title: Simultaneously Detection of 50 Mutations at 20 Sites in the BRAF and RAS Genes by Multiplexed Single-Nucleotide Primer Extension Assay Using Fine-Needle Aspirates of Thyroid Nodules.
abstract: Fine-needle aspiration (FNA) is commonly used for primary evaluation of thyroid nodules. Twenty to 30 percent of thyroid nodules remain indeterminate after FNA evaluation. Studies show the BRAF p.V600E to be highly specific for papillary thyroid carcinoma (PTC), while RAS mutations carry up to 88 percent positive predictive value for malignancy. We developed a two-tube multiplexed PCR assay followed by single-nucleotide primer extension assay for simultaneous detection of 50 mutations in the BRAF (p.V600E, p.K601E/Q) and RAS genes (KRAS and NRAS codons 12, 13, 19, 61 and HRAS 61) using FNA smears of thyroid nodules. Forty-two FNAs and 27 paired formalin-fixed, paraffin-embedded (FFPE) tissues were tested. All BRAF p.V600E-positive FNA smears (five) carried a final diagnosis of PTC on resection. RAS mutations were found in benign as well as malignant lesions. Ninety-two percent concordance was observed between FNA and FFPE tissues. In conclusion, our assay is sensitive and reliable for simultaneous detection of multiple BRAF/RAS mutations in FNA smears of thyroid nodules.
authors.affiliation: Department of Pathology, University of Iowa Hospitals and Clinics, Iowa City, Iowa.
authors.forename: Aaron A
authors.initials: AA
authors.lastname: Stence
authors.affiliation: Department of Pathology, University of Iowa Hospitals and Clinics, Iowa City, Iowa.
authors.forename: Michael P
authors.initials: MP
authors.lastname: Gailey
authors.affiliation: Department of Pathology, University of Iowa Hospitals and Clinics, Iowa City, Iowa.
authors.forename: Robert A
authors.initials: RA
authors.lastname: Robinson
authors.affiliation: Department of Pathology, University of Iowa Hospitals and Clinics, Iowa City, Iowa.
authors.forename: Chris S
authors.initials: CS
authors.lastname: Jensen
authors.affiliation: Department of Pathology, University of Iowa Hospitals and Clinics, Iowa City, Iowa.
authors.forename: Deqin
authors.initials: D
authors.lastname: Ma
mesh_terms: D044963:Biopsy, Fine-Needle; D020022:Genetic Predisposition to Disease / Q000235:genetics; D006801:Humans; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics*; D013961:Thyroid Gland / Q000473:pathology; D016606:Thyroid Nodule / Q000175:diagnosis / Q000235:genetics* / Q000473:pathology; D020663:ras Guanine Nucleotide Exchange Factors / Q000235:genetics*
chemical_list: D020663:ras Guanine Nucleotide Exchange Factors; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf
keywords: BRAF; RAS; cytology smears; single nucleotide primer extension assay; thyroid nodules