Key Points
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Artemisinin is an antimalarial drug precursor that is produced by the plant Artemisia annua. The supply and price of artemisinin have fluctuated substantially throughout the past decade, owing to inconsistencies in harvest.
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Artemisinin-based combination therapies (ACTs) are recommended by the WHO as the first-line treatment for uncomplicated malaria. The Semi-synthetic Artemisinin Project aimed to stabilize the supply and price of artemisinin for the development of artemisinin derivatives for use as part of ACTs.
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Both Escherichia coli and Saccharomyces cerevisiae were engineered using the tools of synthetic biology to produce 25 g per L and 40 g per L, respectively, of the artemisinin hydrocarbon precursor amorphadiene by fermentation.
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Owing to problems using E. coli, S. cerevisiae was used as the chassis for the industrial-scale production of 25 g per L artemisinic acid by fermentation, which was followed by a chemical conversion process to synthesize artemisinin.
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Semi-synthetic artemisinin is now produced at industrial scale and has been approved by the WHO for the preparation of approved pharmaceutical compounds for incorporation into ACTs.
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Lessons learned from the Semi-synthetic Artemisinin Project that are relevant to the development of other pharmaceutical products using metabolic engineering and synthetic biology are summarized.
Abstract
Recent developments in synthetic biology, combined with continued progress in systems biology and metabolic engineering, have enabled the engineering of microorganisms to produce heterologous molecules in a manner that was previously unfeasible. The successful synthesis and recent entry of semi-synthetic artemisinin into commercial production is the first demonstration of the potential of synthetic biology for the development and production of pharmaceutical agents. In this Review, we describe the metabolic engineering and synthetic biology approaches that were used to develop this important antimalarial drug precursor. This not only demonstrates the incredible potential of the available technologies but also illuminates how lessons learned from this work could be applied to the production of other pharmaceutical agents.
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Acknowledgements
The authors thank the Bill and Melinda Gates Foundation for their generous support of this project. They also thank their many colleagues at the University of California Berkeley, Amyris, Sanofi, National Research Council of Canada (NRC) and PATH Drug Solutions for their unstinting efforts that made this work a success.
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C.J.P. and J.D.K. hold stock options and shares in Amyris Inc. (California, USA).
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Glossary
- Chassis organism
-
The host microorganism that is used for the production of a desired product; it is typically subjected to genetic manipulation using the tools of synthetic biology.
- Tunable intergenic regions
-
(TIGRs). Stretches of DNA that are located between genes and that can be modified (for example, by the insertion of hairpins and cleavage sites) to enable each gene in an operon to be varied independently of all others in a random manner.
- Fed-batch fermentation process
-
A batch culture that is fed continuously with nutrient medium. It differs from continuous culture owing to the variation in culture volume from beginning to end.
- Cytochrome P450 enzyme
-
A member of a superfamily of monooxygenase enzymes that catalyse the oxidation of organic substrates. Plant microsomal cytochrome P450 enzymes require a reductase enzyme (CPR) to transfer electrons from NADPH and may also require cytochrome b5 to supply electrons.
- Trichomes
-
Hair-like or glandular structures on the surface of plants. Glandular trichomes are the major biosynthetic site of many natural plant products, including artemisinin.
- Hock fragmentation
-
A reaction of hydroperoxides connected to an unsaturated system, leading to cleavage of the C–C bond and the formation of two carbonyl compounds.
- Photochemical conversion
-
A chemical reaction that is initiated by the absorption of energy in the form of visible, ultraviolet or infrared light.
- Alkaloids
-
Nitrogen-containing natural products, many of which have pharmacological properties. Examples include cocaine, caffeine and the antimalarial drug quinine.
- Chemotypes
-
Plants that are morphologically similar or identical but that are distinguished by differences in the production of secondary metabolites.
- Yeast homologous recombination
-
A method to assemble DNA fragments with overlapping homology regions by transforming yeast and using its innate homologous recombination ability to correctly assemble the fragments.
- Ligase chain reaction
-
(LCR). A method to assemble DNA fragments using bridging oligonucleotides, a thermostable DNA polymerase and multiple denaturation–annealing–ligation temperature cycles.
- Circular polymerase extension cloning
-
(CPEC). A polymerase chain reaction (PCR)-based method for the assembly of multiple DNA fragments.
- Gibson isothermal assembly
-
A single-temperature enzymatic method of assembling multiple DNA fragments.
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Paddon, C., Keasling, J. Semi-synthetic artemisinin: a model for the use of synthetic biology in pharmaceutical development. Nat Rev Microbiol 12, 355–367 (2014). https://doi.org/10.1038/nrmicro3240
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DOI: https://doi.org/10.1038/nrmicro3240
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