The Use of Plants to Make Vaccines: Part-2 is the second of a 3-set series of articles discussing issues involved in making vaccines in plants. In this Part-2 article we will focus on vaccines engineered in potato, tobacco, banana and tomato plants; and the scope for making antibodies in plants.

Charles J Arntzen is regarded as a pioneer in research on vaccines derived from plants. It was his research group that proved HBsAg derived from plants was identical both antigenically and physically to HBsAg particles produced using human serum and recombinant yeast. This was proof that transgenic plants could be used to produce vaccines. Similarly researchers Haq and his group showed that E.Coli LT-B (heat labile enteroxin) made in tobacco and potato plants were identical physically and functionally to E.coli produced protein.

The first ever expression of vaccines in plants was carried out by Curtiss et al with the expression of SpaA surface protein antigen of Streptococcus mutans in tobacco. This happened in 1990. Since then hepatitis B surface antigen (HBsAg) and several other subunits of bacterial toxins have been expressed in plants. Vaccine components have also been expressed in plants


Vaccines in Potato Plant

Edible vaccines were initially made in potato plants, but as potato is not conducive to be eaten in raw form but only after being boiled, it created a problem in that, boiling the potato destroyed almost half the vaccine content. That apart, there was inconsistency in the levels of vaccine expressed in potatoes giving rise to inconsistency of dose in the recipients of those vaccines. A solution to this problem is to make protein extracts from the potatoes in order to enable uniformity of dose -- but this entails additional costs. Clinical trials are underway for plant-derived oral vaccines from potato for HBsAg and LT-B. Norwalk virus and hepatitis B virus antigens have been successfully expressed in potato plants

Other vaccines expressed in potato plants are:
Vibrio cholerae toxin B subunit-human insulin fusion for Autoimmune diabetes has been expressed in potato plants using Agrobacterium mediated transformation expression system.
V. cholerae toxin CtoxA and CtoxB subunits as oral vaccine against cholera can also be made in potato plants using Agrobacterium mediated transformation expression system.

Using tobacco plants to make vaccines

Tobacco plants could be used to make specialized vaccines for cancer patients. First the antibody is isolated from a patient's tumor. Then this antibody is inserted into the modified version of tobacco mosaic virus, which is a retrovirus that has the capacity to cause disease in tobacco plants. When the virus is scratched on the tobacco plant leaves, the plant gets infected with the gene carrying virus. Slowly the virus transfers the gene to the plant cells and the result is generation of antibodies. All it takes then is to cut a few plant leaves after a few days and purify the vaccine protein. A few plants would be enough to make vaccines for a single patient. Speed is of essence as a cancer patient would likely jump to chemotherapy unless a vaccine is quickly developed ---- and in this regard, plants in future would be the saviors of cancer patients.

The tobacco plant has plenty of biomass and produces plenty of seeds as well, which is helpful for growing fresh crops. The other obvious advantage is the fact that tobacco being a non-crop plant the possibility of the transgene entering the human food chain is indeed remote.

Several vaccines can be expressed in the tobacco plant. Some of the specific instances are: Recombinant HBsAg for hepatitis B can be made using Agrobacterium mediated transformation (AMT); Murine hepatitis epitope also for hepatitis B can be made using tobacco mosaic virus expression system; E. coli heat-labile enterotoxin LT-B for Cholera and E. coli diarrhea can be made using Agrobacterium mediated transformation; Rabies virus glycoprotein for rabies can be made using AMT expression system; HIV epitope (gp120) for HIV can be made using cow-pea mosaic virus and AMT expression systems; Mala c-Myc rial B-cell epitope for malaria can be made using tobacco mosaic virus expression system; and c-Myc for cancer can be made using tobacco mosaic virus expression system.

Using banana plants to make vaccines

A genetically altered virus when injected into a banana plant becomes an integral part of the plant cells. So as the plant grows, the banana plant cells start making the specific protein (unrelated to the infectious part of the virus) ---and when plant parts are eaten it can stimulate the immune system to make antibodies. Otherwise the banana plants would be grown and harvested and finally processed to extract the vaccine. In countries with burgeoning population, no production line can cater to the demand excepting plants like banana plants --- and that's a compelling reason for making vaccines in plants. A vaccine to protect against the Norwalk virus has been successfully expressed in the banana plant. Besides that, efforts to express Cholera toxin B subunit in Banana callus culture have also met with success.

Using Tomato Plants to Make Vaccines

Tomatoes can be freshly eaten. So it allows a great possibility of using tomatoes as an oral vaccine vector. Tomato leaves contain plant sugars like tomatine in abundance that are the key to developing new types of vaccine delivery systems. Tomatine is helpful for assisting the impregnation of compounds that combat bacteria directly into the cells. Tomatine traps antigens and increases their efficacy.

But the problem is about the quantum of protein expression levels in tomatoes. This tends to be on the lower side and this creates problems in administering the vaccine. To a certain extent this problem can be resolved by resorting to the following steps.
a) By using stronger promoters
b) By using signal peptide
c) By using plant sourced leader sequences
d) By restricting the expression of proteins only in edible tissue such as the fruit only.

So far Norwalk virus, Vibrio cholerae, and hepatitis B virus antigens have been successfully expressed tomato plants ---- and these tomatoes produced desired blood serum levels and sufficient immunity. When HEV-E2 gene was put into tomatoes using Agrobacterium tumefaciens HEV specific antigenicity could be achieved. This study is reckoned as the basis for development of new types of plant-derived hepatitis E oral vaccine and other oral vaccines. In general, tomato is a fruit-vegetable that can be eaten raw, is nutritious, and can be transformed (easy to manipulate genetically) easily and crop-wise can be grown quickly--- so is an apt plant candidate for expression of oral vaccines, although tomatoes do not grow abundantly in the geographies were edible vaccines are most needed. On the plus side, such genetically modified tomatoes can be freeze dried, or grounded to paste for further processing to conventional form of dosage.

Antibodies in Plants

Instead of priming the immune system to produce antibodies there is another method to achieve immunization. That is by delivering antibodies directly to the body. Transgenic plants can be used to make these functional antibodies or plantibodies too. These recombinant antibodies could be immunoglobulins, antigen-binding fragments of immunoglobulins and synthetic scFv (single-chain variable fragment gene fusions).

Immunoglobulins~~~for example in the tobacco plant, Hybrid sIgA-G specific for S. mutans antigen II could be expressed using Agrobacterium mediated transformation. This is for treatment of dental caries. In the same plant, scFv of IgG from mouse B-cell lymphoma for treatment of non Hodgkin's lymphoma can be made using Agrobacterium mediated transformation. In cereals, tumor-associated marker antigen as for example scFvT84.66 against carcinoembryogenic antigen can be made using particle bombardment expression system.

Conclusion

In the context of immunizing children, the vaccines that are most sought after are the ones required for countering diseases such as measles, tetanus, diphtheria, polio and tuberculosis. It is hoped that plant-derived vaccines will one day address these diseases and the problem of other diseases such as hepatitis, cholera and yellow fever.

That apart, research into plant-based systems to make vaccines could help us source other biopharmaceuticals like glucocerebrosidase which are otherwise difficult to obtain and costly to produce.

About Author / Additional Info:
PART 1: http://www.biotecharticles.com/Agriculture-Article/The-Use-of-Plants-to-Make-Vaccines-Part-1-431.html

PART 3: http://www.biotecharticles.com/Agriculture-Article/The-Use-of-Plants-to-Make-Vaccines-Part-3-435.html