Diagnosis of Fanconi Anemia: Mutation Analysis by Next-Generation Sequencing

Fanconi anemia (FA) is a rare genetic instability syndrome characterized by developmental defects, bonemarrow failure, and a high cancer risk. Fifteen genetic subtypes have been distinguished. The majority of patients (approximate to 85%) belong to the subtypes A (approximate to 60%), C (approximate to 15%) or G (approximate to 10%), while a minority (approximate to 15%) is distributed over the remaining 12 subtypes. All subtypes seem to fit within the "classical" FA phenotype, except for D1 and N patients, who havemore severe clinical symptoms. Since FA patients need special clinical management, the diagnosis should be firmly established, to exclude conditions with overlapping phenotypes. A valid FA diagnosis requires the detection of pathogenic mutations in a FA gene and/or a positive result from a chromosomal breakage test. Identification of the pathogenic mutations is also important for adequate genetic counselling and to facilitate prenatal or preimplantation genetic diagnosis. Here we describe and validate a comprehensive protocol for the molecular diagnosis of FA, based on massively parallel sequencing. We used this approach to identify BRCA2, FANCD2, FANCI and FANCL mutations in novel unclassified FA patients.