An international study led by VHIO within the ENIGMA consortium has analysed 166 variants in and around exon 18 of the BRCA1 gene. Experimental RNA analysis changed the clinical interpretation of 34% of the variants studied and helped resolve uncertainty in around one in ten cases. More accurate classification of BRCA1 variants can help identify those associated with an increased risk of cancer and provide more robust information for genetic counselling of carriers and their families. The study also shows that loss of exon 18 results in a BRCA1 protein that is unable to properly perform its DNA repair function, providing new evidence to support the interpretation of variants that disrupt RNA processing, or splicing.
An international study by the Evidence-based Network for the Interpretation of Germline Mutant Alleles (ENIGMA) consortium has shown that experimental RNA analysis can significantly improve the classification of certain variants in the BRCA1 gene. Led by Dr Sara Gutiérrez-Enríquez, Head of the Translational Laboratory of VHIO’s Hereditary Cancer Genetics Group, the study has been published in The American Journal of Human Genetics.
Understanding the impact of BRCA1 variants for more accurate cancer risk assessment
BRCA1 is a tumour suppressor gene with a key role in DNA repair. Certain inherited variants that impair its function substantially increase the risk of developing cancer. By the age of 80, carriers are estimated to have a risk of around 72% of developing breast cancer and 44% of developing ovarian cancer.
Genetic testing can identify people at increased risk who carry pathogenic variants and may benefit from closer surveillance, preventive strategies and targeted therapies. However, testing can also uncover changes in BRCA1 whose clinical impact remains unclear. These are known as variants of uncertain significance (VUS), and they can make it more difficult to interpret genetic test results and provide clear guidance to carriers and their families.
“Classifying these variants more accurately would help us reduce the uncertainty associated with genetic testing and better distinguish those that are linked to an increased risk of cancer. This is important information for genetic counselling and for guiding surveillance and prevention strategies for carriers and their families,” says Dr Sara Gutiérrez-Enríquez.
From DNA to RNA: what happens during splicing
The information encoded in a gene is not used directly to make a protein. It is first copied into an RNA molecule, which then undergoes a maturation process known as splicing.
During splicing, the cell removes regions called introns and joins together other regions, known as exons, which contain the information ultimately used to produce the protein. Some genetic variants can disrupt this process, causing an exon that should normally be included in the final RNA molecule to be left out.
The researchers focused specifically on exon 18 of BRCA1, which forms part of the protein’s BRCT domain, a region that is essential for its DNA repair function. Certain variants can cause the cell to ‘skip’ this exon during RNA processing, a phenomenon known as exon skipping.
Until now, an important unanswered question was how much of this altered RNA a cell could produce before BRCA1 function became significantly compromised.
A protein that is produced but fails to function properly
“Until now, the consequences of losing exon 18 were unclear because the cell is still able to produce an apparently stable BRCA1 protein. Our results show, however, that this protein cannot properly carry out its DNA repair function, confirming the functional impact of variants that cause this exon to be lost,” explains Dr Joana Domènech-Vivó, from the Clinical and Molecular Genetics Unit at Vall d’Hebron University Hospital and first author of the study.
To reach these conclusions, the researchers combined several experimental approaches, including RNA analysis of samples from people carrying the variants, minigene assays and cellular models that allowed them to directly assess DNA repair capacity.
Reducing uncertainty in genetic diagnosis
Incorporating experimental RNA data improved the interpretation of numerous BRCA1 variants and substantially reduced the number that remained classified as variants of uncertain significance.
Specifically, experimental messenger RNA analysis changed the interpretation of 34% of the variants studied and helped resolve uncertainty in approximately 10% of cases.
“These findings highlight the value of looking not only at the change present in the DNA, but also at how that change affects RNA and, ultimately, the function of the protein,” concludes Dr Gutiérrez-Enríquez.
These findings could contribute to more accurate classification of BRCA1 variants and provide a stronger evidence base for the genetic counselling of individuals and families with an inherited predisposition to cancer.
