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Antibody and Vaccine
production in Plants

Plant biopharming of monoclonal
antibodies
Virus Research 111 (2005) 93–100
 Kisung Ko, Hilary Koprowski


Plant-produced vaccines: promise and
reality

Edward P Rybicki
PLANT-DERIVED BIOMEDICALS FOR TREATMENT OF INFECTIOUS DISEASES
DISEASES
VACCINES
PLANT
Respiratory
Syncytial
Hepatitis B

Tobacco

Lettuce
HIV

Spinach
Rabies

Anthrax

Diphtheria

Spinach
Tobacco
Tobacco
Tomato
Tobacco
SARS

Smallpox

ANTIBODY
PLANT

Tobacco
Tobacco
Tomato
Tobacco
Tomato
PLANT-DERIVED BIOMEDICALS FOR TREATMENT OF CANCER
HUMAN DISEASES
VACCINES
ANTIBODY
Colorectal Cancer
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Epithelial Tumors
(EGF receptor)
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Antibodies
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Magic bullets:
Therapeutic – inhibit target involved in
disease progression
Diagnostic
Prevention of disease
Cause cytotoxic death of target cells
Act as carriers for radioisotopes, toxins and
drugs to site of disease
Anti-rabies virus mAb
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After exposure treated with Ab
Used to be made in horses
First mAb made in transgenic plants
4 genes – 2 H, 2 L
Transgenic plant for each one and
crossing plants
Later used single binary vector with
two promoters
Full-size monoclonal antibodies recently
produced in transgenic plants
Abs expressed in
transgenic plants
Constant
domains
Variable Light chain
Variable Heavy chain
Recombinant hinge region
Functional Abs
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Need to be properly folded and assembled
Need disulfide bond formation and
glycosylation
Down stream processing:
 Purification of Ab mostly with Protein A
or G
Glycosylation is different in plants
 β 1,2 Xylose and α 1,3 fucose
Retain in ER, only mannose is attached
–Shorter half-life of Ab
Glycosylation of Ab
Full-size Ab
Large single-chain Ab
Camelid heavy-chain Ab
Minibodies and Fab fragments don’t get glycosylated
Glycosylation in Golgi
Antibodies: a compelling success story

high specificity: in vitro and in vivo diagnostics
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low toxicity: therapeutic applications
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high drug approval rates (24 approved mAbs)

major products in biotechnology (~240 in clinical
trials)
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inherently stable human proteins
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injectable, topical and oral applications
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applicable for chronic conditions
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potential long-lasting benefits
Production Costs for Antibodies
Production costs
cost in $ /gram
hybridomas
1000
transgenic animals
transgenic plants
100
10
Source: Daniell et al. (2001) TIPS 6, 219-226
E. coli & yeast
Tr. animals and
animal cells
Transgenic
plants
Comparison of Mammalian and Plant-produced
Antibodies

peptide sequence: identical
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correct cleavage of Ig-derived signal peptides
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kinetics & affinity: identical
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stability in seeds > 30 months
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antibody types: plant system more versatile (sIgA)
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post-translational processing: different
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core glycan identical, terminal sugar different plus xylose & fucose
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antigenicity & clearance: apparently identical (shorter half-life)
Vaccines
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Hepatitis B virus surface antigen
HBsAg
Norwalk virus capsid
Vibrio cholerae enterotoxin subunit
Animal vaccines
– Mink enteritis virus - MEV
– Rabbit haemorrhagic disease virus RHDV
– Foot and mouth disease virus - FMDV
Oral vaccines

First thought edible vaccines good idea
– Banana a day
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Oral vaccination
– Might elicit antigenic tolerance
– ProdiGene - Got contamination of maize
and soybean harvest in 2002 with
transmissible gastroenteritis virus TGEV
Past and Future of PMPs
Plant produced Vaccines
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HPV L1 – forms VLPs – can protect
Measles virus haemagglutinin
HBsAg
Porcine TGEV
Tetanus toxin
Vacinia virus B5
Allergy vaccines
HPV
HIV gp41
Newcastle disease virus
Rotavirus VP7
Cancer Vaccines
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LSBC - Large Scale Biology Crop
Individually tailored single chain
variable region Ab fragments from
patients with Non-Hodgkin lymphomas
Colorectal cancer antigen
Cervical Cancer – HPV E7
Vaccines against
biothreat agents
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Biodefence
– Anthrax
– Plague
– Ricin
– Haemorrhagic fever
PMP Development - Highlights

Series of plant-derived vaccines from Arizona State University have
completed clinical trials
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Prodigene has trialled two plant-derived vaccines
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LSBC pipeline of cancer vaccines prior to insolvency
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Guardian Bioscience coccidiosis vaccine, CFIA phase II ongoing
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Fraunhofer CMB, rabies vaccine trialled in humans
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DowAgro Newcastle disease vaccine, approved Feb 2006
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Heberbiovac (Cuba) approved antibody for HepB vaccine
purification
Current Challenges in Molecular Farming

yield of recombinant proteins
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real quantitative comparison (TSP vs pg/cell/24h)
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protein stability (proteases)
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post-translational modification(s)
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backcrossing in elite lines
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extraction and downstream processing
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QA, QC and substantial equivalence
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clinical trials & regulatory approval
Regulatory Challenges for PMP
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loci of transgene insertion
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expression properties and levels, including PTM
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effects of the transgene on the expression of flanking
endogenous genes
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master line banking to ensure product consistency
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contamination with animal excreta, pesticides,
organic fertiliser
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procedures for detection and removal of weeds and pests
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cultivation variables
Roadmap Plants for the Future
Plant production platforms
Health food and
quality
- Amino acids
- Oil
- Starch
Efficient
agriculture
- Bt technology
- Herbicide
resistance
1997
2005
Plant protection
-
Viruses
Nematodes
Fungi
Insects
2015
-
Vitamines
Fatty acids
Enzymes
Bio-polymers
Pigments
Pharmaceutical products
Fibers
Stress resistance
- Cold
- Drought
- Salinization
2025