The primary occurrence of BRAF(V600E) is a rare clonal event in papillary thyroid carcinoma.

CONTEXT: BRAF(V600E) is considered a primary event, a negative prognostic marker, and a site for pharmacological intervention in papillary thyroid carcinoma (PTC). We asked whether BRAF(V600E) can occur as a subclonal event in PTC and whether this and other oncogenes can coexist in the same tumor.

STUDY

DESIGN

We determined by pyrosequencing the percentage of mutant BRAF, NRAS, and KRAS alleles in a series of conventional PTC. We also analyzed the BRAF mutation status in PTC cell clones in culture.

RESULTS

BRAF(V600E) alleles were present in 41 of 72 PTC (56.9%) in the range 44.7 to 5.1% of total BRAF alleles. In four PTC samples, mutant BRAF alleles were about 50%, being therefore compatible with a clonal heterozygous mutation. In 27 PTC samples, BRAF(V600E) alleles were in the range of 25 to 5.1%. This finding was confirmed after exclusion of the presence of a large contamination by lymphoreticular cells and by the analysis of PTC cells selected by laser capture. Analysis of clones derived from a single cell confirmed the presence of two distinct PTC populations with wild-type or mutated BRAF. Simultaneous subclonal BRAF and KRAS mutations were demonstrated in two PTC.

CONCLUSIONS

These data demonstrate that clonal BRAF(V600E) is a rare occurrence in PTC, although frequently this cancer consists of a mixture of tumor cells with wild-type and mutant BRAF. These results suggest that BRAF mutation in PTC is a later subclonal event, its intratumoral heterogeneity may hamper the efficacy of targeted pharmacotherapy, and its association with a more aggressive disease should be reevaluated.