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Case Study Illustrates Benefits, Complexities of NGS Panel Testing for Cancer Risk


NEW YORK (GenomeWeb) – A case study of a 38-year-old woman referred for genetic counseling due to a family history of breast cancer illustrates the benefits of screening multiple genes when estimating cancer risk.

Researchers from the Center for Genomic Medicine at the University of Copenhagen in Denmark described the case in a study published online last month in Familial Cancer.

A Pakistani woman had been referred to genetic counseling because her mother had been diagnosed with breast cancer at age 50 and again at age 52, and her grandmother had been diagnosed with breast cancer at age 60. Running a six-gene hereditary breast and ovarian cancer panel, the study authors identified likely pathogenic mutations in two genes, one of which would have been missed using standard BRCA1 and BRCA2 screening.

The laboratory offers the hereditary panel using Illumina's MiSeq for patients referred by a genetic counselor or oncologist. The test is covered by the government for patients in whom a genetic predisposition is suspected.

The University of Copenhagen's Thomas van Overeem Hansen, corresponding author of the study, told Clinical Sequencing News that the lab began incorporating next-gen sequencing for breast and ovarian cancer risk testing several years ago and is working with university hospital genetic counselors and oncologists to offer the testing. He said that the lab started small with just six genes, but would likely expand the offering in the coming months. Eventually, he anticipated that between 30 and 50 genes could be included in the panel.

In this recent case, sequencing identified two heterozygous mutations — a novel mutation in BRCA2 and a mutation in RAD51C that is involved in the Fanconi anemia pathway, an autosomal recessive disorder that causes hematological abnormalities. RAD51C is also involved in DNA repair by homologous recombination, and the mutation was a missense mutation at a highly conserved amino acid that has been shown to have functional consequences. Other pathogenic missense RAD51C mutations have been found in breast and ovarian cancer families.

Hansen noted that the RAD51C mutation was especially important because breast cancer predisposition testing has predominantly focused on just the BRCA1 and BRCA2 genes. But, there is "growing indication that RAD51C could be highly penetrant," Hansen said.

The BRCA2 mutation had not previously been seen, and Hansen said that the laboratory used a gene prediction tool, in silico analysis, and functional work to try and predict the mutation's consequence. Analysis found that the mutation led to a 46-base in-frame deletion that removed part of the oligonucleotide-binding fold 3 domain, which is in a region of BRCA2 that contains around 27 percent of tumor-derived missense mutations, "indicating the importance of this domain in tumor suppression," the authors wrote.

As such, they reported both mutations as likely pathogenic to the genetic counselor and oncologist, and recommended that her additional family members be tested.

In an ideal world, Hansen said that the laboratory would have also tested both of the patient's parents to determine whether the two mutations came from one parent, or if she received one from each. Additionally, further family testing could determine how the mutations segregate. However, the family members were living abroad in either Pakistan or Great Britain.

Although BRCA1 and BRCA2 are the most commonly tested genes, variants of unknown significance are still found, as illustrated in this case. Hansen said that especially because the patient was of Pakistani origin, the finding of a novel mutation in BRCA2 was not that surprising. The incidences of BRCA1 and BRCA2 mutations in non-European populations have not been characterized as deeply, Hansen said. And as NGS-based testing becomes more widespread, he anticipates that many more novel variants will be found in those genes, particularly in Asian and African populations.

Hansen added that dealing with variants of unknown significance is one of the biggest challenges of running the NGS cancer risk panel. Despite the fact that the lab looks at only six genes, he said that between 10 percent and 15 percent of the lab's cases have a variant of unknown significance.

Others have also reported on the challenges of dealing with variants of unknown significance, including several groups in the August issue of the Journal of Genetic Counseling.

Particularly when making risk predictions where preventative measures might be taken based on the results, clinicians want the results to be as clear as possible. Reporting back a variant of unknown significance is "the worst answer we can give," Hansen said.

He said that the lab will try to classify those variants first by doing in silico and structural studies. Then we "try to select the ones that are the most interesting to do functional studies." The lab does not have enough manpower to do functional studies on every VUS it identifies, Hansen said, so it tries to "select those that have a higher chance of being disease causing."

Hansen said that his lab works closely with the oncologists and genetic counselors and has weekly meetings to discuss cases and recommendations. If further work to understand a VUS leads to a classification of either likely pathogenic or likely benign, that information is reported back the clinician, he said.

Moving forward, he said the lab is taking a cautious approach in adding genes to its panel. "You need to have the geneticists and oncologists willing to use the data, because otherwise it's of no use," he said, except for research purposes. Over the next couple of months, he anticipated adding a few more genes that genetic counselors and oncologists have indicated they will use. The lab is focusing especially on genes that are in the homologous recombination pathway, of which there are around 50, he said.

Hansen said it would likely take time to implement all those genes into clinical practice, and said that one option would be for the lab to screen for all those genes in order to gather more data, but mask the results on all but a handful of them to the clinicians. "The geneticists and oncologists may not use all the information from day one," he said. "They will wait until the data is more convincing for some of the genes."