GNAS Mutations in Fibrous Dysplasia: A Comparative Study of Standard Sequencing and Locked Nucleic Acid PCR Sequencing on Decalcified and Nondecalcified Formalin-fixed Paraffin-embedded Tissues

Appl Immunohistochem Mol Morphol. 2016 Oct;24(9):660-667. doi: 10.1097/PAI.0000000000000242.

Abstract

It is well known that fibrous dysplasia (FD) is characterized by the presence of activating mutations involving G-nucleotide binding protein-α subunit (GNAS) involving codon R201 and rarely codon 227 with a mutation frequency between 45% and 93%. Herein, we investigate the sensitivity of detection of GNAS mutations in exons 8 and 9 using a standard and a highly sensitive locked nucleic acid polymerase chain reaction (LNA-PCR) sequencing in 52 cases of FD. In view of the recent report of GNAS mutations in a small number of low-grade osteosarcomas, we also tested in addition 12 cases of low-grade osteosarcomas. GNAS exon 8 mutations p.R201H (31%), p.R201C (15%), and p.R201S (2%) were identified in 50% of FD cases. LNA-PCR sequencing identified only 1 positive case within the mutation negative cases tested by standard PCR and Sanger sequencing. No mutations were identified in any of the low-grade osteosarcomas by standard and LNA-PCR sequencing. There was no association between age, site, size, specimen type, and mutational status. No exon 9 or codon 227 mutations were identified in any of tested cases. There was a significant difference in the sensitivity of the assay between decalcified and nondecalcified FDs (31% vs. 70%, P=0.002). LNA-PCR has no added value in enhancing detection sensitivity for GNAS mutations in FD. In addition to decalcification, innate somatic mosaicism contributes to the decreased sensitivity in mutation detection.

Publication types

  • Comparative Study

MeSH terms

  • Adolescent
  • Adult
  • Aged
  • Bone Neoplasms / genetics*
  • Bone Neoplasms / pathology
  • Calcinosis
  • Child
  • Child, Preschool
  • Chromogranins / genetics*
  • Female
  • Formaldehyde
  • GTP-Binding Protein alpha Subunits, Gs / genetics*
  • Humans
  • Male
  • Middle Aged
  • Mutation*
  • Oligonucleotides / genetics*
  • Paraffin Embedding*
  • Polymerase Chain Reaction
  • Sequence Analysis / methods
  • Young Adult

Substances

  • Chromogranins
  • Oligonucleotides
  • locked nucleic acid
  • Formaldehyde
  • GNAS protein, human
  • GTP-Binding Protein alpha Subunits, Gs