Charles J. Arntzen [email protected] Co-sponsored by the
ASU Biodesign Institute and the Arizona Farm Bureau Friday, March
27, 2015
Slide 2
After the 9/11/2001 Terrorist Attack on the World Trade Center
in New York, and the Pentagon, military funding of counterterrorism
increased in the U.S. In 2002 the Army funded a project for Mapp
and ASU. Title: Plant Production of Vaccines and Antibodies for
Protection Against Biowarfare Agents. Co-Principle Investigators:
C. Arntzen and L. Zeitlin Goal determine if viral expression
vectors could be used for production of either subunit vaccines or
monoclonal antibodies (mAbs) in tobacco. In our 2005 final report
we said: we have demonstrated that Ebola virus-specific humanized
monoclonal antibodies can be produced using plant
biotechnology.
Slide 3
The tiny company is Mapp BioPharmaceuticals a collaborator of
ASU for the last 12 years. Larry Zeitlin (President) and Kevin
Whaley (CEO) are adjunct ASU faculty via the Biodesign Institute.
August 4, 2014 Bloomberg News broke the story in the U.S. Two
American Missionaries received ZMapp A cocktail of three anti-ebola
antibodies. Ebola Drug Made From Tobacco Plant Saves U.S. Aid
Workers August 4, 2014 7:52pm MT
Slide 4
Zmapp worked quickly and dramatically.
Slide 5
No countermeasures currently exist for the prevention or
treatment of the severe sequelae of Filovirus (such as Ebola virus;
EBOV) infection. To overcome this limitation in our biodefense
preparedness, we have designed monoclonal antibodies (mAbs) which
could be used in humans as immunoprotectants for EBOV, starting
with a murine mAb (13F6) that recognizes the heavily glycosylated
mucin-like domain of the virion-attached glycoprotein (GP). Point
mutations were introduced into the variable region of the murine
mAb to remove predicted human T-cell epitopes, and the variable
regions joined to human constant regions to generate a mAb (h-13F6)
appropriate for development for human use. We have evaluated the
efficacy of three variants of h-13F6 carrying different
glycosylation patterns in a lethal mouse EBOV challenge model. The
pattern of glycosylation of the various mAbs was found to correlate
to level of protection, with aglycosylated h-13F6 providing the
least potent efficacy (ED50 = 33 g). A version with typical
heterogenous mammalian glycoforms (ED50 = 11 g) had similar potency
to the original murine mAb. However, h-13F6 carrying complex
N-glycosylation lacking core fucose exhibited superior potency
(ED50 = 3 g). Binding studies using Fc receptors revealed enhanced
binding of nonfucosylated h-13F6 to mouse and human FcRIII.
Together the results indicate the presence of Fc N-glycans enhances
the protective efficacy of h-13F6, and that mAbs manufactured with
uniform glycosylation and a higher potency glycoform offer promise
as biodefense therapeutics. The media was fascinated by ZMapp The
stories were: How is ZMapp made? Why tobacco? When will more ZMapp
be available? Who will pay for it? Who will decide who gets
it?
Slide 6
1992. Mason, H.D., M.-K. Lam and C. J. Arntzen. Expression of
hepatitis B surface antigen in transgenic plants. Proc. Natl. Acad.
Sci. USA 89:11745-749. Let me give you a personal history of the
work leading up to Zmapp. This starts over 20 years ago with our
work on Plant-Made Pharmaceuticals. Let me give you a personal
history of the work leading up to Zmapp. This starts over 20 years
ago with our work on Plant-Made Pharmaceuticals.
Slide 7
Pre-clinical studies with mice Volunteers eat diced, raw potato
expressing HBsAg Pre-clinical studies with mice Volunteers eat
diced, raw potato expressing HBsAg Milli-International units
(anti-HBsAg) Weeks: 0 1 2 3 4 5 6 7 HBsAg boosting trial Average
mean IgG serum antibody titres for all volunteers 1st Generation
Human Clinical Trials (The Edible Vaccine Concept; Vaccine in
Food)
Slide 8
Virus-based pharmaceutical production the key innovation after
2005 1.Convert RNA virus to DNA sequence, and re- engineer it to
express new gene(s). 2.Inject genetic construct into cellular
spaces within the leaf. 3.Allow the reconstructed virus to
replicate for 4-10 days. 4.Extract and purify the protein
drug.
Slide 9
Let me return to how I became involved in Ebola therapy..
Transgenic plants; minimal processing, oral delivery Viral
(transient) expression, GMP processing 2001 was a turning point for
research funding in the U.S. Terrorism had become real. After the
Sept. 11, 2011 attack on the World Trade Center, Bioterrorism
became a stronger military interest. The U. S. Military asked: Do
we have the technology to respond quickly? 2001 was a turning point
for research funding in the U.S. Terrorism had become real. After
the Sept. 11, 2011 attack on the World Trade Center, Bioterrorism
became a stronger military interest. The U. S. Military asked: Do
we have the technology to respond quickly?
Slide 10
December 20, 2011 PNAS 2011 108 (51) 20690-20694 PNAS 2011 108
(51) 20695-20700 Back to back papers published in 2011: Mapps
antibody protection ASUs vaccine protection Both with a mouse model
of disease Animal studies conducted with US AMRIID Back to back
papers published in 2011: Mapps antibody protection ASUs vaccine
protection Both with a mouse model of disease Animal studies
conducted with US AMRIID
Slide 11
By 2009, there were multiple research centers providing key
elements of the Ebola Antibody Project. Creation of a plant
molecular toolbox (for engineering plant viruses, controlling
protein production, etc.) Antibody engineering (creating mAbs with
human primary structure) Host plant (tobacco) engineering for human
glycoslyation Collaboratio n among groups was essential What was
missing? Scale-up manufacturing expertise. What was missing?
Scale-up manufacturing expertise.
Slide 12
Accelerating Critical Therapeutics Col. Alan McGill, MD.
Project Mngr. Nine member Scientific Advisory Board By 2009, the US
Military (DARPA) had recognized that plant-made pharmaceutical
production was limited by manufacturing capacity. They invested
over $80 Million in bricks and mortar in three U.S. companies to
build non-redundant capacity. By 2009, the US Military (DARPA) had
recognized that plant-made pharmaceutical production was limited by
manufacturing capacity. They invested over $80 Million in bricks
and mortar in three U.S. companies to build non-redundant capacity.
The Blue Angel Project
Slide 13
Genes encoding light and heavy chains Infect plants Redesigned
plant virus How to make monoclonal antibodies (mAbs) at industrial
scale Viral infection (mAb production) for several days; then
harvest. Purify mAbs
Slide 14
In 2012-present, KBP has worked to optimize manufacture of
anti-Ebola mAbs under cGMP. These were used in preclinical drug
characterization. 18 monkeys were infected with high doses of Ebola
virus. All survived when treated with Mapps anti-Ebola mAbs.
Treatment could be given up to 5 days after initial Ebola infection
Dr. Anthony Fauci, Director of NIH