p53 (31 page)

Galina Selivanova at the Karolinska Institute in Stockholm is working on a drug of this design which she has named RITA. She points out that in order to kill cells, it is generally not enough
for p53 simply to be present; to become active, the tumour suppressor needs to receive clear signals that the cell is under stress – signals that are likely to be strongest in cancer cells.
‘My hope is that if you have an Mdm2 inhibitor which is not too strong – maybe it’s enough to release just some p53 from Mdm2 – it will not have very drastic effects in
normal cells. But in tumour cells, where you have all these signals which are activating p53, it will kill.’

MENDING THE MUTANT

When she left Russia in 1992 with a PhD in bacterial genetics from Moscow University, Selivanova intended to spend just three months of summer gaining experience with work on
higher organisms before returning home. However, she joined the lab of Klas Wiman at the Karolinska Institute, discovered p53 and never went back. ‘It was so exciting from the start,’
she told me when I met her at a mutant p53 meeting in Toronto. ‘p53 is
unbelievably
interesting. Everything you do opens new questions, new perspectives.’ She joined the p53
community just as people were beginning to think seriously about translation – how they might use the wealth of knowledge they had accumulated to improve the treatment of people with cancer.
It was a topic with personal significance: Selivanova had seen her own mother die of a brain tumour, and she soon found herself drawn into the quest.

Besides her own work with RITA, she and Wiman have worked together on another drug, known as PRIMA-1, that is turning out to be one of the most exciting p53-based therapies in development. The
drug is designed to work in cancer cells where p53 is mutant and the protein it produces misshapen so that it cannot bind to DNA, as it should, in order to switch on other genes. PRIMA-1 is able to
restore the mutant protein to its normal shape, and serendipity played a large part in its discovery. In 1995, the two scientists were studying small scraps of protein called peptides, looking for
ones that could regulate the activity of p53. They were intrigued to discover one peptide that was able to activate both normal and mutant p53.

This was clear evidence that ‘mending’ mutant p53 was possible, and Selivanova was very excited. ‘It was fantastic,’ she recalled with a smile. ‘I wanted, of
course, to go out and cure tumours – at least in mice!’ But the peptide proved unworkable as a drug: in a living organism these scraps of protein are poor at entering cells and are
quickly broken
down and recycled. What was needed was a chemical compound, a small molecule that would perform the same tricks.

Working with Wiman and a new postdoc, Vladimir Bykov, Selivanova screened thousands of compounds from a library of possible candidates provided by the US National Cancer Institute. In 1999, the
three discovered a molecule they named PRIMA, an acronym for ‘p53 reactivation and induction of massive apoptosis’ that appealed to the scientists because it also implies something that
is first class. Experimenting with the molecule they found, to their gratification, that it is effective with a wide range of p53 mutations, and therefore potentially useful in treating many
different tumour types. They published their results soon afterwards, ‘and PRIMA attracted
a lot
of media attention,’ Wiman, a tall, soft-spoken Swede, told me when I visited
him at the Karolinska Institute. ‘I was on TV and in newspapers and journals around the world, because the concept of having a small molecule that will make the cancer cells commit suicide is
so appealing.’

So how does PRIMA-1 work? Wiman and Bykov discovered, to their surprise, that both PRIMA-1 and a very similar compound known as PRIMA-1 MET are converted to another compound that binds tightly
to p53 protein and refolds it. ‘This was a very important and exciting finding since it gave us a better understanding of how these compounds can reactivate mutant p53,’ said Wiman.

In partnership with the Karolinska, he, Bykov and Selivanova set up a small biotech company to develop PRIMA-1 MET for the market. It has been a steep learning curve. ‘As scientists you
need to work with company people – a completely different culture,’ commented Wiman. ‘Suddenly there are people in suits, board meetings, talk about money . . . And then you
interact with clinicians too. So there are three worlds and you all have to work together all the way through. We had no idea what was involved when we started.’

The company has taken PRIMA-1 MET through a phase 1 clinical trial, which involved 22 patients with cancer of the prostate and blood being given a short course of the
drug by injection. Phase 1 trials are designed to test patients’ tolerance to a potential new drug, and to find out how it disperses in the body and how long it persists. The results,
published in 2012, were promising: they showed that PRIMA-1 MET is not toxic and that side effects – including dizziness and fatigue – are mild.

Phase 2 clinical trials, designed to prove that the drug works in people as it does in the lab and in animal models, are the next step. The Karolinska researchers and their company are hoping to
test PRIMA-1 MET in combination with conventional chemotherapy in cancer patients, where the two drugs are expected to act in synergy: while PRIMA-1 MET restores mutant p53 to its normal shape and
function, the other drug will cause DNA damage that sends clear signals of stress to trigger apoptosis. But this is where the hurdles en route to the clinic really begin: staging a phase 2 trial
for PRIMA-1 MET is likely to cost millions of euros, said Wiman. A tiny biotech company like theirs needs to find a partner with serious money to invest.

A doctor in Denmark who has seen the effects of PRIMA-1 MET on lung-cancer cells in the lab and in mice is so excited by the drug that he has offered to set up a trial himself. The MD Anderson
Center, too, is keen to run a phase 2 trial of PRIMA-1 with cancer patients. But finding the funds for all these activities remains a huge challenge, and so far Big Pharma has shown little interest
in small molecules that restore normal shape and function to mutant p53 because it is still not entirely clear how they work.

SMART THERAPY

A problem that dogs the field of cancer therapy is the issue of drug resistance. The extreme instability of cancer cells
and the terrible speed with
which they pick up mutations mean that they are likely to find a way round a targeted drug before too long, no matter how clever the design, as the cells that survive the initial onslaught of
treatment give rise to equally hardy clones that grow into resistant tumours. To minimise the prospect of failure, oncologists typically treat their patients with a combination of therapies –
either a cocktail of drugs, or a drug together with radiotherapy. With this strategy, cancer cells that are not affected by one drug should be hit by the other.

Researchers are also investigating the use of drug combinations in a novel kind of p53-based treatment called cyclotherapy. One of the biggest shortcomings of conventional chemotherapy, which is
‘cytotoxic’ (meaning that it’s a cell poison) and targets the body’s rapidly dividing cells, is that it is indiscriminate. Cancer cells are by definition fast-dividing, but
so too are the cells in the hair follicles, lining of the gut and bone marrow, which sustain collateral damage during chemotherapy. But hair loss, nausea, diarrhoea, anaemia and depletion of the
immune system are not just distressing side effects for the patient, they are potentially deadly and they limit the dose of cytotoxic drugs the oncologist can administer to attack the cancer.

The principle behind cyclotherapy is that patients be given one drug to ‘protect’ the healthy cells from the chemotherapy by temporarily stopping them from dividing, while their
cancer cells (which continue to divide and therefore remain targets of the chemotherapy) are blasted with a second, cytoxic drug given simultaneously. With healthy cells protected, the theory goes,
the oncologist will be able to increase the dose of the cytotoxic drug and thus maximise its potential to wipe out the tumour. But even if it falls short of clearing the cancer completely,
cyclotherapy will make chemo a lot less unpleasant for the patient because it will limit the side effects by sparing the cells of the hair, gut, bone marrow, etc. from the full force of
treatment.

In laboratory tests, Nutlin is looking the most promising of a number of similar drugs used to protect the healthy cells. However, cyclotherapy is still a few years from
the clinic. Researchers still need to work out which combinations of drugs work best, with what tumour types and in what quantities. The arrest of healthy cells mid-cycle must be reversible: too
high a dose of the protective drug, for example, could cause healthy cells to senesce, but too low a dose might not arrest them for long enough to protect them from the cytotoxic drug. And no one
is sure yet how well cyclotherapy works in living organisms: as of 2012 there was only one published report of an experiment in mice. However, one of the main constraints on cyclotherapy is the
fact that neither Nutlin nor any of the other potential ‘protectors’ has yet been approved for use in the clinic in its own right.

NEW LIGHT ON OLD TREATMENT

Despite the frustratingly slow progress of brand new p53-based therapies, scientists’ understanding of p53 is already beginning to have an impact on the treatment of
cancer patients: it enables oncologists to make more rational decisions about the use of conventional chemo- and radiotherapy.

Chemotherapy has a colourful, if unfortunate, history. Its origins go back to World War I, when the Germans used mustard gas in the trenches of Europe to devastating effect. The use of chemical
weapons was banned by the Geneva Protocol of 1925, but not the possession of such weapons, and the Americans continued to develop and stockpile them. In December 1943, a US cargo ship, the SS
Harvey
, secretly carrying mustard-gas bombs to the Mediterranean war front, was sunk in a German raid on the port of Bari, southern Italy, and a cloud of gas drifted over the city. No one
knows how many civilians were affected, but more
than 600 military personnel were hospitalised and 83 died. During autopsies of the victims, pathologists found evidence that
the normally fast-dividing cells of the bone marrow and lymphoid tissues had been suppressed. From this observation came the idea that perhaps such an agent could be used to attack the rapidly
dividing cells of cancer.

Soon scientists were doing experiments with mustard gas in mice. Encouraged by the results, they moved cautiously on to testing the agent in humans. The first human subject was a patient with
lymphoma – cancer of the lymphoid tissue – and his doctors observed with delight the dramatic shrinkage of his tumours after administration of the drug. Unfortunately the effect was
short-lived, but it galvanised the cancer community: here at last was a new way of treating the disease. For many centuries, surgery had been the only option for getting rid of tumours, and
patients’ long-term survival chances were minimal.

Over the decades since, many different cytotoxic drugs have been developed – all on the same principle, that they are poisonous to cells. But while chemotherapy has been found to work
wonderfully well in some tumours, it does not work at all in others. And in some it works for a while and then stops. Why is the response so varied? Until p53 research began offering clues, no one
had an answer. Today we know that both chemo- and radiotherapy work not by killing cancer cells directly, in a sledgehammer kind of way as had been assumed, but much more subtly: typically, these
therapies work by inducing cancer cells to commit suicide in response to damage of their DNA – the normal response to cell stress, mediated by p53.

Scott Lowe, whom we met in
Chapter 12
creating mouse models and making groundbreaking discoveries about apoptosis and p53, was one of the first to recognise the tumour suppressor’s central
role in conventional therapy. To recap, Lowe subjected the highly sensitive thymus glands of his mice to radiation and discovered that the cells with
normal, functioning p53
died very quickly by apoptosis, but cells with mutant or no p53 were resistant to radiation and survived. Confirming p53’s role in apoptosis in response to radiation set Lowe wondering more
generally: could p53 be responsible for the effect of radiation – and perhaps cytotoxic drugs also – in cancer therapy? As an idea, it was incredibly simple, and so obvious in
retrospect, but revolutionary at the time.

‘Here was a situation where the hypothesis was that if p53 is mutant, the standard chemotherapy drugs are less likely to work,’ explained Lowe. ‘In the case of leukaemias and
lymphomas what we would have predicted holds true. But now, 17 years of subsequent research says of course it’s more complicated than that.’

In leukaemia and lymphoma cells, p53 is almost always normal and, as one would expect, these cancers are highly sensitive to chemo- and radiotherapy. But in solid tumours (cancers of the organs
rather than the blood), the picture is much less predictable – and sometimes it is counter-intuitive. In some types of cancer, cells with mutant p53 are more responsive to cytotoxic drugs
than are cells with normal p53. This is the case with glioblastoma, an aggressive tumour of the brain, for example. So what is going on?

One explanation is that in these cases, the cells with mutant p53 are indeed killed in the sledgehammer way oncologists originally imagined. They sustain severe damage to their DNA that cannot
be repaired because p53 is out of action, nor can cell division be arrested. The cells carry on chaotically through the cycle and eventually succumb to what is called ‘mitotic
catastrophe’ – wholesale failure of the machinery of replication. This scenario implies that it is essential for oncologists to know which way a tumour type will react to conventional
therapy, depending on its p53 status. But things can get even more complicated.

In some cases, giving chemo- or radiotherapy to patients whose cancers have
normal
p53 can actually make things
worse. Cell death, as we know, is just one of
several options chosen by p53 in response to damaged DNA. It can also choose to arrest the cell mid-cycle and send in the repair team before releasing the cell to carry on replication. Or it can
condemn the cell to senescence – permanent arrest, which we know from the chapter on ageing can eventually stimulate cancer in neighbouring cells. Thus cancer cells that are not killed by
chemo- or radiotherapy can be the seed stock for further tumours – and sometimes these new tumours are especially aggressive simply because the cells are survivors of highly toxic treatment,
and bred for resistance.