Genetic Containment Greenhouse Design

1. International Consultation on Plant-based Vaccines Flinn Foundation November 6-7, 2002 Genetic Containment Greenhouse Design Dwayne Kirk Project Manager, Boyce Thompson Institute for Plant Research Adjunct Appointment, Plant Biology, Arizona State University Email: Dwayne.Kirk @ asu.edu

2. Plants as Protein Producers Most efficient producers of protein known to mankind Inexpensive inputs: - airflow - water - sunlight - minerals - right combination of additional genes Cost per gram protein Soymeal $0.15 Corn seed $0.60 Casein hydrolysate $2.00 Yeast hydrolysate $4.00

3. Regulatory Requirement for Containment “Using bioengineered pharmaceutical plants to produce regulated products for use in animals or humans raises a number of environmental concerns that you should address, including confinement measures that may be needed to control the spread of the bioengineered pharmaceutical plants and to keep them from entering the food or feed supply.” FDA/USDA Guidance for Industry – Drugs, Biologics, and Medical Devices Derived from Bioengineered Plants for Use in Humans and Animals (Sept. 2002) “Bioengineered pharmaceutical plants that are grown exclusively in an enclosed building (e.g. greenhouse) generally will be considered to be confined during the growing period if there are control measures in place to eliminate the spread of pollen or seeds outside of the facility.”

4. Essential R&D parameters Antigen Accumulation Palatability Seed Bank Production Antigen yield per acre Gene Expression Geographical Flexibility Potency and Antigen loss Antigen Stability Shelf Life Cost Processing Immunogenic Presentation Available Technology Additive Formulations

5. Advantages and Disadvantages of Physical Containment • Disadvantages: • Cost of production 20-100 fold higher • Scale of production is confined without facility expansion • Advantages: • Maximize yield by optimizing growing conditions • Minimize risk of crop loss from natural disaster • Maximize statistical strength of experimental data • Minimize exposure to insects, animals or plant disease • Minimize bioburden • Avoid genetic drift to environment • Provide security of materials • Satisfy public concern on “safety” during initial trials • Able to be duplicated in time or location

6. Alternative means of pilot production Production facility    bioreactor    refined product

7. Dedicated Biopharmaceutical Facility

8. Practical Design for Containment Concern Solution Pollen drift to environment - Screened barriers at all inlets and outlets (batch contamination) - Caulk sealant on all structural joints - Active integrated pest management process - Limited human traffic into growth cells Concern Solution Plant Destiny - Standard operating procedures - Devitalization (autoclave) of all transgenic matter - Labeling and documentation practices Concern Solution General Security - Limited access, security personnel - Structural building integrity • Oversight by ASU Institutional Biosafety Committee • Inspection and approval from USDA / AZ Dept of Ag

9. Dedicated Biopharmaceutical Facility

10. Dedicated Biopharmaceutical Facility

11. Production under existing capabilities • 3600 sq feet of “manufacturing space” • either hydroponic or potted production will return equivalent yield • projected fruit harvest up to 8,880 kilograms (4035 lbs) • projected antigen production of 88.8 grams (assuming 10ug/g FW) • reduce to 555 kilograms powdered weight • 6g per dose (oral dose of 1mg) • 88,800 doses • fully loaded annual • operating cost •  $1.20 per dose • processing capabilities • storage capabilities • on-site analyticals and QC

12. Production under expanded capabilities • 1 hectare of “manufacturing space” • 5.6 million doses • projected cost under greenhouse production – 15c per dose • projected cost under shadehouse production – 9c per dose • projected cost under field production – 3c per dose

13. Simulating non-contained production

14. Dedicated Biopharmaceutical Facility

15. The Potential of Plants