PLANTIBODIES FARMING PHARMACEUTICALS THROUGH THE USE OF PLANTS

PLANTIBODIES FARMING PHARMACEUTICALS THROUGH THE USE OF PLANTS Presentation by Ethan Novena

WHAT ARE PLANTIBODIES? • The term “plantibody” refers to an antibody that has been produced by genetically engineered plants. Plantibody = a plant derived antibody. • Antibodies are an essential part of the immune system of many organisms. They recognize viral antigens and other dangerous compounds and signal a response. [3] • Genetically modified plants have had antibody-coding genes from mammals inserted into their genome. Plants do not produce antibodies naturally. However, the function of antibodies produced by them through genetic modification has been shown to be the same as natural mammalian antibodies. [2]

MORE GENERAL INFO ON PLANTIBODIES • The endomembrane and secretory systems of these antibody factories allow the production of large quantities of viable proteins which can then be purified from the plant tissue for use in humans and other mammals. [2] • In summary: recombinant DNA technology allows plants to be used as factories for mammalian antibody production.

PLANT TRANSFORMATION / TRANSGENIC PLANT PRODUCTION The Transformation Process (simplified) [4] • Isolate: DNA from a mammal containing an antibody-coding gene. • Insert: the gene of interest into Ti-plasmid from Agrobacterium by cutting with restriction enzymes and joining with DNA ligase. • Introduce: the recombinant Ti plasmid to plant cells in culture. DNA segment of interest is transferred to the plant chromosome. • Regenerate: the transgenic plant in vitro or farms, greenhouses, etc. Figure source: http: //www. sciencedirect. com/science/article/pii/S 0974694313003289

ADVANTAGES OF USING PLANTS FOR ANTIBODY PRODUCTION • The cost of antibody production is significantly lower compared to using animals. • There is a reduced likelihood that pathogens are introduced when using plants for production. • Higher yields in shorter periods of time relative to using animals. Plants that produce lots of biomass (corn, tobacco) especially can produce and hold many of our target products. [2] • Infrastructure for large-scale plant growth and processing is already available. Farms, large greenhouses, and plant factories can be used for cultivating transgenic plants.

DISADVANTAGES OF USING PLANTS FOR ANTIBODY PRODUCTION • Possibility of plantibody strains contaminating food crops. We can get around this by using only plants that do not serve as a food source for people or livestock. • Possibility of transgenic plants to produce allergenic compounds and transfer their antigens into the end products, causing new allergic reactions in the recipient mammal(s).

APPLICATION: NEUTRALIZATION OF RICIN IN MICE • A study by Sully EK et al. In 2014 showed that neutralization of the infamous toxin known as ricin could be achieved in mice, and potentially even in humans. [1] • Extremely toxic CHO-binding protein derived from castor oil plant seeds. • Many assassination attempts and successes have been carried out with it, including an attempt on Barack Obama’s life in 2013. Also seen in “Breaking Bad”. Figure source: https: //www. newyorker. com/tech/elements/the-science-of-ricin

THE STUDY • Four mouse-human chimeric MAb’s against the toxin were generated from c. DNA in the mice B cell hybridomas, then amplified by PCR. These c. DNAs were then sequenced, and the important regions synthesized then fused to human Ig. G 1 and K binding sites to create mouse-human chimeric antibodies. [1] • These chimeric antibodies were then produced using transgenic Nicotiana benthamiana (RAMP platform), purified, and given to BALB/c mice 24 hr before being given 10 x LD-50 of ricin. [1] Figure source: https: //en. wikipedia. org/wiki/Nicotiana_benthamiana

RESULTS [1]

THE CLEAR WINNER: C-PB 10 • Further testing was done using c-PB 10 in which mice were given 25 ug/hr following systemic injection of 10 x LD-50 of ricin. 100% of mice that received the treatment 2 -3 hr after the ricin survived, and 90% survived after being treated 45 hr later. 6 hr later only 30% survival was observed. Therapeutic window in the mice was therefore determined to be 3 -4 hr. [1] • Similar testing was done using aerosolized ricin and a single 250 ug injection of c. PB 10 24 hr before, at the time of exposure, and 4 hr after. All mice survived but experienced some weight loss and body temp. decrease. [1] • Control mice always died within 48 hr during every challenge throughout the study.

CONCLUSION / CLOSING STATEMENTS • C-PB 10 most effective of the four c-MAb’s studied. Lowest IC-50, sufficiently protected mice passively against systemic and aerosolized ricin, and was able to rescue them when administered within 3 -4 hr of exposure. [1] • Large-scale production of plantibodies using Nicotiana benthamiana-based RAMP proved to be readily achievable. [1] • The data collected makes c-PB 10 an “ideal candidate for further development as a fully humanized immunoprotectant against ricin toxin”. [1] • Unfortunately, plantibody use in humans is not yet very well explored.

SOURCES • 1. Sully EK 1, Whaley KJ, Bohorova N, Bohorov O, Goodman C, Kim DH, Pauly MH, Velasco J, Hiatt E, Morton J, Swope K, Roy CJ, Zeitlin L, Mantis NJ. "Chimeric Plantibody Passively Protects Mice against Aerosolized Ricin Challenge. " Clin Vaccine Immunol, vol. 21, no. 5, 2014, pp. 777 -782, http: //cvi. asm. org/content/21/5/777. Accessed 3 Dec. 2017. • 2. Oluwayelu, Daniel O, and Adebowale I Adebiyi. “Plantibodies in Human and Animal Health: A Review. ” African Health Sciences 16. 2 (2016): 640– 645. PMC. Web. 3 Dec. 2017. • 3. Wikipedia contributors. "Plantibody. " Wikipedia, The Free Encyclopedia, 24 Mar. 2017. Web. 3 Dec. 2017. https: //en. wikipedia. org/wiki/Plantibody • 4. S Jhansi Rani, R. Usha. “Transgenic plants: Types, benefits, public concerns and future. ” Journal of Pharmacy Research, vol. 6, no. 8, 2013, pp. 879 -883, http: //www. sciencedirect. com/science/article/pii/S 0974694313003289. Accessed 6 Dec. 2017.