Presence Of BRAFV600E Research Articles

Regulation of CEACAM1 Protein Expression by the Transcription Factor ETS-1 in BRAF-Mutant Human Metastatic Melanoma Cells

BRAF becomes constitutively activated in 50% to 70% of melanoma cases. CEACAM1 has a dual role in melanoma, including facilitation of cell proliferation and suppression of infiltrating lymphocytes, which are consistent with its value as a marker for poor prognosis in melanoma patients. Here we show that BRAFV600E melanoma cells treated with BRAF and MEK inhibitors (MAPKi) downregulate CEACAM1 mRNA and protein expression in a dose- and exposure time–dependent manners. Indeed, there is a significant correlation between the presence of BRAFV600E and CEACAM1 expression in melanoma specimens obtained from 45 patients. Vemurafenib-resistant cell systems reactivate the MAPK pathway and restore basal CEACAM1 mRNA and protein levels. These combined results suggest transcriptional regulation. Indeed, luciferase reporting assays show that CEACAM1 promoter (CEACAM1p) activity is significantly reduced by MAPKi. Importantly, we show that the MAPK-driven CEACAM1p activity is mediated by ETS1, a major transcription factor and downstream effector of the MAPK pathway. Phosphorylation mutant ETS1T38A shows a dominant negative effect over CEACAM1 expression. The data are consistent with independent RNAseq data from serial biopsies of melanoma patients treated with BRAF inhibitors, which demonstrate similar CEACAM1 downregulation. Finally, we show that CEACAM1 downregulation by MAPKi renders the cells more sensitive to T-cell activation. These results provide a new view on a potential immunological mechanism of action of MAPKi in melanoma, as well as on the aggressive phenotype observed in drug-resistant cells.

Colonic perineurioma (benign fibroblastic polyp): case report and review of the literature

BackgroundColorectal perineuriomas are uncommon benign mucosal-based proliferations of mesenchymal cells that express perineurial markers, often associated with colonic crypts displaying a serrated/hyperplastic architecture. The vast majority of cases arise distal to the splenic flexure and have been described as sessile polyps. Using molecular analysis, BRAF mutations have been demonstrated in the serrated crypt epithelium. We report a new case of perineurioma presenting as a pedunculated polyp in the transverse colon, with prominent hemosiderin deposits in the uninvolved lamina propria that separated the perineurial proliferation from the surface epithelium, a previously unreported histological finding. By using immunohistochemistry, we demonstrated the presence of BRAF V600E mutated protein in the serrated crypt epithelium. In addition, a review of the literature on colorectal perineurioma is provided.Case presentationA 5 mm pedunculated polyp was removed from the transverse colon of a 42 year old man who presented with epigastric pain, weight loss and rectal bleeding. A proliferation of uniform plump spindled cells expanded the lamina propria and separated serrated colonic crypts. The epithelial component closely resembled microvesicular hyperplastic polyp. Immunohistochemical stains for epithelial membrane antigen (EMA), glucose transporter 1 (GLUT1) and collagen IV were positive in the stromal proliferation. A mutation-specific monoclonal antibody directed against BRAF V600E showed positive cytoplasmic staining in the serrated crypt epithelium but not in the perineurial proliferation. Conspicuous hemosiderin deposition was seen in the inflamed lamina propria between the perineurial proliferation and the surface epithelium.ConclusionAlthough the majority of colorectal perineuriomas occur in the sigmoid colon and rectum and are described as sessile polyps, colorectal perineurioma can present as a pedunculated polyp proximal to the splenic flexure as described in this case. Conspicuous hemosiderin deposition can be seen in the superficial lamina propria. BRAF mutations are limited to the serrated crypt epithelium.

LKB1 loss cooperating with BRAF V600E promotes melanoma cell invasion and migration by up-regulation MMP-2 via PI3K/Akt/mTOR pathway.

The serine/threonine kinase LKB1, act as a tumor suppressor, has been reported in several sporadic cancers. However, how the loss of LKB1 promotes melanoma invasion and metastasis remains incompletely understood. In this study, we inactivated LKB1expression by RNA interference in BRAF mutation and wild type melanoma cells respectively. We found LKB1 inactivation cooperate with BRAF V600E lead to melanoma cells more aggressive by a series of experiments including wound scratch test, Transwell assay. While single alteration, either LKB1 loss or BRAF V600E, fails to enhance melanoma cells invasion ability. Mechanistically, LKB1 loss synergism with BRAF V600E resulted in the activation of the PI3K/Akt/mTOR signaling pathway and significant up-regulation expression of MMP-2. In addition, LKB1 expression in human melanoma tissues was negatively associated with MMP-2 expression in the presence of BRAF V600E. Thus, our findings indicate a probable explanation on LKB1 function as a tumor suppressor in melanoma and a new therapeutic strategy for melanoma by targeting on BRAF and LKB1 together.

Effects of Coexistent BRAFV600E and TERT Promoter Mutations on Poor Clinical Outcomes in Papillary Thyroid Cancer: A Meta-Analysis.

The presence of a telomerase reverse transcriptase (TERT) promoter mutation has been suggested as a potential prognostic marker for thyroid cancer, and a synergistic association with the BRAFV600E mutation has been demonstrated. The aim of this study was to verify the role of this genetic duet in papillary thyroid cancer (PTC). Studies of the association of BRAFV600E and TERT promoter mutations with clinicopathologic features, recurrence, or PTC-related mortality were included from PubMed and Embase databases (inception to September 2016). Thirteen eligible studies incorporating 4347 patients with PTC were included, and 283 (median 8.3%) of these patients had coexistent BRAFV600E and TERT promoter mutations. The coexistence of the two mutations was far more strongly associated with high-risk clinicopathologic features than either mutation alone was, including advanced TNM stage (vs. BRAFV600E: odds ratio [OR] = 4.19 [confidence interval (CI) 3.07-5.71]; vs. TERT: OR = 4.66 [CI 2.67-8.13]), extrathyroidal extension (vs. BRAFV600E: OR = 3.1 [CI 2.2-4.37]; vs. TERT: OR = 5.66 [CI 3.02-10.6]), lymph node metastasis (vs. BRAFV600E: OR = 1.59 [CI 1.16-2.17]; vs. TERT: OR = 2.03 [CI 1.22-3.38]), and distant metastasis (vs. BRAFV600E: OR = 11.76 [CI 5.63-24.58]). The coexistence of the mutations showed the highest risk of recurrence (coexistence vs. no mutations: hazard ratio [HR] = 6.60 [CI 3.82-11.40]; BRAFV600E vs. no mutations: HR = 1.31 [CI 0.49-3.46]; TERT vs. no mutations: HR = 3.38 [CI 0.85-13.35]). Moreover, PTC-related mortality was significantly higher with coexistent mutations than in the presence of BRAFV600E alone (HR = 20.07 [CI 8.37-48.09]). Coexistent BRAFV600E and TERT promoter mutations have a synergistic effect on clinical outcomes in PTC, whereas each mutation alone has a modest effect. Therefore, molecular testing of BRAFV600E and TERT promoter mutations together is useful in assessing risk stratification of PTC.

Trametinib after disease reactivation under dabrafenib in Erdheim-Chester disease with both BRAF and KRAS mutations

AbstractMajor advances have been made in understanding the pathogenesis of Erdheim-Chester disease (ECD) leading to novel treatment strategies. Targeted therapies such as BRAF inhibition have shown a significant impact on disease management, emphasizing the importance of the activated mitogen-associated protein kinase pathway in this disease. However, incomplete responsiveness, potentially limiting adverse effects, and the occurrence of treatment resistance to BRAF inhibition observed in other BRAF-mutant malignancies imply the importance of therapeutic strategies beyond BRAF inhibition. We report a patient with ECD who carried the BRAFV600E mutation and developed treatment resistance under BRAF inhibition despite initial treatment response. Genetic analyses of a newly developing ECD lesion revealed a somatic KRASQ61H mutation without the presence of BRAFV600E. Accordingly, the addition of MEK-inhibiting trametinib to BRAF-inhibiting dabrafenib was able to overcome acquired partial treatment resistance. This is the first report of treatment resistance as a result of a secondary MAPK pathway–activating mutation during BRAF inhibition in ECD. This case contributes to the ongoing efforts of simultaneous BRAF/MEK inhibition as a promising strategy in ECD.

BRAFV600E Accelerates Disease Progression and Increases Immune Suppression in a Mouse Model of B-Cell Leukemia

Background: Mitogen-activated protein kinase (MAPK) pathway components are frequently mutated in cancer, and abnormal MAPK signaling can drive malignancy by promoting tumor survival and proliferation. Furthermore, work in solid tumors shows that the presence of BRAFV600E further enhances tumor-induced immune suppression, but if and how this occurs in a disseminated disease such as leukemia is unknown. MAPK mutations occur in nearly 100% of classic hairy cell leukemia (HCL) and approximately 10% of chronic lymphocytic leukemia (CLL). Furthermore, up to 30% of B cell lymphomas exhibit activated MAPK signaling via expression of a BRAF pseudogene. However, the pathomechanisms of these abnormalities and the clinical utility of MAPK inhibitors in hematologic malignancies are unclear, and few models are available to evaluate this. To study these aspects, we developed in vitro and in vivo models of BRAF-mutated B cell malignancy.Methods: To mechanistically interrogate the role of BRAFV600E in malignant B cells, we transfected the CLL cell line OSUCLL with doxycycline-inducible constructs containing normal BRAF or BRAFV600E and incubated the cells -/+ doxycycline. These cells were also co-cultured with healthy donor T cells pre-incubated with anti-CD3/anti-CD28, and T cell proliferation was measured by flow cytometry. Cytokines and surface proteins were assessed by flow cytometry. To mimic human HCL, we developed a transgenic mouse model of BRAFV600E B cell leukemia. Mice with Cre-activatable BRAFV600E expression were crossed with mice carrying Cre under the control of the CD19 promoter. CD19-Cre x BRAFV600E or CD19-Cre only mice were then crossed with the well-characterized Eµ-TCL1 model of CLL to generate mice with spontaneous B cell leukemia expressing either wild-type BRAF (CD19-Cre x TCL1) or BRAFV600E (BRAFVE x CD19-Cre x TCL1) under the native BRAF promoter. B cell restricted expression of BRAFV600E was confirmed by immunoblots of purified B and T cells using a BRAFV600E specific antibody. Blood and spleen cells were examined by flow cytometry. For adoptive transfer experiments, leukemia cells (2x10^7) from transgenic mice were introduced intravenously into syngeneic healthy adult animals.Results: OSUCLL cells expressing BRAFV600E showed no significant changes in growth vs. wild-type BRAF expressing cells, but more strongly inhibited anti-CD3/CD28-induced proliferation of normal donor T cells. Transwell assays showed this effect was due both to soluble and contact-dependent factors. Expression of PD-L1 was not different between cells expressing normal and mutated BRAF, indicating this common checkpoint molecule is not the reason for greater immune suppression in this context. TNF levels were higher in BRAFV600E expressing cells and reduced by vemurafenib, but a TNF neutralizing antibody did not alter the inhibitory effect of BRAFV600E expressing cells on T cell proliferation. BRAFV600E x CD19-Cre x TCL1 mice developed B cell leukemia significantly earlier (median 4.9 vs. 8.1 months; P<0.001) and had significantly shorter lifespan (median 7.3 vs. 12.1 months; P<0.001) than their wild-type BRAF counterparts. In contrast to effects of BRAFV600E described in some solid tumors, BRAFV600E expression in B cells had no impact on the rate of B cell proliferation in vivo and only modestly reduced spontaneous apoptosis. To study the effects of this activating mutation on tumor-mediated immune suppression in vivo, leukemia cells from BRAFV600E x CD19-Cre x TCL1 or CD19-Cre x TCL1 mice were adoptively transferred into syngeneic mice. In 3 separate studies, tumor cells from BRAFV600E mice produced leukemia (>10% CD5+/CD19+ cells in blood) sooner than CD19-Cre x TCL1 tumor cells. In mice matched for disease load, BRAFV600E B cells produced a greater negative impact on T cells as evidenced by lower overall percentage of T cells, increased expression of T cell exhaustion markers PD-1, CD244, and CD160, and higher percentage of CD44+ memory T cells. Current studies are investigating the mechanism of these effects in vivo as well as the potential for pharmacologic reversal.Conclusions: Together, these results demonstrate the immune-suppressive impact of BRAFV600E in B-cell leukemias and introduce a novel mouse model to develop rational combination strategies to both directly target the tumor cell and overcome tumor-mediated immune evasion. DisclosuresLozanski:Genentech: Research Funding; Stemline Therapeutics Inc.: Research Funding; Beckman Coulter: Research Funding; Boehringer Ingelheim: Research Funding.

RMEL3, a novel BRAFV600E-associated long noncoding RNA, is required for MAPK and PI3K signaling in melanoma.

Previous work identified RMEL3 as a lncRNA with enriched expression in melanoma. Analysis of The Cancer Genome Atlas (TCGA) data confirmed RMEL3 enriched expression in melanoma and demonstrated its association with the presence of BRAFV600E. RMEL3 siRNA-mediated silencing markedly reduced (95%) colony formation in different BRAFV600E melanoma cell lines. Multiple genes of the MAPK and PI3K pathways found to be correlated with RMEL3 in TCGA samples were experimentally confirmed. RMEL3 knockdown led to downregulation of activators or effectors of these pathways, including FGF2, FGF3, DUSP6, ITGB3 and GNG2. RMEL3 knockdown induces gain of protein levels of tumor suppressor PTEN and the G1/S cyclin-Cdk inhibitors p21 and p27, as well as a decrease of pAKT (T308), BRAF, pRB (S807, S811) and cyclin B1. Consistently, knockdown resulted in an accumulation of cells in G1 phase and subG0/G1 in an asynchronously growing population. Thus, TCGA data and functional experiments demonstrate that RMEL3 is required for MAPK and PI3K signaling, and its knockdown decrease BRAFV600E melanoma cell survival and proliferation.

MGL ligand expression is correlated to BRAF mutation and associated with poor survival of stage III colon cancer patients.

Colorectal cancer (CRC) is the third most prevalent cancer type worldwide with a mortality rate of approximately 50%. Elevated cell-surface expression of truncated carbohydrate structures such as Tn antigen (GalNAcα-Ser/Thr) is frequently observed during tumor progression. We have previously demonstrated that the C-type lectin macrophage galactose-type lectin (MGL), expressed by human antigen presenting cells, can distinguish healthy tissue from CRC through its specific recognition of Tn antigen. Both MGL binding and oncogenic BRAF mutations have been implicated in establishing an immunosuppressive microenvironment. Here we aimed to evaluate whether MGL ligand expression has prognostic value and whether this was correlated to BRAF(V600E) mutation status. Using a cohort of 386 colon cancer patients we demonstrate that high MGL binding to stage III tumors is associated with poor disease-free survival, independent of microsatellite instability or adjuvant chemotherapy. In vitro studies using CRC cell lines showed an association between MGL ligand expression and the presence of BRAF(V600E). Administration of specific BRAF(V600E) inhibitors resulted in decreased expression of MGL-binding glycans. Moreover, a positive correlation between induction of BRAF(V600E) and MGL binding to epithelial cells of the gastrointestinal tract was found in vivo using an inducible BRAF(V600E) mouse model. We conclude that the BRAF(V600E) mutation induces MGL ligand expression, thereby providing a direct link between oncogenic transformation and aberrant expression of immunosuppressive glycans. The strong prognostic value of MGL ligands in stage III colon cancer patients, i.e. when tumor cells disseminate to lymph nodes, further supports the putative immune evasive role of MGL ligands in metastatic disease.

Abstract 4697: Quantitation of BRAF V600E alleles predicts papillary thyroid cancer progression

Abstract BACKGROUND: BRAF V600E is the most common genetic alteration in thyroid cancer. However, there is still controversy regarding its clinicopathological significance and clonal mutation frequency. To clarify these inconsistent results, we investigated the association between the allelic frequency of BRAF V600E and the clinicopathological features in classic papillary thyroid carcinoma (PTC). DESIGN: Tumor tissues from 321 patients with classic PTC were microdissected under a dissecting microscope and analyzed for the presence or absence of the BRAF mutation and mutant allelic percentage using quantitative pyrosequencing. RESULTS: BRAF V600E was found in 264 (82.2%) of 321 classic PTC. In the BRAF V600E-positive tumors, the mutant allelic frequency varied from 8% to 41% of total BRAF alleles (median, 20%). The frequency of BRAF V600E alleles directly correlated with tumor size (R-squared = 0.162, P&amp;lt;0.001) and number of metastatic lymph nodes (R-squared = 0.063, P&amp;lt;0.001). Out of 264 BRAF-positive PTCs, 136 (51%) had ≥20% of BRAF mutant alleles. Extrathyroidal extension and lymph node metastasis were more frequent in PTCs with high (≥20%) abundance of mutant alleles than in those with low abundance of mutant alleles (55% vs 37%, P=0.003 and 62% vs 46%, P=0.012, respectively). However, if the mutant allelic percentage was not considered, the presence of BRAF V600E was only associated with extrathyroidal extension, but not with tumor size and lymph node metastasis. CONCLUSION: High abundance of BRAF V600E alleles predicts tumor progression in classic PTC. We suggest that the clonal, subclonal, or oligoclonal occurrence of BRAF V600E mutation may have different biological and clinicopathological significance in the PTC. Citation Format: Chan Kwon Jung, Min Hee Kim, Ja Seong Bae, Dong Jun Lim, Hyoungnam Lee, So Ra Jeon, Gyeong Sin Park. Quantitation of BRAF V600E alleles predicts papillary thyroid cancer progression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4697. doi:10.1158/1538-7445.AM2014-4697

Quantification of BRAF V600E alleles predicts papillary thyroid cancer progression.

The BRAF V600E mutation is the most common genetic alteration in thyroid cancer. However, its clinicopathological significance and clonal mutation frequency remain unclear. To clarify the inconsistent results, we investigated the association between the allelic frequency of BRAF V600E and the clinicopathological features of classic papillary thyroid carcinoma (PTC). Tumour tissues from two independent sets of patients with classic PTC were manually microdissected and analysed for the presence or absence of the BRAF mutation and the mutant allelic frequency using quantitative pyrosequencing. For external validation, the Cancer Genome Atlas (TCGA) data were analysed. The BRAF V600E mutation was found in 264 (82.2%) out of 321 classic PTCs in the training set. The presence of BRAF V600E was only associated with extrathyroidal extension and the absence of thyroiditis. In BRAF V600E-positive tumours, the mutant allelic frequency varied from 8 to 41% of the total BRAF alleles (median, 20%) and directly correlated with tumour size and the number of metastatic lymph nodes. Lymph node metastases were more frequent in PTCs with a high (≥20%) abundance of mutant alleles than in those with a low abundance of mutant alleles (P=0.010). These results were reinforced by validation dataset (n=348) analysis but were not reproduced in the TCGA dataset. In a population with prevalent BRAF mutations, quantitative analysis of the BRAF mutation could provide additional information regarding tumour behaviour, which is not reflected by qualitative analysis. Nonetheless, prospective studies are needed before the mutated allele percentage can be considered as a prognostic factor.

The presence of B-RAF V600E and K601E mutations in our Ligurian population

The presence of B-RAF V600E and K601E mutations in our Ligurian population

PRDX1 and PRDX6 are repressed in papillary thyroid carcinomas via BRAF V600E-dependent and -independent mechanisms

Many clinical studies highlight the dichotomous role of PRDXs in human cancers, where they can exhibit strong tumor-suppressive or tumor-promoting functions. Recent evidence suggests that lower expression of PRDXs correlates with cancer progression in colorectal cancer (CRC) or in esophageal squamous carcinoma. In the thyroid, increased levels of PRDX1 has been described in follicular adenomas and carcinomas, as well as in thyroiditis, while reduced levels of PRDX6 has been found in follicular adenomas. We studied the expression of PRDX1 and PRDX6, in a series of thyroid tissue samples, covering different thyroid diseases, including 13 papillary thyroid carcinomas (PTCs). Our results show that PRDX1 and PRDX6 are significantly reduced in all PTCs compared to normal tissues, to non-neoplastic tissue (MNG) or follicular neoplasms. This reduction is strongly evident in PTCs harboring BRAF V600E (31% of our cases). Using TPC-1 and BCPAP and FRTL-5 cell lines, we demonstrate for the first time that the presence of BRAF V600E is responsible of the hypoexpression of PRDX1 and PRDX6 both at mRNA and protein levels. Finally, independently of BRAF status, we observe an interesting correlation between the tumor size, the presence of lymph node metastasis and the lowest PRDX1 and PRDX6 levels. Therefore, these data indicate that PRDX1 and PRDX6 expression not only may play a key role in papillary thyroid carcinogenesis via a BRAF V600E-dependent mechanism, but their determination could be considered as potential tumor marker for indicating tumor progression in PTCs, independently of BRAF status.

BRAF Mutations in Metanephric Adenoma of the Kidney

BackgroundMetanephric adenoma (MA) of the kidney is a rare, indolent tumor that may be difficult to differentiate from other small renal masses (SRMs). Genetic alterations associated with MA remain largely unknown. ObjectiveWe aimed at defining genetic events in MA of the kidney and determining their influence in the management of this disease. Design, setting, and participantsMultiplexed mass spectrometric genotyping was performed on 29 MA cases after tumor DNA extraction. We also conducted a mutational screen in an additional 129 renal neoplasms. Immunohistochemistry was performed on the MA cases to assess molecular markers of signaling pathway activation. Patients’ baseline characteristics, as well as follow-up data, were captured. Outcome measurements and statistical analysisWe used descriptive statistics for baseline clinical characteristics and incidence of mutations. The Wilcoxon rank-sum test was used to correlate patient characteristics with mutational status. Results and limitationsWe identified the v-raf murine sarcoma viral oncogene homolog B1 (BRAF) V600E mutation in 26 of 29 MA cases. These results were validated in all cases using the commercially available BRAF Pyro Kit (QIAGEN). In contrast, BRAF mutations were rare in the other 129 non-MA renal neoplasms that were screened. We detected a BRAF mutation (V600E) in only one papillary renal cell carcinoma case. In all MA tumors, we documented expression of phosphorylated mitogen-activated protein kinase and phosphorylated extracellular signal–regulated kinase, accompanied by immunoreactivity for p16 (INK4a). All patients were treated with a partial or radical nephrectomy, and after a median follow-up of 26.5 mo, there were no local or distant recurrences. Limitations include the retrospective nature of this study. ConclusionsBRAF V600E mutations are present in approximately 90% of all MA cases, serving as a potential valuable diagnostic tool in the differential diagnosis of SRMs undergoing a percutaneous biopsy. The presence of BRAF V600E and mitogen-activated protein kinase activation in a largely benign tumor supports the necessity for secondary events (eg, p16 loss) in BRAF-driven oncogenesis.

Correlation of the BRAF V600E mutation in hairy cell leukaemia with morphology, cytochemistry and immunophenotype

Hairy cell leukaemia (HCL) has distinct clinical, morphological and immunophenotypic features with no recurrent cytogenetic or molecular abnormalities reported until the recent description of the BRAF V600E mutation in patients with classical HCL. The incidence of this mutation was sought in 27 patients with either classical HCL or HCL variant by an allele-specific PCR approach and findings related to morphology, cytochemistry and immunophenotype. A high degree of correlation was noted between the presence of BRAF V600E and established diagnostic criteria in 26/27 patients with HCL/HCL variant. Detection of the BRAF V600E mutation is therefore a useful adjunct in the differential diagnosis of HCL and HCL variant and highlights the value of a multifaceted approach to the diagnosis of this malignancy.

Correlation between B-RAFV600E mutation and clinico–pathologic parameters in papillary thyroid carcinoma: data from a multicentric Italian study and review of the literature

Recently, a somatic point mutation of the B-RAF gene (V600E) has been identified as the most common genetic event in papillary thyroid carcinoma (PTC), with a prevalence variable among different series. Since discordant data on the clinico-pathologic features of B-RAF mutated PTC are present in the literature, the aim of the present co-operative study was to establish the prevalence of this genetic alteration and to perform a genotype-phenotype correlation in a large cohort of patients with PTC. To this purpose, a series of 260 sporadic PTCs with different histological variants were included in the study. The mutational analysis of the B-RAF gene was performed either by RT-PCR followed by single-stranded conformational polymorphism or by PCR and direct sequencing. Statistical analyses were obtained by means of chi2/Fisher's exact test and t-test. Overall, a heterozygous T > A transversion at nucleotide 1799 (V600E) was found in 99 out of 260 PTCs (38%). According to the histological type of the tumor, the B-RAF (V600E) mutation was present in 48.3% of cases of classic PTCs (85 out of 176), in 17.6% (nine out of 51) of follicular variants of PTCs, in 21.7% (five out of 23) in other PTC variants and in none of the ten poorly differentiated tumors. B-RAF (V600E) was significantly associated with the classic variant of PTC (P = 0.0001) and with an older age at diagnosis (P = 0.01). No statistically significant correlation was found among the presence of B-RAF (V600E) and gender, tumor node metastasis (TNM), multicentricity of the tumor, stage at diagnosis and outcome. In conclusion, the present study reports the prevalence of B-RAF (V600E) (38%) in the largest series of sporadic PTCs, including 260 cases from three different Italian referring centers. This prevalence is similar to that calculated by pooling together all data previously reported, 39.6% (759 out of 1914 cases), thus indicating that the prevalence of this genetic event lies around 38-40%. Furthermore, B-RAF (V600E) was confirmed to be associated with the papillary growth pattern, but not with poorer differentiated PTC variants. A significant association of B-RAF mutation was also found with an older age at diagnosis, the mutation being very rare in childhood and adolescent PTCs. Finally, no correlation was found with a poorer prognosis and a worse outcome after a median follow-up of 72 months.

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.