Uncommon nucleotide excision repair phenotypes revealed by targeted high-throughput sequencing

Nadège Calmels (1) , Géraldine Greff (1) , Cathy Obringer (2) , Nadine Kempf (1) , Claire Gasnier (1) , Julien Tarabeux (1) , Marguerite Miguet (1) , Geneviève Baujat (3) , Didier Bessis (4) , Patricia Bretones (5) , Anne Cavau (6) , Béatrice Digeon (7) , Martine Doco-Fenzy (7) , Bérénice Doray (8) , François Feillet (9) , Jesus Gardeazabal (10) , Blanca Gener (10) , Sophie Julia (11) , Isabel Llano-Rivas (10) , Artur Mazur (12) , Caroline Michot (6) , Florence Renaldo-Robin (13) , Massimiliano Rossi (14, 15) , Pascal Sabouraud (7) , Boris Keren (16, 17) , Christel Depienne (16, 17) , Jean Muller (1, 2) , Jean-Louis Mandel (1, 18) , Vincent Laugel (19, 2)

1 Laboratoire de Diagnostic Génétique
2 LGM - Laboratoire de Génétique Médicale
3 Service de Génétique Médicale [CHU Necker]
4 CHRU Montpellier - Hôpital Saint Eloi
5 Service d'endocrinologie pédiatrique, Hôpital mère enfant, Bron
6 Hôpital Necker - Enfants Malades [AP-HP]
7 Hôpital universitaire Robert Debré [Reims]
8 Centre hospitalier Félix-Guyon [Saint-Denis, La Réunion]
9 NGERE - Nutrition-Génétique et Exposition aux Risques Environnementaux
10 BioCruces Research Institute
11 CHU Toulouse - Centre Hospitalier Universitaire de Toulouse
12 Rzeszow University
13 Unité de Neurologie, Hôpital Robert Debré
14 CHLS - Centre Hospitalier Lyon Sud [CHU - HCL]
15 CRNL - Centre de recherche en neurosciences de Lyon - Lyon Neuroscience Research Center
16 CHU Pitié-Salpêtrière [AP-HP]
17 ICM - Institut du Cerveau et de la Moëlle Epinière = Brain and Spine Institute
18 Collège de France - Chaire Génétique Humaine
19 CHU Strasbourg - Centre Hospitalier Universitaire [Strasbourg]

Background: Deficient nucleotide excision repair (NER) activity causes a variety of autosomal recessive diseases including xeroderma pigmentosum (XP) a disorder which pre-disposes to skin cancer, and the severe multisystem condition known as Cockayne syndrome (CS). In view of the clinical overlap between NER-related disorders, as well as the existence of multiple phenotypes and the numerous genes involved, we developed a new diagnostic approach based on the enrichment of 16 NER-related genes by multiplex amplification coupled with next-generation sequencing (NGS). Methods: Our test cohort consisted of 11 DNA samples, all with known mutations and/or non pathogenic SNPs in two of the tested genes. We then used the same technique to analyse samples from a prospective cohort of 40 patients. Multiplex amplification and sequencing were performed using AmpliSeq protocol on the Ion Torrent PGM (Life Technologies). Results: We identified causative mutations in 17 out of the 40 patients (43 %). Four patients showed biallelic mutations in the ERCC6(CSB) gene, five in the ERCC8(CSA) gene: most of them had classical CS features but some had very mild and incomplete phenotypes. A small cohort of 4 unrelated classic XP patients from the Basque country (Northern Spain) revealed a common splicing mutation in POLH (XP-variant), demonstrating a new founder effect in this population. Interestingly, our results also found ERCC2(XPD), ERCC3(XPB) or ERCC5(XPG) mutations in two cases of UV-sensitive syndrome and in two cases with mixed XP/CS phenotypes. Conclusions: Our study confirms that NGS is an efficient technique for the analysis of NER-related disorders on a molecular level. It is particularly useful for phenotypes with combined features or unusually mild symptoms. Targeted NGS used in conjunction with DNA repair functional tests and precise clinical evaluation permits rapid and cost-effective diagnosis in patients with NER-defects.

Mots clés: NER; NGS; Cockayne syndrome; xeroderma pigmentosum; ERCC2; ERCC3; ERCC5; ERCC6; ERCC8; POLH

Domaines: Médecine humaine et pathologie

Nadège Calmels, Géraldine Greff, Cathy Obringer, Nadine Kempf, Claire Gasnier, et al.. Uncommon nucleotide excision repair phenotypes revealed by targeted high-throughput sequencing. Orphanet Journal of Rare Diseases, 2016, 11 (1), pp.26. ⟨10.1186/s13023-016-0408-0⟩. ⟨hal-01295286⟩