A pair of researchers at Université Laval, Quebec claim that more effort should be made to develop plant-based vaccines. Hugues Fausther-Bovendo and Gary Kobinger have published a Perspective piece in the journal Science espousing the benefits of plant-based vaccines and suggesting how they might be made.
As the world continues to grapple with the SARS-CoV-2 virus, researchers around the world continue to look for new ways to vaccinate people against similar infections as it appears likely that this will not be the last pandemic. As part of that effort, some in the field have begun to look at alternative vaccine types. In their paper, Fausther-Bovendo and Kobinger suggest that developing plant-based vaccines might be a very good approach.
Vaccines are typically produced in bacterial or eukaryotic systems, and they have proven to be very effective. However, they have high production costs. Plant-based vaccines, the authors suggest, would be far cheaper to produceand could have several other benefits as well.
One is that plant-based vaccines would be far less resource intensive. Instead of bioreactors, vaccines could be grown in fields like crops. Another benefit comes from the very nature of plants – they cannot be infected with the types of human pathogens that lead to the need for vaccines.
Also, prior research has shown that plant-based vaccines tend to produce a stronger immune response than those made in other ways.
And plant-based vaccines have higher yields than other methods. And finally, in some cases, plant-based vaccines could be administered directly as a food product, with no extracting or processing needed.
The authors note that making plant-based vaccines is not unheard of; there is one that is currently being produced and used to treat Gaucher disease. Also, just before the pandemic struck, a plant-based vaccine for influenza made its way through Phase III clinical trials, with promising results. And right now, one team is working on a plant-based vaccine for COVID-19.
They suggest governmental regulating bodies around the world need to become more familiar with the benefits of plant-based vaccines so that guidelines can be written to promote such an approach if plant-based vaccines are ever to become a new standard.
Plant Molecular Pharming to Combat COVID-19
Plants are being used for the production of recombinant vaccines and drugs for more than 30 years and the whole process is described under term ‘molecular farming’ [99,100]. Secondary metabolites have significant biological and ecological functions in plants; particularly advantageous is their role in chemical defense because of their antioxidative and antimicrobial activities. Thus, molecular farming is used for the large-scale production of valuable secondary metabolites.
In addition, metabolic engineering tools can be used to overwhelm the bioactive-compounds availability limitations from medicinal plants and to improve the productivity beneficial from both bioprocessing and molecular farming  The synthesis of desirable recombinant proteins (pharmaceuticals and industrial proteins) using whole plants or in vitro cultured plant tissues/cells in large-scale bioreactors is termed molecular farming.
The advantages of plant-based reactors have been described in a review of molecular farming by Mohammadinejad et al.  as follows: (i) lower cost in maintenance; (ii) lower risks of contamination from animal pathogens; (iii) competence to implement modifications in eukaryotic post-translational machinery function; and (iv) being amenable to the large-scale manufacturing process.
Vaccines generated in plants have been shown to elicit a robust immune response in humans and animals (Figure 3). Plants have a great ability to act as a bioreactor system that supports many important biological processes including virus-like particles (VLPs) and vaccines. Transformation of plants with foreign genes leads to protein drugs, vaccines, and antibodies against different human pathogens hence plants make it easy to deal with safe, inexpensive, and provide trouble-free storage of protein vaccines and drugs .
Many research studies and clinical trials have shown that plant-made vaccines are safe and efficacious [103,104]. Examples of plant-made vaccines and therapeutics produced by molecular pharming include vaccines to combat cholera, Dengue fever virus and Hepatitis B virus, monoclonal antibodies to HIV and Ebola virus, and a therapeutic agent to provide glucocerebrosidase and help Gaucher Disease patients [104,105].
Several plant pharming companies and research labs have taken up the challenge to combat COVID-19. At the same time, there is a dramatic shortage of COVID-19 tests that could be alleviated by producing diagnostic agents in plants . A few examples of vaccines, diagnostics for test kits and antiviral therapeutics are presented in the following section.
Medicago, a biopharmaceutical company based in Canada, has successfully developed a virus-like particle (VLP) of the coronavirus 20 days after obtaining the SARS-CoV-2 genetic sequence. Instead of using egg-based methods to develop vaccines, this technology inserts a genetic sequence encoding the spike protein of COVID-19 into Agrobacterium, a common soil bacterium that is taken up by plants .
The resulting plants that are developed produce a virus like particle that is composed of a plant lipid membrane and COVID-19 spike protein. Medicago is using the plant Nicotiana benthamiana, a close relative of the tobacco plant, to produce VLPs of the SARS-CoV2 virus (COVID-19: Medicago’s Development Programs).
The VLPs are similar in size and shape to actual coronavirus but are lacking in nucleic acid and are, thus, noninfectious. Medicago has successfully completed its Phase 1 clinical trials and is currently working on Phase 2 clinical trials . Previously, Medicago has made VLPs composed of influenza virus haemagglutinin, and have demonstrated their safety and efficacy in animal models as well as in human clinical trials . The cost of producing a plant-made vaccine based on VLPs is a small fraction compared to its conventional counterpart .
In Canada, the University of Western Ontario and Suncor are developing diagnostic test kits for COVID-19 using algae as a production factory to make the viral spike proteins . Algae has long been considered a potential platform for generating pharmaceutical proteins as well as industrial proteins, such as cellulases . Algae is a superior biofactory alternative because it is easy to grow and can be readily modified to produce the viral proteins.
British American Tobacco, through its biotech subsidiary in the US, Kentucky BioProcessing (KBP), is developing a potential vaccine for COVID-19 and is currently undergoing pre-clinical testing . Experts at KBP cloned a part of the genetic sequence of SARS-CoV-2, which they used to develop a potential antigen that was inserted into Nicotiana benthamiana plants for production.
The vaccine has elicited a positive immune response by pre-clinical testing and will be onto Phase 1 human clinical trials soon . BAT could manufacture as much as 1–3 million doses of COVID-19 vaccine per week (they made 10 million vaccines of flu in a month as well as an Ebola vaccine using the same plant-based approach) .
South African company Cape Bio Pharms (CBP) is also responding to the SARS-CoV-2 pandemic through the production of reagents in plants, which could be used for diagnostic kits . CBP is producing SARS-CoV-2 Spike S1 reagents consisting of various regions of the glycoprotein attached to various fusion proteins. The company, based in Cape Town South Africa, is also collaborating with antibody manufacturers to produce antibodies against these proteins .
Another example of a plant molecular pharmed solution to COVID-19 is taking place in the department of nanoengineering at the University of California, San Diego. Researchers in Nicole Steinmetz’ lab have been using Cowpea mosaic virus like particles, with B- and T-cell epitopes from the S protein of SARS-CoV-2 displayed on their icosahedral surfaces . The recombinant virus harboring these COVID-19 epitopes can be applied in the form of an implanted microneedle technology incorporating VLP vaccines to skin and will elicit an immune response to SARS-CoV-2 .
The Steinmetz research group have recently developed positive control probes, composed of Cowpea mosaic virus-like particles, to be used as COVID-19 diagnostics and improve the accuracy of COVID-19 tests. The researchers hope that these positive controls, which are stable at room temperature for prolonged periods of time and cheap to generate, could be useful in resource-poor settings .
Another collaboration between two research groups in Toronto, Canada, has brought about a novel way to fight COVID-19 using a synthetic peptide that binds to the viral deubiquitinase (DUB) and is carried by a plant virus. The work initially began by examining the role of the virus protease, located in ORF 1a of the coronavirus genome responsible for the related MERS virus.
This protease contains a deubiquitinase activity as a means of protecting the virus from the innate immune system of the cell. Ubiquitin is a protein found in eukaryotic cells that play an important role in the regulation of proteins. It labels unwanted proteins (poorly folded proteins, viral proteins) to be degraded by the proteasome into shorter fragments or amino acids that can be recycled for cellular metabolism. Some viruses, such as coronavirus, express deubiquitinases (DUBs) to prevent destruction by the cell.
A synthetic peptide of approximately 80 amino acids and known as a ubiquitin variant (UbV) was created by phage display library design and shown to bind tightly to MERS DUB at its ubiquitin binding site, thus blocking its deubiquitinase activity as well as its proteolytic activity (Figure 4). This synthetic UbV was shown to block MERS virus infection in a human cell line, using a lentivirus vector for cell entry .
An analog to this UbV, which selectively binds to the DUB of SARS-CoV2, has recently been engineered for use in the current pandemic . Both MERS and COVID-19 UbVs have been fused to the N-terminus of the coat protein of a plant virus expression vector known as Papaya mosaic potexvirus (PaMV) (unpublished results). The UbV:CP fusion protein can assemble into virus like particles. PaMV has previously been shown to enter human cells via vimentin, a cytoskeletal protein.
The virus nanoparticle, loaded with COVID-19 UbV, can enter cells and block virus infection. Potexvirus nanoparticles have been shown to successfully enter the epithelial cells of lungs when introduced in the form of an aerosol spray. It is possible that these VLPs can be loaded into an inhaler to treat the lungs of infected and uninfected patients.
The ubiquitin variant is also being produced in a plant geminivirus vector, to be purified as an antiviral for COVID-19 patients (Manuscript in preparation). Geminiviruses, such as Bean yellow dwarf virus, have been engineered to produce large amounts of pharmaceutical proteins from plants in relatively short periods of time . A novel synthetic antibody to COVID-19 that was engineered from a phage display library is also currently being examined using the geminivirus vector system .
The COVID-19 pandemic is a challenge for us all. As a result of the current COVID-19 pandemic, therapies such as Remdesivir, convalescent plasma (CP), and Senna leaf extracts are the best available against COVID-19, until we have vaccine candidates in hand which have successfully undergone laboratory experiments, animal trials, and all phases (1–3) of clinical trials.
Possible targets including the spike, nucleocapsid, membrane, envelope, viral RNA polymerase, and 3-chymotrypsin-like protease (3CLpro), which cleaves the virus polyprotein at 11 distinct sites to generate various non-structural proteins that are important for viral replication, are all being used to develop potential vaccines and antiviral drugs. Virus like particles (VLPs) of SARS-CoV-2 may act as promising vaccines because they have the potential to activate the human immune response in a fashion similar to the original virus.
There is need to explore plant-based systems to check whether VLPs with retained structure and in sufficient quantity can be generated in these systems . On the one hand, this can include the further refinement of herbal extracts, particularly ones that had been used in the past to successfully inhibit SARS-CoV, as they may also function to block SARS- CoV-2.
The use of attenuated viruses and viral vectors in humans as vaccines may pose certain health risks involving the possibility of mutation (in the case of attenuated viruses) and recombination (in the case of viral vectors). The development of monoclonal antibodies against SARS-CoV-2 may also not be a long-term solution due to potential adverse reactions. Thus VLPs of SARS-CoV-2 generated by a plant expression system may act as a viable vaccine for the future.
More information: Hugues Fausther-Bovendo et al, Plant-made vaccines and therapeutics, Science (2021). DOI: 10.1126/science.abf5375