title: ISPD mutations account for a small proportion of Italian Limb Girdle Muscular Dystrophy cases.
abstract: BACKGROUND
  Limb Girdle Muscular Dystrophy (LGMD), caused by defective α-dystroglycan (α-DG) glycosylation, was recently associated with mutations in Isoprenoid synthase domain-containing (ISPD) and GDP-mannose pyrophosphorylase B (GMPPB) genes. The frequency of ISPD and GMPPB gene mutations in the LGMD population is unknown.
  
  
  METHODS
  We investigated the contributions of ISPD and GMPPB genes in a cohort of 174 Italian patients with LGMD, including 140 independent probands. Forty-one patients (39 probands) from this cohort had not been genetically diagnosed. The contributions of ISPD and GMPPB were estimated by sequential α-DG immunohistochemistry (IHC) and mutation screening in patients with documented α-DG defect, or by direct DNA sequencing of both genes when muscle tissue was unavailable.
  
  
  RESULTS
  We performed α-DG IHC in 27/39 undiagnosed probands: 24 subjects had normal α-DG expression, two had a partial deficiency, and one exhibited a complete absence of signal. Direct sequencing of ISPD and GMPPB revealed two heterozygous ISPD mutations in the individual who lacked α-DG IHC signal: c.836-5 T > G (which led to the deletion of exon 6 and the production of an out-of-frame transcript) and c.676 T > C (p.Tyr226His). This patient presented with sural hypertrophy and tip-toed walking at 5 years, developed moderate proximal weakness, and was fully ambulant at 42 years. The remaining 12/39 probands did not exhibit pathogenic sequence variation in either gene.
  
  
  CONCLUSIONS
  ISPD mutations are a rare cause of LGMD in the Italian population, accounting for less than 1% of the entire cohort studied (FKRP mutations represent 10%), while GMPPB mutations are notably absent in this patient sample. These data suggest that the genetic heterogeneity of LGMD with and without α-DG defects is greater than previously realized.
authors.affiliation: Dino Ferrari Centre, Department of Neurological Sciences, University of Milan, I.R.C.C.S. Foundation Cà Granda, Ospedale Maggiore Policlinico, via F. Sforza 35, 20122, Milan, Italy. francescam.magri@gmail.com.
authors.forename: Francesca
authors.initials: F
authors.lastname: Magri
authors.affiliation: Neuromuscular and Rare Disease Unit, Department of Neuroscience, Foundation IRCCS Ca' Granda Ospedale Maggiore Policlinico, University of Milan, via F. Sforza 35, 20132, Milan, Italy. irene_colombo@libero.it.
authors.forename: Irene
authors.initials: I
authors.lastname: Colombo
authors.affiliation: Dino Ferrari Centre, Department of Neurological Sciences, University of Milan, I.R.C.C.S. Foundation Cà Granda, Ospedale Maggiore Policlinico, via F. Sforza 35, 20122, Milan, Italy. roberto.delbo@unimi.it.
authors.forename: Roberto
authors.initials: R
authors.lastname: Del Bo
authors.affiliation: Inspe, Division of Neuroscience, San Raffaele, Via Olgettina 60, Milan, Italy. previtali.stefano@hsr.it.
authors.forename: Stefano
authors.initials: S
authors.lastname: Previtali
authors.affiliation: Dino Ferrari Centre, Department of Neurological Sciences, University of Milan, I.R.C.C.S. Foundation Cà Granda, Ospedale Maggiore Policlinico, via F. Sforza 35, 20122, Milan, Italy. robi.brusa22@gmail.com.
authors.forename: Roberta
authors.initials: R
authors.lastname: Brusa
authors.affiliation: Neuromuscular and Rare Disease Unit, Department of Neuroscience, Foundation IRCCS Ca' Granda Ospedale Maggiore Policlinico, University of Milan, via F. Sforza 35, 20132, Milan, Italy. patrizia.ciscato@policlinico.mi.it.
authors.forename: Patrizia
authors.initials: P
authors.lastname: Ciscato
authors.affiliation: Inspe, Division of Neuroscience, San Raffaele, Via Olgettina 60, Milan, Italy. scarlato.marina@hsr.it.
authors.forename: Marina
authors.initials: M
authors.lastname: Scarlato
authors.affiliation: Dino Ferrari Centre, Department of Neurological Sciences, University of Milan, I.R.C.C.S. Foundation Cà Granda, Ospedale Maggiore Policlinico, via F. Sforza 35, 20122, Milan, Italy. dario.ronchi@gmail.com.
authors.forename: Dario
authors.initials: D
authors.lastname: Ronchi
authors.affiliation: IRCCS E. Medea, Bosisio Parini, Italy. grazia.dangelo@bp.lnf.it.
authors.forename: Maria Grazia
authors.initials: MG
authors.lastname: D'Angelo
authors.affiliation: Dino Ferrari Centre, Department of Neurological Sciences, University of Milan, I.R.C.C.S. Foundation Cà Granda, Ospedale Maggiore Policlinico, via F. Sforza 35, 20122, Milan, Italy. stefania.corti@unimi.it.
authors.forename: Stefania
authors.initials: S
authors.lastname: Corti
authors.affiliation: Neuromuscular and Rare Disease Unit, Department of Neuroscience, Foundation IRCCS Ca' Granda Ospedale Maggiore Policlinico, University of Milan, via F. Sforza 35, 20132, Milan, Italy. maurizio.moggio@unimi.it.
authors.forename: Maurizio
authors.initials: M
authors.lastname: Moggio
authors.affiliation: Dino Ferrari Centre, Department of Neurological Sciences, University of Milan, I.R.C.C.S. Foundation Cà Granda, Ospedale Maggiore Policlinico, via F. Sforza 35, 20122, Milan, Italy. nereo.bresolin@unimi.it.
authors.forename: Nereo
authors.initials: N
authors.lastname: Bresolin
authors.affiliation: Dino Ferrari Centre, Department of Neurological Sciences, University of Milan, I.R.C.C.S. Foundation Cà Granda, Ospedale Maggiore Policlinico, via F. Sforza 35, 20122, Milan, Italy. giacomo.comi@unimi.it.
authors.forename: Giacomo Pietro
authors.initials: GP
authors.lastname: Comi
mesh_terms: D015331:Cohort Studies; D006801:Humans; D007558:Italy; D049288:Muscular Dystrophies, Limb-Girdle / Q000235:genetics*; D009154:Mutation; D009713:Nucleotidyltransferases / Q000235:genetics*; D044465:White People / Q000235:genetics
chemical_list: D009713:Nucleotidyltransferases; C026917:mannose 1-phosphate guanylyltransferase; C573136:CRPPA protein, human