Immortal science

Professor Martin Pera, Program Leader of the ARC-supported national research consortium Stem Cells Australia, reflects on his involvement in the discovery of human embryonic stem cells and where the research field is at today.

Australian Life Scientist: What sparked your interest in science?

Professor Martin Pera: I actually studied English literature as an undergraduate. I grew up in New York and Washington, and in the US you can have a bit more diversity in your program as an undergraduate. So along with poetry and novels, I also studied some science. I very much enjoyed literature but the only thing I was interested in doing with it was writing, and I never thought I’d be a good enough writer, so I decided to try my hand at science instead.

I went back part-time as a student to fill in science courses in chemistry. I decided I wanted to do something where I could apply some of what I’d learnt to human health issues. Then, with a chemistry background I pursued a PhD in pharmacology.

ALS: What led you to come to Australia?

MP: That’s a long story. I did my PhD in the US in pharmacology studying the cancer drug cisplatin. Cisplatin became the mainstay of treatment for human germ cell tumours of the testis and my first couple of postdocs involved studying that drug.

Then I decided the cells we were killing with the drug were more interesting than the actual drug itself.

Germ cell tumours of the testis are the original paradigm for a cancer stem cell. The stem cells of those tumours are very much like embryonic stem cells. They can divide to form more tumour cells but they can also differentiate into normal tissues.

I went on to set up my own lab at the Royal Marsden Hospital in England studying germ cell tumours of the testis and their biology. We isolated pluripotent stem cell lines from those tumours and characterised them.

I spent some time at Oxford University where we first made attempts to derive embryonic stem cells (ESCs) from human embryos. Those attempts didn’t work out.

In a kind of roundabout way I got in touch with Alan Trounson who was interested in recruiting me to really have another go at deriving human ESCs. And that’s how I came to Australia.

I moved out to work with Alan down in Monash University in 1996. By 1998 we had got our first ESC lines going, with the help of Ben Reubinoff, a clinician from Israel, and Ariff Bongso, an embryologist from Singapore. But then we learned of Jamie Thomson’s discovery - he had already derived human ESCs - and that’s how we became the second group in the world to report human ESCs.

ALS: So the work with the germ cell tumours set the scene for isolating ESCs?

MP: Yes, the history of the whole field of ESCs really begins with germ cell tumours both in the mouse and the human. We were isolating these cells in the late ’80s and early ’90s.

Back in those days there were only a few groups around the world working on human pluripotent cells from germ cell tumours.

We showed that we could isolate cells that could differentiate into a whole range of tissues of the body and we used that information as the basis for trying to derive the normal equivalent of these cancer cells directly from embryos.

We made our first attempts to make human stem cells from embryos at Oxford around the mid-1990s. What we did very much paralleled what had gone on before in the mouse. People first studied germ cell tumours in the mouse and from that basis they went on to make mouse ESCs. Essentially we were following that same pathway.

ALS: Were there any eureka moments in the lab?

The lab team at Monash in 2004.

MP: One major breakthrough was when we injected human germ cell tumours into immune-deprived mice.

We’d isolated many cell lines from patients with germ cell tumours but because they were cancer cells many of them didn’t behave normally. But with one or two cell lines, when we grafted them into immune-deprived mice, we removed the cancers that formed and began to see a whole range of cell types like nerves and cartilage developing.

Then we were able to go back and clone individual cell lines from cancers from particular patients and show that they would spontaneously differentiate both in culture and in an animal. That was a pretty big deal because it was probably the most convincing evidence that there were human pluripotent stem cells.

That was when we began working with embryos. And at about that time Jamie Thomson published the first derivation of stem cells from monkey embryos.

This was a real breakthrough because we knew all along that our human cells from the tumours were different from their mouse counterparts - they had different markers and they looked different and grew differently. Although, like the mouse cells, they could turn into all the tissue of the body, we didn’t understand whether those differences were because these were malignant cells and just abnormal or whether there were real species differences.

When Jamie identified the stem cells from monkey embryos they looked exactly like the human cells we’d been isolating from cancers of the testis. And that told us immediately there was no doubt we could make these cells from a human embryo. More importantly, it told us what to look for and how to handle the cells, because they are quite different to handle from mouse ESCs.

ALS: And isolating cells from human embryos was quite controversial back then?

MP: Absolutely it was, yes.

In England the attempts that were made to isolate the cells, although unsuccessful, were approved by the Human Fertilisation and Embryology Authority (HFEA). The regulatory framework to do those experiments was there and we had a licence from the HFEA to do that.

When I came to Australia in 1996, any destructive manipulation of the human embryo for experimental purposes was illegal at that time in Victoria.

We wound up collaborating with Ariff Bongso’s group in Singapore. We would initiate the cell lines there and bring them back to Melbourne for further characterisation. Once we’d turned the embryo into a cell line it was outside of the restrictions, provided it was derived using an acceptable ethical approach.

One of the things we and others did in the early days of the field was to do our best to disseminate this technology, to teach others, to share the cell lines and share reagents so that the field could move forward.

People accused us of going offshore to accomplish something that we couldn’t accomplish in Australia, but there were other states in Australia where this research would have been permissible at that time. Before there was uniform national legislation each of the states had a different approach to the regulation of human embryo research, and Victoria had a particularly restrictive environment.

ALS: What are some of the challenges that have arisen with researching ESCs and understanding their role in repair?

MP: In the early days there was a lot of scepticism about whether true ESCs could be derived from any species other than the mouse. And to this day, there are some people in the field who would still say that the only true ESCs are those described in mouse and rat.

It wasn’t at all clear whether human ESC could ever be isolated or, once it was apparently accomplished, that we’d in fact isolated the right cells. There were enormous issues around just propagating the cells. Now the technology is much better but back then the methodology for growing ESCs was in its infancy - we were developing it and it was always a struggle to keep the cell lines going and to stop them from differentiating.

Then there was the whole question of having made pluripotent stem cells, how to turn them into useful cell types for use in therapy. That’s an ongoing saga.

But the field has progressed. When I was at the University of Southern California, I was part of a group that was doing work toward developing an ESC-based treatment for macular degeneration. I’m very pleased that that work is going forward and heading towards clinical trials. It’s a little over a decade since the early discoveries and it’s remarkable to see that kind of progress.

The key thing for any therapeutic application is that you have to be able to expand the cells first, so you have to be able to keep them going in the undifferentiated state and then you have to turn them into the cell type of interest for a particular disease application.

That’s been an enormous challenge in the field but it’s been one that has been overcome for many types of tissue.

The real challenge now is not making a particular type of cell but how to get it to mature completely - in other words, how to make it fully functional just like the normal cell in the adult tissue.

Making sufficient differentiated cells safely and in a pure form also remains a big challenge. This is applied biotechnology research and I’m not sure that many academic labs are focused on these issues of production, scale-up, cost and the like.

ALS: And understanding the biology of differentiation is still ongoing?

MP: That’s right. We and others have been guided by decades of work on model systems, animal embryos from the fruit fly through to Xenopus to fish to the mouse.

Because many developmental mechanisms are conserved evolutionarily, you can take a lot of that information and apply it to understanding how to manipulate stem cells. But a human isn’t a mouse, and the devil is in the detail, so often there are aspects of stem cell regulation that are specific to human cells that we have to come to understand.

ALS: How do ESCs compare to adult stem cells? Do they have a greater degree of plasticity?

MP: They do.

In the early 2000s, there were a lot of papers published claiming that stem cells from adult tissue could turn into virtually anything - a lot of that work turned out not to be robust in the end.

Now we know that through the process of reprogramming, in the laboratory at least, we can take cells from adult tissues and reset them back to the early embryonic state and turn them into cell lines that very much resemble ESCs. This is the remarkable work of Shinyaya Yamanaka, who received a Nobel Prize last year for this discovery.

So now, more and more, we work with these so-called induced pluripotent stem cells that are made from adult cells. They have opened up a whole new range of possibilities.

ALS: Does the use of adult stem cells help get around the legislative and ethical issues?

MP: It certainly gets around the ethical issues relating to the provenance of the cells because you don’t have to use an embryo anymore to make a stem cell line.

Nevertheless, there are certainly ethical issues around the use of these cells, including genetic privacy issues for the donors of the tissues and what we might ultimately be allowed to do in the laboratory with these cells.

For instance, one day - and we can already do this in the mouse - it may be possible to turn these cells into human gametes; that is, sperm and egg. That might be a very powerful tool for studying and even treating infertility, but it will raise ethical questions along how far we want to go with these cell lines in terms of introducing them into the human reproductive cycle. And these are big questions.

ALS: Where is the legislation for using ESCs now at in Australia?

MP: As a result of the legislation in Australia in the early 2000s, which was recently updated, we can use embryos to make new embryonic stem cell lines - provided we’ve gone through the appropriate review and application process. We can even do somatic cell nuclear transfer, which is the use of cloning technology, to make stem cell lines from particular individuals.

The controversy over the use of embryos will never completely disappear, because there will always be a minority who will be opposed to this work. But its practical implications for the field are much less than they were 10 or 15 years ago.

The reasons for this are that we now have many thousands of ESC lines, which is more than ample for most research purposes. And now we also have this alternate technology - the induced pluripotent stem cell, which really gives us another system to work with. We don’t really know yet whether induced pluripotent stem cells are a replacement for human ESCs, but they are certainly a very promising alternative at the very least.

ALS: What about the unethical use of stem cell therapies, is this happening in Australia?

MP: With the exception of bone marrow transplantation and one or two other applications, almost all stem cell medicine is experimental medicine, and it really should be carried out in the context of a proper controlled clinical trial.

Unfortunately, one of the biggest issues the field faces now is that of clinics providing so-called ‘stem cell therapies’ that are unproven and unfounded.

Clinics are popping up and offering a range of treatments for a staggering array of diseases using cell therapies. These clinics call the treatments stem cell therapies but in many cases it’s not clear whether they are using stem cells at all - many use crude and poorly characterised tissue extracts. And in many cases this is being done without any scientific basis for what is being offered or even appropriate safety evaluations. This is a big concern for us and an enormous issue for the field in Australia and internationally.

There are clinics in Australia offering such unproven stem cell treatments. In general, the regulations we have around cell therapies in this country are very good, and they are enforced by the Therapeutic Goods Association. Unfortunately, there is an exemption to this set of regulations that says that if a clinic or physician is dealing with a patient’s own cells, so-called autologous therapy, then that approach is exempt from these regulations. And that’s a loophole that allows for clinics to offer unproven and untested stem cell treatments. We’re seeing this as an increasing trend.

We’re very concerned that the government go back and take another look at this autologous therapy, as jurisdictions elsewhere are. The International Society for Stem Cell Research, one of the leading international bodies in this field, recently issued a very strong position paper on this because it’s an issue that extends beyond borders. We are working with others to see if we can get this policy reviewed.

One of the biggest challenges we faced from the very first days in the field is getting across the promise of the work and its great potential but not promising too much too quickly. There is understandably a lack of clarity in the public mind about stem cells. We try and educate the public but there is still confusion around what stem cells really are and what they actually can do now versus the long-term promise.

ALS: Do you see a healthy future for stem cell therapies?

The launch of Stem Cells Australia. (R to L) Professor Margaret Sheil, Natasha Stott Despoja, Professor Martin Pera and Professor Glyn Davis.

MP: Oh yes. Remarkably, we now have the first clinical trials on products derived from embryonic stem cells, as I said for macular degeneration, but we are going to see trials for diabetes and possibly other indications soon.

Japan is moving ahead with parallel trials for products derived from induced pluripotent stem cells. I think these early clinical trials will probably wind up raising more questions than they answer and posing more challenges, but it’s remarkable how far we have come today from 1998.

I’ve been personally amazed at the pace of the progress. The potential was always there but it’s surprised me how fast we’ve actually moved.

One of our big challenges in this field in Australia is pushing translational research and making sure that outstanding clinicians, physicians in training, have an opportunity to do research and to have fulfilling careers as academic physicians in this area.

It’s going to be absolutely critical for the future of this field that we have strong communication between the basic science and the physicians - that we have people who understand both the science and the clinical side.

There has to be funding to facilitate that interchange, and there has to be funding for the translational research which, of course, is expensive and high risk.

I think we can do this a lot better. But recognising the problem and doing something about it of course is two different things.

It’s very important that Australia maintain and build its strength in both basic and applied research in this area because in many respects this is a different model to a pharmaceutical model. Cell therapy will be carried out in the context of experimental medicine for a long time, as we constantly refine and improve the technology. In this respect it is similar to cancer medicine, which requires high-level tertiary specialist referral centres steeped in the research enterprise to deliver the latest advances to patients. Unless we have that fundamental strength in basic and applied research in regenerative medicine, Australian patients are going to miss out on the benefits.