title: A Novel Assay for the Identification of NOTCH1 PEST Domain Mutations in Chronic Lymphocytic Leukemia.
abstract: Aims.To develop a fast and robust DNA-based assay to detect insertions and deletions mutations in exon 34 that encodes the PEST domain ofNOTCH1in order to evaluate patients with chronic lymphocytic leukemia (CLL).Methods.We designed a multiplexed allele-specific polymerase chain reaction (PCR) combined with a fragment analysis assay to detect specifically the mutation c.7544_7545delCT and possibly other insertions and deletions in exon 34 ofNOTCH1.Results.We evaluated our assay in peripheral blood samples from two cohorts of patients with CLL. The frequency ofNOTCH1mutations was 8.4% in the first cohort of 71 unselected CLL patients. We then evaluated a second cohort of 26 CLL patients with known cytogenetic abnormalities that were enriched for patients with trisomy 12.NOTCH1mutations were detected in 43.7% of the patients with trisomy 12.Conclusions.We have developed a fast and robust assay combining allele-specific PCR and fragment analysis able to detectNOTCH1PEST domain insertions and deletions.
authors.affiliation: Departments of Hematology and Clinical Pathology, and Research Institute, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil; Department of Hematology, Universidade Estadual de Campinas (Hemocentro-Unicamp), Campinas, SP, Brazil.
authors.forename: Paulo Vidal
authors.identifier: 0000-0001-6005-405X
authors.initials: PV
authors.lastname: Campregher
authors.affiliation: Department of Clinical Pathology, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil.
authors.forename: Roberta Cardoso
authors.initials: RC
authors.lastname: Petroni
authors.affiliation: Department of Clinical Pathology, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil.
authors.forename: Nair Hideko
authors.initials: NH
authors.lastname: Muto
authors.affiliation: Department of Clinical Pathology, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil.
authors.forename: Roberta
authors.initials: R
authors.lastname: Sitnik
authors.affiliation: Department of Clinical Pathology, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil.
authors.forename: Flavia Pereira
authors.initials: FP
authors.lastname: de Carvalho
authors.affiliation: Department of Clinical Pathology, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil.
authors.forename: Nydia Strachman
authors.initials: NS
authors.lastname: Bacal
authors.affiliation: Hematology Service, Hospital das Clínicas, Faculdade de Medicina da Universidade de São Paulo São Paulo, SP, Brazil; Cytogenetics Laboratories, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil.
authors.forename: Elvira Deolinda Rodrigues Pereira
authors.initials: ED
authors.lastname: Velloso
authors.affiliation: Hemocentro, Faculdade de Medicina da Universidade Estadual de Campinas (Hemocentro-Unicamp), Campinas, SP, Brazil.
authors.forename: Gislaine Borba
authors.initials: GB
authors.lastname: Oliveira
authors.affiliation: Department of Clinical Pathology, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil.
authors.forename: João Renato Rebello
authors.initials: JR
authors.lastname: Pinho
authors.affiliation: Bone Marrow Transplantation Center, Instituto Nacional de Câncer (INCA), Rio de Janeiro, RJ, Brazil.
authors.forename: Davi Coe
authors.initials: DC
authors.lastname: Torres
authors.affiliation: Paediatric Haematology-Oncology Program, Instituto Nacional de Câncer (INCA), Rio de Janeiro, RJ, Brazil.
authors.forename: Marcela Braga
authors.initials: MB
authors.lastname: Mansur
authors.affiliation: Bone Marrow Transplantation Center, Instituto Nacional de Câncer (INCA), Rio de Janeiro, RJ, Brazil.
authors.forename: Rocio
authors.initials: R
authors.lastname: Hassan
authors.affiliation: Hemocentro, Faculdade de Medicina da Universidade Estadual de Campinas (Hemocentro-Unicamp), Campinas, SP, Brazil.
authors.forename: Irene Gyongyvér Heidemarie
authors.initials: IG
authors.lastname: Lorand-Metze
authors.affiliation: Department of Hematology, Faculdade de Medicina da Santa Casa de Misericórdia de São Paulo, São Paulo, SP, Brazil.
authors.forename: Carlos Sérgio
authors.initials: CS
authors.lastname: Chiattone
authors.affiliation: Department of Hematology, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil.
authors.forename: Nelson
authors.initials: N
authors.lastname: Hamerschlak
authors.affiliation: Department of Clinical Pathology, Hospital Israelita Albert Einstein, São Paulo, SP, Brazil.
authors.forename: Cristovão Luis Pitangueira
authors.initials: CL
authors.lastname: Mangueira
mesh_terms: D000483:Alleles; D002881:Chromosomes, Human, Pair 12 / Q000235:genetics; D015331:Cohort Studies; D004252:DNA Mutational Analysis / Q000379:methods; D005260:Female; D006801:Humans; D054643:INDEL Mutation; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000235:genetics*; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods*; D000072417:Protein Domains; D051881:Receptor, Notch1 / Q000235:genetics*; D014314:Trisomy / Q000235:genetics*
chemical_list: C497761:NOTCH1 protein, human; D051881:Receptor, Notch1