Author Topic: Delivery Of Vaccines Via Transgenic Plants - Edible Vaccines  (Read 13273 times)

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ASU biologist pioneers edible vaccines
Mandy Redig
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Summary:
A happenstance moment in Thailand forever changed the career and passion of Dr. Charles Arntzen. Watching a mother calm down her fussy baby with a banana, a sudden idea triggered a decade-long pursuit that sounds too good to be true: delivering vaccines via fruit instead of syringes. The dream is quickly becoming reality.

Full Story:As Archimedes could attest, inspiration can strike anywhere. Legend has it that the ancient Greek thinker discovered the mathematical laws governing buoyancy in a bathhouse while idly watching soap float. The nature of scientific research has changed since the 3rd century B.C., but the spirit of observational inquiry that led to Archimedes’ principle is still active.

When Dr. Charles Arntzen of Arizona State University visited Thailand in 1992, he was not expecting a moment of scientific eureka that would redirect his career. However, after observing a young Thai mother soothing her fussy infant with bits of banana, this plant molecular biologist was struck with an idea that is both startling and ingenious. What if, in addition to quieting her child, the mother could also administer a life-saving vaccine – in the banana?

Arntzen's vision is well on its way to becoming reality thanks to a combination of dedication and applied biotechnology. As the president emeritus of the Boyce Thompson Institute for Plant Research, founding director of the Arizona Biomedical Institute, and member of the President’s Council of Advisors on Science and Technology, Arntzen is well equipped to handle the challenge he has undertaken. His breakthrough lies in forging a link between green plants, foreign DNA, and vaccines.

Vaccination stands as one of modern medicine's greatest success stories. Early experiments by Edward Jenner and Louis Pasteur taught physicians that they could prevent disease merely by exposing a patient to a weakened or inactivated pathogen. While his protocols violate today’s clinical trials regulations, Dr. Jenner was able to prevent children from getting smallpox even when he deliberately exposed them to it after first inoculating them with the pus from cowpox.

Today, most new vaccines contain a specific protein or proteins from a pathogen of interest and not the pathogen itself. A protective immune response can result from this more limited (and inherently less risky) exposure. Though materially different from those developed by Jenner and Pasteur, modern vaccines, including Arntzen's, still build upon the same fundamental principle: if the immune system is trained to recognize a pathogen prior to infection, the actual disease can be prevented when the pathogen is encountered.%pagebreak%

Disease-prevention via an edible vaccine is great news for people around the globe. The problem with current vaccination protocols - and the passion behind Arntzen's research is that what works in the developed world is often much more difficult to deliver in the developing world, or simply too costly for them to buy. A vaccine that requires a sterile syringe, refrigeration prior to injection, and repeat booster shots is difficult to implement in many countries. Unfortunately, this often means that the people who most need a vaccine cannot get it. In a discussion of his work, Arntzen points out that “each year diarrhea kills about two and one-half million children under the age of five. Arntzen persuasively uses such horrendous statistics to champion his cause. In his own words, It's hard to be pro-infant mortality.

While Arntzen's edible vaccine is likely to win approval from children everywhere, there are actually significant medical advantages to this route of administration. An oral vaccine incorporated into a plant bypasses the need for sterile syringes, costly refrigeration, or multiple injections. Furthermore, since many of the developing world's most deadly diseases cholera, rotavirus, or E. coli to name a few enter the body through the gastrointestinal tract, a vaccine that is ingested may actually provide the best protection because it mimics the natural route of infection.

The trick with an edible vaccine is convincing a plant to express the genes of a foreign organism. Fortunately, Arntzen's prior work prepared him to face this challenge. As a biochemist, his career focused on unraveling the means by which photosynthetic membranes in plants capture solar energy. What we've done for the last ten years is try to change the cellular machinery of a plant by adding a new gene, cause that gene to make a new protein, and coax the new protein into folding to the desired shape so that it accumulates. I took knowledge about plants proteins under normal circumstances and used that for something new. That something new was vaccine development.

When Arntzen started investigating the vaccine issue in the early 1990's, scientists were already using genetically engineered yeast to produce proteins for injection vaccines. Arntzen's experience with green plants led him to consider other options. He remembers thinking at the time, Would it be possible to use a higher plant instead of a lower plant, something we already know is an agricultural crop? Can we take a potato or tomato and turn it into a green factory?

Indeed, modern technology enables Arntzen to insert specific genes from a foreign organism into the genome of a green plant. Progeny plants will then produce the foreign protein. If the foreign protein happens to be an immunity-inducing pathogen protein, an edible vaccine is in the making.

Yet the challenges of science are not the only obstacles Arntzen faces. This type of project requires a multidisciplinary approach, incorporating the skills of many types of basic and clinical scientists as well as experts in product regulation and distribution. Says Arntzen, “No biochemist can make progress in moving something forward on his own. I need linkages with people who do immunology. I need people in vaccine development. I also have increasingly found that I have to understand the regulatory environment.

In fact, satisfying regulatory policies has been one of the most painstaking elements of Arntzen's work. He is determined to demonstrate that his vaccine passes the rigorous requirements of the Food and Drug Administration, thereby silencing any critics who would accuse him of dumping experimental technology on the world's poor.

His most recent clinical trials are particularly exciting. Human volunteers who enrolled in a study at the University of Maryland in Baltimore started producing antibodies against Norwalk virus (which causes acute bouts of diarrhea) after eating Arntzen's creations – genetically engineered potato. Negotiations are currently in progress to start clinical trials abroad with the International Vaccine Institute in Korea, a new center funded in large part by the Bill and Melinda Gates Foundation. Clinical trials of cholera vaccines are also planned to take place there as well as in Vietnam and Cambodia, regions where cholera is still a serious medical concern.

In addition, during a recent scientific conference held at the Flinn Foundation's Phoenix office, tentative connections were made with company representatives from Egypt and India. This meeting, organized by Arizona Biodesign Institute on behalf of the ProVACS Ceneter (Production of Vaccines from Applied Crop Sciences), highlighted technical advances in plant-based vaccines. Arntzen, the keynote speaker at the meeting, says, "We intend to visit India. They're willing to fund clinical trials there - we could send vaccine materials to them and they're interested in developing the product."

For now, all such clinical trials will involve modified potatoes or tomatoes. Both products can be easily freeze-dried, transported, and reconstituted. Since many target countries have a long history of herbal medicine, Arntzen is interested in working within already-existing ideas. Our goal is not to make the decision for how we want it (the freeze-dried dose) introduced, he says. "We want to work with them."

In the meantime, Arntzen is still working on the banana vaccine. Tomatoes and potatoes have shorter growing seasons and are easier to manipulate in an experimental setting, but Arntzen has not given up on the popular yellow fruit that sparked his initial idea. Currently, there are several transgenic banana trees growing in his greenhouses on the ASU-East campus.

Once the foreign proteins are expressed in the target plant, the same concerns for a traditional vaccine  efficacy, quality control, and dosage regulation  also become an issue. Arntzen's original, utopian vision was of a communal banana tree where villagers could dose themselves. This has been replaced by a more practical vision, perhaps a controlled dose of freeze-dried tomato or, later, a banana chip. Practicality hasn't inhibited Arntzen's idealism however. To this day he keeps a jar of Gerber's baby food  banana of course  on the corner of his desk for inspiration.

Arizon's sunny skies and warm weather are known to attract people from all over the world  who can resist playing golf in short sleeves in December? Arizonans are fortunate that warm weather is also good for growing plants. A combination of ASU's offer of the Florence Ely Nelson Presidential Endowed Chair and greenhouse opportunities brought Arntzen to Phoenix. He sounds remarkably like a winter tourist when he exclaims, The weather is perfect! Yet unlike the tourists, Arntzen is excited about greenhouse horticulture, not golf. In its own way, a functional edible vaccine would indeed be a hole-in-one.

http://www.molecularfarming.com/ediblevaccine.html

MOLECULAR FARMING OF EDIBLE VACCINES. {1998-2001}

{ with special thanks to Charles J. Arntzen, the global expert on edible vaccines }

1998 - FIRST HUMAN TRIAL SHOWS THAT AN EDIBLE VACCINE IS FEASIBLE

Baltimore, Maryland, April 28, 1998 Opening a new era in vaccine delivery, researchers supported by the National Institute of Allergy and Infectious Diseases (NIAID) have shown for the first time that an edible vaccine can safely trigger significant immune responses in people.

The report, by collaborators from the University of Maryland in Baltimore, the Boyce Thompson Institute for Plant Research in Ithaca, N.Y., and Tulane University in New Orleans, appears in the May issue of Nature Medicine. "Edible vaccines offer exciting possibilities for significantly reducing the burden of diseases like hepatitis and diarrhea, particularly in the developing world where storing and administering vaccines are often major problems," says Anthony S. Fauci, M.D., director of NIAID.

The Phase 1 proof-of-concept trial began last fall at the University of Maryland School of Medicine's Center for Vaccine Development under the direction of Carol O. Tacket, M.D., professor of medicine. The goal of the study was to demonstrate that an edible vaccine could stimulate an immune response in humans. Volunteers ate bite-sized pieces of raw potato that had been genetically engineered to produce part of the toxin secreted by the Escherichia coli bacterium, which causes diarrhea. Previously, NIAID-supported in vitro and preclinical studies by John Clements, Ph.D., and colleagues at Tulane University School of Medicine showed that transgenic potatoes containing this segment of the toxin stimulated strong immune responses in animals. The transgenic potatoes were created and grown by Charles Arntzen, Ph.D., and Hugh S. Mason, Ph.D., and their colleagues at the Boyce Thompson Institute for Plant Research, an affiliate of Cornell University. The trial enrolled 14 healthy adults; 11 were chosen at random to receive the genetically engineered potatoes and three received pieces of ordinary potatoes.
   
The investigators periodically collected blood and stool samples from the volunteers to evaluate the vaccine's ability to stimulate both systemic and intestinal immune responses. Ten of the 11 volunteers (91 percent) who ingested the transgenic potatoes had fourfold rises in serum antibodies at some point after immunization, and six of the 11 (55 percent) developed fourfold rises in intestinal antibodies. The potatoes were well tolerated and no one experienced serious adverse side effects.

Encouraged by the results of this study, NIAID-supported scientists are exploring the use of this technique for administering other antigens. Edible vaccines for other intestinal pathogens are already in the pipeline--for example, potatoes and bananas that might protect against Norwalk virus, a common cause of diarrhea, and potatoes and tomatoes that might protect against hepatitis B. Regina Rabinovich, M.D., oversees NIAID's Vaccine and Treatment Evaluation Program, of which the University of Maryland's vaccine center is a part. "This first trial is a milestone on the road to creating inexpensive vaccines that might be particularly useful in immunizing people in developing countries, where high cost and logistical issues, such as transportation and the need for certain vaccines to be refrigerated, can thwart effective vaccination programs," she comments. "The hope is that edible vaccines could be grown in many of the developing countries where they would actually be used."

Details of the Study

The study nurse at the University of Maryland peeled the potatoes just before they were eaten, because potato skin sometimes contains a compound that imparts a bitter taste and can cause nausea and stomach upset. The potatoes were then cut into small, uniform pieces and weighed into 50- gram and 100-gram doses. Each person received three doses of either 50 grams or 100 grams of potato over a three-week period, at 0, 7 and 21 days. The dosage size varied in order to evaluate any side effects from eating raw potatoes. NIAID is a component of the National Institutes of Health (NIH). NIAID conducts and supports research to prevent, diagnose and treat illnesses such as AIDS and other sexually transmitted diseases, malaria, tuberculosis, asthma and allergies. NIH is an agency of the U.S. Department of Health and Human Services.

References: Arntzen CJ. Pharmaceutical foodstuffs-oral immunization with transgenic plants. Nature Medicine (vaccine supplement) 1998;4(5):502-03.

Haq TA, Mason HS, Clements JD, and Arntzen CJ. Oral immunization with a recombinant bacterial antigen produced in transgenic plants. Science 1995;268:714-16.

Mason HS, Haq TA, Clements JD, and Arntzen CJ. Edible vaccine protects mice against E. coli heat-labile enterotoxin (LT): potatoes expressing a synthetic LT-B gene. Vaccine, In Press.

Tacket CO, Mason HS, Losonsky G, Clements JD, Levine MM and Arntzen CJ. Immunogenicity in humans of a recombinant bacterial antigen delivered in a transgenic potato. Nature Medicine 1998;4(5):607-09.

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PLANTS AND HUMAN HEALTH: DELIVERY OF VACCINES VIA TRANSGENIC PLANTS


Tsafrir S. Mor and Charles J. Arntzen 2002
Arizona Biomedical Institute and the Plant Biology Department, PO Box 1601, Arizona State University, Tempe, AZ 85287-1601 (emails: tsafrir.mor@asu.edu, charles.arntzen@asu.edu)


Keywords mucosal immunity, infectious diseases, subunit oral vaccines


1. THE THEORY BEHIND EDIBLE VACCINES

One of major challenges of biotechnology is to reduce clinical innovations to economically viable practices. Plant-derived edible vaccines were first conceived and are continuing to be developed with this prime directive in mind: merging innovations in medical science and plant biology for the creation of efficacious and affordable pharmaceuticals. Since the emergence of the original idea about 10 years ago, it was embraced by a growing number of laboratories in academia and industry. Recent reviews provide detail about progress acheived (Daniell, et al., 2001; Mor, et al., 1998; Tacket and Mason, 1999).

Despite notable successes, traditional vaccine technology has its limitations. Almost all vaccines now commercially available consist of either inactivated or attenuated strains of pathogens which are almost always delivered by injection. (The oral polio vaccine is an exception.) In contrast, many of the currant vaccine development efforts focus on subunit vaccines, and these are being considered for either mucosal or parenteral delivery.

A "subunit vaccine" refers to a pathogen-derived protein (or even just an immunogenic domain of a protein, ie. "an epitope") that cannot cause disease but can elicit a protective immune response against the pathogen. Very often the subunit vaccine candidate is a recombinant protein made in transgenic production-hosts (such as cultured yeast cells), then purified, and injected into vaccinees to immunize against a specific disease. Subunit vaccines are generally considered safer to produce (eliminating the need to culture pathogenic organisms) and more importantly, to use.
However, immunization by injection (parenteral delivery) rarely results in specific protective immune responses at the mucosal surfaces of the respiratory, gastrointestinal and genito-urinary tracts. Mucosal immune responses represent a first line of defense against most pathogens. In contrast, mucosally targeted vaccines achieve stimulation of both the systemic as well as the mucosal immune networks. In addition, mucosal vaccines delivered orally increase safety and compliance by eliminating the need for needles. While subunit vaccines are effective, they currently depend on capital-intensive fermentaion-based technology and a "cold chain" (refrigeration) for delivery. Both of these factors create constraints in use in the developing world, where vaccines are needed the most. Combining a cost-effective production system with a safe and efficacious delivery system, plant edible vaccines, provide a compelling new opportunity.


2. THE THEORY IS PUT TO CLINICAL TRIALS

In 1992 our research team described the expression of hepatitis B surface antigen (HBsAg) in tobacco plants (Mason, et al., 1992). A subsequent succession of papers characterizing the recombinant product which assembled into virus like particles (VLPs, Mason, et al., 1992), and could invoke specific immune responses in mice upon parenteral delivery (Thanavala, et al., 1995). To prove that plant-derived HBsAg can stimulate mucosal immune responses via the oral route, our group switched to potato tubers as an expression system and optimized it to increase accumulation of the protein in the plant tubers (Richter, et al., 2000). The resulting plant material proved superior to the yeast-derived antigen in both priming and boosting of immune responses to oral immunogen in mice (Kong, et al., 2001; Richter, et al., 2000). In parallel with evaluation of plant-derived Hepatitis B surface antigen, Mason and Arntzen explored plant expression of other vaccine candidates including the labile toxin B subunit (LT-B) of entertotoxigenic Escherichia coli (ETEC) and the capsid protein of Norwalk virus (NVCP). The plant derived proteins correctly assembled into functional oligomers that could elicit the expected immune responses when given orally to animals (Haq, et al., 1995; Mason, et al., 1996; Mason, et al., 1998).

Success in mouse experiments provided motivation for conducting Phase I/II human clinical trials to test the safety and immunogenicity of plant-produced LT-B, NVCP and HBsAg (Tacket, et al., 1998; Tacket, et al., 2000, and Thanavala, Mason and Arntzen, unpublished). In the three cases tested, humans who consumed raw potato tubers containing tens of microgram amounts of the antigens developed specific serum and more importantly mucosal immune responses. Significantly, the three antigens in these studies come from three very different pathogens including viral (NV and HBV) and bacterial (E. coli) pathogens, and enteric (NV and E. coli) as well as non-enteric (HBV) disease (Tacket, et al., 1998; Tacket, et al., 2000). Taken together these results provide the basis for wider-scale clinical trials with these antigens which we plan to conduct with the aid of international agencies.

Although mucosal and systemic antibody titers were elevated in vaccinees who received the plant-based oral vaccines, we do not yet have evidence of protection against pathogen challenge. Ethical considerations usually preclude clinical trials from directly assaying protection except in a few cases (e.g. Mason, et al., 1998). In contrast, working with veterinary vaccines provides researchers an opportunity to assess the degree of immune protection more directly. An excellent example of this approach is represented by a series of papers originating from the group of Borca (Carrillo, et al., 2001 and references therein).


3. "SECOND GENERATION" EDIBLE VACCINES

Multicomponent vaccines that provide protection against several pathogens are very desirable. An elegant approach to achieve this goal, based on epitope fusion to both subunits of the cholera toxin (CT), was recently demonstrated by Yu and Langridge (2001). CT provides a scaffold for presentation of protective epitopes of rotavirus and ETEC, acts as as a vaccine candidate by its own right and as a mucosal adjuvant devoid of toxicity. The trivalent edible vaccine elicited significant humoral responses, as well as immune memory B cells and T-helper cell responses, important hallmarks of successful immunization (Yu and Langridge, 2001)..

Commonly, foreign proteins in plants accumulate to relatively low levels (0.01-2% of total soluble protein). In the clinical trials described above, 100 g of raw potato tubers expressing LT-B of ETEC in three doses had to be consumed in order to overcome digestive losses of the antigen and to elicit a significant immune response (Tacket, et al., 1998). Less immunogenic proteins would require even larger doses to be effective. Even with more palatable alternatives to potatoes (e.g. bananas), these accumulation levels may limit the practicality of edible vaccines

Two solutions to overcome this limitation are being explored. First, techniques to enhance antigen accumulation in plant tissues are being explored. These include, optimization of the coding sequence of bacterial or viral genes for expression as plant nuclear genes, and defining the subcellular compartment in which to accumulate the product for optimal quantity and quality. Several laboratories are also developing alternative expression systems to improve accumulation. For example the expression in plastids is advocated by some (Daniell, et al., 2001; Ruf, et al., 2001). Other systems involve plant viruses for expression of foreign genes (e.g. Nemchinov, et al., 2000) or coat-protein fusions (e.g. Modelska, et al., 1998) and even viral assisted expression in transgenic plants (Mor, et al., 2002).

The second approach is to enhance the immunogenicity of the orally delivered antigens by using mucosal adjuvants. One such approach is making use of bacterial entertoxins such as CT or LT (e.g. Yu and Langridge, 2001), mammalian and viral immunomodulators (Matoba, Soreq, Arntzen and Mor unpublished) as well as plant-derived secondary metabolites (Joshi and Arntzen, unpublished).

At doorstep of the 21st century, the fear of a surge in naturally occurring epidemics is heightened by the threat of bio-terrorism. This new reality makes disease prevention through vaccination a necessity in our ever more interconnected world. Any tools we can master and all the tools we can afford will have to be employed. Technical problems and skeptics aside, edible-vaccines have passed the major hurdles of an emerging vaccine technology. We believe production of vaccines in transgenic plants will become an essential component in our disease prevention arsenal.


4. REFERENCES

Carrillo C., A. Wigdorovitz, K. Trono, M.J. Dus Santos, S. Castanon, A.M. Sadir, R. Ordas, J.M. Escribano and M.V. Borca. 2001. Induction of a virus-specific antibody response to foot and mouth disease virus using the structural protein VP1 expressed in transgenic potato plants. Viral Immunol. 14:49-57.
Daniell H., S.J. Streatfield and K. Wycoff. 2001. Medical molecular farming: production of antibodies, biopharmaceuticals and edible vaccines in plants. Trends Plant Sci. 6:219-226.
Haq T.A., H.S. Mason, J.D. Clements and C.J. Arntzen. 1995. Oral Immunization with a recombinant Bacterial antigen produced in transgenic plants. Science 268:714-716.
Kong Q., L. Richter, Y.F. Yang, C.J. Arntzen, H.S. Mason and Y. Thanavala. 2001. Oral immunization with hepatitis B surface antigen expressed in transgenic plants. Proc. Natl. Acad. Sci. U.S.A. 98:11539-11544.
Mason H.S., D.M.K. Lam and C.J. Arntzen. 1992. Expression of hepatitis B surface antigen in transgenic plants. Proc. Natl. Acad. Sci. U.S.A. 89:11745-11749.
Mason H.S., J.M. Ball, J.-J. Shi, X. Jiang, M.K. Estes and C.J. Arntzen. 1996. Expression of Norwalk virus capsid protein in transgenic tobacco and protein and its oral immunogenicity in mice. Proc. Natl. Acad. Sci. U.S.A. 93:5335-5340.
Mason H.S., T.A. Haq, J.D. Clements and C.J. Arntzen. 1998. Edible Vaccine Protects Mice Against E. coli Heat-labile Enterotoxin (LT): Potatoes Expressing a Synthetic LT-B Gene. Vaccine 16:1336-1343.
Modelska A., B. Dietzschold, N. Sleysh, F.Z. Fu, K. Steplewski, D.C. Hooper, H. Koprowski and V. Yusibov. 1998. Immunization against rabies with plant-derived antigen. Proc. Natl. Acad. Sci. U.S.A. 95:2481-2485.
Mor T.S., M.A. Gmez-Lim and K.E. Palmer. 1998. Edible vaccines: a concept comes of age. Trends Microbiol. 6:449-453.
Mor T.S., Y.-S. Moon, K.E. Palmer and H.S. Mason. 2002. Geminivirus Vectors for High Level Expression of Foreign Proteins in Plant Cells. Biotechnol. Bioeng. in press.
Nemchinov L.G., T.J. Liang, M.M. Rifaat, H.M. Mazyad, A. Hadidi and J.M. Keith. 2000. Development of a plant-derived subunit vaccine candidate against hepatitis C virus. Arch. Virol. 145:2557-2573.
Richter L.J., Y. Thanavala, C.J. Arntzen and H.S. Mason. 2000. Production of hepatitis B surface antigen in transgenic plants for oral immunization. Nat. Biotechnol. 18:1167-1171.
Ruf S., M. Hermann, I.J. Berger, H. Carrer and R. Bock. 2001. Stable genetic transformation of tomato plastids and expression of a foreign protein in fruit. Nat. Biotechnol. 19:870-875.
Tacket C.O., H.S. Mason, G. Losonsky, J.D. Clements, S.S. Wasserman, M.M. Levine and C.J. Arntzen. 1998. Immunogenicity in humans of a recombinant bacterial-antigen delivered in transgenic potato. Nat. Med. 4:607-609.
Tacket C.O. and H.S. Mason. 1999. A review of oral vaccination with transgenic vegetables. Microbes Infect. 1:777-783.
Tacket C.O., H.S. Mason, G. Losonsky, M.K. Estes, M.M. Levine and C.J. Arntzen. 2000. Human immune responses to a novel Norwalk virus vaccine delivered in transgenic potatoes. J. Infect. Dis. 182:302-305.
Thanavala Y., Y.-F. Yang, P. Lyons, H.S. Mason and C.J. Arntzen. 1995. Immunogenicity of transgenic plant-derived hepatitis B surface antigen. Proc. Natl. Acad. Sci. U.S.A. 92:3358-3361.
Yu J. and W.H. Langridge. 2001. A plant-based multicomponent vaccine protects mice from enteric diseases. Nat. Biotechnol. 19:548-552.


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2000 --- { AN ARTICLE FROM E.N.N. }

Recently the glare of the media spotlight has fallen on genetically engineered food crops bred to resist herbicides and insects. Meanwhile, plants engineered with human proteins to produce drugs and vaccines for human consumption have escaped notice. Well, take note: At least 350 genetically engineered pharmaceutical products are currently in clinical development in the United States and Canada. Scientists believe that potent drugs and vaccines will soon be harvested just like wheat and corn. Welcome to the new world of molecular farming.

In Canada, a genetically engineered tobacco plant made to produce Interleukin 10 will be tested to treat Crohn's disease, an intestinal disorder. Molecular farming uses the science of genetic engineering to turn ordinary plants into factories for the production of inexpensive drugs and vaccines. Researchers at the London Health Sciences Center in London, Ontario, Canada, are growing potatoes that have been genetically altered to produce a special diabetes-related protein. When the potatoes are fed to diabetic mice, scientists find that most don't develop Type I diabetes, also known as juvenile-onset diabetes. Scientists believe that the low-cost production of this protein may help the 100 million people worldwide affected by diabetes. In the lab, the new transgenic potatoes produce large amounts of a human protein that suppresses the destructive immune response and prevents diabetes from developing.

Molecular biologist Shengwu Ma of the London Health Sciences Center says his team's research has similar potential to combate other autoimmune diseases such as multiple sclerosis, rheumatoid arthritis, lupus and even transplant rejection. "Plants are ideal because they can synthesize and assemble proteins to provide huge quantities of soluble proteins at relatively low cost," says Ma. Many traditional drugs are difficult to make and hence, costly. However, once this technology is perfected, growing transgenic potatoes will cost very little, he notes.

Edible vaccines were first tested on humans in 1997, when scientists asked volunteers to eat anti-diarrheal transgenic potatoes produced by the Boyce Thompson Institute at Cornell University. After consuming the potatoes, almost all the volunteers produced antigens in their bodies just as if they had received a traditional anti-diarrheal vaccination. And they experienced no adverse side effects. Volunteers are also testing raw potatoes engineered to produce a Hepatitis B antigen at the Roswell Park Cancer Institute in Buffalo, New York. Results are expected this summer. Hugh Mason, an associate research scientist in edible vaccines at the Boyce Thompson Institute, hopes to develop "methods to increase production of foreign protein in plant cells and to engineer protein antigens that will enhance their potential as human and animal vaccines." This fall Mason hopes to do human tests on Hepatitis B antigens grown in transgenic tomatoes if the FDA approves. "This technology will be a big plus for the developing world," he says.

copyright for this article - Enviromental News Network 2000 Article by Stephen Leahy
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2001 AUSTRALIAN DEVELOPMENTS
Edible vaccines the key to better immunisation

Monash University scientists have successfully grown a genetically engineered plant containing a measles vaccine in a technique that may eventually lead to simpler and cheaper immunisation programs for a range of viral diseases, including HIV and malaria.

Led by Professor Steve Wesselingh, the research team successfully produced a tobacco leaf containing a viral protein found in the measles virus. When the plant was processed and fed to mice, their immune system responded by producing protective antibodies. Testing has now begun on primates.

The research team is now developing the protein in a range of foods including rice and lettuce and have recognised the potential for the protein to be incorporated into baby food.

"There is no real reason why we couldn't be working with any type of food, but we believe that rice flour, when mixed with breast milk as baby food, is a simple and cheap option even for poor or remote communities," said Professor Wesselingh.

Although measles can be effectively prevented by a 'live' measles vaccine injection, it still causes up to one million deaths each year, mostly among young children in developing countries. In these countries, injectable vaccines are inhibited by many factors, including the need to provide a stable and cold environment during storage and transportation and a lack of trained medical staff to administer the vaccine.

The quest for new and better ways to immunise people against infectious diseases has led to a variety of alternatives to injections, with the food-based vaccine research providing the greatest potential.

Current measles vaccines are made from the actual virus and work by priming the immune system to attack if it becomes exposed to a full assault of the measles virus. In contrast, plant-based vaccines rely on the measles virus gene for the H protein being genetically cloned into the plant.

The H protein sits on the outside of the virus and has a role in provoking the immune response in the body. The edible vaccines, therefore, do not contain the complete 'live' virus - only the key protein to trigger the immune response.

The Monash researchers are working closely with scientists at the CSIRO Plant Industry and at the University of Melbourne

2001 - from TRENDS IN PLANT SCIENCE . Vol.6 No.5 {page 222} [by permission of Elsevier Science]

Proteins with applications for human or animal vaccines and expressed by transgenic plants


KEYS - DISEASE TARGET {Source of the protein} and target species for the VACCINES - PLANT EXPRESSION SYSTEM - Notes and PROTECTIVE CAPACITY of the VACCINES

Enterotoxigenic E. COLI (humans)-TOBACCO - Immunogenic when administered orally
Enterotoxigenic E. COLI {humans}- POTATO - Immunogenic and protective when administered orally
Enterotoxigenic E. COLI {humans}- MAIZE - Immunogenic and protective when administered orally

Vibrio cholerae [CHOLERA] (humans) - POTATO- Immunogenic and protective when administered orally

Hepatitis B virus {humans}- TOBACCO -Extracted protein is immunogenic when administered by injection
Hepatitis B virus {humans} - POTATO - Immunogenic when administered orally
Hepatitis B virus {humans}- LUPIN - Immunogenic when administered orally
Hepatitis B virus {humans}- LETTUCE - Immunogenic when administered orally

Norwalk virus (humans) - TOBACCO - Immunogenic when administered orally
Norwalk virus (humans) - POTATO - Virus-like particles form and immunogenic when administered orally

RABIES virus (humans) -TOMATO - Intact Glycoprotein

Human cytomegalovirus {humans} - TOBACCO - Immunologically related protein

Rabbit hemorrhagic disease virus {rabbits} - POTATO - Immunogenic and protective when administered by injection
FOOT-AND-MOUTH disease {agricultural domestic animals} - ARABIDOPSIS - Immunogenic and protective when administered by injection

FOOT-AND-MOUTH disease {agricultural domestic animals}- ALFALFA - Immunogenic and protective when administered by injection or orally

Transmissible gastroenteritis coronavirus (pigs) - ARIBIDOPSIS - Immunogenic when administered by injection

Transmissible gastroenteritis coronavirus (pigs) - TOBACCO - Intact protein and immunogenic when administered by injection
Transmissible gastroenteritis coronavirus (pigs) - MAIZE - Protective when administered orally

More on edible vaccine from the University of Wisconsin HERE

http://whyfiles.org/166plant_vaccines/biblio.html

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Re: Delivery Of Vaccines Via Transgenic Plants - Edible Vaccines
« Reply #1 on: May 05, 2009, 11:37:45 pm »
absolutely chilling
where will this stop?

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Re: Delivery Of Vaccines Via Transgenic Plants - Edible Vaccines
« Reply #2 on: May 05, 2009, 11:56:30 pm »
Bill Gates funds British scientists in unorthodox health research
http://forum.prisonplanet.com/index.php?topic=103946.0

http://www.timesonline.co.uk/tol/news/uk/science/article6222056.ece

Other projects among the 81 recipients of Grand Challenges Explorations grants, which come from universities, research institutes, non-profit organisations and private companies in 17 different countries, include: giving mosquitoes a head cold to prevent them from detecting and biting humans; using immunised cows as a means of killing or reducing the reproductive abilities of the mosquitoes that bite them; creating therapeutic tomatoes, modified to deliver antiviral drugs targeting particular viruses; and using a laser on skin before an injection to enhance immune responses stimulated by a vaccine.

Read the full article
http://www.timesonline.co.uk/tol/news/uk/science/article6222056.ece

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Re: Delivery Of Vaccines Via Transgenic Plants - Edible Vaccines
« Reply #3 on: May 06, 2009, 12:00:15 am »
http://www.icbps.org/ICBPS_2/Program_2.html

Welcome to the International China Biopharmaceutical Symposium
Sponsored by: ... Battelle Dec. 7, 2008

08:30 am 09:00 am     Opening Ceremony
Chair: Ms. Shaoli Li, Vice President & General Secretary, CPA
Co-chair: Dr. David Robinson, Vice President, Chief Biologist, Battelle Memorial Institute

http://www.icbps.org/ICBPS_2/Abstracts_2.html

"Edible hepatitis B vaccine on the basis of transgenic tomato or carrot plants"
Sergei Shchelkunov, State Research Center of Virology and Biotechnology VECTOR, Russia

http://www.ncsu.edu/extension/news/documents/trumblecv.node1.pdf

http://74.125.95.132/search?q=cache:pcz2CHw9GfsJ:www.ncsu.edu/extension/news/documents/trumblecv.node1.pdf+edible+vaccine+Battelle&cd=5&hl=en&ct=clnk&gl=us

CURRICULUM VITAE
University of New Hampshire
Durham, New Hampshire 03824-3587

NAME: Trumble, William R. DATE: January, 2007

RANK OR TITLE: Dean, College of Life Sciences and Agriculture
Director, New Hampshire Agricultural Experiment Station
Professor of Biochemistry and Molecular Biology
DEPARTMENT: Biochemistry and Molecular Biology E-MAIL: bill.trumble@unh.edu
...
Administrative Summary
...
Project Manager, Battelle N.W. Laboratories, Hanford National Lab, Richland, WA. 1984 -
1987.

...
1993-1999, Associate Professor of Molecular Biology and Biochemistry, Department of
Microbiology, Molecular Biology and Biochemistry, University of Idaho, Moscow, Idaho.


Structure-function studies of muscle calsequestrin. Site-directed mutagenesis of calsequestrin to establish intramolecular interaction sites. Bacterial invasion of mammalian cells in mastitis.

Cardiotoxin as a ligand for Ca2+-binding proteins.

Molecular engineering of an edible vaccine in potato.

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Re: Delivery Of Vaccines Via Transgenic Plants - Edible Vaccines
« Reply #4 on: May 06, 2009, 12:31:56 am »
http://archives.cpmp2005.org/maintemplate.aspx?lang=en

http://archives.cpmp2005.org/About.aspx

The Conference on Plant-made Pharmaceuticals is organized by the Society for Moleculture (formerly known as the international Association of Molecular Farming), and is funded by organizations using plant-factories to produce biopharmaceuticals.

Board of Directors (As of January 2005)

Mr. Franois Arcand, president, Society for Moleculture and ERA Plantech, Spain
Dr. Paul Arnison, FAAR Biotech and Saponin, Canada
Dr. Julio Baez, Fibrogen, USA
Dr. Antony Blanc, Syngenta, Switzerland
Dr. Julian Ma, St. George's Hospital, U.K., Medical School and Planta Pharma, Europe
Mr. Butch Mercer, Dow AgroSciences, USA
Dr. Maurice Moloney, SemBioSys, Canada
Dr. Ulrich Steiner, Bayer CropScience, BioScience

Montral, Qubec, Canada
Tuesday, February 01, 2005

Expression of Recombinant Coagulation Factors in Plants

Dr. Brian Hooker, Chief Engineer
Battelle Memorial Institute, USA


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Battelle is currently developing a variety of recombinant protein products in transgenic plants for the pharmaceutical industry. These include tissue replacement proteins for a variety of therapeutic and reagent applications. Battelle has the ability to produce these proteins using its Transgenic Plant Protein Production Platform (tP4) that allows for the large-scale manufacture of recombinant proteins in greenhouses, thereby facilitating the regulatory and public acceptance of the technology as well as a commercially viable, low cost alternative to other production technologies. Elements of the tP4 technology are generally applicable to both monocot and dicot-based plant systems and available for license. Efforts towards the production of coagulation factor XIII and factor VIII will be presented.

Transgenic rice Power Flour as a powerful seed-based recombinant protein expression system: the Seed as Pill concept.

Prof. ILLIMAR ALTOSAAR, PROFESSOR
UNIVERSITY OF OTTAWA, CANADA

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Given that pills are composed of excipient starch, transgenic rice flour can supply a well characterized source of pharmaceutical ingredients which are gluten-free and incorporated in many drug delivery applications. This unique moleculture system exploits the synthetic capacity of rice endosperm, the basis of rice flour. We have successfully produced many recombinant drug candidates on our rice platform, including insulin-like growth factors 1 and 1B (IGF-1), human CD-14, Hepatitis B Virus core particle protein, bovine serum albumin to explore Type 1 Diabetes links, and even recombinant granulocyte macrophage colony stimulating factor GM-CSF, a cytokine used in treating neutropenia.

Such Plant-Made Pharmaceuticals (PMPs) with potential clinical applications are expressed in rice seeds using a glutelin promoter. Rice grain accumulates 8% protein normally, PMP up to 1-3% of total soluble protein (TSP). Such Power Flour seed-produced proteins are bio-active when assayed on human target cell lines. Protein Farming in the Rice Protein Plant provides safe stable accumulated production (StorUrProtein) in seeds and self-containment of foreign genes due to self-pollination features. Rice proteins enjoy a nutritionally safe status historically. So these rice Power Flour pharma-proteins are novel since this is the first report where additional medical proteins have been produced in transgenic rice seeds. The mature seeds from these plants were found to contain high levels of IGF-1 etc. Levels for e.g. GM-CSF were more than 4-fold higher than the reported expression level in the seeds of tobacco. rth Protein levels in rice seeds may be increased by deploying an even larger Gt1 promoter, one that has raised phaseolin production to 4% in rice endosperm cells.

Basic science developments in the tissue culture and regeneration of commercial rice plants have laid the critical foundations allowing for these productive research collaborations with Genentech Inc., Bayer CropScience, Miles Pharmaceuticals, Red Cross, Monsanto and Health Canada.

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Re: Delivery Of Vaccines Via Transgenic Plants - Edible Vaccines
« Reply #5 on: May 06, 2009, 12:48:31 am »
http://www.bioresearchonline.com/article.mvc/Battelle-Develops-Transgenic-Plant-Protein-Pr-0001?VNETCOOKIE=NO

Battelle Develops Transgenic Plant Protein Production Platform
February 8, 1999

If your company is looking for plant-based protein sources, look into a new transgenic tobacco from Battelle's Pacific Northwest Laboratory. Battelle scientists have developed a new transgenic plant protein production platform, dubbed tP4, and are looking for industrial commercialization partners. They claim that tP4 is a safer and much less expensive route to high-value proteins useful in the manufacture of pharmaceutical, agricultural, chemical, food, diagnostic, and consumer products.

Using tP4, plants are rapidly, genetically transformed, resulting in "plants as bioreactors" that synthesize protein products and crop plants phenotypes that possess unique, beneficial properties. The tP4 system relies on established molecular biology methods for efficient expression of specific proteins.

Examples of specific proteins already produced using tP4 include cellulases (applications in textile, chemical process, and food processing industries) and human growth factors (potential wound care therapeutic).

The tP4 system has the following novel capabilities:

Product quality, safety, and stability. Consistent product quality is essential for all protein product applications. Problems associated with traditional recombinant techniques (microbial fermentation and mammalian cell culture), including improper folding, pyrogenicity, and pathogen transmission, may be circumvented since transgenic plants produce properly processed, biologically active proteins with greatly reduced pathogen transmission risk. Stability of production is also enhanced as transgenic plant seeds may be stored indefinitely and subsequently planted for optimum product delivery.

Whole plant and suspension cell transformation. Both of these systems possess distinctive advantages for production, cost, and regulatory concerns over the current methods. Whole plant production strategies afford ease in scale-up to meet varying product demands while dramatically reducing capital equipment costs. Cell culture production strategies are directly portable to industries currently using fermentation equipment and where strict process validation is crucial for product approval. By possessing capabilities in both systems, the development time required to produce large quantities of single proteins is greatly reduced through 1) rapid transformation and transformant screening (successful transformation and screening activities may require as little as three months), 2) efficient proliferation of plant material, and 3) ease and flexibility in production system scale-up.

Targeted expression of proteins in the non-crop portion of a plant

Ability to target specific plant portions for overexpression. Targeted expression of foreign proteins in designated plant organs allows for the formulation of novel harvesting and separation strategies. Depending on production/formulation constraints of the foreign protein to be over-expressed, synthesis may be limited to plant roots, leaves, or fruit. Additionally, at the cellular level, products may be targeted to specific organelles, affording 1) a protective environment to sequester enzymes that may be deleterious to plants, 2) necessary protein folding and other post-translational modifications, and 3) simplified downstream separations.

Applications-based product delivery. To maintain competitiveness, the product formulation must cost-effectively match its application. For example, an industrial enzyme preparation may require only a fraction of the purification/validation necessary for protein therapeutics and reagents. The tP4 system uses specific strategies to minimize cost in product delivery, while maintaining formulations that effectively deliver the necessary catalytic function.

Design and scale-up of production systems. Past experience in large- and medium-scale cell culture fermentation and product separations allows Battelle scientists to develop both novel transgenic plants and the market-scale production systems needed for either whole plants or cell culture.

For More Information: Brian Hooker, staff scientist, Battelle Pacific Northwest Division, Battelle, P.O. Box 999, Richland, WA 99352. Tel: 509-375-4420. Email: brian.hooker@pnl.gov.

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Re: Delivery Of Vaccines Via Transgenic Plants - Edible Vaccines
« Reply #6 on: June 03, 2009, 06:17:29 pm »
http://www.sciencedaily.com/releases/2009/05/090514084103.htm

HIV Vaccine From Engineered Plants: Mice Form Antibodies Against HIV Protein

ScienceDaily (May 18, 2009) A research team at rebro University in Sweden has succeeded in changing the genes in plants so they can function as a vaccine against HIV. Through gene modification the plants have acquired the capacity to produce a protein that is part of the virus, and the project has taken a giant step forward in that mice that have been fed the plants have reacted and formed antibodies against the protein.

The findings are presented in a new academic dissertation at the university.

To produce drugs with the help of plants is a rapidly growing research field that offers new potential to combat diseases. At rebro University researchers have the goal of developing inexpensive and safe protection against HIV, in the form of plants that contain a vaccine against the virus and can be cultivated all over the world. If they succeed, it will be difficult to exaggerate the significance of this for millions of people around the world, not least in the poorest countries.

A major problem with the HIV virus is that it mutates rapidly and therefore exists in several different variants. In other words, its not possible to create an effective vaccine that is based on the entire virus. Moreover, this would be far too risky. Instead, we have selected a protein, p24, that exists in all HIV viruses and looks roughly the same in the various virus lines, says Ingrid Lindh, author of the dissertation.

To get plants to produce the p24 protein, the gene that underlies the process must be a part of their own genetic make-up, but since its impossible to transfer the gene directly from the virus to the plant, the researchers had to take a detour. This was done by first placing the gene into a bacterium that could then transmit it to the plants. The attempt succeeded; the plants produced p24 and also passed on this ability to their offspring.

In the next phase, mice were fed with the p24 plants, and these trials also proved to be successful. The mices immune defense reacted just as the researchers had hoped, producing antibodies against the protein. In other words, this functioned as a vaccine. This raises hopes that a similar reaction in humans would make them immune to HIV.

It is highly probable that the human immune system will respond in the same manner, but this is not to say that this would be sufficient to provide complete protection.

To increase the potency of the vaccine, these scientists are therefore going to add more HIV proteins together with other compounds that reinforce the bodys reaction to HIV-specific proteins. In parallel with this, they will work to select a suitable vegetable that is easy to cultivate in different climates and is readily accepted in different cultures. Thus far, thale cress (Arabidopsis thaliana) has been used as an experimental plant, a common wild plant that is related to mustard and cabbage and has the great advantage of being well mapped genetically.

The carrot is a good candidate for producing an edible vaccine, not least because it can be eaten raw, which reduces the risk of the proteins being destroyed by heating. Whats more, its a biennial, which means that it doesnt go to seed the first year, making it easier to ensure that it doesnt spread its genes to other plants close by, explains Ingrid Lindh.