p53 (20 page)

‘We started the database because we had this idea that there was going to be a relationship between environmental causes of cancer and the p53 mutation spectrum,’ Harris told me. And
indeed to the disease detectives – the molecular epidemiologists – the database has proved invaluable. Its role in the tobacco story is one of its most notable successes.

TOBACCO’S FINGERPRINT FOUND ON p53

In the early 1990s, Gerd Pfeifer, who heads a lab at the City of Hope medical centre in Duarte, California, was exploring DNA damage in relation to cancer, looking to see if
the causative agents left distinctive patterns of damage, or ‘fingerprints’, that identified them as culprits. His lab had developed a tool that enabled the scientists to home in
on individual genes among the thousands along a strand of DNA – a process akin to finding needles in haystacks – and the general buzz surrounding p53 at the time
persuaded Pfeifer and his fellow researcher, Mikhail Denissenko, that this gene would be an interesting focus for their studies. They would look at the effects of tobacco smoke on p53.

‘The early tobacco work had clearly suggested there is high tumour-causing activity in the tar – the black stuff you can collect on filters when you burn tobacco, and that you can
see in the lungs of heavy smokers when you operate on them. It looks gross,’ Pfeifer commented when I phoned him in his lab. He and Denissenko knew that the most damaging components of tar
are the polycyclic aromatic hydrocarbons, or PAHs, of which one, benzo(a)pyrene (BaP), is particularly noxious. This, they decided, would be an ideal damaging agent to use in their experiments.

Others had already investigated what happens to PAHs when they get inside cells. Their research showed that these substances are not water soluble, so the body has difficulty ridding itself of
them. In an effort to transform the compounds into something that can then be excreted, the machinery of the cells creates a dangerously reactive substance that sticks itself to the DNA. In the
case of BaP, this transformed substance has a mind-numbing formula represented simply as BPDE, which is considered to be one of the most potent carcinogens yet discovered.

Pfeifer and Denissenko took BPDE and added it to various cell types, including human lung, and then left the cells to their fate. After an hour or two they returned to their lab benches to
isolate the DNA and to apply their special technique to identifying the exact positions on the p53 gene that had sustained damage. This, they discovered, was not random. BPDE always attached itself
to the DNA next to the guanine base – one of the four basic building blocks of DNA, represented by G in the DNA code – at three very specific ‘hot spots’ along the gene, at
codons 157, 248 and
273 (to recap, a codon is a segment of a gene just three bases long that codes for one of the building blocks of the protein, and the codon’s
number defines where that building block should go).

Here at last was proof that a defined product of cigarette smoke damages DNA. But the clincher for the case against Big Tobacco came when Pfeifer and Denissenko compared their lab’s
results with the p53 mutation database, which by 1996 had more than 500 entries for lung cancer drawn from the literature worldwide. The great majority of the mutations described in the database
among smokers, but rarely among non-smokers, corresponded precisely with their results: they occurred in the same hot spots targeted by the BPDE and they showed that the building blocks of the gene
had been scrambled such that the guanine (G) was replaced by a thymine (T). Crucially, while codons 248 and 273 are mutation hot spots in many types of cancer, the database revealed that codon 157
is found exclusively in lung tumours. In other words the fingerprint of BPDE was all over the p53 database. Pfeifer and Denissenko’s paper, published in
Science
in October 1996,
concluded, ‘Our study thus provides a direct link between a defined cigarette smoke carcinogen and human cancer mutations.’

BIG TOBACCO QUESTIONS THE SCIENCE

This was bad news for Big Tobacco. It meant not just that smoking was implicated in a generalised threat to public health, but that it could be linked to individual people with
lung cancer. Tobacco companies were now much more vulnerable to lawsuits from customers looking for compensation for ruined lives and, as ever, they set about trying to refute the evidence. In an
address to investors, analysts and journalists soon after the
Science
paper came out, the Chief Executive of British American Tobacco Industries (BAT), Martin Broughton, stated,
‘There is still a lack
of understanding of the mechanisms of disease attributed to smoking . . . The importance of this
Science
magazine study may lie, not
least, in the recognition that there are important missing links in the understanding of causation . . . It may lead to further research . . . into the complex process by which a cell becomes
cancerous – a process we and others have spent millions in trying to understand for many years now.’

The R J Reynolds tobacco company was even more blatantly dismissive. A public statement issued by the company said, ‘That BaP will cause a mutation has been known for a long time . . . The
authors themselves describe these findings as a coincidence. The press release’s conclusion that these findings are the key to lung cancer is an overstatement.’

Interestingly, R J Reynolds’ statement came out on the eve of publication of Pfeifer and Denissenko’s study being published in
Science
– clearly suggesting the company
had been tipped off in advance.

The following year, Pierre Hainaut and a colleague at IARC, Tina Hernandez-Boussard, carried out a detailed analysis of the spectrum of p53 mutations in smokers as recorded in their database.
Their paper for
Environmental Health Perspectives
came to the same conclusion as Pfeifer and Denissenko’s: that these mutations carried the fingerprint of BPDE. ‘We thought at
the time, that settles it,’ said Hainaut as we sat together in the front room of his home in Lyon, with its distant views of the snow-covered Alps, talking about the database’s role in
revealing the causes of cancer. ‘You have experimental data; you know what the mutagenic substance is; you can demonstrate its effect very well in the lab and you can show that the people who
are exposed to the same substances in real life get the mutation at exactly the same place.’

Hainaut and Pfeifer believed their two papers made an irrefutable case, and they were taken aback two years later
to see first one and then a second paper challenging
their results as being ‘over-interpretations’. ‘People had done some analysis themselves of our database to try to prove that we were wrong – that the connection was not
there; that maybe tobacco was
helping
mutations occur, but not actually causing them,’ explained Hainaut. ‘I was shocked. But also I have to say that we’re used to trust
within the scientific community, and you expect people to be fair. So the first reaction when you see a paper like this attacking your work is, oh my God, I’ve missed something very
important! I’ve made a big mistake! In fact when the first of these papers came out my director called me into his office and said, “What’s this about? You have three days to
bring all your data to me for review, because if you’ve mishandled or misrepresented an issue like that it’s a serious matter.” So I was in the hot seat somehow!’

To Hainaut’s relief, his interpretation stood up to review and he began to wonder about the author of the second paper, published in
Mutagenesis
– a scientist named Thilo
Paschke, working for a Munich-based institute, Analytisch-Biologisches Forschungslabor. At this point in our conversation, Hainaut disappeared into his study and returned with a sheaf of yellowing
papers held together in a clear plastic-covered folder – a record of the murky drama he found himself being dragged into. ‘I had never heard of this guy, never seen a paper by him and I
had no idea about the institute he worked for,’ he continued. ‘So I tried looking up the institute on the internet and I didn’t find any website. All I had was an address in
Munich, so I called the German telecommunications people and they told me, “It’s the address of the German Association of Tobacco Manufacturers.”

‘My first reaction was relief. I realised, okay, that’s a completely unfair attack on our work and I can probably forget it. All I have to do is demonstrate that it comes from people
who have a bias they haven’t declared.’

But then the plot began to thicken. As part of the general settlement of lawsuits between the American States and Big Tobacco in 1998, the industry was required to make
publicly available all internal documents used during the trial. These were organised systematically and posted on the internet at
www.tobaccodocuments.org
. In early 2001, Hainaut opened his computer at the website in search of information about the journal
Mutagenesis
, whose
behaviour he and Pfeifer had found ‘strange’ when they raised the issue of Paschke’s conflict of interests and asked for space to respond to his critical paper. Their request had
been turned down and, intrigued to know if anything suspicious lay behind the rejection, Hainaut typed the name of the journal’s founding editor, Jim Parry, into the site’s search
engine and watched wide-eyed as a series of references popped up on screen.

Here was evidence of research and consultancy contracts between Parry and BAT and Philip Morris running over more than a decade. To the fascinated Hainaut, a letter dated 3rd August 1994, from
Parry to his main contact at Philip Morris, the company’s chief scientist Ruth Dempsey, about a proposed research project was particularly revealing. In it Parry, then a Professor of
Biochemistry at the University of Wales at Swansea, advised Dempsey that ‘the overhead figure of 40 per cent I have given in the costs can be overcome if money is given to me as a gift,
“as a contribution to my research” without specifying how it should be spent. My colleagues tell me that some companies are increasingly appreciating the financial advantage of giving
money in this way compared to contracts which specify components of the project.’

Parry ended his letter, ‘Please let me know if your people are interested in supporting the type of work outlined, bearing in mind that we can modify the components in various ways to also
fit in with your interests.’

Curt Harris, original founder of the database and by now editor of the journal
Carcinogenesis,
had been taking a
personal interest in the controversy. Faced with
such clear evidence of a conflict of interest, he blew the whistle on Parry, editor of
Mutagenesis,
with Oxford University Press, which publishes both journals. Although she understood
full well the seriousness of the allegation against Parry, Janet Boullin, the editorial director of journals at OUP, pointed out that she had limited scope for action since OUP was simply the
publisher, not the owner, of
Mutagenesis.
But she immediately strengthened the rules about disclosure of interests for both contributors and editors of all OUP journals. Parry was not
prepared to comply with the new rules and he resigned as editor of
Mutagenesis
soon afterwards. But he remained on the journal’s editorial board, whose members were not required to
sign competing-interest statements.

Among the documents on the tobacco industry website, Hainaut found evidence, too, that Philip Morris was gathering intelligence on his own institution, IARC, home of the p53 database. However,
satisfied for the time being with what he had learnt from his own sleuthing, he handed the evidence he had gathered over to Stanton Glantz, an anti-tobacco campaigner in the US.

SCIENTISTS FIGHT BACK

Stanton Glantz, a heart specialist, is Professor of Medicine at the University of California, San Francisco, director of the Center for Tobacco Control and Education, and a
scientist with a long history of standing up to the tobacco industry. Originally, his focus was passive smoking, which kills tens of thousands of non-smokers in America alone every year. But his
efforts to have smoking banned in public spaces have seen him hassled constantly by industry supporters. ‘After every barrage of personal attacks against me in the
American Smoker’s
Journal
or the American Smoker’s Alliance [set up by Philip Morris and others] or these other publications from these groups, I get a series of hate mail,’ he told
the Public Broadcasting Service (PBS) in an interview for their programme
Frontline
in 1996. ‘I have had hate mail, hate faxes, hate e-mail, hate phone calls. I mean
the police department here intercepts some of my mail now, and it’s a drag.’

Turning up for work on 12th May, 1994, Glantz found dumped on his desk a large box containing several thousand pages of confidential internal documents from the tobacco company Brown and
Williamson, a subsidiary of BAT. There was no clue as to who had sent the parcel, simply the name ‘Mr Butts’ on the ‘return address’ label. The documents covered a period
from the early 1950s to the mid-1980s, and related to such issues as the addictiveness of nicotine, cancer and the company’s public relations and legal strategies. Since these were not
directly relevant to his own research, Glantz intended passing the documents on to a colleague working in these areas. But after flicking through them for 20 minutes or so he found himself unable
to stop reading. ‘The thing that sucked me into them was not their potential political or legal import,’ he told PBS. ‘It was the documents as history, the documents as science.
It was just an unbelievable find. As a professor, it would be like an archaeologist finding a new tomb in Egypt or something . . . I mean it’s an amazing, amazing story of what was going on
inside these cigarette companies during this crucial period.’

Glantz hung on to the documents and worked through them systematically with a team of reviewers, writing a number of papers for journals before pulling them together, in collaboration with
colleagues, into a book,
The Cigarette Papers,
published by the University of California Press in 1998. He remained interested in the tobacco industry and its subversive activities, and a
few years later began investigating its involvement with p53, adding further detail to what Hainaut had discovered and writing up the story in an explosive paper for
The Lancet.