Molecular Farming in Plants: The Long Road to the Market

Fischer, Rainer; Buyel, Johannes F.; Schillberg, Stefan; Twyman, Richard M.

doi:10.1007/978-3-662-43836-7_3

Cited by 10 publications

(6 citation statements)

References 66 publications

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“…The concept of molecular farming was first aired over 20 years ago, but despite its much trumpeted potential (see review by [ 1 ]), the first plant-derived products have only recently reached the market. Examples include recombinant human glucocerebrosidase synthesized in carrot suspension cells [ 2 ], and human growth hormone and cytokines synthesized in the barley grain [ 3 ].…”

Section: Introductionmentioning

confidence: 99%

Transgenic Production of an Anti HIV Antibody in the Barley Endosperm

et al. 2015

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Barley is an attractive vehicle for producing recombinant protein, since it is a readily transformable diploid crop species in which doubled haploids can be routinely generated. High amounts of protein are naturally accumulated in the grain, but optimal endosperm-specific promoters have yet to be perfected. Here, the oat GLOBULIN1 promoter was combined with the legumin B4 (LeB4) signal peptide and the endoplasmic reticulum (ER) retention signal (SE)KDEL. Transgenic barley grain accumulated up to 1.2 g/kg dry weight of recombinant protein (GFP), deposited in small roundish compartments assumed to be ER-derived protein bodies. The molecular farming potential of the system was tested by generating doubled haploid transgenic lines engineered to synthesize the anti-HIV-1 monoclonal antibody 2G12 with up to 160 μg recombinant protein per g grain. The recombinant protein was deposited at the periphery of protein bodies in the form of a mixture of various N-glycans (notably those lacking terminal N-acetylglucosamine residues), consistent with their vacuolar localization. Inspection of protein-A purified antibodies using surface plasmon resonance spectroscopy showed that their equilibrium and kinetic rate constants were comparable to those associated with recombinant 2G12 synthesized in Chinese hamster ovary cells.

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Section: Introductionmentioning

confidence: 99%

Transgenic Production of an Anti HIV Antibody in the Barley Endosperm

et al. 2015

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“…Most of the recombinant molecules currently available for medical use are synthesized in microbial and mammalian cell cultures (Fischer et al, 2014). Although cell lines allow the synthesis of eukaryotic proteins in a similar system to that of their origin, there is a high initial economic cost and great investments scale for protein production due to the cost-intensive bioreactor-based infrastructure (Paul et al, 2013).…”

Section: Introductionmentioning

confidence: 99%

Oral Immunization With a Plant HSP90-SAG1 Fusion Protein Produced in Tobacco Elicits Strong Immune Responses and Reduces Cyst Number and Clinical Signs of Toxoplasmosis in Mice

et al. 2021

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Plant 90kDa heat shock protein (HSP90) is a potent adjuvant that increases both humoral and cellular immune responses to diverse proteins and peptides. In this study, we explored whether Arabidopsis thaliana HSP90 (AtHsp81.2) can improve the immune effects of a Toxoplasma gondii surface antigen 1 (SAG1). We designed two constructs containing the sequence of mature antigen (SAG1m), from aa77 to aa322, and B- and T-cell antigenic epitope-containing SAG1HC, from aa221 to aa319 fused to AtHsp81.2 sequence. When comparing the transient expression in Nicotiana tabacum X-27-8 leaves, which overexpress the suppressor helper component protease HC-Pro-tobacco etch virus (TEV), to that in N. benthamiana leaves, co-agroinfiltrated with the suppressor p19, optimal conditions included 6-week-old N. benthamiana plants, 7-day time to harvest, Agrobacterium tumefaciens cultures with an OD600nm of 0.6 for binary vectors and LED lights. While AtHsp81.2-SAG1m fusion protein was undetectable by Western blot in any of the evaluated conditions, AtHsp81.2–SAG1HC was expressed as intact fusion protein, yielding up to 90μg/g of fresh weight. Besides, the AtHsp81.2–SAG1HC mRNA was strongly expressed compared to the endogenous Nicotiana tabacum elongation factor-alpha (NtEFα) gene, whereas the AtHsp81.2–SAG1m mRNA was almost undetectable. Finally, mice were orally immunized with AtHsp81.2–SAG1HC-infiltrated fresh leaves (plAtHsp81.2–SAG1HC group), recombinant AtHsp81.2–SAG1HC purified from infiltrated leaves (rAtHsp81.2–SAG1HC group), non-infiltrated fresh leaves (control group), or phosphate-buffered saline (PBS group). Serum samples from plAtHsp81.2–SAG1HC-immunized mice had significantly higher levels of IgGt, IgG2a, and IgG2b anti-SAG1HC antibodies than serum from rAtHsp81.2–SAG1HC, control, and PBS groups. The number of cysts per brain in the plAtHsp81.2–SAG1HC-immunized mice was significantly reduced, and the parasite load in brain tissue was also lower in this group compared with the remaining groups. In an immunoblot assay, plant-expressed AtHsp81.2-SAG1HC was shown to react with antibodies present in sera from T. gondii-infected people. Therefore, the plant expression of a T. gondii antigen fused to the non-pathogenic adjuvant and carrier plant HSP90 as formulations against T. gondii can improve the vaccine efficacy, and plant extract can be directly used for vaccination without the need to purify the protein, making this platform a suitable and powerful biotechnological system for immunogenic antigen expression against toxoplasmosis.

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“…The initial absence of regulatory guidance has also been overcome by providing a framework for the industry, especially for biopharmaceutical manufacturing, which should further mitigate uncertainty and thus eliminate the investment risks previously associated specifically with plant-based expression systems (Fischer et al, 2012, 2014).…”

Section: Introduction – Benefits and Drawbacks Of Plant Molecular Farmentioning

confidence: 99%

Plant Molecular Farming – Integration and Exploitation of Side Streams to Achieve Sustainable Biomanufacturing

Buyel

2019

Front. Plant Sci.

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114

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Plants have unique advantages over other systems such as mammalian cells for the production of valuable small molecules and proteins. The benefits cited most often include safety due to the absence of replicating human pathogens, simplicity because sterility is not required during production, scalability due to the potential for open-field cultivation with transgenic plants, and the speed of transient expression potentially providing gram quantities of product in less than 4 weeks. Initially there were also significant drawbacks, such as the need to clarify feed streams with a high particle burden and the large quantities of host cell proteins, but efficient clarification is now readily achieved. Several additional advantages have also emerged reflecting the fact that plants are essentially biodegradable, single-use bioreactors. This article will focus on the exploitation of this concept for the production of biopharmaceutical proteins, thus improving overall process economics. Specifically, we will discuss the single-use properties of plants, the sustainability of the production platform, and the commercial potential of different biomass side streams. We find that incorporating these side streams through rational process integration has the potential to more than double the revenue that can currently be achieved using plant-based production systems.

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Molecular Farming in Plants: The Long Road to the Market

Cited by 10 publications

References 66 publications

Transgenic Production of an Anti HIV Antibody in the Barley Endosperm

Transgenic Production of an Anti HIV Antibody in the Barley Endosperm

Oral Immunization With a Plant HSP90-SAG1 Fusion Protein Produced in Tobacco Elicits Strong Immune Responses and Reduces Cyst Number and Clinical Signs of Toxoplasmosis in Mice

Plant Molecular Farming – Integration and Exploitation of Side Streams to Achieve Sustainable Biomanufacturing

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