Aberrant BRAF splicing as an alternative mechanism for oncogenic B-Raf activation in thyroid carcinoma.

Activating BRAF mutations have recently been reported in 28-83% of papillary thyroid carcinomas (PTCs). However, it is not known whether aberrant BRAF splicing occurs in thyroid carcinoma. To investigate aberrant BRAF splicing and its association with BRAF mutation in thyroid tumours, we studied aberrant BRAF splicing and BRAF mutation from 68 thyroid tumours. BRAF(V600E) mutation was detected in 20 of 43 PTCs and all three anaplastic thyroid carcinomas (ATCs). There is a higher frequency of BRAF mutation in PTC patients with stage III and IV tumours compared with stage I and II. Novel BRAF splicing variants were detected in 12 PTCs, three follicular variants of PTC (FVPTCs), and one ATC, as well as in two thyroid carcinoma cell lines, ARO and NPA. These variants did not have the N-terminal auto-inhibitory domain of wild-type B-Raf, resulting in an in-frame truncated protein that contained only the C-terminal kinase domain and caused constitutive activation of B-Raf. These variants were significantly associated with advanced disease stage and BRAF(V600E) mutation (p < 0.001, Fisher exact test). Furthermore, expression of these variants in NIH3T3 and CHO cells could activate the MAP kinase signalling pathway, transform them in vitro, and induce tumours in nude mice. These data suggest that BRAF splicing variants may function as an alternative mechanism for oncogenic B-Raf activation. Combination of the BRAF(V600E) mutation and its splicing variants may contribute towards disease progression to poorly differentiated thyroid carcinoma.