Discussion with Vitor Martins dos Santos Part 3

Discussion with Professor Vitor Martins dos Santos Part 3

This is the final part of my discussion with Prof Vitor Martins dos Santos.

Have a listen.

Discussion with Professor Vitor Martins dos Santos Part 2 – Laboratory Setup

The setup of a laboratory is not always seen as the most interesting of topics, but I found Vitor Martins do Santos’ description of the way he had different but complementary researchers working together in close proximity very revealing. Have a listen for yourself and see what you think.

Prof Vitor Martins dos Santos on how he runs his laboratory.

Discussion with Professor Vitor Martins dos Santos

Earlier this year I conducted a long and wide ranging interview with Professor Vitor Martins dos Santos of Wageningen University in the Netherlands. I am going to post extracts of that interview over the next few days. The first is relatively short and provides an insight into the relationship between Systems Biology and Synthetic Biology. You can hear it by clicking here.

Prof Vitor Martins dos Santos on Systems Biology and Synthetic Biology.

SYBHEL Conference Podcast

I have just uploaded a podcast recorded at the SYBHEL Conference in London in June 2012. It is in three parts and interviews 5 people who have been involved in producing the final recommendations from the SYBHEL Project. Have a listen!

Part One
Part Two
Part Three

Synthetic Biology and Coronary Heart Disease

Coronary heart disease, CHD, is one of the major global killers. It is a complex condition with multiple risk factors and presentations but the commonest cause of CHD related deaths is the rupture of fatty plaques in the arteries. Known as thin cap fibroatheromas, these are complex structures which have been the focus of a great deal of research. It is now known that there are eight key transcription factors involved in their development – biological molecules that control how genes are expressed. And these eight factors are known to act on around 2000 genes. This makes it fiendishly complex for researchers attempting to work out how the plaques form, and how they might be prevented from accumulating in the first place.

Now Frueh and colleagues from the Department of Bioengineering at Imperial College have published an approach combining systems and synthetic biology that they hope will help unpick exactly what is going on (Frueh, J., et al. Systems and synthetic biology of the vessel wall. FEBS Lett. (2012), http://dx.doi.org/10.1016/j.febslet.2012.04.031).
This is a complex paper featuring a multitude of different techniques. It is, though, of note for the way it goes through a number of different steps using different approaches to achieve its ends. The problem it tackles is fearsome but is also an interesting model for how to approach the many similarly complex problems that lie ahead.

The over-arching principle was to identify new pathways within the cells lining the blood vessels at the points of plaque formation. As blood is pumped through the arteries the vessels expand and contract, subjecting the cells lining the walls to significant mechanical stress. It is known that the sites of greatest stress are most prone to plaque formation so it is here that the researchers focussed their attention. They used a combination of high resolution imaging techniques and computer based analysis to identify the cells at these stress hot spots and then brought in state of the art genetic analysis. The final result was that they identified a number of signalling pathways that were not normally seen in other similar cells elsewhere in the body.

This paper is largely a demonstration of the technique which involves considerable cross discipline expertise and collaboration. The ultimate aim is to identify the genetics of plaque formation and to identify potential targets for treatments. This is the main point of intervention of synthetic biology for human health in this condition. It is still a long way off but this approach holds out the prospect of developing synthetic biology techniques to engineer blood vessels to be less susceptible or even immune to plaque formation.

Directed Evolution as a Tool for Synthetic Biology.

One of the stated aims of SynBio is to design new functionality into living systems for a whole host of reasons that include disease treatment and control, bioremediation, chemical synthesis and the production of biofuels. The basic principle has parallels in engineering, build up a complex system from known simpler elements and this has already borne fruit. However, there are two important unknowns that make this much harder than the equivalent engineering problem. The first is that simple metabolic circuits within the cell do not operate in isolation which can lead to unexpected results. The second is that not every pathway or metabolic process involved is fully understood.

This is where directed evolution comes into play. The principle is relatively straight forward and, as its name suggests, involves mimicking the process of evolution by natural selection but directing that evolution towards a specific outcome. In essence it is just a form of selective breeding, similar to the way that domestic animals have been bred to produce more milk or more meat or crops to produce greater yields. Organisms with the desired characteristics are bred with each other and the offspring with the improved properties selected and cross bred again. At each stage many offspring are rejected and only the best go on to breed further.

The directed evolution approach is to take a biological molecule, a protein or RNA for example, produce many mutated versions in the test tube and then transfer them into cells to grow. This produces a library of cells containing many variations on the original molecule which can then be screened for the desired effects. These could be a more effective drug or a gene that expresses higher levels of a desired protein. The best performers are then extracted, subjected to another round of mutation and screening and so on. The eventual outcome is a biological molecule, metabolic pathway or whole cell which exhibits enhanced desirable properties.

The crucial element here is that the production of a useful molecule can be done without needing to fully understand how it works.
This approach has already borne considerable fruit and is well reviewed in an article currently in press in the journal Methods R.E. Cobb et al., Methods (2012), http://dx.doi.org/10.1016/j.ymeth.2012.03.009. This gives examples of how directed evolution has been used to redesign or alter living systems from the protein to the cellular level. I will give a brief outline of some of these below.

Lovastatin is a common statin prescribed to millions to reduce cholesterol levels. An enzyme from Aspergillus terreus called LovD can convert an inactive precursor molecule into the active drug. However, outside it’s host cell the enzyme is relatively unstable and doesn’t work at maximum efficiency. Gao et al. (X. Gao, et. al., Chem. Biol. 16 (2009) 1064–1074.) applied directed evolution to this enzyme and after seven rounds of mutation and screening produced an version that showed an eleven fold increase in activity.

The lovastatin example is relatively straight forward and is a technique that has been used for some time. However, there are some considerably more complex procedures that have had some significant results.

These involve techniques that are aimed at metabolic pathways, networks or the whole cell. The techniques involved are complex but are based on the same principle, mutate, screen and select the offspring with the most desirable features. This approach has produced a variety of different engineered organisms. A technique called global transcription machinery engineering, or gTME, was used to produce a yeast strain that can more efficiently ferment the sugars xylose and xylose-glucose and has a greater tolerance to ethanol. The target of the mutations here were the proteins that control how genes are transcribed, the first step in producing the proteins they encode. It is a key control step in cellular metabolism and the aim is to alter the way in which genes are read to produce cells with desirable outcomes.

An alternative directed evolution approach called whole genome shuffling has been applied to a number of industrially important organisms. For example, a strain of Streptomyces was engineered to produce increased concentrations of the chemical (2S,3R) hydroxycitric acid (HCA). The significant point here is that the metabolic pathway that produces this chemical is not fully understood. This means that it cannot yet be tackled by most synthetic biology techniques which require the full details of the metabolic processes to be known. Whole genome shuffling has also been used to create a yeast strain that can grow at 55oC and tolerate 25% ethanol. As a keen amateur wine maker I know that most wine yeasts do not cope well above 30oC and struggle when the alcohol levels reach about 15%. This is a dramatic leap in operating conditions.

The flip side of these techniques is that they offer another way to understand cellular metabolism. All the mutant strains produced in this way have been analysed to work out why they behave in the way they do. This offers researchers greater insight into cellular processes and some surprises. For example, one of the yeast strains targeted by gTME had a key protein, sigma, that was full size while another had a significantly truncated one, but both had similar metabolic outcomes.

These approaches are largely still in the laboratory and have yet to make a significant impact on drug production and other processes important to human health. However, the principle of directed evolution fills in a crucial gap in the tools of synthetic biology. Many of the current approaches in synthetic biology require a complete understanding of the metabolic processes to be engineered. There are still many processes that are poorly understood and a complete description of even the simplest cell is some way off. Directed evolution allows synthetic biologists both to target these unknown processes and to understand them better.

David Sprinzak Interview Part 1

I’ve just uploaded this interview with Professor David Sprinzak. This is part one which is a brief overview of his research, part two will follow soon looking at the principles of circuits in synthetic biology.

Between Prevention and Treatment: How Synthetic Biology as a Theragnostic Technology Alters the Concept of Therapy and the Implications for Personal Responsibility

I have just uploaded the talk given by Robin Pierce on 6th Feb 2012 at the SYBHEL conference in Den Haag.

This covers the topic of theragnostics, a combination of therapy and diagnosis, and raises many different issues for the use of synthetic biology in human health. The talk is 20 minutes in total and I have uploaded it in two parts, part 1 is here and part 2 is here.

Dr Robin Pierce is Assistant Professor, Biotechnology and Society, Department of Biotechnology,
Delft University of Technology.

The possible use of synthetic biology in the realization of the capability to detect and intervene on the basis of biochemical markers indicative of pathology will signal a shift in the boundaries of therapy. In essence, as a theragnostic technology, synthetic biology could operate as an internal ?biophysician?, performing both diagnostic and therapeutic functions. However, this development comes with multiple complexities, not the least of which is for the conception of disease and illness. If an internal mechanism effects cure upon manifestation of emerging pathology, the ?disease? never actually materializes. The biochemical markers trigger cure, thus arresting the development to frank onset of disease.

While the development of synthetic chemical structures that are effective in preventing onset and/or recurrence may signal a major development in the promotion and maintenance of health and well-being, it fundamentally alters the concept of therapy, which traditionally has required the onset of disease before the administration of treatment. Moreover, this will also suggest implications for personal responsibility in health care. This paper will explore these potential shifts, their implications, and offer possible policy approaches for dealing with these changes.

Circuits in Synthetic Biology

One of the concepts that runs through synthetic biology is that of circuits. They are often compared to circuits in electronics but the analogy is not perfect and there are significant differences between the two.

To get a handle on the concept in biology I spoke to David Sprinzak, Assistant Professor at Tel Aviv University in the Faculty of Life Science. First, a little about his work. He studies differentiation at the cellular level, the process by which identical cells develop to perform different tasks in the adult organism.

This process happens wherever there are stem cells within an organism but it’s possibly easiest to consider an early embryo consisting of identical cells. For the embryo to develop these cells need to begin the process of differentiation, following different developmental paths to produce the many types of tissues found in the adult organism. At some crucial point adjacent, identical cells will need to head off in different directions. It is this step, the very first, that Professor Sprinzak is exploring.

Professor Sprinzak’s research is primarily investigative, working out what is going on. However, he does foresee significant potential for human health, particularly in the field of tissue engineering. The aim here is to take cells, probably from the patient, and use them to build replacement tissues and organs. A significant challenge is that organs are complex three dimensional structures composed of different types of cells. Stem cell technology provides a way of obtaining the basic cellular building blocks, but to build the organ they need to be directed to differentiate in the correct way. Having all the components of a car is only part of the problem, to build a functioning vehicle you need to put them in the right place. The work David Sprinzak and his group are doing is slowly unpicking the way cells do this naturally. This could be of great help to the tissue engineers of the future.

Audio of this conversation will be posted on this website in the near future if you want to hear more about what he is doing.

Back now to circuits. One of the functions a cell has to perform is to receive information from the outside world and respond to it in an appropriate fashion. The signal could be a signalling molecule like a hormone or a neurotransmitter which the cells receive. The response could be a decision to differentiate into a different type of tissue. In other words it has to do a bit of computation and the analogy with an electrical circuit does stand up reasonably well here.

A biological circuit, however, is made up at the level of genes and proteins and its function is produced by the way these elements interact.

An example of a simple circuit is this toggle switch, which needsjust two genes, and the two proteins produced when those genes are activated. Gene A produces a protein that turns on gene B, and gene B produces a protein that turns off gene A. If you were to monitor the levels of the proteins one would rise as the other falls and vice versa. A classic situation in which this might operate is a cell that is triggered to grow or divide in response to a certain concentration of a growth factor external to the cell. In that case, the cell needs to convert an analogue input (concentration of the growth factor) to a digital all or none switch. This is similar to an electrical switch where the switch will go from off to on only if you press it hard enough.

The circuit-bearing cell would have on its surface growth factor receptors, proteins embedded in its outer membrane to which the growth factor would bind. When they do so, the receptor will send some form of signal into the interior of the cell. This would, in turn, make its way to the nucleus. If the signal was of sufficient strength then it would activate a gene or set of genes that would trigger the cell to divide or grow or whatever. There is a classic example of a toggle switched engineered into E. coli here Timothy S. Gardner, Charles R. Cantor & James J. Collins Nature 403, 339–342. However, this is in a bacterium which, unlike a mammalian cell, has no nucleus.

This is clearly grossly simplified and I’ve glossed over the details of just what sort of signal is sent from the receptor into the nucleus.. Likewise I’ve ignored the particulars of how the cell determines the strength of the signal and the mechanism of how it activates the gene or genes it targets. These could all involve multiple steps including proteins, enzymes and further signalling molecules. The challenge for synthetic biology is to work out what the necessary components of the circuit are and re-engineer them in whatever way is desired without gumming up the system with unwanted by-products of these biological processes.

The electrical analogy, according to Professor Sprinzak, is useful but does not offer a complete explanation. This incompleteness is most noticeable in the potential for interaction between circuits. In a computer the individual circuits added together can be considered as totally separate from each other, (though there are potential unexpected interactions especially when very many circuits are crowded onto a single chip), but this is not necessarily the case for cellular circuits.

Biology has evolved to use a relatively small number of different molecules as chemical signals, but the same molecule can be part of more than one circuit. Also, the circuits are all operating inside a single cell with no physical barriers between discrete circuits. In an electrical chip, in contrast, pathways are separated by insulators. As a result in the biological cell as the concentration of a signalling molecule in one circuit rises, it can easily “leak” into another circuit. Other components may also be involved in other circuits. As a result circuits can not be considered totally in isolation and experiments often reveal unexpected levels of complexity.

This is just an introduction to the concept, if you want to read more about biological circuits in synthetic biology the following papers offer varying degrees of oversight of the subject: Zhang and Jiang Protein Cell 2010, 1(11): 974–978 and Nandagopal and Elowitz, Science p 1244-1248 Vol 333 2011. Also the audio of David Sprinzak discussing these ideas will be posted shortly on this website.

Developing health technologies from synbio: the ethics of experimental treatment

I have just uploaded an interview I conducted with Sarah Chan at the recent Sybhel meeting in Den Haag. A very interesting and thought provoking idea.

Sarah’s abstract is below and you can hear the interview here.

Sarah Chan: Developing health technologies from synbio: the ethics of experimental treatment
The prospect of new health technologies based on synthetic biology raises promising medical possibilities but also a range of ethical considerations. Apart from the issues involved in considering whether synbio health technologies can or should become part of mainstream medical treatment, the process of developing such therapies, from their origins in the laboratory through their progression as forms of experimental treatment to the point of clinical trials and beyond, itself entails particular ethical concerns. In this paper I consider some ways in which synbio therapies are likely to emerge and the ethical challenges these will present. I argue that developments in this new area of health technology will require us to rethink conventional attitudes towards clinical research, the roles of doctors/researchers and patients/participants with respect to research, and the relationship between science and society; and that a broader framework is required to address the plurality of stakeholder roles and interests involved in the development of synbio treatments.

What Synbio Means for our Definition of Health

I have just uploaded a short interview I conducted with Elselihn Kingma, Simon Rippon and Sune Holm at the recent Sybhel meeting in Den Haag. They all spoke on how synthetic biology is changing the concept of “health” and the potential implications of this change. It’s a short listen and a thought provoking introduction to the ideas involved.

To listen, click here.

The three contributors are:

Sune Holm
Sune is a postdoctoral research fellow in the Department of Media, Cognition and Communication at the University of Copenhagen. He did his BA and MA in philosophy at the University of Copenhagen and the University of St. Andrews. In 2006 he received his Ph.D. in philosophy from the University of St. Andrews on a dissertation entitled “The Persistence of Persons.” Since 2009 he has been working on the Ethics and Life project in association with the UNIK Synthetic Biology program launched at the University of Copenhagen. He focuses on questions concerning the moral status of synthetic life, the analogy between engineered machines and evolved organisms, and the ontology and functions of artifacts, organisms, and artifactual organisms.

Elselijn Kingma
Prof Dr Elselijn Kingma is a Research Fellow in the Centre for Humanities and Health & the Department of Philosophy at King‘s College London, and Socrates Professor in Philosophy & Technology in the Humanist Tradition at the Technical University of Eindhoven, the Netherlands. Previously she worked as a Post-Doctoral Research Fellow at the Department of Bioethics, National Institutes of Health, USA.
Elselijn‘s research interests include topics in the philosophy of medicine (concepts of health and disease; evidence based medicine), bioethics (risk, rights & consent, particularly surrounding birth), philosophy of biology (functions), and topics in philosophy of mind. Her PhD thesis entitled ?Health and Disease? was defended in 2008 at the University of Cambridge.

Simon Rippon
Dr. Simon Rippon is a Postdoctoral Research Fellow at the Oxford Uehiro Centre for Practical Ethics. His research interests lie mainly in the fields of bioethics, neuroethics and metaethics. He has recently been working on the nature and normative significance of the distinction between treatment and enhancement, the ethics of procuring organs for transplantation, and the nature of moral expertise. In his philosophy doctoral dissertation at Harvard, he argued that decisive epistemological objections to moral realism should motivate an alternative account which is constructivist, sentimentalist, and response-dependent. (Plain English translation: If we hadn’t, in some sense, made ethics up, then we couldn’t possibly have known what’s right and what isn’t. So ethics is something we have made, not something we have discovered.)


Pathogens and Synthetic Biology

On December 20 2011 the journal Science published an extraordinary editorial. They said that they had been approached by the US Government and asked not to publish the full details of a study on the H5N1 avian‘flu virus on the grounds that it could help anyone who wished to turn the virus into a biological weapon. The denouement came on Jan 20 2012 when the researcher temporarily halted their research over concerns that it could be used by terrorists. (http://www.nature.com/nature/journal/vaop/ncurrent/full/481443a.html).

This is a very rare occurrence, but it raises an important dilemma for anyone applying synthetic biology to any potentially pathogenic organisms. It’s a problem that has been identified for some time and is articulated well by Suk et al. in “Dual-Use Research and Technical Diffusion: Reconsidering the Bioterrorism Threat Spectrum” in PLoS Pathogens, p1-3 Vol 7(1), 2011. Simply put, some research may have a dual purpose in that it can be used to harm as well as to heal. The healing potential is the ability to prepare for possible ‘flu pandemics and maintaining academic freedom. The restriction of academic freedom raises the idea of censorship, either by the state of scientific bodies, and also the value and knowledge, it’s ownership and dissemination. Furthermore, restriction could mean that the benefits of research could be available only to an elite few. These ideas are expanded upon in Calladine, A. M. and R. t. Meulen (forthcoming). Synthetic Biology. Encyclopedia of Applied Ethics, Elsevier. The potential harm is that it might provide terrorists with the knowledge to create a potent bioweapon. The ethical and legal question at the heart of this is to what extent this research can and should be made available to other researchers and the general public.

This piece will look at recent research that, while not all are synthetic biology in practice, they are of direct significance to the field. All focus on pathogens, primarily bacteria. I want to be clear. None of this research has been linked to the best of my knowledge to the dual-use question. However, it is impossible to avoid it when considering research into pathogens.

A brief aside first about the definition of synthetic biology and it’s relationship to systems biology. Calladine and R. t. Meulen (see ref above) in an up and coming publication describe the challenge of defining synthetic biology. It is not straight forward but one point well made is that it aims “toward using a variety of approaches to design, engineer and build new biological systems.” With regard to systems biology, Smolke and Silver (Cell. 2011 March 18; 144(6): 855–859) discuss the relationship between that discipline and synthetic biology. They argue that there are significant synergies between the two disciplines but that these synergies will be the driving force that produces significant advances in biotechnology. Taking these two arguments together it is clear that research that helps identify targets for synthetic biology, while not necessarily synthetic biology themselves, are crucial to the discipline. Which is my justification for talking about this first study of Thiele et. al.

The bacterium Salmonella typhimurium is a human pathogen causing gastroenteritis, and is steadily becoming resistant to the antibiotics normally used to treat it. As such it poses a significant threat, particularly to the young, elderly or immune compromised. Understanding its metabolism would provide synthetic biologists with a host of targets to develop new approaches to killing the bacterium and hence treating infections. Thiele et. al. BMC Systems Biology 2011, 5:8 (http://www.biomedcentral.com/1752-0509/5/8) describe a large scale collaborative approach to producing a knowledge-base and mathematical model of S. typhimurium strain LT2. The paper has 27 authors illustrating the scale of the effort required, and argues that community based projects will be required in order to completely describe the metabolism of similar significant organisms. The research was in the field of systems biology, and was attempt to collate and understand the multiple metabolic pathways of S. typhimurium.

The overall result of the research was to produce a metabolic reconstruction (MR) of the S. typhimurium strain. This includes detailed descriptions of 1,270 genes, 2,200 internal reactions and 1,119 metabolites. These were converted into a mathematical model that represented the metabolism of living bacteria which in turn allowed a further, significant step. The researchers were able to use this model to predict a number of potential drug targets. Again, the use of mathematical modelling is becoming increasingly important in synthetic biology, and computer languages are being developed that can aid synthetic biology design. A future blog will look at this issue in greater detail.

A particularly interesting outcome of this research was the conclusion that combination treatments, using two or more drugs, would be needed to ensure that the bacteria could not easily evolve resistance. Most current antibiotic treatments are single agents, with the notable exception of those for tuberculosis. The researchers argue that while there are many hurdles to overcome with combination therapies, their work provides good evidence that they are necessary to fight antibiotic resistance in bacterial pathogens.

One of the concerns raised about any form of genetic based medicine, including synthetic biology, is how to ensure that the introduced genes do not produce more problems than they solve. How to minimise any unintended consequences. A classic example is the inadvertent production of a lethal strain of the mousepox virus, created by adding a gene that caused the production of large amounts of Interluekin 4. The researchers did not predict this outcome and rapidly communicated their research to the scientific community, warning of the potential for harm in this study. (http://www.newscientist.com/article/dn311-killer-mousepox-virus-raises-bioterror-fears.html)

The targeted and controlled approach of synthetic biology provides an approach that may be able to prevent this type of reaction happening. A study published in 2010 by Bagh et. al. (Biotechnology and Bioengineering, Vol. 108, No. 3 ,p 645-654, 2011) illustrates one approach. The research focussed on the infection of bacterial cells by the bacteriophage ? which infects and kills E. coli by cell lysis, bursting the cells.

The researchers aim was to use synthetic biology to produce an intracellular disease spotting mechanism with the following characteristics. It should lie dormant when no disease is detected; detect the onset of a lethal disease; respond in a way that halts or mitigates the progress of the disease and have a mechanism that can be deactivated by an external mechanism when desired.

When bacteriophage ? infects an E. coli cell it’s DNA becomes integrated into the bacterial genome. From there it follows one of two paths. The first is that it lies dormant, being copied along with the rest of the bacterium’s genes as it divides and replicates, a state called lysogeny. The other route, called lysis, is where the virus replicates rapidly, produces multiple copies of itself, breaks down the cell wall killing the bacterium and releasing fresh virus into the environment. Crucially, there is a gene based switch that controls the change from lysogeny to lysis. This switch can be flicked by exposure to UV light and certain chemicals and was the focus of the synthetic biologists.

The lysogeny-lysis switch is provided by the interaction of a number of genes and proteins with a key role for a protein call CI. High levels of CI keep the bacteriophage ? dormant, low levels switch it to the active, lethal, mode. One simple approach would be to simply ensure that the bacteria always produced an excess of CI, but this would force the cell to spend energy producing excess protein that would not be needed for most of the time. A more sophisticated approach was developed that was modelled on a classical engineering principle. In essence, when there is ample CI in the cell then the synthetic biology construct detected this and turned off. However, when the levels of CI started to drop the synthetic biology circuit registered this and produced additional, lysis suppressing, CI protein. The researchers produced a number of different variants of their controller which demonstrated the key principles they laid out, that of lying dormant when not needed, detecting the onset of disease, halting it and responding to an external off switch.

This, as the authors of the study make clear, does not have any medical applications. However, it is a very interesting proof of principle, establishing some of the criteria that will be developed for future, medical benefits.

It’s also worth noting the research of Saeidi et. al. Molecular Systems Biology 7; Article number 521, which I have discussed in a previous post on this blog http://sybhel.org/?p=730. The technology is interesting, basically E. coli were modified so that they could detect and then destroy pathogenic Pseudomonas aeruginosa. If you want to know more there’s a good comment piece by Andrew Jermy in Nature Reviews Microbiology ( Nature Reviews Microbiology AOP, published online 12 September 2011). I was struck by it’s title “Licensing Bacteria to Kill” accompanied by a James Bond type graphic. While prediction is of course impossible, I would not be surprised if this area of research was the focus of some probing ethical discussions.

These, and related studies, lay the grounds for synthetic biology to tackle pathogenic organisms in a very different way. The potential for an impact on human health is very large and perhaps the most obvious immediate need is to combat the rise in antibiotic resistant bacteria. The flip side of this is that it could also allow synthetic biologists to produce more potent pathogens, perhaps giving them scope to become bio-weapons. The request by the US government to Science to withhold crucial methodology of the ‘flu research shows that they are aware of the issue. It is very likely that this will form part of the future discussions of the uses and abuses of synthetic biology.

Cancer Seek and Destroy

The many different forms of cancer still pose one of the biggest challenges to human health. The fact that cancer cells are, crudely speaking, normal cells replicating out of control poses a number of significant problems. These include detecting cancer cells in vivo; identifying potential drug targets and destroying cancer cells while leaving healthy tissue untouched. Cancer cells have effectively the same genetic makeup as normal cells, but the pattern of gene expression, and hence metabolism, is often different. One of the major potentials of synthetic biology is that it provides the tools with which to spot these metabolic differences, just what’s needed to identify cancer cells in living tissues. This approach is being applied in many different ways to cancer treatments and this entry will look at a couple of recent, diverse examples which act as interesting proofs of principle.

The first is the use of a synthetic biology approach to find out what drives metastasis of some breast cancer cells. Preventing metastasis, the movement of cancer cells from the original tumour to elsewhere in the body, is a key goal of cancer treatment. Around 90% of deaths from breast cancer are caused by metastases The approach taken by Yagi et. al. Science Signaling 4 (191), ra60, 2011, has shown how the cancer cells co-opt a set of chemicals used to control the movement of white blood cells around the body.

White blood cells are part of the immune system and move freely around the body, congregating at wound sites and other regions as and when they are needed. The signals that draw them to these sites are chemicals called chemokines, and it’s been known for some time that metastatic cancer cells respond to them. What was unclear was exactly what mechanism was used by the cancer cells to detect and follow these chemokine trails. To find out the researchers constructed a synthetic biology system that would produce a protein important in this process, called G?13, that could be turned on with an artificial signal. This meant that the researchers could control where and when the G?13 was present. The result was they found that the way the cancer cells responded to the chemokines was different to the way white blood cells did. They then went on to experiments in mice in which they inhibited this signalling pathway and found that this significantly reduced the rate at which breast cancer cells metastasised.

The significance of this research is that a synthetic biology approach allowed a very specific, detailed exploration of a complex metabolic pathway involved in cancer spread and mortality.

This study of metastatic breast cancer by Yagi et al demonstrated how synthetic biology can be used to identify potential therapeutic targets, but once identified the challenge is to exploit them. A paper by Zhen Xie, et al. in Science 333, 1307 (2011), demonstrates how synthetic biology can be used to identify the unique metabolic fingerprint of cancer cells, and a possible mechanism for killing them.

The researchers developed a synthetic biology circuit that can be inserted into a cell, detect elements of that cell’s metabolism and trigger it to self destruct if it matches a pre-determined profile. The key to this study are small RNA molecules, called endogenous microRNA’s (miRNA), that are produced as part of the cells metabolism. Different cells produce different variants and amounts of these miRNA’s and this profile can act as a form of cellular fingerprint.

The important feature of all DNA or RNA molecules is that their properties are determined by the order of the 4 different types of bases arranged along their length. That order is the sequence of the molecule. This makes each one unique and allows them to be identified in a mixture of other similar DNA or RNA molecules. The synthetic biology approach described here utilised the unique sequence of some miRNA’s produced by cultured HeLa cancer cells. They identified six that were either expressed at high or low levels by HeLa cells in a pattern of expression very different from normal, non-cancerous, cells. Then a series of genes were strung together in an artificial sequence in such a way that they would only react in the presence of the correct amounts of all six HeLa miRNA’s Basically, they had a genetic tool that could, when inserted into a cell, detect whether it was a HeLa cell or not.

Experiments determined that this DNA detective could indeed discriminate between HeLa and other types of cells. Furthermore, the researchers coupled it to the production of a protein that would prompt apoptosis, programmed cell death. So they had a tool that would, if inserted into a human cell in culture, determine whether it was a HeLa cell and if so tell it to commit suicide.

The potential for this as a cancer treatment is clear. However, this approach would have to be modified significantly before it could be used in this way. Firstly, it involves the insertion of DNA into the cells and this is both currently impossible in humans and raises major risk and ethical questions. However, as a proof of principle it demonstrates that the different metabolism of cancer cells could be targeted to kill the malignancies.

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