Synthetic biology entered the fields of engineering and molecular biology as an innovative approach to study and manipulate biological systems rationally. Its novelty relied on the combination of genomics and systems biology and aimed to surpass the previous focus on reductionist biology (Van Regenmortel, 2004). Nowadays, it is widely known as the specialization that employs modern molecular biology techniques to engineer organismal behaviour (Cameron, Bashor and Collins, 2014). Currently, scientists are discussing the potential of translational medicine, defined in this case as the transition of basic synthetic research into clinical applications for the public benefit (Feidt et al., 2017), which had always been a laborious and troublesome task. Moreover, synthetic biology has direct applications in clinical research through biomedicine, with ongoing clinical research targeting cancer, infectious diseases and autoimmune disorders; and metabolic engineering discussed below using codeine as an example (Cameron, Bashor and Collins, 2014).
According to (Balas and Boren 2000), translation of 14% of scientific discoveries into clinical applications takes 17 years (Feidt et al., 2017). One of the actual models of translational research proposes an initial translation of laboratory findings into applied human research and a posterior translation into clinical trials, taking into consideration ethical and social values. It is important to stress the fact that it is a continuous bidirectional process since discoveries can affect basic research and clinical observations could open the doors to new research. Other models agree that basic research, practical and clinical research, and health studies are essential but differ on the number of phases required. Previous issues faced by early molecular laboratory techniques that rendered them not suitable for clinical use included: transient transfections leading to short term duration of treatment and non-site-specific integration into host genome causing off-target effects (Yvonne Y. Chen and Christina D. Smolke, 2011). With the advent of synthetic biology and the tools associated (i.e. CRISPR-Cas9, TALENs and ZF nucleases) efficient and precise integration of robust systems is now possible.
At present, synthetic biology is becoming an essential player in the biomedical field, with special relevance in the areas of cancer, infectious diseases and autoimmune disorders. Table 1 shows the common main constraint faced by the above areas of research: inadequate detection systems able to selectively trigger a response against cancerous cells, pathogens and autoantigens respectively. With the new technologies, 108, 109 and 110 have been able to tackle cancerous cells showing differential expression due to the hypoxic environment. Besides, Escherichia coli and bacteriophages have been engineered by 110 and 115 as vectors to detect a detrimental microbial presence and deliver specific treatments against biofilm formation. These new ways of targeting pathogens without the use of antibiotics will lead to advances towards antibiotic resistance prevention and improved antibiotic efficiency. On top of that, phage display libraries have been designed by 117 to detect autoantigens inducing immune responses in immunochallenged patients. Nonetheless, we must have in mind that most of these novel technologies are still under development and have not been implemented(Cheng and Lu, 2012). Local release of microbials in vulnerable patients is one of the main controversies encountered by these approaches.
Main challenges faced by Synthetic Biology
From the technical point of view, even though there is a high rate of advances under the label of synthetic biology, their translation into products or clinical applications is currently the lagging strand (Feidt et al., 2017). Biological systems are comprised of genetic networks that interact and respond to their surrounding signals. As such, assembling genes de novo into new entities could have unpredictable consequences, since biological systems can replicate and the interaction pathways are complex and non-linear (Fu, 2013).
Legal, ethical, environmental and economic disciplines have opened the debate to discuss safety and regulation procedures attaining Synthetic Biology and its applications in Translational Medicine. Despite being regarded as the birth of a new technology, some of the current projects are just employing leading-edge molecular biology and genetic engineering techniques and might use this denomination for marketing purposes. J. Robienski and J.Simon (de Miguel Beriain and Romeo Casabona, 2014) criticized the absence of a common scientific definition, and henceforth the impossibility of having a consistent international legal definition. On the environmental and ethical side, biosafety and biosecurity are major concerns (Gómez-Tatay and Hernández-Andreu, 2019). The first one entangles the risk of disrupting the ecosystem’s equilibrium and reducing biodiversity if synthetic organisms are released into the environment. In the second case, the European Commission fears the inclusion of synthetic biology in the production of chemical and biological weapons (de Miguel Beriain and Romeo Casabona, 2014). Moreover, biosynthesized pharmaceuticals could threat natural local production (Feidt et al., 2017). Researchers have additionally evaluated the danger of a few bigger companies dominating the field, which could hinder the further evolution of this technique due to limited access to publications and results. This concept is defined as economic fairness (de Miguel Beriain and Romeo Casabona, 2014) and addresses the importance of regulating intellectual property (IP) and other legal questions without forgetting global health and safety to have a sustainable market.
Applications in Metabolic Engineering: Codeine
Metabolic engineering: synthetic biology on metabolic engineering focuses on an increasing profit of current and future production. From improved photosynthesis and increased biofuel tolerance to the production of artificial amino acids and directed evolution, synbio plays an essential role in industrial bioprocesses such as microbial production of biofuels and chemical synthesis (Cameron, Bashor and Collins, 2014).
Codeine or 3-methylmorphine is a drug chemically derived from morphine, an opium derivative. Opium is a polymer emulsified from the seed capsules of Papaver somniferum (Van Hout et al., 2017). Together with its derivatives, they interact with the opioid receptors MOP (μ), KOP (κ), and DOP (δ) and induce analgesia through blockage of the pain transmission pathways. These drugs, especially morphine, have been used since 1799 to treat pain and diarrhoea (McDonald and Lambert, 2015). In the 1950s, different approaches were attempted to synthesize codeine, including the Grewe-type pathway (electrophilic cyclization of 1-benzylhezahydroisoquinoline) and codeine-precursors biomimetic approaches, but none of them produced high enough yields to be commercially competitive (REFERENCE mol li). Codeine is currently synthesized from poppy-derived morphine (Thorn, Klein and Altman, 2009), which results in 100,000 hectares of P. somniferum cultivated annually. Crop’s susceptibility to pests and climate fluctuations in addition to an increasing market demand rises the need for finding new marketable and profitable synthetic approaches. Galanie et al., 2015 considered codeine production using Saccharomyces cerevisiae due to its rapid growth (only a few days compared to annual plants) and the possibility of a more controlled and stable system (growth occurs inside vessels, not directly exposed to the outside climatic conditions). The research group engineered genetic modules coding for enzymes required to produce Benzylisoquinoline alkaloids (BIA), in other words: codeine precursors from cheap substrates such as carbon or nitrogen-based substrates. Despite being the most promising achievement up till now, yields decreased every time a new enzymatic step was added. As a result, a 100,000-fold increase in fermentation titer would be needed to use yeast for opioid production as a commercial alternative to the current poppy farming. One year later, an E.coli opiate production system was refined for thebaine production (Nakagawa et al., 2016). Higher bacterial enzymatic activity might be the cause of increased yields compared to previous yeast systems (Galanie et al., 2015), however these results are not profitable for commercial purposes. On a more positive note, Nakagawa et al., 2016 suggested that the four engineered E.coli strains were suitable hosts for future industrial opiate production from a simple carbon source, including codeine. The recent discovery of an enzyme involved in the yeast-based opiate synthesis might favour future codeine production in eukaryotic systems over prokaryotic (Chen et al., 2018). More recently, Zhang et al., 2019 moved away from synthetic biology and designed an alternative greener chemical pathway for codeine synthesis using 4,4′-di-tert-butylbiphenylide (LiDBB).
On the other hand, it is imperative to consider the social consequences of a new and more economic synthetic codeine production method. Measures must be developed to avoid illicit drug production (i.e. home-brewed) and that also consider a potential increase in drug misuse if access to codeine becomes easier.
Synthetic biology is a promising field in the fields of therapeutics, diagnostics and medical research focused on the understanding and design of biological systems (Cameron, Bashor and Collins, 2014). Synthetic eukaryotic and prokaryotic strains have been designed to tackle the increasing need for an alternative codeine production pathway (Galanie et al., 2015) (Nakagawa et al., 2016). In both cases, stepwise culture methods allowed for optimization of each individual step, reduction of undesirable side reactions and production of multiple compounds. Moreover, chemically synthesized alternatives are becoming increasingly (Zhang et al., 2019). Nevertheless, yields were not acceptable for commercial use in any of the cases above and further research is required to achieve a more economic, sustainable and profitable alternative.