title: A combined nucleic acid and protein analysis in Friedreich ataxia: implications for diagnosis, pathogenesis and clinical trial design.
abstract: BACKGROUND
  Friedreich's ataxia (FRDA) is the most common hereditary ataxia among caucasians. The molecular defect in FRDA is the trinucleotide GAA expansion in the first intron of the FXN gene, which encodes frataxin. No studies have yet reported frataxin protein and mRNA levels in a large cohort of FRDA patients, carriers and controls.
  
  
  RESULTS
  We enrolled 24 patients with classic FRDA phenotype (cFA), 6 late onset FRDA (LOFA), all homozygous for GAA expansion, 5 pFA cases who harbored the GAA expansion in compound heterozygosis with FXN point mutations (namely, p.I154F, c.482+3delA, p.R165P), 33 healthy expansion carriers, and 29 healthy controls. DNA was genotyped for GAA expansion, mRNA/FXN was quantified in real-time, and frataxin protein was measured using lateral-flow immunoassay in peripheral blood mononuclear cells (PBMCs). Mean residual levels of frataxin, compared to controls, were 35.8%, 65.6%, 33%, and 68.7% in cFA, LOFA, pFA and healthy carriers, respectively. Comparison of both cFA and pFA with controls resulted in 100% sensitivity and specificity, but there was overlap between LOFA, carriers and controls. Frataxin levels correlated inversely with GAA1 and GAA2 expansions, and directly with age at onset. Messenger RNA expression was reduced to 19.4% in cFA, 50.4% in LOFA, 52.7% in pFA, 53.0% in carriers, as compared to controls (p<0.0001). mRNA levels proved to be diagnostic when comparing cFA with controls resulting in 100% sensitivity and specificity. In cFA and LOFA patients mRNA levels correlated directly with protein levels and age at onset, and inversely with GAA1 and GAA2.
  
  
  CONCLUSIONS
  We report the first explorative study on combined frataxin and mRNA levels in PBMCs from a cohort of FRDA patients, carriers and healthy individuals. Lateral-flow immunoassay differentiated cFA and pFA patients from controls, whereas determination of mRNA in q-PCR was sensitive and specific only in cFA.
authors.affiliation: Department of Neurological Sciences, University Federico II, Naples, Italy. francesco.sacca@unina.it
authors.forename: Francesco
authors.initials: F
authors.lastname: Saccà
authors.forename: Giorgia
authors.initials: G
authors.lastname: Puorro
authors.forename: Antonella
authors.initials: A
authors.lastname: Antenora
authors.forename: Angela
authors.initials: A
authors.lastname: Marsili
authors.forename: Alessandra
authors.initials: A
authors.lastname: Denaro
authors.forename: Raffaele
authors.initials: R
authors.lastname: Piro
authors.forename: Pierpaolo
authors.initials: P
authors.lastname: Sorrentino
authors.forename: Chiara
authors.initials: C
authors.lastname: Pane
authors.forename: Alessandra
authors.initials: A
authors.lastname: Tessa
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Brescia Morra
authors.forename: Sergio
authors.initials: S
authors.lastname: Cocozza
authors.forename: Giuseppe
authors.initials: G
authors.lastname: De Michele
authors.forename: Filippo M
authors.initials: FM
authors.lastname: Santorelli
authors.forename: Alessandro
authors.initials: A
authors.lastname: Filla
mesh_terms: D000328:Adult; D002986:Clinical Trials as Topic; D003710:Demography; D005621:Friedreich Ataxia / Q000175:diagnosis* / Q000235:genetics*; D005786:Gene Expression Regulation; D005838:Genotype; D006579:Heterozygote; D006801:Humans; D033862:Iron-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D008875:Middle Aged; D016011:Normal Distribution; D017354:Point Mutation / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000096923:Frataxin
chemical_list: D033862:Iron-Binding Proteins; D012333:RNA, Messenger; D000096923:Frataxin