Medicago has developed a proprietary plant-based manufacturing platform that produces vaccines made of influenza antigen scaffolds
(i.e., viruslike particles) that are currently in clinical trials in Canada and the United States. This platform produces
vaccine doses within a month of the disclosure of hemagglutinin sequences from emerging strains. It is now being automated
and scaled up in North Carolina to bring its surge capacity to more than 10 million doses/month. The platform will be housed
in a facility that will be built within 12 months for less than $35 million.
The recent swine H1N1 influenza pandemic revealed the limitations of the current influenza-vaccine manufacturing technologies.
Although Wyman and colleagues had predicted in 2007 that egg-based manufacturing would be able to supply at least 60 million
vaccines doses within five months of the declaration of a pandemic, the actual vaccine output in the 2009–2010 porcine H1N1
(pH1N1) pandemic was much lower (1). In fact, only three million doses were available five months after the identification
of the causal strain. The first doses of split vaccine became available shortly thereafter, but at levels far below expectations.
Fortunately, the pH1N1 strain had a low mortality rate. Had the pH1N1 pandemic had a higher severity index, the global human
cost of the delays in egg-based vaccine production could have been catastrophic. Nevertheless, this outbreak has underscored
the need for a faster, simpler, and more flexible manufacturing technology to respond to an emerging threat and to protect
as many people as possible rapidly.
To meet the surge capacity required for the production of pandemic vaccines, recombinant technologies are being actively developed
with various expression hosts, such as bacteria, yeast, insect, or mammalian cells. Medicago developed a plant-based expression
system designed to provide great surge capacity and safety, at low cost. This article will describe how this new manufacturing
platform is combined with a nanoparticle antigen-presentation scaffold (i.e., viruslike particles or VLPs) that is synthesized
in plants, with an extraction approach that separates the nanoparticles from the host cell components early in the process,
and with a purification scheme that yields highly purified particles as drug substance.
The authors have demonstrated that this platform efficiently produces VLPs from the sole expression of recombinant influenza
hemagglutinin (HA), and that administration of these purified influenza VLPs was well tolerated and triggered an immune response
of great strength and breadth in animals and humans (2, 3). The speed at which influenza VLPs can be produced following the
identification of a new HA gene sequence was assessed in the context of the pH1N1 pandemic. The first doses of a plant-made
H1 VLP candidate vaccine against the pH1N1 strain were produced within three weeks of the disclosure of the new HA sequence
by the Centers for Disease Control and Prevention (see Figure 1).
Figure 1: H1 viruslike particle (VLP) experimental vaccine production timeline. (FIGURES ARE COURTESY OF THE AUTHORS)
The manufacturing platform based on transient expression, and the VLP-based antigen-presentation platform were developed in
the context of the pandemic threat. They now have been applied successfully to the development of a trivalent seasonal vaccine.
Vaccine products for these two indications have been produced under good manufacturing practice (GMP) conditions and currently
are in clinical trials in Canada and the United States.
THE TRANSIENT EXPRESSION SYSTEM
The transient expression system uses the capacity of a bacterium, Agrobacterium tumefaciens, to infect plant cells and transfer essential genetic information in the form of a mobile DNA fragment to the nucleus of the
infected cell. Because this mobile DNA fragment only remains viable for a few days in the nucleus and is not integrated in
the plant genome, the cells are only transiently transformed. Despite the transient nature of the transformation, this technology
results in extremely high expression of the gene of interest for days after cell infection, and enables synthesis of large
quantities of the targeted protein.
Medicago's transient expression system starts with the assembly of an expression cassette where the target gene is positioned
within transcription and translation components adapted for expression in plants. The expression cassette is inserted within
mobile DNA borders of a plasmid, which is transferred into the bacterial transfection vector A. tumefaciens. Transfection of the mobile DNA copy containing the expression cassette to Nicotiana benthamiana leaf cells is performed by soaking healthy plants upside down in a vacuum tank containing a liquid suspension of A. tumefaciens, the inoculum. As vacuum is applied, the air-filled cavities inside the leaves are emptied, thus leaving space for infiltration
of the bacterial inoculum upon release of the vacuum. As the inoculum is infiltrated, the A. tumefaciens establish contact with parenchyma cells, which constitute the vast majority of the leaf active cells. Because vacuum infiltration
is instantaneous and highly invasive, infection of leaf cells is also instantaneous, which helps to synchronize cell infection
and the resulting transfer of mobile DNA copies containing the expression cassette. Transient expression is extremely fast
and results in high accumulation of the recombinant protein within three to six days.