title: A two-step approach for sequencing spliceosome-related genes as a complementary diagnostic assay in MDS patients with ringed sideroblasts.
abstract: Our study aimed to analyze the presence of mutations in SF3B1 and other spliceosome-related genes in myelodysplastic syndromes with ringed sideroblasts (MDS-RS) by combining conventional Sanger and next-generation sequencing (NGS) methods, and to determine the feasibility of this approach in a clinical setting. 122 bone marrow samples from MDS-RS patients were studied. Initially, exons 14 and 15 of the SF3B1 gene were analyzed by Sanger sequencing. Secondly, they were studied by NGS covering besides SF3B1, SRSF2, U2AF1 and ZRSR2 genes. An 86% of all patients showed mutations in the SF3B1 gene. Six of them, which were not identifiable by conventional sequencing in the first diagnostic step, were revealed by NGS. In addition, 19.5% of cases showed mutations in other splicing genes: SRSF2, U2AF1, and ZRSR2. Furthermore, 8.7% of patients had two mutations in SF3B1, SF3B1 and SRSF2, and SF3B1 and U2AF1, while 5.7% showed no mutations in the four spliceosome-related genes analyzed. The combined use of conventional Sanger and NGS allows the identification of mutations in spliceosome-related genes in almost all MDS patients with RS. This two-step approach is affordable and could be useful as a complementary technique in cases with an unclear diagnosis.
authors.affiliation: IBMCC, Centro de Investigación del Cáncer, Universidad de Salamanca-CSIC, Spain; IBSAL, Instituto de Investigación Biomédica de Salamanca, Spain.
authors.forename: Kamila
authors.initials: K
authors.lastname: Janusz
authors.affiliation: IBMCC, Centro de Investigación del Cáncer, Universidad de Salamanca-CSIC, Spain; IBSAL, Instituto de Investigación Biomédica de Salamanca, Spain.
authors.forename: Mónica
authors.initials: M
authors.lastname: Del Rey
authors.affiliation: IBMCC, Centro de Investigación del Cáncer, Universidad de Salamanca-CSIC, Spain; IBSAL, Instituto de Investigación Biomédica de Salamanca, Spain.
authors.forename: María
authors.initials: M
authors.lastname: Abáigar
authors.affiliation: Hospital General Universitario de Valencia, Spain.
authors.forename: Rosa
authors.initials: R
authors.lastname: Collado
authors.affiliation: Hospital General Universitario de Valencia, Spain.
authors.forename: David
authors.initials: D
authors.lastname: Ivars
authors.affiliation: IBMCC, Centro de Investigación del Cáncer, Universidad de Salamanca-CSIC, Spain; IBSAL, Instituto de Investigación Biomédica de Salamanca, Spain.
authors.forename: María
authors.initials: M
authors.lastname: Hernández-Sánchez
authors.affiliation: Complejo Hospitalario de Navarra, Servicio de Genética, Spain.
authors.forename: Alberto
authors.initials: A
authors.lastname: Valiente
authors.affiliation: IBMCC, Centro de Investigación del Cáncer, Universidad de Salamanca-CSIC, Spain; IBSAL, Instituto de Investigación Biomédica de Salamanca, Spain.
authors.forename: Cristina
authors.initials: C
authors.lastname: Robledo
authors.affiliation: IBMCC, Centro de Investigación del Cáncer, Universidad de Salamanca-CSIC, Spain; IBSAL, Instituto de Investigación Biomédica de Salamanca, Spain.
authors.forename: Rocío
authors.initials: R
authors.lastname: Benito
authors.affiliation: IBSAL, Instituto de Investigación Biomédica de Salamanca, Spain; Hospital Universitario de Salamanca, Spain.
authors.forename: María
authors.initials: M
authors.lastname: Díez-Campelo
authors.affiliation: Hospital Virgen Blanca, León, Spain.
authors.forename: Fernando
authors.initials: F
authors.lastname: Ramos
authors.affiliation: AstraZeneca, Personalized Healthcare and Biomarkers, Innovative Medicines and Early Development, Cambridge, UK.
authors.forename: Alexander
authors.initials: A
authors.lastname: Kohlmann
authors.affiliation: IBSAL, Instituto de Investigación Biomédica de Salamanca, Spain; Hospital Universitario de Salamanca, Spain.
authors.forename: Consuelo Del
authors.initials: CD
authors.lastname: Cañizo
authors.affiliation: IBMCC, Centro de Investigación del Cáncer, Universidad de Salamanca-CSIC, Spain; IBSAL, Instituto de Investigación Biomédica de Salamanca, Spain; Hospital Universitario de Salamanca, Spain. Electronic address: jmhr@usal.es.
authors.forename: Jesús María
authors.initials: JM
authors.lastname: Hernández-Rivas
mesh_terms: D000756:Anemia, Sideroblastic / Q000175:diagnosis / Q000235:genetics*; D001853:Bone Marrow; D006801:Humans; D008722:Methods; D009154:Mutation; D009190:Myelodysplastic Syndromes / Q000175:diagnosis* / Q000235:genetics*; D010750:Phosphoproteins / Q000235:genetics; D000072260:RNA Splicing Factors / Q000235:genetics; D017422:Sequence Analysis, DNA; D000068103:Serine-Arginine Splicing Factors / Q000235:genetics; D017381:Spliceosomes / Q000235:genetics*; D000072668:Splicing Factor U2AF / Q000235:genetics
chemical_list: D010750:Phosphoproteins; D000072260:RNA Splicing Factors; C112491:SF3B1 protein, human; D000072668:Splicing Factor U2AF; C076753:U2AF1 protein, human; C073288:SRSF2 protein, human; D000068103:Serine-Arginine Splicing Factors
keywords: Myelodysplastic syndrome; Next generation sequencing; Ringed sideroblasts; SF3B1; Spliceosome