Diagnostic value of pyrosequencing for the BRAF V600E mutation in ultrasound-guided fine-needle aspiration biopsy samples of thyroid incidentalomas.

Context: Dideoxy sequencing is the most commonly used method for detecting the BRAF(V600E) mutation in thyroid cancer and melanoma. However, this gold standard method often makes less definite results in detecting the BRAF(V600E) mutation when there are relatively low amounts of the mutant template in biopsy specimens, which are invariably contaminated with normal tissues. Pyrosequencing, which measures the incorporation of each of the four nucleotides at each template position and indicates the amounts of mutant template present, may be more useful in such situations.

Objective: To investigate the diagnostic efficiency of pyrosequencing for the mutant BRAF allele in ultrasound (US)-guided fine needle aspiration biopsies (FNABs) of thyroid incidentalomas.

Design, Setting And Subjects: A total of 101 thyroid incidentaloma cases were included prospectively. Cytological diagnoses of the FNAB samples were made according to the American Thyroid Association (ATA) guidelines, 2006. The presence of the BRAF(V600E) mutation was investigated by pyrosequencing and dideoxy sequencing.

Results: On the basis of cytological analysis, the thyroid incidentalomas were classified into benign (n = 43), malignant (n = 30), indeterminate or suspicious neoplasm (n = 24), and nondiagnostic (n = 4) categories. Pyrosequencing detected the BRAF(V600E) mutation in 30 cases: 22 malignant cases, 7 indeterminate cases, and 1 nondiagnostic case. Dideoxy sequencing also detected the BRAF(V600E) mutation in 28 of the same cases but failed to clearly distinguish the mutant allele from the wild-type allele in one indeterminate case and one nondiagnostic case. Histopathological analysis ascertained that all BRAF(V600E)-positive cases were papillary thyroid carcinomas.

Conclusions: Pyrosequencing may be suitable for detecting the BRAF(V600E) mutation in thyroid incidentaloma and may be superior to dideoxy sequencing when low amounts of the mutant template are present in the biopsy.