Clinical Significance Of BRAF Gene Mutation And Expression In Papillary Thyroid Carcinoma

Papillary thyroid carcinoma is the most common thyroid neoplasm, which accounts for 80% of all thyroid cancers. It is demonstrated that RET/PTC and TRK rearrangements have been characterized by papillary thyroid carcinoma (PTC) and have been found to be involved in about 20-40% of cases.In contrast to this, proto-oncogene Ras mutations and oncogenic PAX8-PPARγfusions have been found in follicular thyroid carcinoma (FTC). In the past several years, many researchers were interested in the BRAF mutation status in papillary thyroid carcinoma and corresponding cell lines. Several studies have demonstrated that BRAF mutation was restricted in papillary thyroid carcinoma and had a high prevalence in PTC. However, BRAF mutation was not detected in benign thyroid lesions and other malignant thyroid carcinomas.RAF gene is a vital component in the RET/PTC-RAS-RAF-MEK/ERK-MAPK signaling pathway, which plays a central role in mediating the cellular response to extracellular signals that regulate cell growth, division, and proliferation. There are three isoforms of the RAF kinase: ARAF, BRAF, and CRAF. BRAF is a cytoplasmic serine/threonine kinase. Among three isoforms of RAF kinase, the BRAF is the strongest activator of the downstream MAPK signaling pathway. Lots of studies indicated that this pathway played an important role in formation and progression of papillary thyroid cancer. When this pathway was constitutively activated, it caused tumorigenesis. Recent studies have examined the role of BRAF gene mutation in PTC and found that there was an activating mutation. BRAF mutation is a point mutation with thymine-to-adenine transversion at nucleotide position 1799 of the 15 exon, which resulted in a valine to-glutamate substitution and led to the activation of downstream genes in MAPK signaling pathway or elevated phosphorylated protein expression. BRAF mutation occurred in papillary thyroid cancer (PTC) with a prevalence ranging from 29 to 70%. This mutation was not detected in benign thyroid lesions, such as nodular goiter and thyroid adenoma, and follicular thyroid carcinoma (FTC).Materials and MethodsIn this experiment, 112 cases of paraffin-embeded thyroid lesions were collected, which were made up of 63 cases of PTC, 21 cases of FTC, 2 cases of thyroid adenomas, 26 cases of nodular goiters. B-raf protein expression was immunohistochemically examined in 112 cases of thyroid lesions using Envision method. At the same time, 65 from these samples were also analyzed for the point mutation of BRAF gene, consisting of 46 cases of PTC, 6 cases of FTC and 13 cases of nodular goiters, by sequencing the PCR-amplified products of exon 15 of the BRAF gene. BRAF mutation status was determined by comparison of the sequencing results and the gene bank. The results of immunohistochemistry were determined by the positive cytoplasmic staining and the percentage of the positive cells.ResultsBRAF mutation was detected in 21 of 46 PTC (45.7%). All mutations were heterozygous BRAF mutation that lied in nucleotide position 1799 of exon 15. The mutation was not found in 6 FTC and 13 nodular goiters. Our result was consistent with the studies of published English literatures. Statistical analysis demonstrated that the difference between PTC and nodular goiters in BRAF mutation was significant (P<0.05) . However, there was no significant difference between PTC and FTC (P>0.05) (showed in table 1). BRAF mutation in PTC was not correlated with age, gender, histological type, regional lymph node metastasis and TNM staging (showed in table 2). Table 1 BRAF gene mutation in 65 thyroid lesionsTable 2 Correlation between clinicopathological characteristics and BRAF mutation status in PTC(B-raf protein expression was not detected in one of the 46 PTC)There were few reports about B-raf protein expression in thyroid cancer and correlation between B-raf expression and BRAF gene mutation. We examined B-raf protein expression in 112 cases of thyroid lesions using immunohistochemistry. The positive rate of B-raf expression in PTC,FTC, nodular goiters, thyroid adenoma and normal thyroid tissues were 65%,47.6%,15.4%,0 and 0, respectively. The difference between PTC and nodular goiters, adenoma, normal thyroid tissues were statistically significance (P<0.05). But there was no statistically significance referring to PTC and FTC. (showed in table 3). The prevalence of BRAF gene mutation in positive B-raf expression group was much higher than in negative group. Statistical analysis showed that there was correlation between BRAF gene mutation and B-raf protein expression (P<0.05) (showed in table 3)。Table 3 B-raf protein expression in 112 thyroid lesionsConclusion1,BRAF gene mutation was only detected in papillary thyroid carcinoma, while it was not found in follicular thyroid carcinoma and nodular goiter. It is suggested that BRAF gene mutation should be specific for PTC and plays an important role in the tumorigenesis of papillary thyroid cancer.2,B-raf protein expression in papillary and follicular thyroid cancinomas was much higher than that in benign thyroid lesions, which showed that the detection of B-raf expression should be a diagnostic marker in the pathological diagnosis of papillary and follicular thyroid cancers.3,There was a significant correlation between B-raf protein expression and BRAF gene mutation in papillary thyroid cancer, but no relationship between B-raf protein expression and BRAF gene mutation was found in follicular thyroid cancer, which indicated that the immunohistochemistry for B-raf antibody commercially used in the present study could not be substituted for the direct detection of BRAF gene mutation.