title: Significance of allelic percentage of BRAF c.1799T > A (V600E) mutation in papillary thyroid carcinoma.
abstract: BACKGROUND
  Somatic BRAF mutation is frequently observed in papillary thyroid carcinoma (PTC). Recent evidence suggests that PTCs are heterogeneous tumors containing a subclonal or oligoclonal occurrence of BRAF mutation. Conflicting results have been reported concerning the prognostic significance of the mutant allele frequency. Our present aim was to investigate the association between the percentage of BRAF c.1799T > A (p.Val600Glu) alleles and clinicopathological parameters in PTC.
  
  
  METHODS
  Genomic DNA was extracted from fresh-frozen specimens obtained from 50 PTC patients undergoing total thyroidectomy. The BRAF mutation status was determined by Sanger sequencing. The percentage of mutant BRAF alleles was quantified by mass spectrometric genotyping, pyrosequencing, and competitive allele-specific TaqMan PCR (castPCR).
  
  
  RESULTS
  Positive rate of BRAF mutation was 72 % by Sanger sequencing, 82 % by mass spectrometric genotying, and 84 % by pyrosequencing or castPCR. The average percentage of mutant BRAF alleles was 22.5, 31, and 30.7 %, respectively. There was a good correlation among three quantification methods (Spearman's rho = 0.87-0.97; p < 0.0001). The mutant allele frequency was significantly correlated with tumor size (rho = 0.47-0.52; p < 0.01) and extrathyroidal invasion. The frequency showed no difference in pathological lymph node metastasis.
  
  
  CONCLUSIONS
  The percentage of mutant BRAF alleles is positively associated with tumor burden and extrathyroidal invasion in PTC. Relatively good correlations exist among mass spectrometric genotyping, pyrosequencing, and castPCR in quantification of mutant BRAF allele frequency.
authors.affiliation: Mackay Junior College of Medicine, Nursing, and Management, Taipei, Taiwan.
authors.forename: Shih-Ping
authors.initials: SP
authors.lastname: Cheng
authors.forename: Yi-Chiung
authors.initials: YC
authors.lastname: Hsu
authors.forename: Chien-Liang
authors.initials: CL
authors.lastname: Liu
authors.forename: Tsang-Pai
authors.initials: TP
authors.lastname: Liu
authors.forename: Ming-Nan
authors.initials: MN
authors.lastname: Chien
authors.forename: Tao-Yeuan
authors.initials: TY
authors.lastname: Wang
authors.forename: Jie-Jen
authors.initials: JJ
authors.lastname: Lee
mesh_terms: D000328:Adult; D000483:Alleles; D002277:Carcinoma / Q000235:genetics* / Q000473:pathology*; D002291:Carcinoma, Papillary; D005260:Female; D005787:Gene Frequency; D005838:Genotype; D060005:Genotyping Techniques; D006801:Humans; D008297:Male; D013058:Mass Spectrometry; D008875:Middle Aged; D009154:Mutation; D009361:Neoplasm Invasiveness / Q000235:genetics; D016133:Polymerase Chain Reaction; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics*; D017422:Sequence Analysis, DNA / Q000379:methods; D000077273:Thyroid Cancer, Papillary; D013964:Thyroid Neoplasms / Q000235:genetics* / Q000473:pathology*; D047368:Tumor Burden / Q000235:genetics
chemical_list: C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf