One Biopsy Not Enough to Reveal Genetic Landscape of a Tumor

Zosia Chustecka

March 08, 2012

March 8, 2012 — A small study that shows a surprising complexity of genetic changes within a single tumor has far-reaching implications for the march toward personalized cancer therapy, according to researchers.

A single biopsy from a tumor might not be sufficient to give a full picture of its genetic landscape, a team from the United Kingdom reports.

Dr. Charles Swanton

When the researchers examined 10 biopsies taken from a single kidney cancer, they found "an extraordinary amount of diversity" in the genetic changes that had taken place in different parts of the tumor. "There were more differences between biopsies from the same tumor at the genetic level than there were similarities," said senior author Charles Swanton, MD, PhD, from the Cancer Research UK, London Institute, and the University College London, United Kingdom.

The findings, published in the March 7 issue of the New England Journal of Medicine, were highlighted at a London press conference organized by Cancer Research UK, which funded the study.

The team also found differences in genetic changes between the primary tumor and metastases that developed both locally and at a distant site (in the chest wall). Similar findings have been documented by other research groups. Specifically, marked changes have been demonstrated between the primary tumor and metastases in breast cancer, which have led to calls for biopsies to be taken of the metastases.

What is surprising in this study is the extent of intratumoral heterogeneity, the researchers note.

Far-Reaching Implications

The findings have far-reaching implications for the efforts currently being directed toward personalized cancer therapy, in which therapy is targeted at genetic changes identified in tumor tissue. An example of this is the SnaPshot broad genetic screen program being used in routine clinical practice at the Massachusetts General Hospital in Boston.

If you take only 1 biopsy, you could be misled clinically.

However, Dr. Swanton cautioned that "if you take only 1 biopsy, you could be misled clinically."

"The simple view of directing therapy on the basis of genetic tumor markers is probably too simple," agrees Dan Longo, MD, deputy editor of the New England Journal of Medicine, in an accompanying editorial.

"A whole new world has been anticipated in which patients will undergo a needle biopsy of a tumor in an outpatient clinic, and a little while later, an active treatment will be devised for each patient on the basis of the distinctive genetic characteristics of the tumor," Dr. Longo writes.

However, a serious flaw in this imagined future of oncology is the underestimation of tumor heterogeneity, he notes. "Not just heterogeneity between tumors, which is a central feature of the new image of personalized medicine, but heterogeneity within an individual tumor," he adds.

Differences Within in a Single Tumor

The study evolved out of research into kidney cancer and the E-PREDICT trial, which was aiming to find a biomarker that would predict response to everolimus (Afinitor, Novartis), explained coauthor James Larkin, MD, PhD, consultant medical oncologist at the Royal Marsden Hospital in Surrey, United Kingdom. In that trial, patients underwent a 6-week course of treatment with everolimus; after a week-long washout period, they underwent a nephrectomy. Several biopsies were taken from the primary tumor and from metastases both before and after treatment with everolimus.

The team analyzed the cancer genome in biopsy samples taken from 4 patients. For 2 patients, the analysis involved whole-exome multiregional spatial sequencing of DNA extracted from the biopsy samples; for the other 2 patients, a less expensive process was used, Dr. Swanton explained.

In total, 30 biopsies were taken from 4 primary tumors, and the genome analysis revealed that 26 of these 30 biopsies were different, he said.

In total, 118 different genetic mutations were identified. Some of these were "ubiquitous" mutations (n = 40), which were found in all of the biopsies from the primary tumor and from the metastases, but there were also some shared mutations (n = 53), which were found only in some primary tumors and/or only in some of the metastases. In addition, the researchers found some mutations that they described as "private mutations" (n = 25), which were found only in a single biopsy.

The researchers also noted that gene-expression signatures associated with a good prognosis and those associated with a bad prognosis were detected in different regions of the same tumor.

Evolutionary Changes

When the researchers modeled the genetic changes, they found an evolution pattern that resembles the trunk and branches of a tree. The tumor began with a number of genetic changes that developed early on in the "trunk"; over time, different groups of cells evolved different genetic changes and formed different "branches" of the cancer's evolutionary tree.

Another key finding was that different regions of the tumors had different faults in the same genes, corresponding to separate evolutionary changes (as occurs in convergent evolution).

At the press conference, Dr. Swanton likened this evolutionary tree pattern of genetic changes within the tumor to Darwin's descriptions of the evolution of species, and showed his team's diagrams alongside a diagram from Darwin's notes — both showed similar tree patterns with branches.

Evolutionary tree patterns of genetic changes within the tumor (left) and Darwin's description of the evolution of species (right). Source: Charles Swanton


"For the first time, we've been able to use the pattern of genetic faults in a tumor to trace the origins of certain populations of cancer cells in much the same way that Darwin used his 'tree of life' theory to show how different species are related," Dr. Swanton explained.

This theory is in stark contrast to current cancer theories being taught in medical schools, Dr. Swanton said. A mainstay of the medical school curriculum is that cancer is a clonal disease that evolves in a linear fashion, with mutations arising in a sequential fashion. "This is guiding our research, but this theory is driven by a single biopsy from a tumor," he said.

However, this revolutionary way of thinking about cancer is based on a very small sample size. "We only analyzed 4 patients," Dr. Swanton acknowledged. More work is needed, and is already underway as part of Cancer Research UK's Genomics Initiative.

Evolutionary Trees

Nevertheless, the theory could explain a number of clinical observations, Dr. Swanton said. Targeted agents that have already succeeded in the treatment of cancer could be acting on genetic mutations that occur early in the evolution — in the trunk of the tree, he explained. Examples of such agents are trastuzumab (Herceptin) for HER2-positive breast cancer and the EGFR inhibitors erlotinib (Tarceva) and gefitinib (Iressa) in the treatment of nonsmall-cell lung cancer.

Dr. Swanton is concerned that some of the newer drugs targeting new mutations will be acting only on the branches, so will not have a big impact on the cancer. "Just because a mutation is there doesn't mean that you are going to see a robust response when you target it," he told Medscape Medical News.

Also, different cancers could be imagined as having different tree shapes. Some would have a long trunk and short branches, like a palm tree; these could be dealt a deadly blow by a hit to the trunk, perhaps like vemurafenib (Zelboraf) acting on BRAF mutations in some cases of melanoma. "It may also explain why surgery to remove the primary kidney tumor can improve survival by decreasing the likelihood that resistant cells will...regrow the tumor after treatment," Dr. Swanton said.

Other cancers would have a very short trunk and many branches, perhaps like pancreatic cancer, which is notoriously difficult to treat and has not responded well to any targeted therapies.

"This research is going to change the way that we analyze cancers," predicted Peter Johnson, MD, chief clinician at Cancer Research UK, who also spoke at the press conference. "We are clearly only at the beginning of this process." The findings highlight the extreme complexity of cancer genetics, but they also point to a way forward despite this complexity, he said. "They suggest a way to improve the success rate of personalized medicine."

Impact on Personalized Medicine?

Approached for comment, James Hsieh, MD, PhD, from the Memorial Sloan-Kettering Cancer Center in New York City, told Medscape Medical News that "after reading this exciting paper, we should take a deep breath and think again what we can do to make personalized medicine better, focusing on drivers, assistant drivers, and supporters of individual cancer. It is not going to be easy."

"The existence of diverse genetic blueprints in the primary and metastatic kidney cancer within the same patient, as demonstrated by this study, seems to have greatly dampened the hope for personalized medicine at first glance," Dr. Hsieh said.

"However, the idea of intratumor heterogeneity is not new and can be commonly observed in many different cancers simply under the microscope (e.g., clear cell renal cell carcinoma with sarcomatoid changes).... Should we be discouraged, or can we take this knowledge into consideration to improve the prospect of personalized medicine?"

Dr. Hsieh believes that the latter is the case. He noted that this "elegant study" has taught several lessons. First, there is ongoing intratumor evolution giving rise to multiple cancer clones, some of which have the capacity to metastasize. Second, a single biopsy of the tumor might miss mutations that have clinical significance.

Extrapolating to a clinical scenario, Dr. Hsieh focused on kidney cancers, with which he has extensive clinical experience.

For patients with primary kidney cancer who have no evidence of metastasis at presentation, nephrectomy alone cures a large percentage. Therefore, extensive biopsy of multiple sites for whole-exome sequencing might not be necessary, he explained. "Having said that, this multiregional approach might help predict which patients will later develop metastasis, under the premise that we know the critical subset of genes involved in kidney cancer metastasis. For these patients, we may wish to initiate systemic treatment in addition to surgery before the full manifestation of frank metastasis."

However, for patients presenting with metastatic disease, either with the primary kidney cancer in place or after nephrectomy, biopsy and extensive study of metastatic sites could be high yield, Dr. Hsieh said. Cases presented in this study indicate that cancers at different metastatic sites of the same patient carry shared mutations and are likely evolved from the same original clone. Therefore, biopsy and analysis of one metastatic site is likely to provide pertinent genetic information on the other sites for these treatment-naïve patients. However, clonal evolution constantly takes place, leading to the mixed response that we commonly witness when patients are on targeted treatment. When this happens, biopsy and analysis of the nonresponsive tumors likely provide the most critical information for individual patients.

As for treatment strategies, what we need to focus on are the so-called driver mutations, not the passenger mutations that are likely the private mutations described in this paper. Driver mutations underscore the success of targeted therapy. These key drivers are present at the trunk and common branches of the individual tumor evolution tree, and should serve as targets. When key drivers are being eliminated, disguised drivers (drivers with drug-resistant mutations) or assistant drivers (collateral players or pathways) might emerge and jeopardize the treatment response. Finding these drivers by studying resistant tumors will certainly guide the refinement of targeted therapy, Dr. Hsieh noted.

Dr. Swanton and several of his coauthors report receiving research grants from Novartis.

N Engl J Med. 2012;366:883-892,956-957. Abstract, Editorial


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