Keywords
Chikungunya, Zika, Molecular farming, Vaccine
This article is included in the Emerging Diseases and Outbreaks gateway.
This article is included in the Neglected Tropical Diseases collection.
Chikungunya, Zika, Molecular farming, Vaccine
Plant expression systems have been used for the past 26 years for the production of human or animal proteins of biopharmaceutical interest. Antigens, antibodies, and enzymes have been produced, and some of them commercialized using several plant expression platforms1. While certain impediments remain such as community reluctance to accept transgenic products and the strict regulations for approval2,3, the FDA approval of ELELYSO® (alfa taliglucerase), a recombinant cerebrosidase for the treatment of Gaucher’s disease, has motivated plant-based biopharmaceutical protein production. These methods are all the more attractive because they are cost-effective, safe and scalable4.
After the arrival of Zika and chikungunya viruses to Latin American countries, they quickly became endemic diseases. They currently pose an acute and chronic burden for health systems and represent a diagnostic challenge in areas where those infections co-circulate with dengue and other febrile-illnesses5,6. Clinical diagnosis frequently is difficult given the similar clinical features with other viral infections such as dengue. However, the laboratory confirmation of Zika, dengue or chikungunya infection is important because each one has different implications for follow-up both in the short and long term. Diagnosis of acute, symptomatic infection is typically achieved through pathogen detection by virus isolation or qRT-PCR, Serology may be helpful later in the acute illness, but requires convalescent sampling in many cases and comes at a significant cost for healthcare systems. For this reason, confirmation is not recommended for the general population and has been restricted to specific cases. On the other hand, prevention of infection has been in the spotlight for policy makers. There are Zika and chikungunya vaccines under development, but current vaccine production is compromised by reduced capacity of vaccine manufacturers and substantial unmet needs for investment7.
Developing countries have been the most affected worldwide with these vector-borne diseases, and plant-based expression platforms have been proposed as a biotechnological tool to address the vaccine development challenge8. Plants could be used as bio-factories for the production of antigens, for both rapid diagnostic test design and vaccine production. Plant platforms operate at a small fraction of the cost (0.1% to 10%) of other expression systems like bacteria or mammalian cells9. Additionally, it has higher protein yield, lower contamination risk, lower storage cost, ability to assemble complex proteins with minor glycosylation differences, as well as high product quality, safety, and scalability9,10.
Although post-translational modifications have been a concerning issue, plant-derived vaccines can elicit protective immune responses11,12. Glycoengineering allows modification of protein glycosylation patterns in order to improve immunogenicity. Additionally, plant derived polysaccharides have been proposed as adjuvants and vehicles, further highlighting plants as a biofactory for antigen production10,11. Finally, viral antigens produced in plants have been used to target other arboviruses like the West Nile virus11.
In this context, a research agenda to assess the production of pharmaceutical proteins through plant molecular farming seems like a possible scenario to deal with current arboviruses epidemics. At first, candidate proteins should be defined. These proteins should be highly conserved and highly immunogenic. Importantly, antigen similarity between flaviviruses like dengue, yellow fever and Zika viruses has limited target antigen selection, for both vaccine and diagnostic test design for Zika, because of cross-reactivity and the risk of antibody dependent enhancement of infection13. Regarding chikungunya virus, the envelope glycoprotein E2 has been studied for both vaccine and rapid diagnostic test design, and even the use of plant produced virus like particles has been proposed as candidate for vaccine production14 (Table 1).
Target protein | Protein characteristics | Rapid diagnostic test | Vaccine development | Advantages | Disadvantages | Reference |
---|---|---|---|---|---|---|
E1 | Protein type: Surface glycoprotein; Function: Virus entry | X | X | Produce neutralizing antibodies response | Plant glycosilation pattern could modify immune response* | 15 |
E2 | Protein type: Surface glycoprotein; Function: Viral attachment | X | X | Produce neutralizing antibodies response | Plant glycosilation pattern could modify immune response* | 16,17 |
nsP2 | Cysteine protease with two separate domains with helicase and protease activity | X | Could be used as adjuvant for glycoproteins based vaccines | Reduced immune response induction. Greater genetic diversity | 18 | |
E3-E2-6K-E1 | Envelope poly-proteins | X | Produce neutralizing antibodies | Large cloning vector size, complex assembly and purification* | 19 | |
C-E3-E2-6K-E1 | Virus like particles | X | High immunogenicity. Produce neutralizing antibodies that have proved protection against wild-type virus | Large cloning vector size, complex assembly and purification* | 20–22 | |
Neutralizing Monoclonal Antibodies | IgG monoclonal antibodies against E1, E2 or C | X | Could be used in passive immunization | Difficult production in commercial scale, complex cloning vector assembly | 23,24 |
In addition, the plant expression system should be carefully selected. It is important to note that not all plants can be transformed, and phenolic compounds produced by plants and some of its secondary metabolites could make the purification of the desired protein difficult. Furthermore, the risk of contamination of other crops by the spread of transgenic pollen must be monitored according to the transformation method and plant species used for it. Transformation protocols in Nicotiana benthamiana, N. tabacum, and Solanum tuberosum have been used most commonly9.
Because of its scalability, efficiency and effectiveness, transformation using A. tumefaciens has been the preferred method for biopharmaceutical protein production. This method does not require special equipment like the gene gun, it allows a more precise and selective transgene insertion, and results in lower tissue damage and thus higher available biomass for protein production. Using this method both transient and stable transformation is obtained. In recent years, the method of agroinfiltration for transient plant transformation is preferred because of its potential to be systematized and provide an adequate yield of protein in the short-term4,9.
In conclusion, plant expression systems of heterologous proteins are a feasible strategy for vaccine development and rapid diagnostic kit design. Additionally, it could enable developing countries to address the challenge of current arboviruses epidemics, both in improving diagnostics as well as increasing primary prevention. The development of a molecular plant farming research agenda seems as a worthy solution to empower research in developing countries. It will permit every country to take advantage of its own natural resources in an individualized manner to deal with its own epidemiologic challenges.
JACO and JCSA conceived the idea. JACO, JCSA, LM and LGGL carried out the literature search. JACO and JCSA prepared the first draft. LM and LGGL contributed to its revision. All authors were involved in the revision of the final draft of the manuscript and have agreed to the final content.
This study was funded by Sistema General de Regalías de Colombia and Universidad Tecnológica de Pereira, assigned to Jaime A. Cardona-Ospina in the framework of the project "Development of scientific and technological skills in biotechnology applied to health and agro-industry sectors in Risaralda" (BPIN 2012000100050).
We thank Dr. Matthew Collins (Division of Infectious Diseases, University of North Carolina) for assisting us with language style.
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Competing Interests: No competing interests were disclosed.
Competing Interests: No competing interests were disclosed.
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