BRAF mutations are associated with some histological types of papillary thyroid carcinoma

Mutations in the BRAF gene have recently been detected in a wide range of neoplastic lesions with a particularly high prevalence in melanoma and papillary thyroid carcinoma (PTC). The hot-spot mutation BRAF(V599E) is frequently detected in PTC (36-69%), in contrast to its absence in other benign or malignant thyroid lesions. In order to unravel whether there is any association between the occurrence of the BRAF mutation and the histological pattern of PTC, in this study a previous series of 50 PTCs was extended to 134 cases, including ten cases of PTC-related entities-hyalinizing trabecular tumour (HTT) and mucoepidermoid carcinoma (MEC). Using PCR/SSCP and sequencing, the BRAF(V599E) mutation was detected in 45 of the 124 PTCs (36%). No mutations were detected in any case of HTT and MEC. BRAF(V599E) was present in 75% of Warthin-like PTCs and 53% of conventional PTCs, whereas no BRAF(V599E) mutations were detected in any of the 32 cases of the follicular variant of PTC. BRAF(V599E) was also detected in 6 of 11 cases of the oncocytic variant of PTC that displayed a papillary or mixed follicular-papillary growth pattern and in none of the four oncocytic PTCs with a follicular growth pattern. A distinct mutation in BRAF (codon K600E) was detected in three cases of the follicular variant of PTC. This study has confirmed the high prevalence of BRAF(V599E) in PTC and has shown that the mutation is almost exclusively seen in PTC with a papillary or mixed follicular-papillary growth pattern, regardless of the cytological features of the neoplastic cells. The results support the existence of an oncocytic variant of PTC that should be separated from the oncocytic variant of follicular carcinoma and suggest that the follicular variant of PTC may be genetically different from conventional PTC.     

The p75 neurotrophin receptor is widely expressed in conventional papillary thyroid carcinoma

Papillary thyroid carcinomas (PTCs) are associated with alterations in several proto-oncogenes related with nervous system development and function, such as TrkA and RET, which are commonly rearranged in these carcinomas. The other oncogenic event recently identified in PTC is the BRAF V600E mutation. Because the role of TrkA was not completely elucidated in thyroid cancer ethiopathogenesis, we decided to study the expression of active, phosphorylated TrkA and of its coreceptor p75 neurotrophin receptor (p75 NTR) in a series of 92 PTC (37 lesions of conventional PTC, 28 of follicular variant of PTC [FVPTC], and 27 of other variants of PTC) as well as in 21 samples of normal thyroid and nonneoplastic thyroid lesions used as a controls. We observed neoexpression of p75 NTR in PTC, particularly in conventional PTC and in other variants of PTC displaying a papillary growth pattern, rather than in FVPTC. No immunoexpression of p75 NTR was observed in normal thyroid nor in nonneoplastic thyroid lesions. The cellular localization of p75 NTR immunoexpression was also significantly associated with the growth pattern of PTC, being much more frequently detected in an apical localization in PTC with papillary architecture than in PTC with a follicular or solid growth pattern. This apical localization of p75 NTR was significantly associated with the presence of BRAF V600E. No significant differences were detected between normal thyroid, nonneoplastic lesions, and PTC (or any PTC variant) regarding expression/activation of TrkA, thus suggesting that by itself and in contrast to p75 NTR, TrkA is not altered during PTC development.     

分化型甲状腺癌中BRAF V600E突变分析

目的探讨分化型甲状腺癌中BRAF V600E突变与临床病理特征的关系。方法收集甲状腺乳头状癌(papillary thyroid carcinoma,PTC)80例(其中经典型67例、滤泡亚型8例、嗜酸细胞亚型3例、高细胞亚型2例)、滤泡癌5例,其中30例PTC取相应癌旁组织,全部送基因检测室检测BRAF V600E突变情况。结果 80例PTC中BRAF V600E突变率为65.0%,5例滤泡癌及30例癌旁组织中未发现BRAF V600E突变;BRAF V600E突变与患者年龄、肿瘤包膜侵犯、淋巴结转移及TNM分期有关。PTC亚型中,经典型和高细胞亚型的BRAF V600E突变率较高(70.1%、100.0%),滤泡亚型的突变率较低(33.3%)。结论PTC中BRAF V600E突变可能与患者年龄有一定相关性,还与包膜侵犯、淋巴结转移及TNM分期有关,经典型和高细胞亚型的BRAF V600E突变率较高,明显高于滤泡亚型。     

<Emphasis Type="Italic">BRAF</Emphasis> mutations typical of papillary thyroid carcinoma are more frequently detected in undifferentiated than in insular and insular-like poorly differentiated carcinomas

Somatic mutations of the BRAF gene (BRAF^V599E and BRAF^K600E) were found to be closely associated with different histotypes of papillary thyroid carcinoma (PTC). The V599E mutation is highly prevalent in PTC with a papillary or mixed papillary follicular growth pattern, and the K600E mutation is apparently restricted to the follicular variant of PTC. It is usually accepted that thyroid malignancies may follow a progression path from well-differentiated to poorly differentiated (PDC) and undifferentiated (UC) carcinomas. One would expect that at least some of the less differentiated carcinomas would harbour the genetic alterations of pre-existing well-differentiated tumours. In order to find the prevalence of BRAF mutations in PDC and UC, we screened a series of 19 PDCs and 17 UCs, as well as 3 UC-derived cell lines, for both mutation types. The group of PDCs was restricted to the so-called insular and insular-like PDCs, thus excluding PTCs with solid, insular or trabecular foci of growth and PDCs displaying typical PTC nuclei. No BRAF mutations were detected in any of the 19 cases of PDC, whereas 6 of the UCs (35%) and one UC-derived cell line presented the BRAF^V599E mutation. The BRAF^K600E mutation was not detected in any case. We conclude that UC may progress from BRAF^V599E-mutated PTC. The absence of BRAF mutations in our series of PDC supports the assumption that pure insular and insular-like PDCs are more closely related to follicular carcinoma than to PTC.Paula Soares and Vítor Trovisco contributed equally to this work.     

Stomatin‐like protein 2 overexpression in papillary thyroid carcinoma is significantly associated with high‐risk clinicopathological parameters and BRAFV600E mutation

Stomatin‐like protein 2 (SLP‐2), a member of the stomatin protein family, has emerged as a potential molecular hallmark of tumor progression in several human malignancies. The aim of this study was to analyze SLP‐2 expression pattern in benign and malignant thyroid tumors (n = 210) and to examine its relationship with clinicopathological parameters and BRAFV600E mutation in thyroid cancer. SLP‐2 immunohistochemical expression was not detected in benign adenomas and was absent/weak in follicular and anaplastic carcinomas. High expression levels of SLP‐2, found only in papillary thyroid carcinoma (PTC), particularly in the classical variant, were significantly associated with adverse clinicopathological parameters: lymph node metastasis (p = 0.002), extrathyroid invasion (p < 0.001), pT status (p < 0.001), and advanced tumor stage (p = 0.001). Additional genotyping of PTC cases for the BRAFV600E mutation revealed for the first time a close relation between SLP‐2 overexpression and the presence of BRAF mutation (p = 0.02) with high positive rates of lymph node metastasis (70%) and extrathyroid invasion (80%) in these cases. The significant association of SLP‐2 overexpression with unfavorable clinicopathological characteristics and BRAFV600E mutation indicates that SLP‐2 may have a role in aggressiveness of BRAF‐mutated PTC and that SLP‐2 evaluation could be clinically useful in identification of high‐risk PTC patients.     

A Novel Complex BRAF Mutation Detected in a Solid Variant of Papillary Thyroid Carcinoma

BRAF gene mutations are identified in about 45% of papillary thyroid carcinomas (PTC) and represent the most common genetic event in this tumor. Here, we report a case of PTC, solid variant, with a complex BRAF mutation that involves one nucleotide substitution, C1796T, and a CTT triplet insertion, 1798_1799insCTT, located on the same allele. This mutation leads to the replacement of a threonine with an isoleucine, T599I, and replacement of a valine with an alanine and a leucine, V600delinsAL. This mutation was identified both in the preoperative fine needle aspirate sample and in the surgical specimen after total thyroidectomy. Other rare BRAF mutations in PTC are reviewed.     

The Relationship of the BRAFV600E Mutation and the Established Prognostic Factors in Papillary Thyroid Carcinomas

It has been shown that BRAF(V600E) mutation in papillary thyroid carcinomas (PTC) is associated both with pathogenesis and poor prognosis. In this study, we aimed to investigate the relationship of the BRAF(V600E) mutation and the established prognostic factors in a cohort of Turkish patients with PTC. Forty-six cases of papillary thyroid carcinoma have been evaluated for the presence of BRAF(V600E) mutation. BRAF(V600E) has been examined by restriction fragment length polymorphism. BRAF(V600E) mutation status has been compared with well-known histopathological and clinical prognostic parameters such as invasion of thyroid capsule, extrathyroidal extension, and the presence of lymph node and/or distant metastasis. We have found that BRAF(V600E) mutation was present in the majority of our cases (40/46). Considering the stage of the disease, five of the negative cases were in stage 1 while the remaining one was in stage 2. Only one BRAF(V600E) negative case has shown extrathyroidal extension and lymph node metastasis. All four patients with distant metastasis had BRAF(V600E) mutation. Statistical analyses revealed that there are no significant relationship between the BRAF(V600E) mutation and the established prognostic factors. We found a relatively higher BRAF(V600E) mutation rate in classical type PTC than in other similar studies. We think that the limited number of our cases may either weaken or mask some potentially important relationship between BRAF(V600E) mutation and the established prognostic factors.     

Correlation between BRAF mutation and the clinicopathological parameters in papillary thyroid carcinoma with particular reference to follicular variant

Mutation of the BRAF gene is common in thyroid cancer. Follicular variant of papillary thyroid carcinoma is a variant of papillary thyroid carcinoma that has created continuous diagnostic controversies among pathologists. The aims of this study are to (1) investigate whether follicular variant of papillary thyroid carcinoma has a different pattern of BRAF mutation than conventional papillary thyroid carcinoma in a large cohort of patients with typical features of follicular variant of papillary thyroid carcinoma and (2) to study the relationship of clinicopathological features of papillary thyroid carcinomas with BRAF mutation. Tissue blocks from 76 patients with diagnostic features of papillary thyroid carcinomas (40 with conventional type and 36 with follicular variant) were included in the study. From these, DNA was extracted and BRAF V600E mutations were detected by polymerase chain reaction followed by restriction enzyme digestion and sequencing of exon 15. Analysis of the data indicated that BRAF V600E mutation is significantly more common in conventional papillary thyroid carcinoma (58% versus 31%, P = .022). Furthermore, the mutation was often noted in female patients (P = .017), in high-stage cancers (P = .034), and in tumors with mild lymphocytic thyroiditis (P = .006). We concluded that follicular variant of papillary thyroid carcinoma differs from conventional papillary thyroid carcinoma in the rate of BRAF mutation. The results of this study add further information indicating that mutations in BRAF play a role in thyroid cancer development and progression.     

Molecular genetic study comparing follicular variant versus classic papillary thyroid carcinomas: association of N-ras mutation in codon 61 with follicular variant

Although the follicular variant of papillary thyroid carcinoma (FVPTC) has been classified as a papillary cancer based on nuclear features, its follicular growth pattern and potential for hematogenous spread are more characteristic of follicular carcinoma. To gain insight into the biologic nature of FVPTC, we compared genetic alterations characteristic of papillary and follicular thyroid carcinomas in 24 FVPTCs and 26 classic PTC (CPTCs). In FVPTCs, we observed ras mutation in 6 of 24 cases (25%), BRAF mutation in 1 of 13 cases (7.6%), and ret rearrangement in 5 of 12 cases (41.7%). In CPTCs, we found ras mutation in no case, BRAF mutation in 3 of 10 cases (30%), and ret rearrangement in 5 of 11 cases (45%). One FVPTC exhibited simultaneous ras mutation and ret/PTC1 rearrangement, and one CPTC harbored simultaneous BRAF mutation and ret/PTC3 rearrangement. Based on these findings, we concluded that ras mutation correlates with follicular differentiation of thyroid tumors whereas ret activation is associated with papillary nuclei but not with papillary architecture. ret activation is not exclusive of ras or BRAF mutation, whereas ras and BRAF mutations are mutually exclusive. The implications of these results for follicular and papillary carcinogenesis are discussed.     

Alterations of the BRAF gene in thyroid tumors

BRAF belongs to the RAF family of protein kinases that are important components of the MAPK signaling pathway mediating cell growth, differentiation and survival. Activating point mutation of the BRAF gene resulting in V600E (previously designated as V599E) is a common event in thyroid papillary carcinoma, being found in approx 40% of this tumor. It has strong association with classical papillary carcinoma and tall cell and possibly Warthin-like variants. This mutation also occurs in thyroid poorly differentiated and anaplastic carcinomas, usually those containing areas of papillary carcinoma. Alterations in the BRAF gene do not overlap with RAS mutations and RET/PTC rearrangement, indicating that activation of one of the effectors of the MAPK pathway is sufficient for papillary thyroid carcinogenesis. Recently, another mechanism of BRAF activation has been identified, which involves chromosome 7q inversion that results in the AKAP9-BRAF fusion. It is rare in sporadic papillary carcinomas and is more common in tumors associated with radiation exposure. Yet another mechanism of BRAF activation may involve copy number gain, which is seen in a significant portion of thyroid follicular tumors of both conventional and oncocytic (Hürthle cell) types.     

Occult oncocytic papillary thyroid carcinoma with lymphoid stroma (Warthin-like tumor): report of a case with concomitant mutations of BRAF V600E and V600K

Warthin-Like tumor of the thyroid is a recently described rare variant of papillary thyroid cancer. The distinct histological feature of this variant is papillary architecture lining oncocytic epithelial cells with nuclear characteristics of papillary carcinoma, accompanied by prominent lymphocytic infiltration in the papillary stalks. Here, we present a case of occult Warthin-like papillary thyroid carcinoma, 0.5-cm in maximum dimension, underwent left thyroid lobectomy in a 65 years old Chinese woman. In this case, there was no extrathyroid extension, vascular invasion and lymphatic metastasis, as well as no complication of lymphocytic thyroiditis. Immunohistochemistry staining revealed that the tumor cells were positive for Leu-M1, HBME-1, 34βE12, and MIB-1 labeling index was low. RET/PTC expression was absent in tumor cells. Furthermore, activated point mutations of BRAF V600E and V600K were concurrently detected by DNA sequencing. Further studies are needed to elucidate the prevalence and role of BRAF^V600K mutation in papillary thyroid carcinoma, and long-term follow-up for the patient is needed to clarify the biological behavior of this variant with dual BRAF mutations.     

TGF-beta/Smad pathway and BRAF mutation play different roles in circumscribed and infiltrative papillary thyroid carcinoma

Poorly circumscribed growth pattern, extra-thyroid extension and high intratumoural lymph vessel density are significantly associated to nodal metastatization in papillary thyroid carcinoma (PTC). It was also shown that transforming growth factor beta (TGF-beta)/Smad-dependent pathway activity is associated with local invasion, nodal metastatization and BRAF-mutated PTCs. We analysed the immunoexpression of TGF-beta, Smad2/Smad3, Smad4 and Smad7 in a series of 42 cases of classic PTC and 33 cases of follicular variant of PTC with known clinico-pathological and follow-up data, as well as BRAF and RAS status. The 75 PTCs were divided into poorly circumscribed (PCPTC) (n?=?53) and well circumscribed (WCPTC) (n?=?22) according to their borders. Nodal metastases were not detected in any WCPTC regardless of the presence of immunoexpression for TGF-beta, Smad2/Smad3, Smad4 and Smad7 and occurrence of BRAF mutation (in 20?% of WCPTCs). Increased cytoplasmatic expression of TGF-beta at the periphery of PCPTC was associated to morphological features of invasiveness, featuring the so-called epithelial-to-mesenchymal transition (EMT), and presence of nodal metastases, as well as to the occurrence of BRAF mutation which did not significantly alter, per se, the frequency of nodal metastases. The nuclear expression of Smad7 was more frequent in WCPTCs than in PCPTCs and was associated with unicentricity and absence of extra-thyroid extension, vascular invasion and nodal metastases. We conclude that nodal metastases are associated to poorly circumscribed, locally invasive PTCs that exhibit low levels of nuclear Smad7 and a peripheral EMT phenotype displaying TGF-beta overexpression, regardless of the occurrence of BRAF mutation.     

Mutations of the BRAF gene in papillary thyroid carcinoma and in Hashimoto's thyroiditis

The purpose of the present study was to investigate the frequency of BRAF mutations in human papillary thyroid carcinoma (PTC) and Hashimoto's thyroiditis (HT) and to evaluate the association of the BRAF mutation with the clinicopathological features of both of these thyroid disorders. A total of 51 PTC with no HT, 28 PTC with HT and 27 HT with no PTC were evaluated using DNA extracted from paraffin-embedded specimens. BRAF mutations were analyzed by direct DNA sequencing of the polymerase chain reaction (PCR)-amplified exon 15. The BRAF missense mutation at codon 599 (T1796A) was present in 46 of 51 PTC (90%) with no HT, 18 of 28 PTC (64%) with HT, four of 28 HT (14%) with PTC, and zero of 27 HT with no PTC. The BRAF mutation at codon 600 (A1798G) was not detected in any case. Clinicopathological examination of 106 patients with either PTC or HT showed that the BRAF mutation was significantly correlated with patient age. These data indicate that the BRAF mutation is associated with a valuable biological property of PTC and may participate in the pathogenesis of PTC arising in HT. These results indicate that the detection of the BRAF mutation in HT can be helpful for prediction of progress to PTC.     

Investigation of BRAF V600E detection approaches in papillary thyroid carcinoma

Objective

The detection of BRAF V600E mutation in papillary thyroid carcinoma (PTC) may be helpful to offer diagnostic confirmation. Additionally, such detection may provide a targeted therapeutic approach for the radioactive iodine resistant patients to predict adverse outcomes. To compare the results of immunohistochemistry (IHC) method using the anti-BRAF V600E (VE1) antibody with the Quantitative real-time polymerase chain reaction (qPCR) approach in examining BRAF V600E mutation in PTC, we investigated the sensitivity and specificity of BRAF V600E (clone VE1) mouse monoclonal antibody in detecting the BRAF V600E mutation and correlated BRAF V600E mutation with clinicopathologic features in PTC.

Immunohistochemistry is highly sensitive and specific for detecting the BRAF V600E mutation in papillary thyroid carcinoma

The V600E mutation in the B-type Raf kinase (BRAF) gene is a common genetic change in cases of papillary thyroid carcinoma (PTC) that appears to play a key role in the development and progression of this disease. We sought to assess the sensitivity and specificity of immunohistochemical detection of this mutation with a V600E mutated BRAF antibody in a Chinese PTC cohort. In this study, we used fully automated immunohistochemistry (IHC) assay with a BRAF V600E (VE1) mouse monoclonal primary antibody to screen for the BRAF V600E mutation in 556 cases of PTC. Moreover, to verify the IHC staining results, real-time PCR was applied to detect this mutation in the same patient cohort. Among the 556 cases in the examined primary PTC cohort, 414 (74.5%) cases and 419 (75.4%) cases were positive for the BRAF V600E mutation by IHC staining and by real-time PCR, respectively. The real-time PCR results indicated that the sensitivity and specificity of IHC staining for the BRAF V600E mutation were 98.8% and 100%, respectively. The BRAF V600E mutation was common among Chinese patients with primary PTC, and was strongly correlated with older patient age and the conventional subtype of PTC but was not associated with parameters of clinicopathological aggressiveness. The fully automated IHC is a reliable technique that can serve as an alternative to molecular biological approaches for the routine detection of the BRAF V600E mutation in PTC patients.     

Synchronous Metastatic Medullary and Papillary Thyroid Carcinomas in a Patient with Germline RET Mutation: Case Report,Molecular Analysis,and Implications for Pathogenesis

The occurrence of medullary thyroid carcinoma (MTC) in patients with germline RET mutation is well established, but the occasional occurrence of papillary thyroid carcinoma (PTC) in this setting remains unexplained. We report a case of synchronous MTC and PTC in a patient with germline RET mutation, and investigate the molecular pathogenesis of these two tumors. A 48-year-old man presented with cervical lymphadenopathy and was found to have metastatic MTC involving cervical and mediastinal lymph nodes. He underwent thyroidectomy and debulking of metastatic disease. The resected thyroid showed bilateral MTC and a 1-cm PTC. His lymph node metastases were predominantly MTC along with a small focus of metastatic PTC. Molecular testing using peripheral blood revealed a germline RET point mutation (Val804Met). Both tumors were analyzed for the BRAF (Val600Glu) mutation and the RET/PTC1 translocation. The PTC was positive for the BRAF mutation but negative for RET/PTC1. The MTC was negative for both abnormalities. We conclude that the MTC was caused by the germline RET mutation, while the PTC was caused by a somatic BRAF mutation. The occurrence of PTC in patients with germline RET mutations appears to be purely coincidental.     

甲状腺乳头状癌BRAF基因突变的检测及临床意义

目的 了解甲状腺乳头状癌(papillary thyroid carcinoma,PTC)BRAF基因T1799A点突变的情况与临床病理学特征的关系.方法 应用聚合酶链反应及DNA直接测序法对43例PTC患者,20例非PTC甲状腺病变患者及40份正常甲状腺组织对照的新鲜标本进行BRAF基因检测.分析BRAF基因突变与性别、发病年龄、原发灶大小、甲状腺包膜外浸润、颈淋巴结转移及远处转移等临床病理学特征的关系.结果 43例PTC中1 7例检出BRAF基因T1799A点突变.检出率为39.5%,而在非PTC甲状腺病变患者和40份正常甲状腺组织未发现T1799A点突变.BRAF基因突变与PTC甲状腺包膜外浸润及颈淋巴结转移密切相关(P<0.05及P<0.05),与性别、发病年龄、原发灶大小及远处转移无关.结论 BRAF基因突变与颈淋巴结转移和甲状腺包膜外浸润密切相关,突变可能增加PTC的侵袭性并影响预后.     

甲状腺乳头状癌BRAF基因突变的检测及临床意义

目的 了解甲状腺乳头状癌(papillary thyroid carcinoma,PTC)BRAF基因T1799A点突变的情况与临床病理学特征的关系.方法 应用聚合酶链反应及DNA直接测序法对43例PTC患者,20例非PTC甲状腺病变患者及40份正常甲状腺组织对照的新鲜标本进行BRAF基因检测.分析BRAF基因突变与性别、发病年龄、原发灶大小、甲状腺包膜外浸润、颈淋巴结转移及远处转移等临床病理学特征的关系.结果 43例PTC中1 7例检出BRAF基因T1799A点突变.检出率为39.5%,而在非PTC甲状腺病变患者和40份正常甲状腺组织未发现T1799A点突变.BRAF基因突变与PTC甲状腺包膜外浸润及颈淋巴结转移密切相关(P<0.05及P<0.05),与性别、发病年龄、原发灶大小及远处转移无关.结论 BRAF基因突变与颈淋巴结转移和甲状腺包膜外浸润密切相关,突变可能增加PTC的侵袭性并影响预后.     

甲状腺乳头状癌BRAF基因突变的检测及临床意义

目的 了解甲状腺乳头状癌(papillary thyroid carcinoma,PTC)BRAF基因T1799A点突变的情况与临床病理学特征的关系.方法 应用聚合酶链反应及DNA直接测序法对43例PTC患者,20例非PTC甲状腺病变患者及40份正常甲状腺组织对照的新鲜标本进行BRAF基因检测.分析BRAF基因突变与性别、发病年龄、原发灶大小、甲状腺包膜外浸润、颈淋巴结转移及远处转移等临床病理学特征的关系.结果 43例PTC中1 7例检出BRAF基因T1799A点突变.检出率为39.5%,而在非PTC甲状腺病变患者和40份正常甲状腺组织未发现T1799A点突变.BRAF基因突变与PTC甲状腺包膜外浸润及颈淋巴结转移密切相关(P<0.05及P<0.05),与性别、发病年龄、原发灶大小及远处转移无关.结论 BRAF基因突变与颈淋巴结转移和甲状腺包膜外浸润密切相关,突变可能增加PTC的侵袭性并影响预后.