title: The impact of sensitive KIT D816V detection on recognition of indolent Systemic Mastocytosis.
abstract: Patients with Systemic Mastocytosis (SM) need a highly sensitive diagnostic test for D816V detection of the KIT receptor gene. Along with histology/cytology and flow cytometry evaluation, bone marrow (BM) from 110 consecutive adult patients referred with a suspicion of SM to Multidisciplinary Outpatient Clinic for Mastocytosis in Verona were tested both by Amplification Refractory Mutation System Reverse Transcriptase quantitative real time Polymerase Chain Reaction (ARMS-RT-qPCR) and RT-PCR+Restriction Fragment Length Polymorphism (RFLP) followed by Denaturing-High Performance Liquid Chromatography (D-HPLC) and Sanger sequencing. ARMS-RT-qPCR identified D816V mutation in 77 patients, corresponding to 100% of cases showing CD25(+) mast cells (MCs) whereas RT-PCR+RFLP/D-HPLC+sequencing revealed D816V mutations in 47 patients. According to the 2008 WHO criteria 75 SM, 1 Cutaneous Mastocytosis (CM), 1 monoclonal MC activation syndrome (MMAS), and 1 SM Associated with Haematologic Non-Mast Cell Disorder (SM-AHNMD) were diagnosed. Seventeen out 75 SM patients (23%) would have not satisfied sufficient WHO criteria on the basis of the sole RT-PCR+RFLP: these patients had significantly lower serum tryptase levels and amount of CD25(+) MCs. Therefore, ARMS-RT-qPCR might result particularly useful, in patients that do not fulfil major BM histological criterion, for the recognition of indolent SM with a very low MC burden.
authors.affiliation: Section of Clinical Biochemistry, Department of Life and Reproduction Sciences, University of Verona, Verona, Italy; Multidisciplinary Outpatients Clinics for Mastocytosis, Verona, Italy. Electronic address: giovanna.dematteis@ospedaleuniverona.it.
authors.forename: Giovanna
authors.initials: G
authors.lastname: De Matteis
authors.affiliation: Multidisciplinary Outpatients Clinics for Mastocytosis, Verona, Italy; Section of Haematology, Department of Medicine, University of Verona, Verona, Italy.
authors.forename: Roberta
authors.initials: R
authors.lastname: Zanotti
authors.affiliation: Department of Haematology/Oncology, L. and A. Seragnoli, University of Bologna, Bologna, Italy.
authors.forename: Sabrina
authors.initials: S
authors.lastname: Colarossi
authors.affiliation: Department of Haematology/Oncology, L. and A. Seragnoli, University of Bologna, Bologna, Italy.
authors.forename: Caterina
authors.initials: C
authors.lastname: De Benedittis
authors.affiliation: Spanish Network on Mastocytosis (REMA), Toledo, Salamanca, Spain; Centro de Investigacion del Cancer/IBMCC (USAL/CSIC) and IBSAL, Departamento de Medicina and Servicio General de Citometrıa, University of Salamanca, Salamanca, Spain.
authors.forename: Andrès
authors.initials: A
authors.lastname: Garcia-Montero
authors.affiliation: Multidisciplinary Outpatients Clinics for Mastocytosis, Verona, Italy; Section of Haematology, Department of Medicine, University of Verona, Verona, Italy.
authors.forename: Massimiliano
authors.initials: M
authors.lastname: Bonifacio
authors.affiliation: Section of Clinical Biochemistry, Department of Life and Reproduction Sciences, University of Verona, Verona, Italy.
authors.forename: Marta
authors.initials: M
authors.lastname: Sartori
authors.affiliation: Section of Clinical Biochemistry, Department of Life and Reproduction Sciences, University of Verona, Verona, Italy.
authors.forename: Fiorenza
authors.initials: F
authors.lastname: Aprili
authors.affiliation: Section of Clinical Biochemistry, Department of Life and Reproduction Sciences, University of Verona, Verona, Italy.
authors.forename: Beatrice
authors.initials: B
authors.lastname: Caruso
authors.affiliation: Section of Clinical Biochemistry, Department of Life and Reproduction Sciences, University of Verona, Verona, Italy.
authors.forename: Elisa
authors.initials: E
authors.lastname: Paviati
authors.affiliation: Multidisciplinary Outpatients Clinics for Mastocytosis, Verona, Italy; Section of Haematology, Department of Medicine, University of Verona, Verona, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Carli
authors.affiliation: Multidisciplinary Outpatients Clinics for Mastocytosis, Verona, Italy; Section of Haematology, Department of Medicine, University of Verona, Verona, Italy.
authors.forename: Omar
authors.initials: O
authors.lastname: Perbellini
authors.affiliation: Section of Pathological Anatomy, University of Verona, Verona, Italy.
authors.forename: Alberto
authors.initials: A
authors.lastname: Zamò
authors.affiliation: Multidisciplinary Outpatients Clinics for Mastocytosis, Verona, Italy; Allergy Unit, Azienda Ospedaliera Universitaria Integrata, Verona, Italy.
authors.forename: Patrizia
authors.initials: P
authors.lastname: Bonadonna
authors.affiliation: Multidisciplinary Outpatients Clinics for Mastocytosis, Verona, Italy.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Pizzolo
authors.affiliation: Section of Clinical Biochemistry, Department of Life and Reproduction Sciences, University of Verona, Verona, Italy.
authors.forename: Giancesare
authors.initials: G
authors.lastname: Guidi
authors.affiliation: Department of Haematology/Oncology, L. and A. Seragnoli, University of Bologna, Bologna, Italy.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Martinelli
authors.affiliation: Department of Haematology/Oncology, L. and A. Seragnoli, University of Bologna, Bologna, Italy.
authors.forename: Simona
authors.initials: S
authors.lastname: Soverini
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000235:genetics*; D001853:Bone Marrow / Q000378:metabolism / Q000473:pathology*; D002851:Chromatography, High Pressure Liquid; D005260:Female; D005500:Follow-Up Studies; D006801:Humans; D008297:Male; D034721:Mastocytosis, Systemic / Q000097:blood / Q000175:diagnosis* / Q000235:genetics*; D008875:Middle Aged; D017354:Point Mutation / Q000235:genetics*; D012150:Polymorphism, Restriction Fragment Length; D011379:Prognosis; D019009:Proto-Oncogene Proteins c-kit / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055815:Young Adult
chemical_list: D014408:Biomarkers, Tumor; D012333:RNA, Messenger; D019009:Proto-Oncogene Proteins c-kit
keywords: D-HPLC; D816V; KIT; Real time PCR; Sequencing; Systemic mastocytosis