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It’s flu season. Again. Which means it’s time for the flu vaccine. Again. The reason we need a shot year after year is that different strains of influenza circulate annually, so a vaccine that protects against last year’s strains likely won’t confer immunity to this year’s. This is particularly true because the seasonal flu vaccine spurs the production of antibodies that recognize an especially mutable part of the virus.

For decades, scientists have been trying to develop a universal vaccine that would protect people against seasonal flu for years, and also against pandemics, which emerge when viral strains completely novel to people’s immune systems start spreading. “A universal flu vaccine is often referred to as ‘The Holy Grail’ of influenza research, and like the Holy Grail, it is challenging to achieve,” Tamar Ben-Yedidia, chief scientific officer of BiondVax, whose universal flu vaccine is now in Phase 3 clinical trials, tells The Scientist in an email.

It has been about 20 years in the making to get to the point of a Phase 3 study—the first universal flu vaccine to have progressed to that stage—and there are numerous others following behind. All of these vaccines employ variations on a similar strategy, which is to generate immunity to parts of the virus that are the least variable from strain to strain.

In February 2018—in the midst of the worst flu season in a decade—the National Institute of Allergy and Infectious Diseases (NIAID) announced that it would prioritize the development of a universal flu vaccine. It also published a strategic plan for achieving that goal, which includes learning more about the influenza virus and its pathogenesis, characterizing the human immune response to the flu, and supporting the development of universal influenza vaccines. To that end, NIAID is funding many universal vaccine trials.

“I think what we’re really interested in is seeing a wide array of products that have promise moving into the clinic and to see the different kinds of strategies being used,” Jennifer Gordon, NIAID’s Influenza Vaccines Program officer, tells The Scientist. “I think right now there are a number of good candidates.”

The Phase 3 study

Influenza has two surface glycoproteins: hemagglutinin, which helps the virus enter host cells, and neuramidase, which helps the virus spread among cells. Hemagglutinin is commonly likened to a mushroom because it has a round head and a stalk. The highly variable head is what seasonal flu vaccines tend to generate antibodies against. The stalk, in contrast, is more conserved, and so is the target of many efforts to generate a universal flu vaccine.

BiondVax’s M-001 vaccine—the furthest along in the pipeline toward a universal flu vaccine—contains no viruses, dead or alive; rather, it is a peptide vaccine. It contains nine, highly conserved viral epitopes, those parts of an antigen recognized by the immune system, that are common to 40,000 influenza viruses listed in a National Institutes of Health database, Ben-Yedidia tells The Scientist. These epitopes come from three different viral proteins: M1 (a matrix protein that lines the viral envelope), NP (a nucleoprotein that wraps around the viral RNA), and conserved parts of the hemagglutinin head.

BiondVax’s trials so far have found that M-001 and a standard flu vaccine have a synergistic effect. While M-001 by itself does not stimulate antibodies against hemagglutinin, M-001 followed by an inactivated flu vaccine does. Further, the hemagglutinin antibodies produced after vaccination with both M-001 and a standard flu shot have broader immunity than those stimulated by the seasonal vaccine alone.

I think right now there are a number of good candidates. —Jennifer Gordon, NIAID

M-001 has already had six clinical trials in Israel and Europe: two Phase 1/2 trials and four Phase 2 trials with a total of 698 participants. Those experiments looked at the vaccine’s safety and immunogenicity, finding that the vaccine was well-tolerated and generated a broad immune response (which researchers measure by drawing participants’ blood, evaluating antibodies, and measuring the levels of molecules, such as interferons and other cytokines, that indicate a T-cell response). In May, a NIAID-sponsored Phase 2 trial of M-001 in adults began in the US.

It’s only with a Phase 3 trial that researchers can examine how well a vaccine protects against influenza. In August, BiondVax launched such a study of M-001 in Eastern Europe with the goal of recruiting about 10,000 participants in two cohorts. The first group will be monitored for two flu seasons—this year’s and next year’s—while the second group will be tracked for one flu season, and the results are expected in late 2020. Volunteers will receive two doses of M-001 or a placebo about three weeks apart. Afterward, the investigators will monitor subjects for any influenza-like illness and compare the number of flu cases in each group.

Normally, a vaccine would only need to show that it reduced the number of cases of the specific influenza strains it targeted. “In contrast, since BiondVax’s M-001 is designed as a universal flu vaccine, we intend to show M-001 reduces illness caused by any influenza strain,” Ben-Yedidia says.

M-001 is by no means the only universal vaccine currently in development, and that’s important, says Gordon. “You can put a lot of candidates in the beginning of the pipeline, but they don’t all succeed in the pipeline for reasons that are various. . . . You want to make sure that you have a lot of candidates going into the pipeline so that we have a higher probability of success coming out.”

Chimeras spark immunity to conserved antigens

For the last several years, Florian Krammer and his colleagues at the Icahn School of Medicine at Mount Sinai have been developing vaccines against the stalk part of hemagglutinin. But they found that when they immunized animals with vaccines containing the entire hemagglutinin protein, the resulting antibodies tended to target only the head. So instead they’ve designed so-called chimeric hemagglutinin proteins to include in vaccines, where the head came comes from one variety of hemagglutinin (often an exotic kind, such as one from a bird flu) and the stalk from a human influenza A subtype (such as H1 or H3).

These constructs are doing something special. —Florian Krammer, Icahn School of Medicine at Mount Sinai

When they vaccinated animals twice over the course of a few weeks using vaccines that included chimeric hemagglutinin proteins with identical stalks but different heads, the immune system produced many more antibodies against the stalk common to both flu strains than to the heads.

“These constructs are doing something special,” says Krammer. “They redirect the immune response to the stalk domain, which is more conserved, so at least in animal models, they work much better than the regular vaccines that we used.”

See “How to Build a Better Flu Shot”

Vaccines based on this strategy are the basis of two Phase 1 clinical trials. In the first, adults ages 18 through 39 are given a live-attenuated influenza vaccine (LAIV) containing one chimeric hemagglutinin protein followed three months later by an inactivated vaccine with a different hemagglutinin chimera. The second trial also involves sequential vaccinations and chimeric hemagglutinin, but all of the vaccines are inactivated. This second trial also tests the benefits of adjuvants, compounds added to the mix that are intended to boost vaccine effectiveness.

LAIVs and inactivated vaccines both have their pros and cons, according to Krammer. Live-attenuated vaccines delivered through the nose replicate a small amount in the upper respiratory tract, stimulating a mucosal immune response involving not only antibodies but also T cells, which is a plus, Krammer says. The inactivated vaccine, on the other hand, typically does not induce mucosal immunity but provides a stronger antibody response in the blood. “It’s hard to say right now which one would work better,” Krammer says. “That’s why we run both clinical trials.”

Another kind of LAIV with the potential to elicit universal immunity, called RedeeFlu, is being tested in a NIAID-sponsored, early-stage clinical trial in kids ages 9 to 17. RedeeFlu’s mechanism for achieving broad effectiveness is that, like other LAIVs, it stimulates both antibody and T-cell responses, but RedeeFlu does those things better than other LAIVs, according to FluGen cofounder Yoshihiro Kawaoka, who holds joint appointments at the University of Wisconsin and the University of Tokyo.

For instance, a study Kawaoka and his colleagues published earlier this year in ferrets compared the commercial LAIV FluMist with RedeeFlu. Ferrets previously exposed to one or more strains of flu (to mimic people’s previous exposure to influenza), then immunized with either RedeeFlu or FluMist, a commercial LAIV, were better protected from a new virus that was related but not identical to the one in the vaccine if they received RedeeFlu than if they got FluMist.

The alchemy of adjuvants

If a universal flu vaccine is the Holy Grail, then perhaps it’s not surprising that one approach to creating it almost sounds like alchemy—turning one thing into another.

It turns out that adjuvants aren’t just something you add to a universal vaccine to make it work better. They can also transform a seasonal vaccine into a universal one, at least in theory.

Jay Evans, director of the Center for Translational Medicine at the University of Montana and president of the immunotherapeutics company Inimmune, and his colleagues are working with an adjuvant called TRAC-478, which stimulates a suite of toll-like receptors (TLRs) on antigen-presenting cells. TRAC-478 combines properties of two individual adjuvants that target TLR4 and TLR7/8 individually, about which Evans and colleagues have published studies (for example here and here). TLR4 recognizes bacterial infections, while TLR7 and TLR8 recognize viral infections, Evans says. “By combining TLR4 with the TLR7/8, we’re stimulating an immune response in two very different ways: one that the body would recognize as potentially a bacterial infection; and the other body would recognize as a viral infection. So those two are very synergistic and drive a much stronger immune response when combined with a traditional vaccine.” No studies of TRAC-478 have been published thus far, but Evans says that the data are encouraging and that he expects to publish results in 2019.

The adjuvant TRAC-478 has multiple effects, Evans says. According to research in mice and in pigs, the adjuvant in combination with the seasonal vaccine leads to the production of higher levels of antibodies and a stronger T-cell response than the unadjuvanted vaccine, Evans says. The seasonal vaccine adjuvanted with TRAC-478 also confers immunity to viruses not included in the vaccine—hence the universality, Evans says. “Not only do we make that immune response stronger, but we can actually drive responses against epitopes that wouldn’t normally have been recognized without the adjuvant. And by broadening that response against other epitopes, you can now develop vaccines that are cross-seasonally protective, or universal.”