Genetic Diagnosis of Neurofibromatosis type 2

Introduction Neurofibromatosis type 2 is an autosomal dominant disease caused by mutations in the NF2 gene on 22q12.2. Its protein product, merlin, supposedly plays an important role in connecting membrane proteins with the cytoskeleton by coordinating growth-factor signalling. The most common mutations are truncating and splice site mutations, showing a genotype-phenotype correlation. A high rate of somatic mosaicism may account for the difficulty of molecular genetic diagnosis. The main clinical feature is bilateral vestibular schwannomas and multiple neural tumours. Furthermore patients present tinnitus, hearing loss and dysequilibrium, subcapsular cataracts and cutaneous manifestations. The management of the disease includes surgical removal and radiological staging. Objectives It was intended to develop a genetic testing protocol for patients with clinical criteria for this condition comprising sequence analysis and exon dosage study, allowing the observation of genotypephenotype correlations. Methodology Primers were designed to cover the entire coding region, flanking intronic sequences and 5’ and 3’ UTR. The obtained sequences were aligned with the reference sequence and checked against SNPs and documented mutations. Multiplex Ligation-dependent Probe Amplification was also performed covering the 17 exons and the gene promoter. Two newly diagnosed cases and one individual with an identified mutation along with his mother were studied. Results It was possible to validate our mutation screening procedure through the detection of a documented mutation in exon 11, although his mother had no identifiable mutations. In a patient with bilateral schwannomas was detected a novel mutation, being a nonsense frameshift insertion in exon 5. In another patient with several neural tumours it was impossible to identify any mutation. Conclusions The main purpose of this study was fulfilled despite the impossibility of drawing conclusions on genotype-phenotype correlation due to sample shortage. The importance of genetic testing in both lymphocyte and tumour DNA with sequencing and Multiplex Ligation-dependent Probe Amplification should not to be neglected.