MOLECULAR DIAGNOSIS OF DYSTROPHINOPATHIES AND DMD GENE CHARACTERIZATION

CARCIONE, MICAELA; MAZZANTI, CHIARA; GILIBERTO, FLORENCIA; LUCE, LEONELA

Workshop; XIV Escola Latino-Americana de Genética Humana e Médica (ELAG) Course; 2018

Dystrophinopathies are X-linked diseases caused by mutations in DMD gene, associated with thedevelopment of Duchenne Muscular Dystrophy (DMD), Becker Muscular Dystrophy (BMD) and X-linked Dilated Cardiomyopathy (XLDC). DMD, the most frequent and severe dystrophy affects 1:3500born males, BMD is less severe and affects 1:18000. De-novo mutations and germline mosaicism areresponsible for 1/3 of cases, while 2/3 are familial cases. Molecular alterations in DMD gene are grossdeletions/duplications in 80% of cases and small mutations in 20%. Hitherto there is no effectivetreatment for these pathologies, which enhances the importance of performing genetic assessment tothe affected families in order to detect mutation carriers and prevent diseased newborns. However,two mutation-specific gene therapies for DMD were recently approved: Exon Skipping of exon 51 andPremature Stop Codon Read-through. Therefore, accurate detection and characterization of thecausing mutation is essential to allow genetic counseling, patient follow-up and determine the suitablegene therapy. In order to achieve an accurate molecular diagnosis in patients and in at risk familymembers, we performed an algorithm based on several techniques: MLPA, simplex PCR, Sangersequencing, Whole Exome Sequencing (WES) and STRs segregation analysis. In the last 6 years wecould determine the DMD/BMD diagnosis in the 73,5% of patients, 22 resulted candidates for Atalurenand 12 for the exon skipping treatments. The reading frame rule was corroborated in the 78.9% ofpatients. The molecular alterations we found accounted for: 54% deletions, 11% duplications, 29%small mutations (14% nonsense), 1% del/dup and 5% were not detected. We have classified anduploaded to public databases the 148 small variants we have found in the DMD gen, as benign, VUSor pathogenic. We also, could establish an association between the most frequent deletion intronbreakpoints and the abundance of dinucleotide microsatellites loci. Interestingly, could demonstratedthat dystrophin expression is altered in non-myogenic tumors, corroborating the newphysiopathological implication of the DMD gene in tumor development. In conclusion, the work wehave been performing since 1992 and the analysis of more than 1500 samples allow thecharacterization of the DMD/BMD argentine population and contribute to the understanding of thegenetic/molecular basis of these pathologies.