Detection of BRAFV600E Mutations in Papillary Thyroid Carcinomas by Peptide Nucleic Acid Clamp Real-Time PCR: A Comparison with Direct Sequencing

BACKGROUND Papillary thyroid carcinoma (PTC) of the thyroid is the most common endocrine malignancy. High prevalence of an activating point mutation of BRAF gene, BRAF(V600E), has been reported in PTC. We assessed the efficiency of peptide nucleic acid clamp real-time polymerase chain reaction (PNAcqPCR) for the detection of BRAF(V600E) mutation in PTC in comparison with direct sequencing (DS). METHODS A total of 265 thyroid lesions including 200 PTCs, 5 follicular carcinomas, 60 benign lesions and 10 normal thyroid tissues were tested for BRAF(V600E) mutation by PNAcqPCR and DS. RESULTS The sensitivity and accuracy of the PNAcqPCR method were both higher than those of DS for the detection of the BRAF(V600E) mutation. In clinical samples, 89% of PTCs harbored the BRAF(V600E) mutation, whereas 5 follicular carcinomas, 50 benign lesions and 10 normal thyroid tissues lacked the mutation. The mutation was associated with aggressive clinical behaviors as extrathyroid invasion (p=0.015), lymph node metastasis (p=0.002) and multiple tumor numbers (p=0.016) with statistical significance. CONCLUSIONS The PNAcqPCR method is efficiently applicable for the detection of the BRAF(V600E) mutation in PTCs in a clinical setting.