Beyond classic molecular alteractions: non-contiguous mutations in DMD gene

LEONELA LUCE; CARCIONE, MICAELA;; MAZZANTI, CHIARA; IRENE SZIJAN; MENAZZI, SEBASTIÁN; FRANCIPANE LILIANA,; NEVADO JULIAN; LAPUNZINA, PABLO; LILIANA ROSSETTI; PAMELA RADIC; ABELLEYRO MIGUEL M; CARLOS DE BRASI; FLORENCIA GILIBERTO

Congreso; REUNIÓN ANUAL DE SOCIEDAD DE BIOCIENCIAS SAIC-SAI-SAFIS 2020 LXV REUNIÓN ANUAL DE LA SOCIEDAD ARGENTINA DE INVESTIGACIÓN CLÍNICA; 2020

93. (406) BEYOND CLASSIC MOLECULAR ALTERATIONS: NON-CONTIGUOUS MUTATIONS IN DMD GENELeonela Luce1,2, Micaela Carcione1,2, Chiara Mazzanti1,2, Irene Szijan1, Sebastian Menazzi3, Liliana Francipane3, Julián Nevado4,5, Pablo Lapunzina4,5, Liliana Rossetti6 , Pamela Radic6,Martin Abelleyro6, Carlos De Brasi6, Florencia Giliberto1,21 Universidad de Buenos Aires; Departamento de Microbiología, Inmunología, Biotecnología y Genética; Cátedra de Genética; Laboratorio de Distrofinopatías, Buenos Aires, Argentina.2 CONICET - Universidad de Buenos Aires; Instituto de Inmunología, Genética y Metabolismo (INIGEM), Buenos Aires, Argentina.3 Universidad de Buenos Aires, Hospital de Clínicas ?José de San Martín?, División de Genética, Buenos Aires, Argentina. 4 Hospital Universitario La Paz, IdiPAZ, Instituto de Genética Médica y Molecular (INGEMM), Madrid, España. 5 Centro de Investigaciones Biomédicas en Red para Enfermedades Raras (CIBERER), Madrid, España.6 CONICET - Academia Nacional de Medicina, Instituto de Medicina Experimental (IMEX), Buenos Aires, Argentina.Introduction: Dystrophinopathies are neuromuscular X-linked recessive diseases caused by DMD mutations. Molecular alterations in this gene are large deletions/duplications in 80% of cases and small variants in the remaining. Several authors reported the occurrence of non-contiguous rearrangements within the same DMD allele, with frequencies up to 4%. The present work aims to characterize the incidence of complex rearrangements in an Argentinian dystrophinopathycohort and unravel the causing molecular mechanisms. Materials and Methods: We analyzed 437 boys with clinical diagnosis of Dystrophinopathy. The following techniques were implemented: MLPA, WES, WGS, PCR-Sanger Sequencing, CGH Array and HUMARA assay. In 2 cases, breakpoints were precisely determined, so we performed a bioinformatic screening of microhomologies, interspersed repeats, secondary structures and recombinogenic motifs 50pb surrounding each breakpoint. Results: We detected 6 patients carrying complex rearrangements in DMD: 2 deletions-duplications, 3 non-contiguous duplications and 1 large deletion plus a 20pb insertion. These accounted for 1.4% of our cohort. In a deletion-duplication case, familial segregation and bioinformatics analysis suggested that the duplication was the first mutagenic event caused by Fork Stalling and Template Switching (FoSTeS), while the deletion occurred secondly by Non-homologous end joining. Furthermore, bioinformatic screening of the deletionplus insertion propose that the deletion was due to Microhomology-mediated end joining, while the insertion arose by FoSTeS. Conclusions: Our findings widen the understanding of the molecular events that may take place in DMD and characterize the occurrence of complex rearrangements in our dystrophinopathy cohort.