaper and less likely to produce adverse side effects in people.”
While the reported results are preliminary, they are promising for a future where patients could have access to stockpiled therapeutics to be deployed rapidly to fight seasonal outbreaks or pandemics. Currently, flu vaccines are updated seasonally because the virus continuously mutates, making immunity to prior strains irrelevant. “We need something that is off the shelf when we don’t necessarily have the time to make a new vaccine if we do have an outbreak or pandemic where lethality is high, so this type of therapy could be readily available for anyone in any situation,” Paust said.
Potential for prophylactic and therapeutic use
Paust further commented to GEN, “We believe our non-neutralizing monoclonal antibody cocktail has both prophylactic and therapeutic potential. In the therapeutic setting, infusion of antibodies, similar to what was done during the COVID-19 pandemic, could reduce disease severity and mortality, which would be the desired outcome. As a prophylactic, while non-neutralizing antibodies are unlikely to block infection entirely, they may prevent progression to severe disease, thereby reducing hospitalization and death. Moving forward, we will explore the M2e antibody cocktail in both contexts.”
The team is working on designing antibodies for clinical trials. The idea is to make a “humanized” antibody with the same specificity to target the M2 protein, but without triggering an immune response against the therapy itself or diminishing its efficacy in humans. The team envisions a future where the cocktail could work as a standalone prophylactic for the elderly, immunocompromised, and other high-risk groups, in addition to serving as a therapy for those severely ill with flu. They concluded, “Our data will critically shape future M2e-mAb–based IAV-therapeutic development strategies and provide precedence for embracing non-neutralizing M2e-mAbs for clinical development.”
Humanizing antibody candidates
Paust acknowledged to GEN that there is still significant work ahead before clinical studies might start. “The critical next step is the humanization of the antibodies, preserving their therapeutic efficacy while making them effectively ‘invisible’ to the human immune system. This process will take several years and must be completed before moving into clinical testing. Beyond that, we need to confirm that the robust effector functions observed in vivo in mice are conserved in humans, which is no small task. It will be several years, if all milestones are met throughout the humanization, before clinical trials can be applied for.”
Paust continued, “The Influenza Antibody Therapeutics Project is one of the oldest efforts in my lab. It began from my concern that we remain poorly prepared to face a serious influenza pandemic like the one in 1918, which killed 50 million people when the world’s population was only 1.8 billion. I am drawn to problem solving and to understanding how things work, but I also want my research to have direct relevance to human health.”
The project is one centered on collaboration and persistence, GEN was told. “… Over more than a decade, my lab has published two papers on this topic, with the project carried forward by multiple trainees who were willing to try new approaches, put in the extra time, and pursue good science. Collaborators such as Dr. Mark Tompkins at UGA, the co-corresponding author on our latest paper, shared that spirit, writing grants and conducting research with me before we had even met in person. Together, we refused to be discouraged, even when people were sceptical of the idea that non-neutralizing antibodies could be developed into a therapeutic.
“This combination of community, collaboration, and personal commitment has brought us to this point. The next steps are the humanization of the antibodies, followed in time by an IND application and, we hope, successful clinical trials. My ultimate goal is to see this work become an effective influenza drug available to everyone.”
