Using powerful DNA sequencing technology to decode the genomes of cancer patients, scientists at Washington University School of Medicine in St. Louis are getting an unprecedented look at the genetic basis of a highly lethal breast cancer that disproportionately affects younger women and those who are African-American.
Scientists have decoded the entire genome — all the DNA — of a 44-year-old African-American woman with so-called “triple negative” breast cancer, along with the genomes of her breast tumor and a metastatic tumor that quickly developed in her brain. By comparing the DNA sequences of the three genomes, the scientists identified 20 genetic changes from a subset of cells in the woman’s breast tumor that likely played a role in the metastasis that killed her within months, they report April 15 in the journal Nature.
The research suggests that sequencing the complete genomes of cancer patients, as well as the genomes of their primary and metastatic tumors, can reveal clues to the way tumors spread and can set the stage for developing more effective drugs that target key genetic errors.
“We are getting an intimate look at the lethal spread of a breast cancer, which is now possible because we can sequence entire genomes quickly at a reasonable cost,” says senior author Elaine Mardis, PhD, co-director of Washington University’s Genome Center. “This work lays the foundation for understanding the genetic basis of tumor progression and metastasis and for identifying new drug targets that can improve the outlook for women with this disease.”
The patient, the first African-American to have her genome sequenced, had an aggressive, “basal-like” subtype of breast cancer. These genetically unstable tumors account for about 10 percent of all breast cancers and often are called triple-negative because they lack any receptors to which treatment can be targeted. They grow rapidly and typically do not respond well to chemotherapy.
That was the experience of the woman in this study. She initially received chemotherapy to shrink a large breast tumor and then had a mastectomy. When surgeons found evidence of treatment-resistant tumor cells in her lymph nodes, she received radiation therapy, but it did not halt the cancer. The tumor spread to the woman’s brain, and she died within a year of her diagnosis.
WUSTL researchers also sequenced the genome of the patient’s breast tumor after growing it in a mouse to determine whether the animal model can reproduce the genetic profile of the human cancer.
“The tumor grown in the mouse was remarkably similar to the patient’s own breast tumor,” Mardis says. “The human tumor and the mouse counterpart shared all of the same mutations and the same fateful course: Both spread to other sites.
"This similarity suggests that mouse models can be valid preclinical surrogates of metastatic disease to evaluate new cancer drugs,” Mardis says.
After decoding the genomes of the patient and her tumors, the researchers, led by Li Ding, PhD, research associate professor of genetics, painstakingly sifted through many billions of bases of DNA, identifying 48 single base-pair changes, or mutations, that were shared among the breast tumor, the metastatic tumor in the brain and the tumor grown in the mouse. Then, they looked to see how many cells in the breast tumor and brain metastasis carried each of these mutations.
Surprisingly, they found 20 mutations that occurred at relatively low levels in the patient’s breast tumor that also were present in a high percentage of the metastatic tumor cells, suggesting that these genetic alterations in the primary tumor likely drove the spread of the disease.
“This indicates that a small subset of cells with a lethal mutation repertoire break free from the primary tumor, circulate in the body, set up residence in other organs and grow aggressively,” says co-lead author Matthew Ellis, MD, PhD, the Anheuser-Busch Endowed Professor in Medical Oncology and a breast cancer specialist with the Siteman Cancer Center at Barnes-Jewish Hospital.
“Mutation enrichment is likely to be important for tumor spread since the mutation enrichment pattern in the brain metastasis and the tumor grown in the mouse strongly overlapped, indicating that the process was not random,” Ellis says.
For years, scientists studying cancer have asked whether metastasis is driven by mutations that occur after tumor cells arrive at a distant site or whether the primary tumor carries the genetic instructions that unleash the spread of the disease. The WUSTL researchers found that the metastatic tumor contained only two additional mutations that were not present in the patient’s breast tumor, as well as a missing chunk of DNA. But none of these genetic alterations appeared to be important to the cancer’s spread, they said.
They are now decoding the genomes of additional women with basal-like breast cancer to determine whether mutations in their primary tumors also influence the spread of the disease.
Whole genome sequencing involves spelling out all 3 billion letters of an individual’s DNA. In this study, the researchers sequenced each of the patient’s four DNA samples about 30 times to ensure accuracy.
Unlike sequencing studies that focus only on genes, which make up just 1 percent of the entire genome, whole genome sequencing captures the full breadth of genetic alterations in the DNA, including large insertions, deletions and other structural changes. Indeed, the researchers found numerous structural changes in the tumor genomes they analyzed, including 28 large deletions, six inversions and seven translocations, an indication of the “genomic chaos” underlying the patient’s cancer.
Researchers sequencing only genes miss many of these alterations because they either occur beyond the reach of genes in long stretches of non-coding DNA or in large chunks of missing DNA. For example, the investigators found two large, overlapping deletions in all three tumor samples that rendered both copies of a gene called CTNNA1 non-functional. Because both copies were knocked out, looking only at protein-coding genes would not have revealed the change. Interestingly, loss of this gene has been linked in previous studies to a loss of cell adhesion in human breast cancers, a precursor to metastasis.
The researchers also found a single mutation in the CSMD1 gene, which has been associated with poor survival in invasive ductal breast cancer and is frequently deleted in some colorectal and head and neck cancers. Alterations in the patient’s JAK2 and NRK genes also have been observed in other breast cancers.
The new research builds on earlier work by the same team of researchers, who less than two years ago became the first to sequence the entire genome of a cancer patient, a woman with leukemia. That research also was published in Nature. Last year, they reported sequencing the genome of a second leukemia patient in the New England Journal of Medicine.
Since then, the WUSTL scientists have decoded the normal and tumor genomes of about 150 cancer patients, including those with breast, lung and ovarian cancer as well as glioblastoma, a brain tumor. Some of this work is part of The Cancer Genome Atlas Project, funded by the National Human Genome Research Institute and the National Cancer Institute. The researchers also have begun sequencing the genomes of 600 pediatric cancer patients as part of a joint project with St. Jude Children’s Research Hospital.
"We've learned some significant lessons about cancer from sequencing the genomes of individual patients and their tumors, but it's clearly just the tip of the iceberg,” says Richard K. Wilson, PhD, director of The Genome Center and a co-author of the new research. “Moving forward, we'll be comparing tumor genomes from many patients with the same type of cancer to find common genetic alterations. This comprehensive understanding of cancer can aid in the development of new approaches to cancer diagnosis and treatment."
The research was funded by the National Human Genome Research Institute, the National Cancer Institute, the Susan G. Komen for the Cure®, the Fashion Footwear Charitable Foundation and Washington University’s Institute for Clinical and Translational Sciences.
Ding L, Ellis MJ, Weinstock GM, Aft R, Watson M, Ley TJ, Wilson RK, Mardis ER et al. Cancer remodeling in a basal-like breast cancer metastasis and xenograft. Nature. April 15, 2010.
Washington University School of Medicine's 2,100 employed and volunteer faculty physicians also are the medical staff of Barnes-Jewish and St. Louis Children's hospitals. The School of Medicine is one of the leading medical research, teaching and patient care institutions in the nation, currently ranked third in the nation by U.S. News & World Report. Through its affiliations with Barnes-Jewish and St. Louis Children's hospitals, the School of Medicine is linked to BJC HealthCare.
Siteman Cancer Center is the only NCI-designated Comprehensive Cancer Center within a 240-mile radius of St. Louis. Siteman Cancer Center is composed of the combined cancer research and treatment programs of Barnes-Jewish Hospital and Washington University School of Medicine.