Green Biologics: Harnessing the Power of Plants to Produce Pharmaceuticals
Abstract
:1. Introduction
2. Types of Plant-Derived Biologics
2.1. Antibodies and Antibody Fragments
2.2. Vaccines and VLPs
2.3. Therapeutic Enzymes
2.4. Receptor Modulators
2.5. Small Molecules
2.6. Bioactive Proteins from Plants
3. Strengths, Weaknesses, Opportunities, and Threats (SWOT) Analysis of Biologics
4. Manufacturing of Plant-Based Production Systems
5. Challenges in Producing and Using Plant-Derived Biologics
5.1. Complex Protein Expression
5.2. Post-Translational Modifications
5.3. Post-Translational Gene Silencing (PTGS)
5.4. Proteolytic Degradation of Recombinant Proteins
5.5. Downstream Processing
6. Novel Strategies for Improving Production and Efficacy
6.1. Protein Engineering
6.2. Synthetic Biology Approaches
6.3. Metabolic Engineering
6.4. Advancing Plant Molecular Biology
6.5. Field Scale-Up and Commercialization
7. Applications of Plant-Derived Recombinant Proteins
7.1. Therapeutic Applications of Plant-Derived Recombinant Proteins for Human and Animal Health
7.2. Biologics Produced in Space
7.3. Industrial and Agricultural Applications of Plant-Derived Recombinant Proteins
8. Regulatory Considerations for Plant-Derived Biologics
8.1. Current Regulatory Landscape
- Source of Plant Expression System: Detailed scrutiny of plant species, genetic modifications, and expression vectors used in production.
- Characterization of Biologics: Thorough evaluation of structural attributes, purity, potency, and stability.
- Variability in Expression: Implementation of measures to ensure consistent production and quality despite inherent variability within plants.
- Co-expression of Plant-Specific Proteins/Allergens: Rigorous characterization is required to identify and mitigate potential risks associated with unintended co-expression.
8.2. Future Regulatory Requirements
9. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Biologic/Application | Plant/Expression System | Status/Research Findings/Reference/Website Links | Company |
---|---|---|---|
Therapeutics | |||
β-Glucocerebrosidase Gaucher’s disease, enzyme replacement | Daucus carota, carrot cell culture, stable gene expression ProCellEx® | Elelyso™ has been approved by the USDA and EMA as the first biologic on the market [18]. https://protalix.com/ (accessed 10 October 2023) | Protalix BioTherapeutics Inc., Karmiel, Israel/Pfizer |
Pegunigalsidase alfa Fabry disease | ProCellEx® Stable gene expression | ELFABRIOTM has been approved for the USA and EU [19]. https://protalix.com/ (accessed on 10 October 2023) | Protalix BioTherapeutics Inc., Israel |
Clinical-grade plant material of the virus-trapping proteins CTB-ACE2 SARS-CoV-2 | Lactuca sativa, Lettuce stable chloroplast expression | Clinical trial phase I/II of chewing gum containing proteins CTB-ACE2 (angiotensin-converting enzyme 2 fused to the non-toxic cholera toxin subunit B) [20]. | University of Pennsylvania |
Uricase (PRX 115) Severe Gout | ProCellEx® Stable gene expression | Clinical trial phase I https://protalix.com/( accessed on 10 October 2023) | Protalix BioTherapeutics Inc., Carmiel, Israel |
Insulin Diabetes | Helianthus annuus, (sunflower)/stable gene expression | Clinical trial phase I/II [21] http://www.sembiosys.com/ (accessed on 9 October 2023) | SemBioSys Genetics Inc., Calgary, Alberta, Canada; in 2012, SemBioSys terminated its operation |
Lactoferrin VEN120 Inflammatory bowel disease VEN BETA E. coli gastroenteritis | Oryza sativa, Transgenic rice seeds, cell culture media | Products on the market https://ventria.com/ (accessed on 9 October 2023) | Ventria Bioscience, Junction City, KS, USA |
Allergens bioparticles | N. benthamiana Transient expression | Production of high-quality (“natural-like”) allergens and other sophisticated proteins for pharmaceutical purposes https://angany.com/ (accessed on 9 October 2023) | Angani Inc., Québec, QC, Canada |
Vaccines | |||
Influenza VLPs vaccine Seasonal flue | N. benthamiana Transient expression | Clinical trial phase III [22] | Medicago Inc., Quebec City, QC, Canada; the company closed in 2023. |
Covifenz® SARS-CoV-2 vaccine | N. benthamiana Transient expression | Authorized for use by Canada Health after successfully completed clinical trials [23]. | Medicago Inc., Quebec City, QC, Canada |
KBP-201 with CpG oligonucleotides SARS-CoV-2 vaccine | N. benthamiana Transient expression | Clinical trial phase 1/2 https://kbio.com/ (accessed on 10 October 2023) | Kentucky BioProcessing, Owensboro, KY, USA |
IBIO-201 IBIO-202 SARS-CoV-2 vaccine | N. benthamiana Transient expression | Preclinical studies [24] | iBio Inc., Bryan, TX, USA |
Baiya SARS-CoV-2 Vax 1 SARS-CoV-2 vaccine | N. benthamiana Transient expression | Clinical trial phase 1 ongoing [25] https://baiyaphytopharm.com/ (accessed on 10 October 2023) | Baiya Phytopharm Co., Ltd., Bangkok, Thailand |
SARS-CoV-2 RBD vaccine | Chlamydomonas reinhardtii, algae | SARS-CoV-2 RBD was evaluated as an oral immunogen in mice. The test immunogen was stable in freeze-dried algae biomass and able to induce mucosal responses [26]. | - |
HERBAVAC™ CSF Green Marker Classical swine fever virus (CSFV) | N. benthamiana Transient expression | Registered by the World Organization for Animal Health (WOAH) http://bioapp.co.kr/eng/ (accessed on 10 October 2023) | BioApplications Inc., Pohang, Republic of Korea |
Newcastle disease vaccine (in poultry) | N. tabacum sell suspension culture, stable gene expression | The first vaccine produced in plants approved by the US Food and Drug Administration for application in poultry [27]. | Dow AgroSciences LLC, Benton County, IN, USA |
Oral delivery platform of vaccines | Chlamydomonas reinhardtii, TransAlge technology | Edible vaccine https://www.transalgae.com/( accessed on 9 October 2023) | TransAlgae, Rehovot, Israel |
Others | |||
Growth factors, cytokines, lectins anti-CD25 antibody | N. benthamiana Transient expression | Research reagents on the market https://ibioinc.com// (accessed on 9 October 2023) | IBio Inc., Bryan, TX, USA |
Antibodies, viral proteins, VLPs | N. benthamiana Transient expression | Research reagents on the market https://capebiologix.com (accessed on 9 October 2023) | Cape Bio Pharma, South Africa, Cape Town, Africa |
Antibodies, enzymes, cytokines VLPs, viral proteins | N. benthamiana Transient expression | Research reagents on the market https://www.leafexpressionsystems.com/ (accessed on 10 October 2023) | Leaf Expression Systems, Norwich, UK |
Diagnostic antibodies, cytokines, growth factors | N. benthamiana Transient expression | Research reagents on the market https://www.agrenvec.es/ (accessed on 9 October 2023) | Agrenvec, Madrid, Spain |
Plant virus-like particles, Alternanthera Mosaic Virus | N. benthamiana Transient expression | Research reagents on the market https://www.diamante.tech/ (accessed on 9 October 2023) | Diamante Società Benefit, Verona, Italy |
Growth factors | Hordeum vulgare/Barley grains, stable gene expression | Cosmetics [28] https://www.orfgenetics.com/ (accessed on 10 October 2023) | ORF Genetics, Kópavogur, Iceland |
Enzymes | Zea mays, Corn, stable gene expression | Industry https://www.infiniteenzymes.com/ (accessed on 10 October 2023) https://www.greenlab.com/#in-production (accessed on 10 October 2023) | Infiniteenzyme Inc., Jonesboro, AR, USA Greenlab, Inc., Jutland, Denmark |
Therapeutic antibodies | |||
Anti-Human rAntibody (BLX-301) Non-Hodgkin’s lymphoma | Lemna minor, Duckweed, LEX system Stable gene expression | Phase II; The product and duckweed production system has been abandoned. | Biolex Inc., Pittsboro, NC, USA; the company declared bankruptcy in 2012 [29] |
ZMapp™ Anti-Ebola monoclonal antibodies | N. benthamiana Transient expression | In randomized, controlled trial, although the estimated effect of ZMapp appeared to be beneficial, the result did not meet the prespecified statistical threshold for efficacy [30]. https://mappbio.com (accessed on 10 October 2023) | Mapp Biotherapeutics, Inc., San Diego, CA, USA |
P2G12 HIV-neutralizing human monoclonal antibody 2G12 | Nicotiana tabacum cv. Petit Havana cv. SR1, stable gene expression | Phase I clinical trial [31] | Pharma-Planta consortium, Fraunhofer IME, Schmallenberg, Germany |
Anti-Spike antibody (mAb B38, H4) SARS-CoV-2 | N. benthamiana Transient expression | Both mAb B38 and H4 demonstrated specific binding to receptor binding domain (RBD) of SARS-CoV-2 and exhibited efficient virus neutralization activity in vitro [32].https://baiyaphytopharm.com/ (accessed on 10 October 2023) | Baiya Phytopahrn, Bangkok, Thailand |
Plant-made monoclonal antibody against ricin exposure | vivoXPRESS® plant-based manufacturing system | www.antoxacorp.com (accessed on 10 October 2023) www.swiftpharma.eu (accessed on 10 October 2023) | AntoXa Corporation, Toronto, Ontario, Canada and SwiftPharma, Belgium |
Strengths | Weaknesses |
Low cost: Plants can be grown at a relatively low cost compared to other expression systems. Profit can be made if the production of recombinant protein is high, the downstream processing is efficient, and there is an opportunity to scale up the production for a short period of time. | Time consuming: The production process of plant-derived biologics can be time-consuming. |
Scalability: Plant-based biologic production can be easily scaled up to meet demand. | Variable yields: The yields of plant-derived biologics can be highly variable. |
Complex molecules: Plants can produce complex biological molecules with post-translational modifications. | Regulatory considerations: Plant-derived biologics are subject to regulatory scrutiny and require approval from regulatory agencies. |
Safety: Plant-derived biologics are considered safe for human consumption and do not pose a risk of contamination. | Protein degradation: Proteases present in plants can degrade proteins during production. |
Opportunities | Threats |
Alternative to traditional expression systems: Plant-derived biologics have clinically improved profiles. | Intellectual property: Intellectual property rights can be a barrier to development and commercialization. |
Unmet medical needs: Plant-derived biologics can address unmet medical needs, such as low-cost vaccines for developing countries. | Competition: The field of plant-derived biologics is highly competitive. |
Diversification: Using plant-derived biologics diversifies biological production sources. | Public perception: The use of genetically modified plants may face skepticism. |
Method | Description | Advantages | Disadvantages |
---|---|---|---|
Stable nuclear transformation [151,152] | Stable integration of the gene of interest into the plant genome, enabling long-term and consistent protein production. Agrobacterium or biolistics-mediated delivery of transgenes. | Potential for large-scale production. Suitable for biologics with high demand or requiring complex PTMs. The use of edible plant species for oral delivery and cereals for long storage at ambient temperature. | The time-consuming process of plant transformation and regeneration. Relatively lower protein yield. Potential position effect and gene silencing. Regulatory considerations and public concerns regarding GMOs. |
Stable chloroplast transformation [153,154,155] | Each plant cell has 10,000 copies of the chloroplast genomes, which can stably integrate the gene of interest using a biolistic method of delivery. | The recombinant protein can be expressed at very high levels, up to 45% of the TP; there is no reported gene silencing; toxic proteins can be expressed successfully; more than one gene can be expressed, facilitating the production of complex proteins; and no gene flow. | A time-consuming process with low transformation frequencies, the formation of inclusion bodies, and challenges during the purification of recombinant proteins. Regulatory considerations. |
Viral Vectors [156] | Utilization of viral vectors, such as TMV or CPMV, to enhance protein expression levels by leveraging the viral replication machinery within plants. | Increased protein yields compared to non-viral expression systems. Compatibility with both transient and stable expression approaches. | Risk of viral contamination and potential biosafety concerns. Additional steps are required for viral vector construction and handling. Potential for adverse effects on plant growth. |
Transient Expression [157,158,159] | Rapid production of target proteins by introducing the gene of interest into plants using binary vector-based plasmids and agroinfiltration or viral vectors. | Quick and high-yield protein production. Suitable for rapid response scenarios. Flexibility and versatility in terms of the biological molecules that can be produced. | The transient nature of expression requires repeated plant agroinfiltration for continuous production. |
Plant cell cultures [160] | Production of recombinant proteins in plant cell suspension cultures. | Potential for easy scale-up for manufacturing under aseptic conditions using classical fermentation technology. Low risk of contamination. The same regulatory requirements as mammalian cell production systems. | Slower growth and lower yields compared to microbes and mammalian cells; overall cost is medium. Plant cell cultures are characterized by heterogenicity. |
Hairy roots Rhizobium rhizogenes [161] | Rhizosecretion of recombinant proteins in the hydroponic medium. | Secretion of the proteins into medium, facilitated purification, and improved product homogeneity. | Protein degradation, high proteolytic activity, and GMO regulatory considerations. |
Gene Editing [162] | Precise modification of plant genomes using gene editing technologies, such as CRISPR/Cas9, to optimize protein production and reduce proteolytic degradation. | Targeted modification of specific genes or regulatory elements to enhance protein expression. Potential for multiplex gene editing to improve multiple traits simultaneously. | Technical complexity and optimization required for gene editing experiments. Potential for off-target effects and unintended genomic modifications. Regulatory considerations for GMOs. |
Glycoengineering [163] | Elimination of unwanted glycan modifications and expression of glycosylation enzymes to provide the required specific glycans. | Production of recombinant glycoproteins with human-type glycans that resemble natural glycosylation. Eliminate unwanted glycan modifications. | Some plant species do not tolerate the engineering of glycan processing pathway, N-glycan heterogenicity, or GMO safety risks. |
Downstream Processing [164,165] | Implementation of purification strategies to effectively remove plant-specific contaminants, ensuring stability and quality of the final product. | Improved purity and removal of unwanted plant-specific contaminants. Optimization of downstream processing for specific biological molecules. | Additional processing steps, costs, and requirements. Need for customized purification methods for different biological molecules. |
Formulation and Delivery [166,167] | Development of innovative formulation and delivery methods to improve stability, bioavailability, and targeted delivery of plant-derived biologics. | Improved stability during storage and transportation. Enhanced bioavailability and efficacy in the target tissues or cells. Targeted delivery to specific organs or cellular compartments. | Additional costs associated with formulation and delivery systems. Potential challenges in achieving targeted delivery to specific sites. |
Regulatory Considerations for Plant-Derived Biologics |
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1. Source of plant expression system: Detailed information on plant species, genetic modifications, and expression vectors used in production. |
2. Characterization of biologics: Thorough evaluation of structure, purity, potency, and stability. |
3. Variability in expression: Measures to ensure consistent production and quality of biologics despite inherent variability within plants. |
4. Co-expression of plant-specific proteins/allergens: Identification and mitigation of potential risks associated with unintended co-expression. |
5. Environmental impact: Risk assessment to evaluate the potential environmental effects of cultivation and production processes. |
6. Case-by-case evaluation: Tailoring regulatory requirements based on the specific characteristics of each plant-derived biologic. |
7. Global harmonization: Collaboration between regulatory authorities and industry stakeholders to establish international guidelines and standards. |
8. Good Manufacturing Practice: Compliance with GMP guidelines to ensure consistent quality and safety during manufacturing processes. |
9. Preclinical and clinical data: Submission of comprehensive preclinical and clinical data to establish safety and efficacy profiles of plant-derived biologics. |
10. Post-marketing surveillance: Monitoring and reporting of adverse events and safety data following the commercialization of plant-derived biologics. |
11. Intellectual property rights: Consideration of intellectual property protection for novel plant-derived biologics and their manufacturing processes. |
12. Labeling and product information: Clear and accurate labeling to provide information on indications, dosage, administration, and potential risks associated with the use of plant-derived biologics. |
13. Risk management plan: Development of a risk management plan to identify and address potential risks throughout the lifecycle of plant-derived biologics. |
14. Regulatory updates and advancements: Staying informed about evolving regulations, guidelines, and advancements in the field of plant-derived biologics. |
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Share and Cite
Zahmanova, G.; Aljabali, A.A.A.; Takova, K.; Minkov, G.; Tambuwala, M.M.; Minkov, I.; Lomonossoff, G.P. Green Biologics: Harnessing the Power of Plants to Produce Pharmaceuticals. Int. J. Mol. Sci. 2023, 24, 17575. https://doi.org/10.3390/ijms242417575
Zahmanova G, Aljabali AAA, Takova K, Minkov G, Tambuwala MM, Minkov I, Lomonossoff GP. Green Biologics: Harnessing the Power of Plants to Produce Pharmaceuticals. International Journal of Molecular Sciences. 2023; 24(24):17575. https://doi.org/10.3390/ijms242417575
Chicago/Turabian StyleZahmanova, Gergana, Alaa A. A. Aljabali, Katerina Takova, George Minkov, Murtaza M. Tambuwala, Ivan Minkov, and George P. Lomonossoff. 2023. "Green Biologics: Harnessing the Power of Plants to Produce Pharmaceuticals" International Journal of Molecular Sciences 24, no. 24: 17575. https://doi.org/10.3390/ijms242417575