Technique Pinpoints Oncogenic BRCA1 Mutations Free

Certain germline BRCA1 mutations increase the risk of developing breast or ovarian cancer, but the significance of many of these alterations remains unclear. In a recent study, however, scientists evaluated the functional consequences of nearly 4,000 BRCA1 single-nucleotide variants (SNV) in a human cell line and found that 21% of them are likely oncogenic. The research may aid in the clinical interpretation of BRCA1 variants.

The work “could be transformative,” says Dezheng Huo, MD, PhD, of the University of Chicago in Illinois. The approach the researchers used is an improvement over previous methods because “it's a more systematic experimental way” to classify BRCA1 mutations, he says.

Even though BRCA1 has been sequenced for more than 2 million people, most SNVs are so rare that researchers don't have enough information to reach a conclusion about their impact—these mutations are known as variants of unknown significance. For many other SNVs, interpretations of their oncogenic potential have been contradictory.

To gauge whether these SNVs might cause cancer, Gregory Findlay, PhD, of the University of Washington in Seattle, and colleagues applied a technique called saturation genome editing, which involves mutating each nucleotide base in a gene or a section of a gene. Using CRISPR/Cas9, the scientists induced mutations in 13 exons that are crucial for BRCA1′s function and in neighboring introns. Overall, they analyzed 3,893 SNVs, more than 96% of the possible number in these sections of the gene.

The researchers introduced the mutations into a line of human leukemia cells known as HAP1, which are haploid and lack a backup copy of BRCA1, making it easier to identify the functional consequences of mutations. Because HAP1 cells die without working BRCA1 proteins, the impact of a particular mutation is easier to detect. If HAP1 cells carrying a particular mutation died, patients with the same mutation likely have a nonfunctional version of BRCA1 that promotes cancer.

After tracking 260 million mutated cells, the scientists calculated that 21% of the SNVs resulted in nonfunctional BRCA1, making them likely to increase cancer risk. The team then checked their results against ClinVar, an NIH database that records evaluations from clinical testing labs of variants’ clinical significance. ClinVar categorized 169 SNVs in BRCA1 as likely to promote cancer, and the results from Findlay and colleagues agreed for 162 of them. The team also came to the same conclusion as ClinVar for 20 of the 22 variants the database lists as benign. “Our data correlates pretty well,” says Findlay.

The researchers are the first to analyze so many mutations across such a large portion of the gene, says Fergus Couch, PhD, of the Mayo Clinic in Rochester, MN, who was not connected to the study. “This is a giant leap forward.”

However, Couch cautions that the results of the study are not ready for clinical use. Future work needs to determine the sensitivity and specificity of the technique compared with known standards before doctors can rely on it to make decisions, he says.

The study's findings “will have to be integrated with other data when drawing conclusions about whether these variants are pathogenic,” adds William Foulkes, MBBS, PhD, of McGill University in Montreal, Canada. Although the results alone are insufficient to determine whether a variant is associated with increased cancer risk, they could provide the decisive evidence if combined with other sources of information, he says. –Mitch Leslie