I come from a family in which there have been no scientists or doctors. I was interested,
however, in biology at school and started my scientific career by training in medicine
at Oxford University and Guys Hospital, London. Practising as a doctor reinforced
my curiosity about the biological processes underlying human disease. As a consequence,
I pursued a clinical vocation in histopathology, a discipline that couples exposure
to the sights and smells of the autopsy room with a daily journey into the often beautiful,
sometimes ugly world of healthy and diseased human tissues under the microscope. After
an introduction to general histopathology in Nick Wright's department at the Hammersmith
Hospital, London, I completed my postgraduate medical training in neuropathology with
Peter Lantos at the Maudsley Hospital, London.
»Peering at the nuclei of cancer cells under the microscope, for me it was a matter
of fascination that hidden within them were the key events converting normal cells
into cancer cells, and frustration because they were out of reach.«
Many of the tissue samples examined by pathologists are from cancers. The clonal theory
of cancer development and the general role of DNA mutations in generating cancer cell
clones had been established by 1986 when I was working as a junior pathologist. Indeed,
the first mutated cancer gene, HRAS, had recently been identified through application
of the, then new, technologies of recombinant DNA technology. Peering at the nuclei
of cancer cells under the microscope, for me it was a matter of fascination that hidden
within them were the key events converting normal cells into cancer cells, and frustration
because they were out of reach.
So, I took 3 years break from medicine to study for a PhD, learning the methods and
thinking of molecular oncology in Colin Cooper's laboratory at the Institute of Cancer
Research, London. Many Southern blots later I had experienced the joys and insights
brought by detecting abnormalities of DNA in cancer cells. Essentially similar experiments
characterize the science I have done since, at increasing scale, using different strategies
and technologies, but with the same underlying aim.
On the trail of cancer susceptibility genes
Having accredited as a pathologist, I returned to the Institute of Cancer Research
as a Group Leader, with the intent of developing studies on the genetics of breast
cancer susceptibility in collaboration with epidemiologists Doug Easton and Julian
Peto. It had been recognized for many years that breast cancer cases clustered in
some families. When I started, the field had just been set alight by the discovery,
through genetic linkage analysis of such families, of the genomic location of the
first high risk (10- to 20-fold) breast cancer susceptibility gene, BRCA1, on chromosome
17q. This discovery was a magnet for many of the major international groups in human
disease genetics to engage in a highly publicised cloning race typical of the era
to identify BRCA1. My fledgling team was certainly not in that league, and anyway,
the job was clearly being done. Instead, I decided to explore the possibility that
a further high-risk breast cancer susceptibility gene existed, although the evidence
for this was by no means definitive. Nevertheless, after a couple of years acquiring
breast cancer families that did not seem to be due to BRCA1, Richard Wooster (then
a post doc with me) and the group embarked on another genome-wide search by genetic
linkage analysis (in collaboration with David Goldgar and others) and ultimately located
BRCA2 in 1994 to chromosome 13q, in the process of course, also proving that the second
gene existed (Wooster et al, 1994).
The next stage was to identify BRCA2 itself. In this undertaking, we ourselves were
now ensnared in a cloning race because we disagreed with the gene patenting and monopolization
policies of Myriad Genetics, a biotechnology company from Utah, who became our competitors.
Although the outlook initially did not look optimistic, we forged a collaboration
with many participants, notably including Alan Ashworth, Andy Futreal and David Bentley
(at the then Sanger Centre) and won the race at the end of 1995 (Wooster et al, 1995).
Since then, analysis of BRCA2 has entered routine clinical genetics practice to diagnose
women at high risk of developing breast cancer.
Following the identification of BRCA2, we set off on a search for yet another high
risk breast cancer gene, ‘BRCA3’. Unfortunately, this transpired to be a fruitless
endeavour and the general conclusion now is that only two such genes exist. Nevertheless,
a few years later with Nazneen Rahman, my erstwhile PhD student who was now leading
the breast cancer genetics group at the Institute of Cancer Research, we started systematically
sequencing candidate genes that were part of the DNA damage and repair pathways that
include BRCA1/2 and identified a number of intermediate risk (two- to fourfold) breast
cancer susceptibility genes including CHEK2, ATM, BRIP1 and PALB2.
Towards the end of the last millennium, it was becoming apparent that most high-risk
cancer susceptibility genes had been found. The era of genome-wide association studies
to look for low risk susceptibility alleles was not yet upon us. Anyway, to me it
felt like that enterprise would be more the domain of statisticians and epidemiologists
than scientists with my background.
Entering the uncharted jungle of cancer DNA
All cancers, however, are thought to arise through somatically acquired mutations.
During the course of 1998–1999, in discussions with Richard Wooster and then Andy
Futreal, the notion developed of using the reference human genome sequence, which
was rapidly emerging from sequencing machines around the world, as a template against
which we could sequence genome-wide for somatic mutations in cancer. It was clear
that sequencing technology was not yet fit-for-purpose to do this, but perhaps the
time had come to make a start.
Whichever way one looked at it, this experiment was big, full of risks, largely unpiloted
and expensive. Not the sort of description that finds favour with most funding bodies.
The Wellcome Trust, however, has a different approach to such matters. The Trust was
already funding the sequencing of one-third of the reference human genome sequence
and is accustomed to, indeed, has an appetite for, large-scale expeditions into the
scientific darkness. Our Cancer Genome Project was approved for funding and started
work in 2000 at the Wellcome Trust's genome facility, the Sanger Institute, near Cambridge,
UK.
We started by sequencing coding exons in cancer genomes through PCR amplification
and conventional Sanger sequencing. The amounts of DNA required were substantial,
so we had to restrict ourselves to cancer cell lines, from which large quantities
could be made. Unfortunately, very few of these lines had the available normal tissue
DNA sample from the same individual that was necessary for us to call somatic mutations.
By necessity, therefore, we embarked using a somewhat ad hoc assortment of 20–30 cancers,
a few breast, a few lung, a few melanoma and odd examples of additional types. With
the available technology, we were unable to plough through large numbers of genes
and therefore assembled a shortlist with which to start. These genes were from pathways
in which a gene was already known to be mutated and implicated in cancer development
and/or genes encoding protein kinases, since the recent success of the targeted drug
imatinib against the rearranged BCR-ABL protein in chronic myeloid leukaemia dramatically
demonstrated how mutated kinases could be tractable drug targets.
Occasional jewels among the sands of random mutations
No more than a few weeks into this early screen we began to see somatic mutations
in the BRAF gene. BRAF encodes a cytoplasmic serine threonine protein kinase that
is part of the well-studied RAS-RAF-MEK-ERK-MAP kinase signalling pathway. Further
work demonstrated that BRAF mutations are present in approximately 60% malignant melanoma,
15% colorectal cancer, 30% papillary thyroid cancer and others. Biological studies
conducted by Richard Marais and Chris Marshall confirmed that the mutations usually
activate the BRAF kinase conferring transforming activity upon it (Davies et al, 2002).
Metastatic malignant melanoma is generally a remorseless disease unresponsive to conventional
chemotherapy or radiotherapy. However, the nature of BRAF and its mutations recommended
it as a drug target. In work conducted by others over the subsequent decade, small
molecule inhibitors of mutated BRAF have been derived and shown to be effective in
patients with metastatic malignant melanoma. Although these remarkable responses represent
a major advance in treatment of the disease, resistant clones generally emerge and
patients still succumb. Therefore, this is the beginning, rather than the end, of
the story of a new approach to treating malignant melanoma (Flaherty et al, 2010).
Meanwhile, back at the cancer genome, we were able to implement PCR-based conventional
Sanger sequencing of exons to much higher throughput and apply it to primary cancer
samples. This provided us with a bird's eye view of somatic mutations in cancer genomes.
A small minority are ‘driver’ mutations in cancer genes, which convert normal cells
into cancer cells, while the large majority are ‘passengers’ (Fig 1). A small number
of cancer genes are mutated frequently, but many appear to contribute infrequently
to cancer development (Greenman et al, 2007).
Figure 1
The cellular lineage between a fertilized egg and a fully malignant cancer cell
Coloured symbols represent the accumulation of somatic mutations over a lifetime.
The number of driver mutations reflects the number of biological processes that need
to be subverted to convert a normal cell into a cancer cell. The genes in which driver
mutations occur (cancer genes) can present tractable targets for new drug development.
The number of passenger mutations reflects the number of mitoses between the fertilized
egg and the cancer cell and the mutation rate at each mitosis. Passenger mutations
provide insights into the underlying mutational processes operative in each case.
The tangled and tattered strands of cancer DNA revealed
The advent of next generation sequencing technologies around 2007 transformed our
studies, and the field in general. Using these approaches, with Peter Campbell joining
us at Sanger, we have been able to explore cancer genomes at sequence-level resolution
revealing their extraordinarily contorted architecture (Fig 2; Stephens et al, 2009),
producing essentially complete catalogues of somatic mutations from individual human
cancers (Pleasance et al, 2010) and yielding many new mutated cancer genes (Stephens
et al, 2012).
Figure 2
The genome-wide rearrangements in six breast cancer genomes (three cell lines, top
and three primary tumours, below)
In each case, the genome is represented as a circle and the lines represent somatic
rearrangements, green are intrachromosomal and purple interchromosomal. First published
in Stephens et al (2009).
In an initiative reminiscent of the original Human Genome Project, with colleagues
worldwide, the International Cancer Genome Consortium was constituted to coordinate
the burgeoning sequencing activities across the range of human cancer types. Mutated
in the appropriate manner, approximately 500 of the ∼20,000 protein coding genes in
the human genome now appear to be causally implicated in the genesis of one or other
of the 100–200 types of cancer. Given this diversity, enabling new drugs to be assessed,
prior to starting clinical trials, for the cancer class and genome configuration that
is most sensitive would be advantageous. With colleagues at the Massachusetts General
Hospital, this has been set in train using 1000 genomically characterized cancer cell
lines (Garnett et al, 2012).
Exploring cancer genomes continues to provide new intriguing dimensions of insight.
Recently, we have shown that multiple underlying somatic mutational processes are
operative in cancer, each of which can leave its own distinctive mutational signature
on the genome (Nik-Zainal et al, 2012). Some may be due to exogenous exposures, others
to abnormalities of DNA maintenance. Some operate genome-wide, others are targeted
to small regions of the genome. Furthermore, it has allowed detailed analysis of the
subclonal evolution of cancers, both within the primary cancer and in metastasis formation.
Cancer genomes entering the clinic
Cancer genomics is already established in the clinical management of patients, as
tests for mutations in certain genes, for example EGFR, BRAF and HER2, are required
before drugs targeting the encoded protein can be prescribed. It is likely that this
position will consolidate further and it is not unreasonable to speculate that in
a decade whole cancer genome sequences may be routine for patients requiring cancer
treatment. Over the next few years, the complete repertoire of mutated cancer genes
across the spectrum of cancer types will be identified. Some are promising direct
targets for new therapeutics and novel drugs are likely to emerge quickly. Others
are not as tractable. However, they remain potential Achilles' heels of cancer cells
and researchers will be exploring ways to somehow exploit them to develop new therapies.
At the same time, deeper understanding of the mechanisms underlying the processes
generating somatic mutations will lead to new insights into cancer causation and,
potentially, new preventive strategies.