"edible vaccines": Vaccines or candidate vaccines derived from edible plants. Transgenic plants are used as recombinant protein production systems and the edible plant tissue functions as an oral vaccine.

1. EDIBLE VACCINESEDIBLE VACCINES
PRESENTED BY :- PRIYA DEVIPRESENTED BY :- PRIYA DEVI
MSc. BIOTECHNOLOGY (4MSc. BIOTECHNOLOGY (4thth
sem)sem)
DEPARTMENT OF BIOTECHNOLOGYDEPARTMENT OF BIOTECHNOLOGY

2. CONTENTSCONTENTS
 Introduction to vaccinesIntroduction to vaccines
 Ideal vaccineIdeal vaccine
 Introduction to edible vaccinesIntroduction to edible vaccines
 Concept of edible vaccineConcept of edible vaccine
 Why edible vaccines are used ?Why edible vaccines are used ?
 Mechanism of actionMechanism of action
 Methods for transformation of DNA/gene into plantsMethods for transformation of DNA/gene into plants
 Candidates for edible vaccinesCandidates for edible vaccines
 Consideration factors/factors affecting efficacy of edibleConsideration factors/factors affecting efficacy of edible
vaccinesvaccines
 Regulatory aspects/issuesRegulatory aspects/issues
 Current status/Applications of edible vaccinesCurrent status/Applications of edible vaccines
 Advantages & DisadvantagesAdvantages & Disadvantages
 Future aspectsFuture aspects

3. VaccinesVaccines
 A vaccine is aA vaccine is a biologicalbiological
preparationpreparation that improvesthat improves
immunity to a particular disease.immunity to a particular disease.
 It contains an agent that resemblesIt contains an agent that resembles
a disease-causing microorganisma disease-causing microorganism
and is often made from weakenedand is often made from weakened
or killed forms of the microbes, itsor killed forms of the microbes, its
toxins or one of its surfacetoxins or one of its surface
proteins.proteins.
 The process of distributing andThe process of distributing and
administrating vaccines is referredadministrating vaccines is referred
to as vaccination. Vaccination is ato as vaccination. Vaccination is a
form of immunization.form of immunization.
 Routes of administration, includingRoutes of administration, including
oral, nasal, intramuscular,oral, nasal, intramuscular,
subcutaneous, and intradermal.subcutaneous, and intradermal.
VACCINES
PROPHYLACTIC
(e.g. to prevent the
effects of a future
infection by any
Natural or “ wild”
pathogen )
THERAPEUTIC
(e.g. vaccines against
cancer)

4. IDEL VACCINEIDEL VACCINE
It should not be toxic or pathogenic.It should not be toxic or pathogenic.
Low levels of side effect.Low levels of side effect.
Long lasting humoral and cellular immunities.Long lasting humoral and cellular immunities.
It should not contaminate the environment.It should not contaminate the environment.
It should not cause problems in individual.It should not cause problems in individual.
Technique of vaccination should be simple.Technique of vaccination should be simple.
It should be cheap.It should be cheap.

5. EDIBLE VACCINESEDIBLE VACCINES
 A genetically manipulated food, containing organisms or relatedA genetically manipulated food, containing organisms or related
antigens that may provide active immunity against infection.antigens that may provide active immunity against infection.
 In the edible vaccines Transgenic plants are used as vaccineIn the edible vaccines Transgenic plants are used as vaccine
production systems.production systems.
 The genes encoding antigens of bacterial and viral pathogens canThe genes encoding antigens of bacterial and viral pathogens can
be expressed in the plants in a form in which they retain nativebe expressed in the plants in a form in which they retain native
immunogenic properties.immunogenic properties.
 Initially thought to be useful only for preventing infectiousInitially thought to be useful only for preventing infectious
diseases, it has to be found application in prevention ofdiseases, it has to be found application in prevention of
autoimmune diseases, birth control, cancer therapy, etc.autoimmune diseases, birth control, cancer therapy, etc.
 Edible vaccines are currently being developed for a number ofEdible vaccines are currently being developed for a number of
human and animal diseases.human and animal diseases.
 As Hippocrates said, Let “ thy food be thy medicine”.As Hippocrates said, Let “ thy food be thy medicine”.

6. Concept of edible vaccineConcept of edible vaccine
Developed by Arntzen in the 1990s.Developed by Arntzen in the 1990s.
Introduce gene of interest into plants (Introduce gene of interest into plants (TransformationTransformation))
Gene expressed in the plant tissues edible parts (Gene expressed in the plant tissues edible parts (Transgenic plantsTransgenic plants))
Genes encode putatively protective vaccine antigens form viral, bacterial, andGenes encode putatively protective vaccine antigens form viral, bacterial, and
parasitic pathogens that cause disease in humans and animalsparasitic pathogens that cause disease in humans and animals
Ingestion of the edible part of the transgenic plant (Ingestion of the edible part of the transgenic plant (oral delivery of vaccineoral delivery of vaccine))

7. Why edible vaccines used?Why edible vaccines used?
 Oral vaccines provide “mucosal immunity” at various sites by
secreting antibodies.
 Don’t need to worry about re-use, misuse and lack of
sterilization. Thus, low risk of infection .
 Estimated cost of $0.005 to grow antigen for one dose of
hepatitis B vaccine in an unprocessed form.
 Administering oral vaccines would require little or no training at
all.
 Heat-stable; do not require cold-chain maintaince.
 If the local/native crop of a particular area is engineered to
produce the vaccine, then the need for transportation and
distribution can be eliminated.
 Most importantly, they trigger the first line of defence.
 Needs no purification.
 They activates both mucosal and systemic immunity.
NEEDLE FREE
CHEAP
STORAGE
SAFE

8. Mechanism of actionMechanism of action
☼ The goal of oral vaccination is to stimulate the mucosal and systemicThe goal of oral vaccination is to stimulate the mucosal and systemic
immunity against pathogen.immunity against pathogen.
☼ Edible vaccine when taken orally undergoes the mastication process and theEdible vaccine when taken orally undergoes the mastication process and the
majority of plant cell degradation occur in the intestine as a result of action ofmajority of plant cell degradation occur in the intestine as a result of action of
digestive or bacterial enzyme on edible vaccine.digestive or bacterial enzyme on edible vaccine.
☼ Peyer’s patches (PP) are an enriched source of IgA producing plasma cells andPeyer’s patches (PP) are an enriched source of IgA producing plasma cells and
have the potential to populate mucosal tissue and serves as mucosal immunehave the potential to populate mucosal tissue and serves as mucosal immune
effector site.effector site.
☼ The breakdown of edible near PP, consisting of the 30-40 lymphoid nodulesThe breakdown of edible near PP, consisting of the 30-40 lymphoid nodules
on the outer surface of intestine and contain follicles.on the outer surface of intestine and contain follicles.
☼ These follicles act as the site from which antigen penetrates the intestinalThese follicles act as the site from which antigen penetrates the intestinal
epithelium, thereby accumulating antigen within organized lymphoid structure.epithelium, thereby accumulating antigen within organized lymphoid structure.
☼ The antigen then comes in contact with M-cell.The antigen then comes in contact with M-cell.
☼ M-cell passes the antigen to macrophages and B-cell.M-cell passes the antigen to macrophages and B-cell.
☼ These B cell activates the T cell to provide immune response.These B cell activates the T cell to provide immune response.
☼ In this way the immunity is activated by the edible vaccine.In this way the immunity is activated by the edible vaccine.

10. How edible vaccines provideHow edible vaccines provide
protectionprotection

11. Methods for transformation of genesMethods for transformation of genes
into plantsinto plants
TRANSIENT EXPRESSIONSTABLE INTEGRATION
VECTOR CARRIER
METHOD
MICROPROJECTILE
BOMBARDMENT
METHOD
ELECTRO-
PORATION
CHIMERIC VIRUSES

12. Plasmid/Vector carrier system:Plasmid/Vector carrier system:
Agrobacterium tumifaciensAgrobacterium tumifaciens methodmethod

13. Micro projectile bombardment/geneMicro projectile bombardment/gene
gun methodgun method
Selected DNA sequences are precipitatedSelected DNA sequences are precipitated
onto metal (e.g. Gold, tungsten) micro-onto metal (e.g. Gold, tungsten) micro-
particleparticle
Bombarded against the vegetable tissueBombarded against the vegetable tissue
with a particle gun at an acceleratedwith a particle gun at an accelerated
speedspeed
Micro-particles penetrate the walls andMicro-particles penetrate the walls and
release the exogenous DNA inside therelease the exogenous DNA inside the
cell where it will be integrated in thecell where it will be integrated in the
nuclear genomenuclear genome
Produce large number of geneticallyProduce large number of genetically
identical cropidentical crop

14. Chimeric viruses methodChimeric viruses method

15. Electro- porationElectro- poration
• Introduction of DNA into cells by exposingIntroduction of DNA into cells by exposing
them for brief period to high voltage electricalthem for brief period to high voltage electrical
pulse which is thought to induce transient porespulse which is thought to induce transient pores
in the plasma lemma.in the plasma lemma.
• The cell wall presents an effective barrier toThe cell wall presents an effective barrier to
DNA. Therefore, it has to be weakened by mildDNA. Therefore, it has to be weakened by mild
enzymatic treatment so as to allow the entry ofenzymatic treatment so as to allow the entry of
DNA into cell cytoplasm.DNA into cell cytoplasm.

16. Candidate plantsCandidate plants
POTATO
POTATO
Advantages:
Do not need cooking.
Proteins not destroyed even if cooked.
Inexpensive
Growth widely in developing countries.
Disadvantages:
Trees take 2-3 years to mature.
Spoil rapidly after ripening.
Advantages:
Easily transformed.
Easily propagated.
Stored for long periods without refrigeration.
Disadvantages:
Cooking denature antigen.
BANANA

17. Advantages:
Grow quickly.
Cultivate broadly.
High content Vitamin-A may boost immune response.
Disadvantages:
Spoils readily.
Advantages:
Commonly used in baby food low allergenic potential.
High expression of proteins.
Vaccines does not dissolve when exposed to stomach acids.
Less risk of contaminating than normal crop.
Disadvantages:
Grow slowly.
Specialized glasshouse conditions.
Advantages:
Cheaper.
Not need to be refrigerated.
Disadvantages:
Need cooking to use take a time to reach.
TOMATO
RICE
MAIZE

18. Advantages:
Good model for evaluating recombinant proteins.
Easy purification of antibodies.
Stored in the seeds.
Large harvests, number of time/year.
Disadvantages:
Produces high level of toxic alkaloids.
Advantages:
Large number of seeds help in increased harvest.
Disadvantages:
Need cooking.
Rich in beta-carotene, production of Insulin.
WHEAT
TOBACCO
CARROT

19. Factors affecting efficacy of edibleFactors affecting efficacy of edible
vaccinesvaccines
FACTORS
AFFECTING
EDIBLE
VACCINES
Antigen selection (safe, suitable, stable)
Efficacy in model systems (small qty)
Choice of plant species (suitable, easy grown, storage,
cost)
Delivery and dosing issues
Safety issues (allergic & toxic potential)
Public perceptions and attitudes to genetic modification
Quality control and licensing (consistent)

20. Regulatory AspectsRegulatory Aspects
It has to be decided whether edible vaccines would beIt has to be decided whether edible vaccines would be
regulated under food, drugs or agricultural products.regulated under food, drugs or agricultural products.
Undecided that licensing shall be required for theUndecided that licensing shall be required for the
antigen or genetically engineered fruit or transgenicantigen or genetically engineered fruit or transgenic
seeds.seeds.
Transgenes may spread by pollen, sucking insects,Transgenes may spread by pollen, sucking insects,
transfer to soil microbes during plant wounding ortransfer to soil microbes during plant wounding or
breakdown of roots and may pollute surface andbreakdown of roots and may pollute surface and
ground water.ground water.
Ethical considerations usually restrict clinical trials fromEthical considerations usually restrict clinical trials from
directly assessing protection in humans.directly assessing protection in humans.

21. Current statusCurrent status
Measles:-
Mice fed with tobacco expressing MV-H (measles virus
Heamagglutinin from Edmonston strain) could attain antibody titers
five times the level considered protective for humans and they also
demonstrated secretory IgA in their faeces.
Carrot, banana and rice are the potential candidates.
Malaria:-
three antigens are currently being investigated for the
development of a plant- based malaria vaccine, merozoite surface
protein (MSP) 4 and MSP 5 from Plasmodium falciparum, and MSP 4/5
from P. yoelli. Wang et al have demonstrated that oral immunization
of mice with recombinant MSP4, MSP4/5 and MSP1, co-administered
with CTB as a mucosal adjuvant, induced antibody responses effective
against blood-stage parasite.

22. Hepatitis B:-
for hepatitis B, parenteral VLPs could invoke specific
antibodies in mice.
First human trails of a potato-based vaccine against hepatitis B
have reported encouraging results.
The amount of HBsAg needed for one dose could be achieved
in a single potato.
When cloned into CaMv, plasmid HBsAg subtype showed higher
expression in roots as compared to leaf tissue of the transgenic
potato.
Cholera:-
transgenic potato with CT-B gene of Vibrio cholerae was
Shown to be effective in mice.
Eating one potato a week for a mouth with periodic boosters was
said to provide immunity.

23. ETEC:-
Boyce Thompson Institute, USA accomplished the first published
successful human trial in 1997.
Eleven volunteers were fed raw transgenic potatoes expressing
LT-B.
Ten (91%) of these individuals developed neutralizing antibodies,
and six (55%)developed a mucosal response.
Norwalk virus:-
transgenic potato expressing norwalk virus antigen
showed seroconversion.
Nineteen out of 20 people fed with transgenic potato
expressing norwalk virus antigen showed seroconversion.
Attempts are underway to engineer bananas and
powdered tomatoes expressing norwalk virus.

24. AdvantagesAdvantages
 Do not require administration by injection.Do not require administration by injection.
 Possible production of vaccines with low costs.Possible production of vaccines with low costs.
 Do not require separation and purification of vaccinesDo not require separation and purification of vaccines
from plant materials.from plant materials.
 Necessary syringe & needles not required.Necessary syringe & needles not required.
 Economical in mass production and transportation.Economical in mass production and transportation.
 Heat stable, eliminating the need for refrigeration.Heat stable, eliminating the need for refrigeration.

25. DisadvantagesDisadvantages
Development of immunotolerance to vaccine peptideDevelopment of immunotolerance to vaccine peptide
or protein.or protein.
Consistency of dosage from fruit-to-fruit, plant-to-plantConsistency of dosage from fruit-to-fruit, plant-to-plant
and generation-to-generation is not similar.and generation-to-generation is not similar.
Stability of vaccine in fruit is not known.Stability of vaccine in fruit is not known.
Dosage of vaccines would be variable.Dosage of vaccines would be variable.
Selection of best plant is difficult.Selection of best plant is difficult.
Certain foods like potato are not eaten raw, andCertain foods like potato are not eaten raw, and
cooking the food might weakens the medicine presentcooking the food might weakens the medicine present
in it.in it.
Not convenient for infants.Not convenient for infants.

26. FutureFuture
☻ Edible vaccine holds a great potential.Edible vaccine holds a great potential.
☻ It reduces the cost of transportation and refrigeration.It reduces the cost of transportation and refrigeration.
☻ It neglect the needle and complicated way of vaccineIt neglect the needle and complicated way of vaccine
administration.administration.
☻ For many disease the research in going on in many countriesFor many disease the research in going on in many countries
funded by their government or industry.funded by their government or industry.
☻ Significant challenges are still to be overcome before vaccineSignificant challenges are still to be overcome before vaccine
crop can become a reality.crop can become a reality.
☻ There are some safety concerns which need to be overcome inThere are some safety concerns which need to be overcome in
near future.near future.
☻ In another concern, scientists need to be sure that vaccinesIn another concern, scientists need to be sure that vaccines
meant to enhance immune response do not backfire andmeant to enhance immune response do not backfire and
suppress immunity instead.suppress immunity instead.