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Delicious Vaccines

                Delicious Vaccines
                Axis Genetics, plc 

                by William A. Wells

                (BioMedNet, Posted April 30, 1999  Issue 53) 


                If the developing world could grow vaccines, they
                could afford them. But will edible vaccines work?

                The vaccine industry is turning babies into pincushions.
                Although some vaccines are combined, there is still
                more than one injection per baby, and plenty of
                screaming in pediatric offices. How soothing, then, to
                think of the injection being replaced by toothless
                munching on some mashed banana.

                For developing countries, an edible
                vaccine could stop far more than a baby's
                screaming. If the transgenic plant
                containing the vaccinating material can
                be grown on site, vaccination logistics
                could be simplified, and costs reduced.

                Several large companies such as Dow and DuPont are
                moving into this area, but it is a small company named
                Axis Genetics, plc (Cambridge, United Kingdom) that
                is leading the way. The next few years will be crucial
                for Axis, as it moves beyond the promising early
                science, and into the real world of vaccine production. 

                First the Virus 

                The quickest way to make lots of
                a foreign protein in a plant is to
                infect the plant with a virus. Axis
                started with the cowpea (known in
                the United States as the black-eyed pea) and its
                fast-growing virus CPMV (cowpea mosaic virus). The
                coat of CPMV is derived from the splice and cleavage
                products of a single gene, and up to 38 amino acids can
                be inserted into a loop of the coat protein with no
                adverse effects on coat assembly. Axis is using this
                system, dubbed Epicoat, for vaccines against
                Pseudomonas aeruginosa, Staphylococcus aureus,
                and cancer (the latter directed against a mucin
                expressed on breast cancer cells). 

                Epicoat is an alternative to peptide vaccines, which
                were first proposed in the 1980s and have struggled to
                establish themselves ever since. Axis CEO Iain Cubitt
                sees the lack of conformational stability in these
                peptides as a crucial flaw, and says that the Axis
                technology solves this by putting the peptide in a
                structured loop. 

                Epicoat is a purified injectable,
                but purified inhalant and
                non-purified edible forms may
                follow. Inhaled or ingested
                vaccines encounter the sites that are used by most
                infecting pathogens, and these vaccines should induce
                more effective immunity - mucosal immunity and
                secretory IgA antibodies, in addition to serum
                antibodies. This approach is not practical for peptide
                vaccines made by chemical synthesis because of the
                large amounts of material necessary, but the quantity is
                reasonable for a plant-based vaccine. 

                But with Epicoat the worst defect of peptide vaccines
                remains. "Epicoat can make a lot of material but you
                can only display a single epitope," says Hugh Mason of
                the Boyce Thompson Institute for Plant Research, Inc.
                (BTI; Ithaca, New York). "There is not a lot of data so
                far that single epitope vaccines will be very useful."
                With single epitope vaccines, it takes but a single
                mutation for the infectious agent to avoid the
                vaccine-induced immunity. This has led some
                investigators to string together several peptides in the
                same insertion site, an approach that is the subject of an
                Axis patent application. "It is likely that several
                peptides represents a more realistic approach than a
                single peptide," says Harry Greenberg of Stanford
                University (Palo Alto, California), who helped develop
                the newly released rotavirus vaccine. 

                I Say Potato, Dan says Potatoe 

                If the limiting factor is the amount of
                protein sequence that can be inserted
                in the virus, why not insert the whole
                protein into the plant? This thought,
                or something similar to it, occurred
                to Roy Curtiss and Guy Cardineau (Washington
                University, St. Louis, Missouri) in 1990, and to Charles
                Arntzen and Mason of BTI a year later. Arntzen and
                Mason were heavily involved in subsequent
                developments in edible vaccine technology. In 1998,
                Axis concluded deals with both Mycogen (San Diego,
                California), who own the Curtiss patents, and BTI. (Yet
                another approach involves using plant-derived

                In the intervening years, the plant vaccine field had
                demonstrated the successful production of foreign
                proteins in an immunogenic state (1992), the production
                of antibodies in mice fed with plant material (1995),
                protection against disease in animals fed plant material
                (1997), and finally the production of antibodies in
                humans fed plant material (1998). The latter trials
                involved the ingestion of "bite-sized chunks" of
                delicious raw potato from a plant producing LT-B - the
                binding subunit of a toxin from enterotoxigenic
                Escherichia coli. A pentamer of LT-B binds gut
                epithelial cells and allows the active LT-A toxin
                subunit to enter the cell. The trial participants ate 50 or
                100 g of raw potato on each of three occasions, for a
                mean of 0.75 mg of LT-B per dose. Ten of eleven
                showed a fourfold rise in serum antibodies, and six of
                eleven showed a fourfold rise in secretory (IgA)

                Potential Pitfalls 

                The LT-B trials directed by
                Arntzen are encouraging, but no
                one is relaxing yet. "People said
                edible vaccines would never
                work," says Cubitt. "Then when LT-B elicited a good
                response they said of course it works because [the
                bacterium and the protein are] normally in the gut. For
                non-enteric diseases it is not so clear. There it depends
                on how we present the protein." 

                When given orally, a particulate virus or even
                bacterium is efficiently sampled by the immune system.
                Although LT-B may be a special case, single proteins
                are usually not efficiently taken up. One solution that
                the BTI is working on is to produce complete, empty
                virus-like particles in plants simply by expressing the
                coat protein. "We've shown that we can produce these
                structures with Norwalk virus in plants," says Mason.
                "The efficiency is questionable and variable, but I think
                it's largely a problem with the level of expression." 

                Virus-like particles or not, expression levels remain a
                challenge. The solution may be inducible expression at
                a particular time, to limit toxicity to the plant. 

                The flip side - containing expression -
                is also a concern. We do not make
                antibodies to all of our food because
                of tolerance - the damping down of the
                immune system in the face of
                overwhelming amounts of antigen. The escape of a
                vaccine plant into the general food chain could be a
                disaster if it induced tolerance to a major surface
                protein of a virus. "One can't say at this point that it's
                not a possibility," says William Langridge (Loma Linda
                University, California). "We hope [Axis] will fund
                further research into this question." 

                Langridge stresses, however, that tolerance looks to be
                an unlikely prospect. His experience stems from the
                successful induction of tolerance to a diabetes
                autoantigen in mice (thus preventing further destruction
                of the pancreas). This was achieved with plant
                material, but only when the autoantigen was fused to
                LT-B, so that the complex entered gut epithelial cells
                efficiently. In earlier work an unfused autoantigen
                induced tolerance only after two months of continuous

                The safety issue is particularly sensitive
                following the public uproar in Britain
                over genetically modified (GM) foods.
                This curious incident was sparked by
                one researcher's description of his
                unpublished and questionable results on
                a TV show. "It's bizarre what has happened in the
                U.K.," says Cubitt. "Science is being ignored in the
                formation of the public perception here." In the United
                States, there is an equally distressing state of affairs -
                complete apathy - and it is in that country that Axis
                grows its crops (in greenhouses). But even in Britain,
                Cubitt does not anticipate any problems. "We are not
                involved because we are in pharmaceutical products
                not food products - we are already highly regulated and
                not planning to grow large areas of crops," he says.
                "There may be a moratorium on growing in the field or
                growing agricultural products, but we grow in the U.S.
                and we don't do agriculture. It's nothing to do with us." 

                Business Basics 

                Vaccines used to be a treacherous business, filled with
                interminable trials with thousands of participants, and
                endless lawsuits from the few of the vaccinated
                millions with an adverse reaction. No more. 

                "The vaccine field has changed out of all recognition in
                the last ten years," says Cubitt. "The reason it has
                changed is that, if you don't use the whole organism you
                can't cause the disease, so you get away from the
                liability issues." 

                "It's turned from being the Cinderella
                of the business to being very
                attractive," he says. "It's now one of
                the most profitable parts of the
                pharmaceutical industry." 

                Protection from liability has also come from
                legislation, and the size of some trials has been scaled
                back if the protective levels of antibody are known
                from an earlier vaccine (as is the case with Axis's
                hepatitis B vaccine). Axis is further protecting itself by
                drawing extensively on the expertise of academic
                collaborators. It uses this expertise, for example, to
                identify the proteins or epitopes for use in vaccines. 

                The Promise to the Developing World 

                The dream of a vaccine-laden
                banana tree in every backyard is
                not going to happen - for starters,
                there is the containment issue.
                Cubitt says the final vaccine
                "won't be fresh material; it will be a powdered
                formulation that can be stored at room temperature."
                How the temperature stability will be achieved has not
                been disclosed but, he says, "we believe that our
                approaches will lead to temperature stability." 

                Could this material be produced in the countries that
                need it most? "The type of technology we are using is
                more related to food processing technology than
                pharmaceutical technology, because we are not taking
                out the food material or doing a purification," says
                Cubitt. "Theoretically this could be produced around
                the world, but we need to know that it is produced
                under pharmaceutical controls, not agricultural
                controls. These are pharmaceutical products, not
                agricultural products." 

                In the final stages of production, edible vaccines may
                resemble the pharmaceuticals that they most certainly
                are. But if the vaccines work, the wonder of their
                source will remain: protection from disease using only
                sunlight, dirt, and water as the primary ingredients. 

                    William A. Wells is a freelance science
                    writer based in San Francisco. 

                    Caleb Brown is an illustrator and biologist
                    living in Montana. By day he drives a
                    delivery van, and by night he draws pictures
                    with his computer.


                             Tell us what you think.


                Edible Vaccines - an outline of principles, plus a
                bibliography of research papers.

                DNA Vaccine Web - a rich resource of information on
                vaccines including new clinical updates, research
                articles, Web sites, and protocols.

                World Health Organization: Vaccines - information on
                all major vaccine-preventable diseases, dealing with
                the disease itself, the vaccine, and research and policy
                for each.

                Understanding Vaccines - a downloadable brochure on
                vaccines and the immune system. In PDF format;
                requires Adobe Acrobat Reader. From the National
                Institute of Allergy and Infectious Diseases.

                Plant Biotechnology - an overview of plant
                biotechnology at Boyce Thompson Institute for Plant
                Research as it relates to new pharmaceutical products.

                Living in a Genetically Modified World - a New
                Scientist special feature on genetically modified food.

                                     1999 BioMedNet Ltd. All rights reserved.