p53

by Sue Armstrong

  But this was not what he had come to say. His genetically engineered animals were fine. Not only had they survived without the protection of the guardian the period of explosive growth, cell division and differentiation that turns an embryo into a pup, but they had no signs of physical deformities or cancerous growth. Donehower’s audience was stunned.

  David Lane remembered the occasion vividly when I interviewed him for this book on the fringes of a big conference in Liverpool. ‘We were all in a very triumphant mood as a community. “p53 is now the most important bloody protein in the world and all you guys can get stuffed!”, you know? “Who’s been telling us that we’ve been wasting our time for the last 10 years?” It felt exciting and good,’ he recalled with a grin. ‘We were having this big p53 workshop in the US, and instead of just 20 people coming, 200 were coming. There were lots of positive data . . . loads of people finding mutations, and everything was now very convincing. Then Larry stands up and says, “I’ve made a knock-out mouse and there are some interesting things about this mouse . . . First of all it’s completely viable. There are no defects that I can see – and it certainly hasn’t got any cancer!” Everybody went, “Uh oh!”’ Lane leaned back in his chair and gave a huge laugh of incredulity. ‘Of course, I guess it was about two months after the conference, Larry started to see massive development of tumours in all the animals and there was a collective sigh of relief . . . Poor mice, but lucky us!’ Lane paused to reflect over the 20 years since that meeting. ‘It was incredible, actually. And, of course, having the knock-out as a tool has made an enormous difference to everything.’

  Donehower had been as stunned as everyone else by his initial results, which had ramifications beyond the purely scientific. The colossal effort that goes into such research and the money poured in to support it generate high expectations and heavy pressure for exciting results. When ‘nothing’ happens, panic sets in and doubt ripples across the whole community. In time, however, all Donehower’s knock-out mice did indeed succumb to cancer, surviving for less than five months compared with around 30 months for normal mice of the same genetic background. Further research also revealed that p53 knock-out mice have much smaller litters than normal, suggesting the gene plays a part somewhere in reproduction. And it has flagged up a vital role for p53 in regulating metabolism, which is one of the hottest topics of investigation at the moment.

  But back to the story of ageing. In 2002, Larry Donehower and his team dropped their second bombshell when they made a mistake with one of their experiments and got a mighty surprise. They were trying to make a knock-out mouse using a different technique from before, but they ended up instead with a mouse in which the still-present p53 gene was hyperactive. Sure enough, the creatures proved well protected from cancer, as the researchers would have predicted. What none of them expected to see, however, was that they aged exceptionally fast. In just a few months, they looked like very old mice. ‘They had hunchback spines, ruffled fur, grey hair; things like that. And they lived only about two-thirds of their normal life span,’ Donehower told me when I spoke to him at a p53 meeting in New York. ‘Some of the most interesting findings in science are accidental, actually. They’re not what you’re looking for or expecting, and this was very surprising. Nature published it in 2002. Now this accidental finding is opening up a whole new area of research about how this very important cancer gene can also modify the ageing process.’

  People have known for a long time that ageing and cancer are related, in that the chances of getting cancer increase with age. But not even the scientists suspected they might be two sides of the same coin, sharing a common mechanism in which the scales can be tipped either way. In other words, that wrinkled skin, thinning bones and failing organs may be the price we pay in the long run for holding cancer at bay. Donehower’s findings, however, left room for a smidgen of doubt about the role of p53, since the ‘accident’ that produced the hyperactive version also knocked out a stretch of DNA upstream of the tumour-suppressor gene. The possibility could not be ruled out that something here might be responsible for the premature ageing. But soon another lab, run by Heidi Scrable of the University of Virginia at Charlottesville, provided new evidence that Donehower’s original hunch was right. She and her team created a mouse model in which the only change to its DNA was the replacement of one allele of p53 with a naturally occurring hyperactive version of the gene, and found the same thing – premature ageing and death.

  Donehower, Scrable and others working in this compulsively intriguing field have gradually pieced together the picture of how this can happen. A hormone known as insulin-like growth factor 1 (most often represented as IGF-1) that, not surprisingly, plays a central role in the growth and proliferation of cells, has long been known to promote ageing too in all manner of organisms, from fruit flies and nematode worms to mice. By tinkering with the strength of the signals this hormone sends out to the cells, researchers have managed to manipulate the life span of these creatures. The effect is most obvious in flies and worms, which live considerably longer when IGF-1 signalling is dampened down and shorter when it is amplified.

  Scrable and her team found that the hyperactive p53 in their engineered mice stimulated hyperactivity of the growth hormone too. The amplified signals from IGF-1 in turn triggered the mechanism designed to bring runaway cells under control by driving them into senescence, or irreversible arrest. This, of course, is tumour suppression at work, and is orchestrated by ‘regular’ p53. To that extent it was an appropriate, and clearly beneficial, response. But senescent cells can become dysfunctional and as they accumulate in the tissues they begin to cause trouble of their own.

  Unlike cells that have been driven to suicide by apoptosis, senescent cells remain alive and active – and, significantly, they alter the micro-environment of the tissues by secreting proteins that communicate with neighbouring cells and even with distant organs. Some of these proteins are important for tumour suppression – for example, they inhibit the development of new blood vessels which might feed a developing tumour. But as they metabolise in the normal course of events, senescent cells also produce large amounts of material that seeps into the surrounding tissue. ‘This begins to chew up the extra-cellular matrix – you know, the stuff that keeps cells glued together,’ said Judith Campisi, who studies senescence at the Buck Institute for Research on Aging in Berkeley, California, when I spoke to her at the same New York meeting as Larry Donehower. ‘The major extra-cellular molecule that keeps your skin looking young is collagen. And sure enough, senescent cells produce molecules that destroy collagen.’ Hence wrinkles.

  Campisi, dark haired and petite, with dangly earrings and the graceful posture of a ballet dancer, uses her hands and eyes expressively as she speaks. She began her research career focusing on cancer, and it was here she first encountered senescent cells, in the context of tumour suppression. But she soon became fascinated by their possible role in the normal processes of ageing – an idea that all but a small ‘crazy contingent’ of scientists had dismissed for a long time. ‘I didn’t buy [the theory] for a minute,’ she said in an interview with one of her colleagues at the Buck Institute in early 2013. ‘[But] sure enough, we started working on this problem and I had to realise that this “crazy contingent” was actually correct!’

  To the uninitiated, the scientists’ original doubts seem strange, since the very term ‘senescence’ implies ageing. This was clearly intended by Leonard Hayflick, the man who discovered and named the cells in 1961 – but he was putting himself out on a fragile limb scientifically, Campisi told me. ‘Len Hayflick was a cell biologist, and he was studying cell proliferation for a very specific reason: he was interested in growing viruses in human cell cultures as opposed to animal cell cultures, and virologists were having a helluva time. They would get these cultures and initially they’d do great, and then eventually they wouldn’t do so great, and they’d throw them out and start again.’ Hayflick decided to study this in much more detail, and he
made the startling discovery that, in contrast to most cancer cells, normal human cells have a finite ability to undergo cell division in culture. ‘Now that finite ability is huge,’ said Campisi. ‘For stem cells taken from a human embryo we’re talking 40, 50, 60 population doublings. So you can see why you’d be fooled – you’d do an experiment for three months, the cells are growing great and then they start not to do so great and then they stop.

  ‘Hayflick made two interesting observations,’ she continued. ‘One of them was obvious and was immediately accepted, and that is, “My God, tumour cells don’t do this. Maybe this is a way of stopping cancer.” It fuelled a whole area of research to think about the senescence process, which is controlled by this famous suppressor gene, p53, as stopping cancer, and I think there’s very little controversy about that now. But he also made another observation that was totally unscientific, totally intuitive, based only on his sense as a cell biologist. He looked in the microscope; he looked at these cells and said, “They look old.” Now what on earth does that mean, that a cell looks “old”? I mean it’s an unquantifiable observation. But he said, “Maybe what’s also happening is that we are recapitulating some aspects of ageing in a culture dish.” That observation, that comment, went largely unnoticed except for a few, again pretty imaginative, people in the field who picked this up and began to study senescence, not as a tumour-suppressive mechanism but as an ageing process. It is still somewhat controversial – less so than it was 50 years ago, but still controversial, though it’s also gained a lot of momentum.’

  The number of divisions a normal cell can undergo before becoming senescent is known today as the Hayflick Limit, and it is measured by the telomeres on the ends of the chromosomes. Telomeres are protective tips to the chromosomes that are rather like the little plastic caps put on the ends of shoelaces to stop them unravelling. Every time a cell divides, the telomeres shorten, until they are no longer able to protect the chromosomes and the cell goes into permanent arrest. And though this is not the only route to cell senescence, dangerously shortened telomeres are one of the stressors that trigger the p53 response.

  Today Judith Campisi is a world leader in the field of cell senescence, and she is in no doubt that these cells are at the pivotal point of a mechanism that can tip either way. ‘What my work tries to do is to reconcile the two very different views of senescence. One says it’s really good for you, it stops cancer. The other says it happens during ageing and it looks like it’s bad for you because the cells look kind of old and ragged.’

  Her lab has discovered recently that senescent cells provoke inflammation – a condition that underlies almost every major age-related disease, she told her interviewer at the Buck Institute. ‘We’ve shown now very clearly that one senescent cell sitting in a sea of non-senescent cells will provoke an inflammation that will spread to other cells. So it’s a very appealing hypothesis that you don’t need very many senescent cells to be able to drive the degenerative changes that are a characteristic of ageing organisms.’

  Senescent cells are also highly resistant to apoptosis and the ultimate irony is that, with time, they themselves become a cancer risk, helping to drive the process of uncontrolled growth. But how? ‘In the last couple of years we’ve learnt that the senescence response has another life, and that is to promote tissue repair when needed,’ said Campisi. ‘That’s where it begins to be problematic in later life.’ As senescent cells become dysfunctional with time, she explained, they can start to send out signals to initiate tissue repair and proliferation of cells in the absence of real injury, thus driving the development of tumours.

  This, of course, is not inevitable. As researchers have found with every aspect of tumour suppression, context is all important: different cell types and tissues follow different paths on activation of p53. Scott Lowe, another mouse-model man, whom we met in the chapter on apoptosis, is also at the cutting edge of cell-senescence research; he discovered that, although these cells are indeed resistant to killing by apoptosis, they don’t always hang around in the tissue to become toxic. In some tissues they communicate with the immune system, which sends in the scavenger cells to clear them away.

  ‘We had this study in 2007, in which if you had no p53 you had a cancer cell; if you flipped p53 on, the cancer cell went senescent,’ Lowe explained. ‘We could see this in the Petri dish and also in the animal. But whereas in the Petri dish the cells just sat there – they never divided, they didn’t die but they didn’t grow – in the animal the tumour went away.’ This was perplexing: surely, the researchers figured, if the cells weren’t dividing, nor were they dying, the tumour should stay the same size. So what was happening here? Digging more deeply into the mechanism, they found that proteins secreted by the senescent cells were triggering an immune response that was removing them as effectively as apoptosis. What’s more, they could watch this happening in the lab, watch the scavenger cells engulf senescent cells, if they put the two together in the Petri dish and allowed them to communicate – in effect creating a simplified version of the community of cells you would find in a living body.

  As with their discovery of dead cells and apoptosis, this was a surprising result, not what they were expecting to see at all, but Lowe found it strangely pleasing intellectually. ‘For 15, 20 years, we studied how p53 affects the cell that it’s turned on in, but now we realised it does more than that . . . It also can send signals that affect the surrounding tissue.’

  Still today no one knows exactly why the stress response leads to different outcomes under different circumstances. ‘Part of it is tissue-dependent: lymphoid cells will, by and large, apoptose, and connective tissue will senesce,’ said Lowe. ‘But it isn’t completely that. There are other factors that influence it, some of which we know, but none in a way that you can satisfyingly say is decisively the answer . . . And it’s not even that p53 is turning on different sets of genes when the cells die versus when they arrest, so it’s also something about how the cell interprets the genes that p53 turns on. This is a really interesting question for what we now call “systems biology” – how the cell integrates multiple signals to make a yes or no decision to go down a certain path – and it’s stuff we study to this day.’

  Researchers interested in the gene’s role in ageing believe that both apoptosis and senescence are significant to the process – senescence for all the reasons discussed above, and apoptosis because it gradually depletes the pool of stem cells our bodies need for repair and maintenance. ‘The simplest model would be that you’re born with a limited number of stem cells,’ explained David Lane. ‘Those stem cells are very easily killed off by DNA damage, so they’re the ones most tightly controlled by p53. If you set a stress-response threshold where they’re too easily killed, then you don’t get cancer but you run out of stem cells more quickly. If you set the threshold such that they’re hard to kill, then you could live a long time, but you’re more likely to get cancer.’

  Age researchers also have a theory, drawn from evolutionary biology, to explain the paradox of why a system designed to preserve life by protecting us from cancer should also drive the mechanism that leads inexorably to our decline. Basically, nature only cares about perpetuating the species, so evolutionary pressures to select for advantageous traits – and weed out ones that are harmful – operate only up to and across our reproductive years. Beyond that we are living on borrowed time: nature no longer has a use for us, and natural selection is a spent force.

  Ageing is anyway a modern phenomenon. During the great majority of our time on Earth, humans didn’t die of ageing: we didn’t die of cancer or Alzheimer’s disease, or even cardiovascular disease; we died of accidents and predators and infection and starvation. Thus for millennia, ageing operated below the radar of evolution by natural selection. It’s only today, as we’ve conquered infection, hunger and predation in much of the world, that the damaging flip-side of tumour suppression has been able to play itself out to full effect. But understanding the ro
ots of ageing holds promise for the future. ‘People are beginning to ask: can I manipulate the system to get the best of both worlds?’ commented David Lane. ‘Can I sensitise the gene for short periods (to eliminate cancerous cells) and can I suppress it (to keep ageing at bay)? I think one can imagine really quite extraordinary results as we begin to be able to control this system.’

  CHAPTER TWENTY

  The Treatment Revolution

  In which we hear of p53’s place at the cutting edge of gene therapy and personalised medicine, which are revolutionising the treatment of cancer – and, some predict, will remove the threat of ever dying of cancer from today’s young people.

  ***

  [p53] already, with no help from doctors, stops incipient cancer millions of times every day. Scientists do not have to top the elegant system that nature has engineered. They just have to harness it.

  Sharon Begley

  ‘If you peer into cancer cells – and we’ve got amazing technologies now to catalogue all the things that are happening – and you look at them from an evolutionary point of view, it turns out that even within a single tumour there are many, many different subspecies of cells that have evolved in slightly different ways,’ says Gerard Evan, whose remarks about the rarity of cancer opened this book. ‘And here we are, basically trying to wipe out the entire tumour while keeping the patient alive at the same time. It’s a tough deal!’

  But Evan is not disheartened. Indeed, as evidence mounts that cancer is an even more complex disease than anyone realised – a hotbed of evolution that makes of tumours a constantly moving target for therapy – he remains decidedly optimistic. Why? ‘Let me give you an analogy,’ he says. ‘Absent of cancer, human beings have been subject to terrible diseases of cells that grow inside them and invade and spread. These cells are genetically very heterogeneous, they exchange genetic information one with the other, and they grow like crazy . . . They’re called bacteria, right?