Cellular Alchemy: the science of reprogramming cells
This essay was written by Ben Martynoga and was first published in the 2013 Mill Hill Essays.
Alchemy is the art that separates what is useful from what is not by transforming it into its ultimate matter and essence.
Philippus Aureolus Paracelsus (1493-1541)
From heart attacks and strokes to Alzheimer’s and diabetes, diseases which result in the irretrievable death of cells are common, life- shattering and, in most cases, still uncurable. An enduring dream of modern medicine is to be able to provide new cells, to order, which can be used to repair damaged organs and stop disease progression.
In 2012 two scientists, John Gurdon and Shinya Yamanaka, were honoured with a Nobel Prize in celebration of work that has provided the first steps toward making this dream a reality. What these pioneers have achieved bears some resemblance to alchemy. Whilst alchemists of yore strove to transform base metals into gold, Gurdon and Yamanaka found ways to transform specialized, mature cells into cellular gold: stem cells, with the potential to develop into any bodily organ, from the aorta to the zygomatic bone.
These acts of cellular alchemy have given rise to the new science of reprogramming. In recent years reprogramming has caught the imagination of scientists, doctors and policy makers who are enthusiastically channeling huge amounts of funding and research time into this area. Whilst reprogramming has huge potential, there are significant technical and conceptual hurdles to clear before we can be sure it will be of practical use in the clinic and not just of academic interest in the lab.
What is so special about cellular reprogramming?
Transplanting cells and organs to replace those lost in accidents and disease is not a new idea. From 17th Century attempts at blood transfusion, to the first successful full face transplant in 2010, to promising recent clinical trials for the treatment of motor neuron disease, doctors have been saving and transforming lives with transplants for many years.
However, whilst organ, cell and stem cell transplants are powerful techniques, they share some common problems. Foremost are the difficulty of obtaining the correct cells and tissues when and where they are needed; the risk of rejection by the immune system; and the unpleasant side effects of longterm treatment with the immunosuppressive drugs that are needed to reduce this risk.
Reprogramming has the potential to side-step all of these problems. A patient’s very own cells could be biopsied, reprogrammed precisely into the cell type that is needed and then transplanted back into their own body, without the risk of rejection. This ability to take cells from a tissue that can regenerate itself (such as the skin) and customize them for use in a tissue with limited ability to repair itself (such as the brain or the heart) is undoubtedly an exciting possibility. But what sounds so simple on the page can often be much harder to achieve in reality.
Why our cells are resistant to reprogramming
The radical change of identity required for cellular reprogramming is not easy to achieve. Each cell in an immature embryo has a huge amount of potential and many possible paths it could follow. These immature cells which are yet to commit themselves are what biologists call stem cells. Just as humans tend to become more fixed in their professional specialization and habits: from the freedom of pre-school, to the relative focus of high school, to the more intensive training of further education, to the specific role of a day job – so it is for cells. As embryonic development proceeds and the embryo grows, the stem cells and their progeny become ever more specialized and less malleable in their identity. By the time all the organs are formed, most cells are highly committed to their own tissue and in many cases few stem cells remain.
The great polymath and biologist Conrad Waddington came up with another beautiful way of visualizing this process, a metaphor that fits very well with what we know of the actual biology. The early stem cells of an embryo can be thought of as a group of balls at the top of a hill. As development proceeds, the balls start to roll downhill. Eventually they will roll into a river valley and that valley will in turn split and split again repeatedly as the river navigates through the landscape. At each fork in the river, each ball must choose a route to follow: eg. “The Valley of the Brain” or “The Dale of Skin” (Figure 1A). Once they have chosen a route, the balls are committed. They cannot roll back uphill to the fork and they cannot climb over the land in between the valleys.
In the same way, the commitment of cells to specific tissues and organs is a one-way process.This is how the body ensures cells are precisely tailored to their role. Hair follicles need to grow hair, whilst brain cells need to conduct electric nerve impulses. Furthermore, reversal of the commitment process can have hugely destructive effects. This is precisely what leads to the development of cancer: rare cells acquire mutations that let them turn back the clock, acquire the properties of stem cells (which include the ability to reproduce themselves indefinitely), and start to wreak havoc. The one-way process of commitment ensures cancer is as rare as it is and that it tends to strike mainly later in life.
Making frogs jump backwards: the inefficiency of reprogramming
The development of cancer shows that cellular commitment need not always be a one-way process. John Gurdon showed, in important experiments he did as long ago as the 1950s, that all cells contain all of the genetic information in their DNA to make an entire organism and that, under the correct circumstances, they can be forced to use it. This may seem obvious now, but before Gurdon’s proof, it was perfectly feasible that as cells committed to their new identity they could shed, or at least irreversibly shut-off, the genes needed for the other cell types of the body.
In his breakthrough experiments Gurdon took a cell that had already committed to being part of a tadpole’s tail and removed the genetic material from it. He then injected this material into a frog egg cell that had in turn had its own genetic material removed. From this curious hybrid he was able to grow entire new tadpoles. The tail cell hadn’t lost any DNA and, crucially, some properties of the egg cell were able to re- activate all the genes that the tail cell had shut-off, reprogramme it into a stem cell and re-start development from scratch.
The reprogramming technique that Gurdon pioneered in the 1950s is exactly the same one that was used to clone Dolly the sheep in 1996. Since then the practice of cloning has become widespread and even has some applications in commercial agriculture. For example, some companies have decided to sidestep the genetic lottery of sexual reproduction and make exact clones of their most productive or disease-resistant cattle.
However, the idea of cloning entire human beings opens up a whole set of ethical debates that go beyond the scope of this essay. Nevertheless, some people think that this cloning method could still make an important contribution to regenerative medicine. This is because stem cells can be extracted and grown from cloned embryos, just as they can from normal embryos. Therefore, cells from a human patient could be cloned to make a source of stem cells that match the patient’s genetic make-up and could then be used for transplantation and tissue repair.
However, whilst cloned meat is deemed safe to eat, it remains something of a mysterious art. Scientists still don’t fully understand how it works, nor do they understand why many cloned animals develop unexpected defects. Also, since even using cloning as a source of new stem cells still requires the creation and destruction of a new human embryo, many will never find the process ethically viable. For these reasons the cloning of human cells has not taken off and may never be deemed acceptable practice.
Transcription factor reprogramming: from black art to hard science
10 years after Dolly was cloned our other Nobel prize winner, Shinya Yamanaka, made the next significant leap in the reprogramming field, a step that really does make the use of reprogramming in medicine a more realistic prospect.
Yamanaka focused on proteins in our cells called transcription factors. Transcription factors have a critical function in controlling what cells are and how they work because they function as tiny switches that turn genes on and off. Although all cells contain the entire set of genes in their DNA, each type of cell only uses the specific subset that they need to get their job done. It is transcription factors that turn nerve cell genes on and skin genes off in a nerve cell. It is transcription factors that activate the genes required to make a stem cell.
Following this logic, Yamanaka and his colleagues set about trying to identify a set of transcription factors that could be plugged into an adult cell that would transform it into an early embryonic stem cell. After trying many combinations, they eventually hit the jackpot. A set of just four transcription factors could do the trick. Returning to Waddington’s landscape analogy, Yamanaka’s factors are able to forcibly push a committed cell, in Yamanaka’s case a skin cell, back up to the peak of the landscape, where the whole range of possibilities for specialization are open to it once again (Figure 1B). In its final effect, this is exactly what Gurdon’s cloning technique also achieved, although via a quite different route.
In the flurry of excited research activity that followed Yamanaka’s breakthrough scientists have learnt much more about the reprogramming process. Another key step came in 2010 when Marius Wernig in Stanford University identified a distinct group of transcription factors that could reprogramme skin cells directly into brain cells. This type of transformation had already been achieved for cells that are more similar to each other, such as different types of blood cell, but never for such distinct cells as skin and brain. Returning to the landscape model again, this direct reprogramming approach catapults cells directly over the hill that separates two valleys to change identity without revisiting the stem cell state (Figure 1C). As we shall see later, this could be important when we consider the safety of reprogramming since stem cells, if not controlled properly, can cause tumours. Since Wernig’s feat, other researchers have used different transcription factors to reprogram skin cells into heart cells, liver cells, muscle cells, pancreatic cells and more. Such is the progress in this field that every month appears to yield a new recipe for a new type of reprogramming.
Crucially, whether direct from cell a to cell b, or indirect from cell a, to stem cell, to cell b, the transcription factor reprogramming approach comes with fewer ethical and practical strings attached when compared to Gurdon’s cloning. It does not require the harvesting of human egg cells, nor the creation of a new human embryo. Nevertheless, these techniques are still somewhat hit and miss. As discussed before, most cells are intrinsically resistant to reprogramming and it can be difficult to be certain that the reprogrammed cell really has wholeheartedly adopted its new identity and suppressed its old state.
How reprogrammed cells could actually be used
Even before scientists overcome the technical barriers to making the reprogramming process sufficiently specific, efficient and reliable enough to be useful, we need to consider how reprogrammed cells might actually be used. There are three main options, as outlined below and summarized in Figure 2.
1. Cells for transplantation
The most obvious application for reprogramming is to use it as a way to generate new, exactly genetically matched, cells to replace cells lost to damage or disease. For example if neurons in a specific brain region are killed by a stroke, the patient’s skin could be biopsied and the cells reprogrammed into replacement neurons, either directly or via a stem cell intermediate, and those neurons could be engrafted in the hope of repairing brain function.
2. Correction of genetic abnormalities
The direct transplant strategy described above sounds great when a one-off injury has occurred, but when a disease process continues to kill cells it will, at best, be a temporary fix. Where the disease has a genetic cause, an intriguing possibility is to combine reprogramming with genetic engineering to fix the defect. For example, some forms of motor neuron disease are caused by a single defect in a single gene that makes motor neurons more likely to die. If, as part of the reprogramming process, the gene could be repaired, then the transplanted neurons could be expected to survive and thrive when returned to the brain. Repairing the defective gene is much easier to achieve in immature cells than in adult cells, so this strategy is only a realistic option when reprogramming all the way to the stem cell state, according to Yamanaka’s process. Amazingly, scientists have already used this technique to “cure” mice that carry a mutation causing the blood disorder Sickle Cell Anaemia. In principle at least this strategy is feasible.
3. Finding new drugs to treat diseases
Even if it proves too difficult to efficiently and safely reprogram cells for human transplant, the reprogramming process can have an important impact on the development of medicine. Many human diseases are extremely difficult to study because it is hard to access the affected cells and, by the time the disease has progressed, there is little to learn about how the cells came to be diseased. For example, it would be ethically dubious, practically dangerous and likely uninformative for doctors to delve into the brain of a patient with Alzheimer’s disease. Instead, if they can take a sample from the patient’s skin and reprogramme those cells into neurons, scientists can observe, in the petri dish, when the Alzheimer’s-prone neurons deviate from normal and thus gain new insights into how cells accumulate the defects that eventually result in the disease itself.
An exciting extension of this idea is to use reprogrammed cells in the lab to find new medicines that can tackle the precise cellular defects that cause disease.Whereas clinical trials in patients are hugely expensive and safety is always of the utmost importance, in the lab huge numbers of disease relevant, genetically appropriate cells can be used to test thousands of potential medicines in a short time. Although the conditions in the petri dish are inescapably distant from reality, this type of approach is a new and powerful way to speed up the drug discovery cycle and bring promising drugs to the clinical trial stage and into the clinics.
Is reprogramming safe and if so when can I have my cells reprogrammed?
John Gurdon, Shinya Yamanaka and their colleagues around the globe have given us a glimpse of a world where cells can be forced to make transformations that would have impressed even the alchemists of medieval times. There are, of course, lots of practical details to work out and safety issues to address before any of these techniques can be used to treat human patients. For example, where reprogrammed stem cells are transplanted, there is always a risk that tumours could develop if the stem cells are not well controlled. It is also worrying that the most effective ways to introduce reprogramming transcription factors into cells use viruses, which can cause unexpected genetic problems. Nevertheless, none of these are intractable problems. Unlike medieval alchemy it seems that the new science of cellular reprogramming will eventually have a significant impact on human health and prosperity.