nirupama's blog

a.Rabies

Tomato plants expressing rabies antigens could induce antibodies in mice. Instead, TMV (Tobacco Mosaic Virus) may also be used. Transformed tomato plants using CaMV (Cauliflower Mosaic Virus) with the glycoprotein (G-protein) gene of rabies virus (ERA strain) were shown to be immunogenic in animals.

Scientists at the University of Agricultural Sciences (UAS), Bangalore, have also developed an edible vaccine for rabies. The vaccine can be injected into musk melons, lettuce and other vegetables. UAS has received a pre-patent and is awaiting a patent, according to Dr T.K. Siddarame Gowda, Head, Biotechnology Department. He said tests have been completed on mice at the National Institute of Mental Health & Neuro Sciences (NIMHANS), the country's only accredited centre for research on rabies vaccine.

The first human clinical trial of an edible vaccine took place in 1997, when volunteers ate raw potatoes genetically engineered against diarrhoea-causing E. coli. Ten of the 11 volunteers who received the vaccine had fourfold rises in serum antibodies.

 a.Diarrhoea

 Researchers from the Boyce Thompson Institute (BTI) at Cornell University conducted a clinical trial of an edible vaccine in 1999. Potatoes containing the Norwalk virus (which causes vomiting and diarrhoea) fed to volunteers elicited an immune response in 19 out of 20 volunteers.

Arntzen and John Clements, of Tulane University Medical School, have also worked with tobacco plants to provide immunity against diarrhoea caused by E. coli.

b. Measles

Antigen expression in plants has been successfully shown in the past, like LT-B (ETEC) in tobacco and potato, rabies virus-G protein in tomato, HBsAg in tobacco and potato, norwalk virus in tobacco and potato; CT-B (Vibrio cholerae) in potato.

a.ETEC

Charles Arntzen at Boyce Thompson Institute, USA, accomplished the first published successful human trial in 1997. Eleven volunteers were fed raw transgenic potatoes expressing LT-B. Ten of these individuals developed neutralizing antibodies and six developed mucosal responses.

b.Norwalk virus

Seroconversion is the development of detectable specific antibodies to microorganisms in the serum as a result of infection or immunization.
Nineteen out of 20 people fed with transgenic potato expressing Norwalk virus(cause of dysentery) antigen showed seroconversion.

c.Cholera

Regulatory concerns include lot-to-lot consistency, uniformity of dosage and purity.

Controversies still exist as to whether edible vaccines are to be regulated under food, drugs or agricultural products and what vaccine component should be licensed - antigen itself, genetically engineered fruit or transgenic seeds. They will be subjected to very close scrutiny by the regulatory bodies in order to ensure that they never enter the food supply. This will include greenhouse segregation of medicinal plants from food crops to prevent cross-breeding and will necessitate separate storage and processing facilities. Although edible vaccines fall under "GM" plants(genetically modified), it is hoped that these vaccines will avoid serious controversy, because they are intended to save lives. This is because of lack of investors' confidence in returns on investments in GM foods.

Three successful human clinical trials have shown that adequate doses of antigen can be achieved with plant-based vaccines.
To determine the right dosage, one must consider the person's weight, age; fruit/plant's size, ripeness and protein content. The amount to be eaten is critical, especially in infants, who might spit it, eat a part or eat it all and throw it up later. Too low a dose would fail to induce antibodies and too high a dose would, instead, cause tolerance.
Plants may also get contaminated by foreign proteins which accumulate in low amounts (0.01-2% of total protein) and are less immunogenic; therefore the oral dose exceeds the intranasal/parenteral dose.

We have so far seen expression of antigens in plants.
Expression of antibodies in plants is an example of passive immunity. So now, let us see how this works.

Introduction of foreign DNA into plant's genome can either be done by
a. bombarding embryonic suspension cell cultures using gene-gun (biolistic method)- pellets of metal (usually tungsten) coated with the desirable DNA are fired at plant cells. Those cells that take up the DNA (again, this is confirmed with a marker gene) are then allowed to grow into new plants, and may also be cloned to produce more genetically identical crop.
b. more commonly through Agrobacterium tumefaciens, a naturally occurring soil bacterium, which has the ability to get into plants through some kind of wound (scratch, etc.).

Let’s go back to what I discussed in “Concept behind edible vaccines”.

The gene which codes the active antigenic protein is isolated from the pathogen and is incorporated in a suitable “gene vehicle”. This gene vehicle is integrated into the genome of the plant and is allowed to express to produce the corresponding antigen. Then these plant parts are fed to animals and humans to run their course.

A "subunit vaccine" refers to a pathogen-derived protein that cannot cause disease but can elicit a protective immune response against the pathogen.
Conventional subunit vaccines as well as edible vaccines are composed of antigenic proteins and do not contain pathogenic genes(because obviously they use attenuated strains). Thus, they have no way of establishing infection and safety is assured.

Edible vaccines are created by introducing selected desired genes into plants and inducing these genetically modified plants to manufacture the encoded proteins. This process is known as "transformation," and the altered plants are called "transgenic plants." 

When an antigen of a pathogen can produce an immunogenic response when delivered orally, it is considered as a likely candidate for an edible vaccine. The gene encoding the orally active antigenic protein is isolated from the pathogen, and a suitable vehicle for constitutive or tissue-specific expression of the gene is prepared.