Impact of clinical risk scores and BRAF V600E mutation status on outcome in papillary thyroid cancer Severine M. Niederer-W€ ust, MD,a Wolfram Jochum, MD,b Diana F€ orbs,b Michael Br€ a ndle, MD, MS,c c d e e Stefan Bilz, MD, Thomas Clerici, MD, Ren e Oettli, MD, Joachim M€ uller, MD, Sarah R. Haile, PhD,f g a Silvia Ess, MD, Sandro J. Stoeckli, MD, and Martina A. Broglie, MD,a St. Gallen, Switzerland
Background. To evaluate the relationship between the BRAF V600E mutation and clinicopathologic parameters and to assess the impact of the BRAF V600E mutation and established risk scores on survival in patients with papillary thyroid carcinoma (PTC). Methods. Retrospective analysis of a consecutive, single-institutional cohort of patients with PTC larger than 1 cm. Clinical risk scores according to the Metastases, Age, Completeness of Resection, Invasion, Size (MACIS), European Organisation for Research and Treatment of Cancer (EORTC), and tumor, node, metastases (TNM) scoring systems were determined. BRAF exon 15 mutation analysis was performed by polymerase chain reaction and Sanger sequencing. Results. BRAF V600E mutations were found in 75/116 (65%) PTC. The rates for 5- and 10-year overall survival (OS), disease-specific survival (DSS), and recurrence-free survival (RFS) were 92% and 87%, 98% and 96%, and 96% and 94%, respectively. Low MACIS scores were associated with longer OS (10 y 95% vs 75%, P = .008), DSS (10 y 100% vs 89%, P = .02) and RFS (100% vs 85%, P = .006). Comparable survival advantages were observed for patients with early EORTC scores and low TNM stage. BRAF V600E mutation status was not associated with clinicopathologic characteristics of aggressive behavior such as extrathyroidal extension, lymph node metastases, higher T-categories, male sex, and greater age. Furthermore, BRAF V600E mutation status was not correlated with clinical risk scores and decreased survival. Conclusion. In concordance with other studies, we did not find a negative prognostic impact of a positive BRAF V600E mutation status on survival. In contrast, the risk algorithms MACIS, EORTC score, and TNM stage were associated with impaired prognosis. Therefore, clinical staging systems represent better tools for risk stratification than BRAF V600E mutation status. (Surgery 2015;157:119-25.) From the Department of Otorhinolaryngology, Head and Neck Surgery,a Institute of Pathology,b Division of Endocrinology and Diabetes, Department of Internal Medicine,c Departments of Surgeryd and Nuclear Medicine,e and Clinical Trials Unit,f Kantonsspital St. Gallen, St. Gallen, Switzerland; and Cancer Registry Eastern Switzerland, St. Gallen, Switzerlandg
PAPILLARY THYROID CARCINOMA (PTC) is the most common type (>70%) of thyroid cancer.1 PTC shows increasing incidence rates in many countries and has a high cure rate with an estimated 10-year survival of 80–90%.2,3 A small proportion of patients, however, develops locoregional recurrences or distant metastases, and eventually dies Supported by the MFZ Fonds of the Kantonsspital St. Gallen (MFZ 2010_003). Accepted for publication July 17, 2014. Reprint requests: Martina A. Broglie, MD, Department of Otorhinolaryngology, Head and Neck Surgery, Kantonsspital St. Gallen, Rorschacherstrasse 95, 9007 St. Gallen, Switzerland. E-mail: [email protected]. 0039-6060/$ - see front matter Ó 2015 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.surg.2014.07.015
of the disease. Identification of PTC patients at increased risk for an adverse course would raise the opportunity for aggressive therapy or followup. Various prognostic factors have been identified and incorporated in currently applied staging classifications, such as age, sex, extrathyroidal extension, and distant metastasis. The most commonly used staging systems are the European Organisation for Research and Treatment of Cancer (EORTC)4 scoring system, the Age, Grade, Extent, Size (AGES),5 the Age, Metastases, Extent, Size,6 the Metastases, Age, Completeness of resection, Invasion, Size (MACIS),7 and the Union Internationale Contre le Cancer (UICC) tumor, node, metastases (TNM)8 score. The risk factor of incomplete resection is unique in the MACIS risk score, whereas the UICC TNM staging system is based on the size and extent of the primary SURGERY 119
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tumor, and on the presence of regional lymph node and distant metastases. The understanding of molecular genetics in thyroid cancer has expanded dramatically in recent years and this knowledge has started to be translated into clinical practice. The aim of all these scientific efforts is to improve preoperative diagnosis, to better assess prognosis, and to potentially individualize treatment.9 BRAF mutations occur in 30–80% of PTC.10,11 Most mutations affect BRAF V600 in exon 15 of the BRAF gene. The BRAF V600E mutation is more frequent in PTC with classic histology than in the follicular variant of PTC. Recent reports suggest an association between BRAF V600E mutation and aggressive clinicopathologic characteristics, including extrathyroidal extension, lymph node metastasis, histologic subtypes with poor prognosis (tall cell variant PTC), advanced stages, and recurrence.11-15 Most studies including metaanalyses have shown a correlation with at least one poor prognostic factor.16-19 Conversely, other investigations were not able to confirm these correlations, even by analyzing a large number of patients.11,20Therefore, the clinical significance of BRAF V600E mutation in PTC is still controversial.21 The aim of this study was to assess the prognostic importance of established clinical risk scores and the BRAF mutation status in a consecutive single institutional cohort of patients with PTC in Switzerland. METHODS Patients and clinicopathologic factors. A cohort of 147 consecutive PTC patients treated between 1990 and 2011 at the Kantonsspital St. Gallen was analyzed. Patients with microcarcinoma (size #1 cm) were excluded. All patients were treated with total thyroidectomy. In 120/147 (81%), a central neck dissection was performed, of which 20/147 (13%) received a therapeutic neck dissection because of a clinically positive neck and 100/ 147 (67%) a prophylactic neck dissection. Prophylactic central compartment dissection revealed positive nodes in 55/100 (55%) patients. Postoperatively, 99% of the patients were treated by ablative radioiodine ablation (RAI) therapy in hypothyroid conditions after hormone withdrawal (thyroid-stimulating hormone >30 mU/L), stopping levothyroxine approximately 5–6 weeks before RAI treatment. Thyroid-stimulating hormone suppression therapy was continued in all patients for at least 10 years. Patient charts were assessed for the following clinicopathological parameters: age, sex, extrathyroidal extension, lymph node metastasis, multifocality,
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histologic subtype, tumor size at the time of diagnosis, UICC TNM stage, treatment modality (surgery, radiotherapy, RAI, and/or combination therapy), and follow-up data (locoregional control, metastasis, survival). Tumor stage at the time of diagnosis was determined based on medical data and the records of the radiologist, nuclear medicine physician, surgeon, and the pathologist according to the TNM classification edition 2011 for thyroid carcinoma.8 The survival status of all patients was last updated on December 31, 2012. Overall survival was defined as the period from the day of diagnosis to the last date of follow-up or death. Recurrent PTC was defined as a detectable thyroglobulin, positive radiographic findings, and/or pathologic confirmation of disease >6 mo from thyroidectomy and RAI. MACIS and EORTC scores for PTC were calculated as previously described.4,7 BRAF mutation analysis. Formalin-fixed, paraffin-embedded PTC tissues were retrieved from the archive of the Institute of Pathology, Kantonsspital St. Gallen. Tumor areas were marked on hematoxylin and eosin–stained sections by a trained pathologist (W.J.). PTC tissue was microdissected manually from unstained 4-mm thick serial sections. Tumor tissue was deparaffinized and lysed with proteinase K. Polymerase chain reaction (PCR)-based amplification was performed using 50-100 ng DNA and primers flanking exon 15 of the BRAF gene (forward: 59-TGCTTGCTCTGA TAGGAAAATG-39, reverse: 59-AGCATCTCAGGGC CAAAAAT-39). The following amplification protocol was used: initial denaturation at 958C for 5 minutes, 34 cycles of denaturation at 958C for 30 seconds, annealing at 558C for 20 seconds, extension at 728C for 1 minute, followed by a final extension at 728C for 5 minutes. Successful amplification was confirmed by electrophoresis of PCR products (228 bp) in a 2% agarose gel containing GelRED fluorescent dye Direct sequencing of PCR products was performed by automated capillary electrophoresis on the ABI PRISM 3100 Genetic Analyzer instrument (Applied Biosystems) using the primers described and the BigDye Terminator v1.1 Cycle Sequencing Kit (Applied Biosystems). BRAF mutational status was assessed retrospectively and had no impact on treatment decisions or follow up policy of study patients. Statistical analysis. SPSS 20.0 for windows was used to calculate the summary odds ratio (OR), Cox regression model, and the chi-square test. The OR was determined with 95% confidence intervals (CIs). Therefore, a chi-square test (Pearson) was performed for the analysis of the relationship between the BRAF V600E mutation and clinicopathologic
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Surgery Volume 157, Number 1 parameters for univariate analysis. A P < .05 was considered statistically significant. Cox proportional hazard modelling was used to relate risk factors to overall survival (OS), recurrence-free survival (RFS), and disease-specific survival (DSS). For statistical analysis, MACIS scores, EORTC scores, and the TNM stages were grouped in low-risk (MACIS 1, EORTC 1 + 2, and TNM stage I) and high-risk categories (MACIS 2–4, EORTC 3–5, and TNM stage II–IVc). To rank and compare each staging system’s ability to predict OS, RFS, and DSS, Kaplan-Meier curves were generated to visualize the data. Ethics committee. The study was approved by the local ethics committee of St. Gallen. RESULTS Cohort characteristics. BRAF exon 15 analysis was successful in 116/147 (79%) PTC. In the other 31 PTC tissue samples BRAF exon 15 analysis failed due to insufficient DNA quality. These patients were excluded from the subsequent analysis resulting in a final study cohort of 116 patients. BRAF V600E mutations were found in 75/116 (65%) PTC, whereas 41/116 PTC contained the wild type gene. Patient and tumor characteristics of the final cohort are summarized in Table I. Follow-up information was available for all patients. The median observation time of the patient cohort was 79 months (range 2–252 months), during which 9/116 patients (8%) developed recurrent disease. The median time to recurrence was 67 months (range 7–184 months). In 1 patient, the recurrence was local, in 4 (45%) regional, in 2 (22%) locoregional, and in 2 (22%) distant. In total, 12 patients (10%) died after a median of 63 months (range 12–174 months), 3 of whom died of disease (median 44 months, range 12–80 months). Of the remaining 9 patients 1 died of unknown cause, 5 due to a second primary cancer, and 3 due to non-cancer related causes. The 5- and 10-year OS, DSS, and RFS rates were 92% and 87%, 98% and 96%, and 96% and 94%, respectively. Clinicopathologic factors. For statistical analysis, patients with MACIS 1 (n = 71) were compared to patients with MACIS scores 2–4 (n = 45). Patients with MACIS score 1 had better OS (10 y 95% vs 75%, P = .008), DSS (10 y 100% vs 89%, P = .02) and RFS (100% vs 85%, P = .006) than patients with high MACIS scores as shown by KaplanMeier analysis. The EORTC groups (EORTC 1 + 2 ‘‘early score,’’ n = 73 vs EORTC 3–5 ‘‘advanced score,’’ n = 43) were also pooled and compared and statistically significant differences could be demonstrated in all three survival estimates between the groups. In the TNM groups (stage I
Table I. Clinicopathologic characteristics of the patient cohort (n = 116) Sex Mean age T category
N category
Focality Histology
Extrathyroidal extension TNM stage (UICC)
MACIS
EORTC
Female Male 50 y pT1b pT2 pT3 pT4a pT4b pN0 pN1a pN1b Unifocal multifocal Classic variant Follicular variant Oxyphil variant Tall cell variant No Yes I II III IVa IVb IVc 1 2 3 4 1 2 3 4 5
78 38 range 24 25 56 10 1 51 33 32 70 46 69 40 6 1 55 61 49 13 33 17 1 3 71 18 12 15 46 27 26 16 1
(67%) (33%) (10–86 y) (21%) (22%) (48%) (8%) (1%) (44%) (28%) (28%) (60%) (40%) (60%) (34%) (5%) (1%) (47%) (53%) (42%) (11%) (28%) (15%) (1%) (3%) (61%) (16%) (10%) (13%) (40%) (23%) (22%) (14%) (1%)
EORTC, European Organisation for Research and Treatment of Cancer; MACIS, Metastases, Age, Completeness of Resection, Invasion, Size; TNM, tumor, node, metastases; UICC, Union Internationale Contre le Cancer.
‘‘low stage,’’ n = 50 vs stage II–IV ‘‘high stage,’’ n = 60), the differences between the low and high stage TNM groups in OS and RFS reached statistical significance, but not in 10-y DSS (100% vs 91%, P = .08). BRAF V600E mutation status. In comparison to the subgroup without a BRAF V600E mutation (41/116, 35%) the subgroup with mutation (75/ 116, 65%) did not show greater rates of known, aggressive, clinicopathologic characteristics such as extrathyroidal extension, lymph node metastases, higher T-categories, male sex, or a greater age. Furthermore, there was no correlation between the BRAF V600E mutation status and the clinical risk scores MACIS, EORTC, or TNM stage. V600E mutations were, however, more frequent in the
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subgroup of classic variant PTC (50/69, 73%) than in the subgroup of follicular, oxyphil, and tall cell variant PTC (25/47, 53%; P = .03). When comparing the BRAF V600E positive subgroup to the BRAF V600E-negative subgroup the 10-year OS, DSS, and RFS rates were 92% and 79%, 98% and 91%, and 98% and 88%, respectively. None of these differences were statistically significant (P > .05). Furthermore, we performed a separate analysis of BRAF mutation effects in medium and greater risk tumors based on clinical risk scores but could not find any statistical differences or trends in survival between the groups either. Cox proportional hazard regression analysis revealed significantly lesser OS rates for age greater than 45 years. Other factors such as BRAF mutation status, sex, extrathyroidal extension, lymph node metastasis, multifocality, or histologic were not associated with decreased survival. Moreover, lesser OS rates were found when comparing MACIS scores (>1 vs 1), TNM stages (stage >1 to stage 1), and EORTC scores (>2 vs 1–2). Furthermore, differences in RFS could be demonstrated for EORTC scores (>2 vs 1–2). No significant differences in RFS were observed for the other parameters (Table II). DISCUSSION A major clinical challenge in the management of patients with PTC is how to reliably distinguish those patients who need aggressive treatments to decrease potentially treatment-associated morbidity and disease mortality, especially in view of the low overall mortality rate in PTC. Both the European and the American Thyroid Associations proposed practical management guidelines in which the clinical stage was integrated by additional parameters such as the tumor histologic variant, the results of postablative whole-body scan, and serum thyroglobulin levels.22,23 Despite that, patients included in the same risk group still show heterogeneous recurrence and survival rates. In addition, risk stratification according to the guidelines is not accurate in predicting long-term patient outcome and have a very low positive predictive value.24 The increasing understanding of the molecular basis of thyroid cancer has started to be translated into clinical practice and may help to better assess the prognosis and to individualize treatment.9 The data in the literature are conflicting, and the clinical importance of molecular markers in PTC is still controversial.21 The BRAF V600E mutation is the most frequent genetic alteration in PTC.19,25 In various reports, this mutation has been associated with aggressive tumor characteristics,11-13 whereas
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Table II. Hazard ratio for OS of different risk factors Covariates T-Category >T1a N-Category N+ ETE Female sex BRAF V600E negative UICC TNM >I MACIS >1 EORTC >2
OS hazard ratio (95% CI) 2.52 1.13 2.74 0.99 0.46 5.38 4.99 14.62
(0.32–19.73) (0.34–3.48) (0.74–10.15) (0.30–3.31) (0.14–1.49) (1.76–13.19) (1.34–18.52) (2.99–71.30)
P value .3 .8 .1 .3 .1 .05* .01* .001*
*Statistically significant. EORTC, European Organisation for Research and Treatment of Cancer; ETE, extrathyroidal extension; MACIS, Metastases, Age, Completeness of Resection, Invasion, Size; OS, overall survival; TNM, tumor, node, metastases; UICC, Union Internationale Contre le Cancer.
other recent studies20,26-28 questioned its prognostic impact. The frequency of the BRAF V600E mutation is high (>50%) compared with that of the poor outcomes (around 20%).29 The aim of our study was to evaluate the relationship between BRAF V600E mutational status and clinicopathologic parameters, including three established risk scores (MACIS score, EORTC score, TNM stage) and to analyze the impact of BRAF V600E mutation on survival in comparison with established risk scores in a consecutive single institutional patient cohort with PTC in Switzerland, excluding patients with papillary microcarcinoma to avoid a selection bias. This is the first study to investigate the prevalence of BRAF V600E mutational status in PTC patients in Switzerland. In line with previous studies13,14 as well as reports from other European countries,30 the BRAF V600E mutation was detected with a prevalence of 65% in our cohort. We did not find any difference in aggressive clinicopathologic characteristics between the subgroup with the BRAF V600E mutation in comparison to the BRAF wild type subgroup. This finding is in line with a recent study by Li et al31 but in contrast to other data in the literature.11-13 Li et al found no significant associations between BRAF V600E mutation and aggressive clinicopathologic features in a consecutive patient cohort who underwent total thyroidectomy and routine lateral neck dissection. Moreover, multivariate analysis revealed that the BRAF V600E status was not an independent predictor of lymph node metastases. Similar results were demonstrated by Ito et al.20 Furthermore, Nam et al32 did not find any correlation between BRAF V600E mutation and high-risk MACIS score, lymph node metastasis, age distribution, sex, or
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cancer size; in contrast, extrathyroidal extension and multifocality were associated with the BRAF V600E mutation. We found a greater frequency of the BRAF V600E mutations in the subgroup of classic variant PTC compared to the subgroup of nonclassic variant PTC. This finding is in line with previous studies which reported an association between the BRAF mutation status and the histologic PTC subtype and was significantly related to the classic and tall cell variant PTC but not to the follicular variant.12 There is no prognostic difference between the follicular and the classic variant type,33-36 whereas the tallcell variant PTC is characterized as having a more aggressive behavior usually presenting at an older age and higher stage than their classic PTC counterparts.37-40 The subgroups of oxyphil and tall cell variants could not be analyzed separately in our study due to low number of patients. In a multi-institutional, retrospective study addressing the relationship between the BRAF V600E mutation and mortality, Xing et al30 found an association between the BRAF V600E mutation and greater cancer-related mortality. The authors suggested that the BRAF V600E mutation may promote aggressive tumor behavior and risk of PTC-related mortality by up-regulation of classic angiogenic and tumor-promoting molecules, such as vascular endothelial growth factor, matrix metalloproteinases, and nuclear transcription factors.16,30,41 Furthermore, the BRAF V600E mutation seems to influence the tumor microenvironment such as tumor adhesion, migration, invasion, and metastasis.42 Mesa et al43 demonstrated that BRAF V600E activated normal rat thyroid cells to express genes possibly promoting tumor invasion by degradation of different noncellular components of the extracellular matrix. Furthermore, BRAF V600E activation in thyroid cells appears to lead to altered function of the sodium iodide symporter and other genes metabolizing iodide, which could be responsible for the decreased ability of PTC with BRAF V600E mutations to trap radioiodine leading to treatment failure.9,44-46 In contrast, the proposed primary tumorigenic role of BRAF V600E in PTC has been questioned. BRAF V600E seems to be a secondary subclone, rather than a primary event in thyroid tumorigenesis.47 Moreover, the BRAF V600E mutation has been found in lymph node metastases, but not in the primary tumor.16 The survival estimates in our patient cohort achieved with a 10-year OS of 87% and a 10-year DSS of 96% excellent rates comparable to the literature.2,3,30 A number of formal scoring systems have been developed to estimate prognosis among
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patients diagnosed with PTC. Lang et al48 conducted a comprehensive study to determine the most predictive staging system for PTC. The relative importance of each staging system was determined by calculating the proportion of variation in survival time explained (PVE). The three highest ranked staging systems by proportion of variation in survival time explained were the MACIS, the 6th edition TNM, and EORTC. They considered the MACIS score to be the stratification system of choice for PTC. In our cohort, high clinical risk scores (MACIS, EORTC, and TNM) were associated with lesser survival rates according to Kaplan-Meier survival analysis and the importance of these clinical risk scores for OS was also confirmed by Cox proportional hazard regression. We could demonstrate that individual components of the clinical risk scores (presence of regional or distant metastasis, age at diagnosis, completeness of resection, and presence of invasion, tumor size) had no impact on survival, whereas the combination in risk score was associated with all three survival estimates (OS, DSS, RFS). In contrast BRAF V600E mutational status did not influence survival in our cohort. According to Prescott et al,49 the addition of BRAF V600E mutational status to established risk algorithms improved the discrimination of risk recurrence in patients undergoing total thyroidectomy for PTC. In their retrospective study, the 5-year, cumulative recurrence incidence among BRAF V600E patients was 20% versus 8% among BRAF wild type. Moreover, the BRAF V600E mutation was significantly associated with time to recurrence when added to different current risk classification systems (Age, Metastases, Extent, Size; MACIS; UICC-TNM). Unfortunately, the authors did not analyze the additional impact of BRAF V600E mutational status on OS or DSS. We did not find any correlation between the BRAF V600E mutational status and the clinical risk scores MACIS, EORTC, or TNM. Moreover, neither a separate analysis of the effects of the BRAF mutation status in medium and higher risk tumors nor in combination with clinical risk scores as proposed by Prescott et al49 had any influence on survival estimates. In line with our findings, no impact of the BRAF V600E mutation on survival could be demonstrated in various recent studies20,50-53 leading to a great debate concerning the prognostic role of BRAF V600E mutations in PTC. In fact, when Xing et al30 adjusted for aggressive tumor behaviors, the association with BRAF V600E was no longer statistically significant. In multivariable analyses, BRAF V600E status was not independently
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associated with mortality. It seems correct that BRAF V600E plays an important biologic role in promoting aggressive tumor behavior; however, the BRAF V600E mutation may only be a marker of aggressiveness and not a true prognostic factor. According to Sancisi et al,54 analyzing the frequency of the BRAF V600E mutation within a group of highly aggressive PTC, which had developed distant metastases and a control group BRAF V600E mutation, was not associated with the development of distant metastases or fatal outcome. In conclusion, we could not confirm a negative prognostic impact of BRAF V600E mutational status on survival. The established risk algorithms of MACIS, EORTC, or TNM offered benefit for survival estimates. Based on our results, the usefulness of BRAF V600E mutation in the management algorithm of patients of PTC remains questionable, and meanwhile, we recommend using the current staging systems for risk stratification in PTC patients.
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carcinoma: data from a multicentric Italian study and review of the literature. Endocr Relat Cancer 2006;13:455-64. Lee JH, Lee ES, Kim YS. Clinicopathologic significance of BRAF V600E mutation in papillary carcinomas of the thyroid: a meta-analysis. Cancer 2007;110:38-46. Xing M, Westra WH, Tufano RP, Cohen Y, Rosenbaum E, Rhoden KJ, et al. BRAF mutation predicts a poorer clinical prognosis for papillary thyroid cancer. J Clin Endocrinol Metab 2005;90:6373-9. Lassalle S, Hofman V, Ilie M, Butori C, Bozec A, Santini J, et al. Clinical impact of the detection of BRAF mutations in thyroid pathology: potential usefulness as diagnostic, prognostic and theragnostic applications. Curr Med Chem 2010;17:1839-50. Handkiewicz-Junak D, Czarniecka A, Jarzab B. Molecular prognostic markers in papillary and follicular thyroid cancer: current status and future directions. Mol Cell Endocrinol 2010;322:8-28. Xing M. BRAF mutation in papillary thyroid cancer: pathogenic role, molecular bases, and clinical implications. Endocr Rev 2007;28:742-62. Kim TH, Park YJ, Lim JA, Ahn HY, Lee EK, Lee YJ, et al. The association of the BRAF(V600E) mutation with prognostic factors and poor clinical outcome in papillary thyroid cancer: a meta-analysis. Cancer 2012;118:1764-73. Kebebew E, Weng J, Bauer J, Ranvier G, Clark OH, Duh QY, et al. The prevalence and prognostic value of BRAF mutation in thyroid cancer. Ann Surg 2007;246:466-70; discussion 470-1. Elisei R, Ugolini C, Viola D, Lupi C, Biagini A, Giannini R, et al. BRAF(V600E) mutation and outcome of patients with papillary thyroid carcinoma: a 15-year median follow-up study. J Clin Endocrinol Metab 2008;93:3943-9. Ito Y, Yoshida H, Maruo R, Morita S, Takano T, Hirokawa M, et al. BRAF mutation in papillary thyroid carcinoma in a Japanese population: its lack of correlation with high-risk clinicopathological features and disease-free survival of patients. Endocr J 2009;56:89-97. Lim JY, Hong SW, Lee YS, Kim BW, Park CS, Chang HS, et al. Clinicopathologic implications of the BRAFV600E mutation in papillary thyroid cancer; a subgroup analysis of 3130 cases in a single center. Thyroid 2013;23:1423-30. Cooper DS, Doherty GM, Haugen BR, Kloos RT, Lee SL, Mandel SJ, et al. Revised American Thyroid Association management guidelines for patients with thyroid nodules and differentiated thyroid cancer. Thyroid 2009;19:1167-214. Pacini F, Schlumberger M, Dralle H, Elisei R, Smit JW, Wiersinga W. European consensus for the management of patients with differentiated thyroid carcinoma of the follicular epithelium. Eur J Endocrinol 2006;154:787-803. Castagna MG, Maino F, Cipri C, Belardini V, Theodoropoulou A, Cevenini G, et al. Delayed risk stratification, to include the response to initial treatment (surgery and radioiodine ablation), has better outcome predictivity in differentiated thyroid cancer patients. Eur J Endocrinol 2011;165:441-6. Namba H, Nakashima M, Hayashi T, Hayashida N, Maeda S, Rogounovitch TI, et al. Clinical implication of hot spot BRAF mutation, V599E, in papillary thyroid cancers. J Clin Endocrinol Metab 2003;88:4393-7. Barbaro D, Incensati RM, Materazzi G, Boni G, Grosso M, Panicucci E, et al. The BRAF V600E mutation in papillary thyroid cancer with positive or suspected pre-surgical cytological finding is not associated with advanced stages or worse prognosis. Endocrine 2013;45:462-8.
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27. Zoghlami A, Roussel F, Sabourin JC, Kuhn JM, Marie JP, Dehesdin D, et al. BRAF mutation in papillary thyroid carcinoma: predictive value for long-term prognosis and radioiodine sensitivity. Eur Ann Otorhinolaryngol Head Neck Dis 2014;131:7-13. 28. Fugazzola L, Mannavola D, Cirello V, Vannucchi G, Muzza M, Vicentini L, et al. BRAF mutations in an Italian cohort of thyroid cancers. Clin Endocrinol (Oxf) 2004;61:239-43. 29. Baldini E, Sorrenti S, Tuccilli C, Prinzi N, Coccaro C, Catania A, et al. Emerging molecular markers for the prognosis of differentiated thyroid cancer patients. Int J Surg 2014;12(Suppl 1):S52-6. 30. Xing M, Alzahrani AS, Carson KA, Viola D, Elisei R, Bendlova B, et al. Association between BRAF V600E mutation and mortality in patients with papillary thyroid cancer. JAMA 2013;309:1493-501. 31. Li C, Aragon Han P, Lee KC, Lee LC, Fox AC, Beninato T, et al. Does BRAF V600E mutation predict aggressive features in papillary thyroid cancer? Results from four endocrine surgery centers. J Clin Endocrinol Metab 2013; 98:3702-12. 32. Nam JK, Jung CK, Song BJ, Lim DJ, Chae BJ, Lee NS, et al. Is the BRAF(V600E) mutation useful as a predictor of preoperative risk in papillary thyroid cancer? Am J Surg 2012; 203:436-41. 33. Zidan J, Karen D, Stein M, Rosenblatt E, Basher W, Kuten A. Pure versus follicular variant of papillary thyroid carcinoma: clinical features, prognostic factors, treatment, and survival. Cancer 2003;97:1181-5. 34. Sebastian SO, Gonzalez JM, Paricio PP, Perez JS, Flores DP, Madrona AP, et al. Papillary thyroid carcinoma: prognostic index for survival including the histological variety. Arch Surg 2000;135:272-7. 35. Mendelsohn AH, Elashoff DA, Abemayor E, St John MA. Surgery for papillary thyroid carcinoma: is lobectomy enough? Arch Otolaryngol Head Neck Surg 2010;136: 1055-61. 36. Lin HW, Bhattacharyya N. Clinical behavior of follicular variant of papillary thyroid carcinoma: presentation and survival. Laryngoscope 2010;120:712-6. 37. Ganly I, Ibrahimpasic T, Rivera M, Nixon I, Palmer F, Patel SG, et al. Prognostic implications of papillary thyroid carcinoma with tall-cell features. Thyroid 2014;24:662-70. 38. Michels JJ, Jacques M, Henry-Amar M, Bardet S. Prevalence and prognostic significance of tall cell variant of papillary thyroid carcinoma. Hum Pathol 2007;38:212-9. 39. Jalisi S, Ainsworth T, Lavalley M. Prognostic outcomes of tall cell variant papillary thyroid cancer: a meta-analysis. J Thyroid Res 2010;2010:325602. 40. Ito Y, Hirokawa M, Uruno T, Kihara M, Higashiyama T, Takamura Y, et al. Prevalence and biological behaviour of variants of papillary thyroid carcinoma: experience at a single institute. Pathology 2008;40:617-22. 41. Xing M. Prognostic utility of BRAF mutation in papillary thyroid cancer. Mol Cell Endocrinol 2010;321:86-93.
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42. Nucera C, Lawler J, Parangi S. BRAF(V600E) and microenvironment in thyroid cancer: a functional link to drive cancer progression. Cancer Res 2011;71:2417-22. 43. Mesa C Jr, Mirza M, Mitsutake N, Sartor M, Medvedovic M, Tomlinson C, et al. Conditional activation of RET/PTC3 and BRAFV600E in thyroid cells is associated with gene expression profiles that predict a preferential role of BRAF in extracellular matrix remodeling. Cancer Res 2006;66:6521-9. 44. Ricarte-Filho JC, Ryder M, Chitale DA, Rivera M, Heguy A, Ladanyi M, et al. Mutational profile of advanced primary and metastatic radioactive iodine-refractory thyroid cancers reveals distinct pathogenetic roles for BRAF, PIK3CA, and AKT1. Cancer Res 2009;69:4885-93. 45. Riesco-Eizaguirre G, Gutierrez-Martinez P, Garcia-Cabezas MA, Nistal M, Santisteban P. The oncogene BRAF V600E is associated with a high risk of recurrence and less differentiated papillary thyroid carcinoma due to the impairment of Na+/I- targeting to the membrane. Endocr Relat Cancer 2006;13:257-69. 46. Durante C, Puxeddu E, Ferretti E, Morisi R, Moretti S, Bruno R, et al. BRAF mutations in papillary thyroid carcinomas inhibit genes involved in iodine metabolism. J Clin Endocrinol Metab 2007;92:2840-3. 47. Guerra A, Sapio MR, Marotta V, Campanile E, Rossi S, Forno I, et al. The primary occurrence of BRAF(V600E) is a rare clonal event in papillary thyroid carcinoma. J Clin Endocrinol Metab 2012;97:517-24. 48. Lang BH, Lo CY, Chan WF, Lam KY, Wan KY. Staging systems for papillary thyroid carcinoma: a review and comparison. Ann Surg 2007;245:366-78. 49. Prescott JD, Sadow PM, Hodin RA, Le LP, Gaz RD, Randolph GW, et al. BRAF V600E status adds incremental value to current risk classification systems in predicting papillary thyroid carcinoma recurrence. Surgery 2012;152:984-90. 50. Eloy C, Santos J, Soares P, Sobrinho-Simoes M. The preeminence of growth pattern and invasiveness and the limited influence of BRAF and RAS mutations in the occurrence of papillary thyroid carcinoma lymph node metastases. Virchows Arch 2011;459:265-76. 51. Cheng S, Serra S, Mercado M, Ezzat S, Asa SL. A highthroughput proteomic approach provides distinct signatures for thyroid cancer behavior. Clin Cancer Res 2011; 17:2385-94. 52. Paulson L, Shindo M, Schuff K, Corless C. The role of molecular markers and tumor histological type in central lymph node metastasis of papillary thyroid carcinoma. Arch Otolaryngol Head Neck Surg 2012;138:44-9. 53. Sassolas G, Hafdi-Nejjari Z, Ferraro A, Decaussin-Petrucci M, Rousset B, Borson-Chazot F, et al. Oncogenic alterations in papillary thyroid cancers of young patients. Thyroid 2012;22:17-26. 54. Sancisi V, Nicoli D, Ragazzi M, Piana S, Ciarrocchi A. BRAFV600E mutation does not mean distant metastasis in thyroid papillary carcinomas. J Clin Endocrinol Metab 2012;97:E1745-9.
Background. To evaluate the relationship between the BRAF V600E mutation and clinicopathologic parameters and to assess the impact of the BRAF V600E mutation and established risk scores on survival in patients with papillary thyroid carcinoma (PTC). Methods. Retrospective analysis of a consecutive, single-institutional cohort of patients with PTC larger than 1 cm. Clinical risk scores according to the Metastases, Age, Completeness of Resection, Invasion, Size (MACIS), European Organisation for Research and Treatment of Cancer (EORTC), and tumor, node, metastases (TNM) scoring systems were determined. BRAF exon 15 mutation analysis was performed by polymerase chain reaction and Sanger sequencing. Results. BRAF V600E mutations were found in 75/116 (65%) PTC. The rates for 5- and 10-year overall survival (OS), disease-specific survival (DSS), and recurrence-free survival (RFS) were 92% and 87%, 98% and 96%, and 96% and 94%, respectively. Low MACIS scores were associated with longer OS (10 y 95% vs 75%, P = .008), DSS (10 y 100% vs 89%, P = .02) and RFS (100% vs 85%, P = .006). Comparable survival advantages were observed for patients with early EORTC scores and low TNM stage. BRAF V600E mutation status was not associated with clinicopathologic characteristics of aggressive behavior such as extrathyroidal extension, lymph node metastases, higher T-categories, male sex, and greater age. Furthermore, BRAF V600E mutation status was not correlated with clinical risk scores and decreased survival. Conclusion. In concordance with other studies, we did not find a negative prognostic impact of a positive BRAF V600E mutation status on survival. In contrast, the risk algorithms MACIS, EORTC score, and TNM stage were associated with impaired prognosis. Therefore, clinical staging systems represent better tools for risk stratification than BRAF V600E mutation status. (Surgery 2015;157:119-25.) From the Department of Otorhinolaryngology, Head and Neck Surgery,a Institute of Pathology,b Division of Endocrinology and Diabetes, Department of Internal Medicine,c Departments of Surgeryd and Nuclear Medicine,e and Clinical Trials Unit,f Kantonsspital St. Gallen, St. Gallen, Switzerland; and Cancer Registry Eastern Switzerland, St. Gallen, Switzerlandg
PAPILLARY THYROID CARCINOMA (PTC) is the most common type (>70%) of thyroid cancer.1 PTC shows increasing incidence rates in many countries and has a high cure rate with an estimated 10-year survival of 80–90%.2,3 A small proportion of patients, however, develops locoregional recurrences or distant metastases, and eventually dies Supported by the MFZ Fonds of the Kantonsspital St. Gallen (MFZ 2010_003). Accepted for publication July 17, 2014. Reprint requests: Martina A. Broglie, MD, Department of Otorhinolaryngology, Head and Neck Surgery, Kantonsspital St. Gallen, Rorschacherstrasse 95, 9007 St. Gallen, Switzerland. E-mail: [email protected]. 0039-6060/$ - see front matter Ó 2015 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.surg.2014.07.015
of the disease. Identification of PTC patients at increased risk for an adverse course would raise the opportunity for aggressive therapy or followup. Various prognostic factors have been identified and incorporated in currently applied staging classifications, such as age, sex, extrathyroidal extension, and distant metastasis. The most commonly used staging systems are the European Organisation for Research and Treatment of Cancer (EORTC)4 scoring system, the Age, Grade, Extent, Size (AGES),5 the Age, Metastases, Extent, Size,6 the Metastases, Age, Completeness of resection, Invasion, Size (MACIS),7 and the Union Internationale Contre le Cancer (UICC) tumor, node, metastases (TNM)8 score. The risk factor of incomplete resection is unique in the MACIS risk score, whereas the UICC TNM staging system is based on the size and extent of the primary SURGERY 119
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tumor, and on the presence of regional lymph node and distant metastases. The understanding of molecular genetics in thyroid cancer has expanded dramatically in recent years and this knowledge has started to be translated into clinical practice. The aim of all these scientific efforts is to improve preoperative diagnosis, to better assess prognosis, and to potentially individualize treatment.9 BRAF mutations occur in 30–80% of PTC.10,11 Most mutations affect BRAF V600 in exon 15 of the BRAF gene. The BRAF V600E mutation is more frequent in PTC with classic histology than in the follicular variant of PTC. Recent reports suggest an association between BRAF V600E mutation and aggressive clinicopathologic characteristics, including extrathyroidal extension, lymph node metastasis, histologic subtypes with poor prognosis (tall cell variant PTC), advanced stages, and recurrence.11-15 Most studies including metaanalyses have shown a correlation with at least one poor prognostic factor.16-19 Conversely, other investigations were not able to confirm these correlations, even by analyzing a large number of patients.11,20Therefore, the clinical significance of BRAF V600E mutation in PTC is still controversial.21 The aim of this study was to assess the prognostic importance of established clinical risk scores and the BRAF mutation status in a consecutive single institutional cohort of patients with PTC in Switzerland. METHODS Patients and clinicopathologic factors. A cohort of 147 consecutive PTC patients treated between 1990 and 2011 at the Kantonsspital St. Gallen was analyzed. Patients with microcarcinoma (size #1 cm) were excluded. All patients were treated with total thyroidectomy. In 120/147 (81%), a central neck dissection was performed, of which 20/147 (13%) received a therapeutic neck dissection because of a clinically positive neck and 100/ 147 (67%) a prophylactic neck dissection. Prophylactic central compartment dissection revealed positive nodes in 55/100 (55%) patients. Postoperatively, 99% of the patients were treated by ablative radioiodine ablation (RAI) therapy in hypothyroid conditions after hormone withdrawal (thyroid-stimulating hormone >30 mU/L), stopping levothyroxine approximately 5–6 weeks before RAI treatment. Thyroid-stimulating hormone suppression therapy was continued in all patients for at least 10 years. Patient charts were assessed for the following clinicopathological parameters: age, sex, extrathyroidal extension, lymph node metastasis, multifocality,
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histologic subtype, tumor size at the time of diagnosis, UICC TNM stage, treatment modality (surgery, radiotherapy, RAI, and/or combination therapy), and follow-up data (locoregional control, metastasis, survival). Tumor stage at the time of diagnosis was determined based on medical data and the records of the radiologist, nuclear medicine physician, surgeon, and the pathologist according to the TNM classification edition 2011 for thyroid carcinoma.8 The survival status of all patients was last updated on December 31, 2012. Overall survival was defined as the period from the day of diagnosis to the last date of follow-up or death. Recurrent PTC was defined as a detectable thyroglobulin, positive radiographic findings, and/or pathologic confirmation of disease >6 mo from thyroidectomy and RAI. MACIS and EORTC scores for PTC were calculated as previously described.4,7 BRAF mutation analysis. Formalin-fixed, paraffin-embedded PTC tissues were retrieved from the archive of the Institute of Pathology, Kantonsspital St. Gallen. Tumor areas were marked on hematoxylin and eosin–stained sections by a trained pathologist (W.J.). PTC tissue was microdissected manually from unstained 4-mm thick serial sections. Tumor tissue was deparaffinized and lysed with proteinase K. Polymerase chain reaction (PCR)-based amplification was performed using 50-100 ng DNA and primers flanking exon 15 of the BRAF gene (forward: 59-TGCTTGCTCTGA TAGGAAAATG-39, reverse: 59-AGCATCTCAGGGC CAAAAAT-39). The following amplification protocol was used: initial denaturation at 958C for 5 minutes, 34 cycles of denaturation at 958C for 30 seconds, annealing at 558C for 20 seconds, extension at 728C for 1 minute, followed by a final extension at 728C for 5 minutes. Successful amplification was confirmed by electrophoresis of PCR products (228 bp) in a 2% agarose gel containing GelRED fluorescent dye Direct sequencing of PCR products was performed by automated capillary electrophoresis on the ABI PRISM 3100 Genetic Analyzer instrument (Applied Biosystems) using the primers described and the BigDye Terminator v1.1 Cycle Sequencing Kit (Applied Biosystems). BRAF mutational status was assessed retrospectively and had no impact on treatment decisions or follow up policy of study patients. Statistical analysis. SPSS 20.0 for windows was used to calculate the summary odds ratio (OR), Cox regression model, and the chi-square test. The OR was determined with 95% confidence intervals (CIs). Therefore, a chi-square test (Pearson) was performed for the analysis of the relationship between the BRAF V600E mutation and clinicopathologic
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Surgery Volume 157, Number 1 parameters for univariate analysis. A P < .05 was considered statistically significant. Cox proportional hazard modelling was used to relate risk factors to overall survival (OS), recurrence-free survival (RFS), and disease-specific survival (DSS). For statistical analysis, MACIS scores, EORTC scores, and the TNM stages were grouped in low-risk (MACIS 1, EORTC 1 + 2, and TNM stage I) and high-risk categories (MACIS 2–4, EORTC 3–5, and TNM stage II–IVc). To rank and compare each staging system’s ability to predict OS, RFS, and DSS, Kaplan-Meier curves were generated to visualize the data. Ethics committee. The study was approved by the local ethics committee of St. Gallen. RESULTS Cohort characteristics. BRAF exon 15 analysis was successful in 116/147 (79%) PTC. In the other 31 PTC tissue samples BRAF exon 15 analysis failed due to insufficient DNA quality. These patients were excluded from the subsequent analysis resulting in a final study cohort of 116 patients. BRAF V600E mutations were found in 75/116 (65%) PTC, whereas 41/116 PTC contained the wild type gene. Patient and tumor characteristics of the final cohort are summarized in Table I. Follow-up information was available for all patients. The median observation time of the patient cohort was 79 months (range 2–252 months), during which 9/116 patients (8%) developed recurrent disease. The median time to recurrence was 67 months (range 7–184 months). In 1 patient, the recurrence was local, in 4 (45%) regional, in 2 (22%) locoregional, and in 2 (22%) distant. In total, 12 patients (10%) died after a median of 63 months (range 12–174 months), 3 of whom died of disease (median 44 months, range 12–80 months). Of the remaining 9 patients 1 died of unknown cause, 5 due to a second primary cancer, and 3 due to non-cancer related causes. The 5- and 10-year OS, DSS, and RFS rates were 92% and 87%, 98% and 96%, and 96% and 94%, respectively. Clinicopathologic factors. For statistical analysis, patients with MACIS 1 (n = 71) were compared to patients with MACIS scores 2–4 (n = 45). Patients with MACIS score 1 had better OS (10 y 95% vs 75%, P = .008), DSS (10 y 100% vs 89%, P = .02) and RFS (100% vs 85%, P = .006) than patients with high MACIS scores as shown by KaplanMeier analysis. The EORTC groups (EORTC 1 + 2 ‘‘early score,’’ n = 73 vs EORTC 3–5 ‘‘advanced score,’’ n = 43) were also pooled and compared and statistically significant differences could be demonstrated in all three survival estimates between the groups. In the TNM groups (stage I
Table I. Clinicopathologic characteristics of the patient cohort (n = 116) Sex Mean age T category
N category
Focality Histology
Extrathyroidal extension TNM stage (UICC)
MACIS
EORTC
Female Male 50 y pT1b pT2 pT3 pT4a pT4b pN0 pN1a pN1b Unifocal multifocal Classic variant Follicular variant Oxyphil variant Tall cell variant No Yes I II III IVa IVb IVc 1 2 3 4 1 2 3 4 5
78 38 range 24 25 56 10 1 51 33 32 70 46 69 40 6 1 55 61 49 13 33 17 1 3 71 18 12 15 46 27 26 16 1
(67%) (33%) (10–86 y) (21%) (22%) (48%) (8%) (1%) (44%) (28%) (28%) (60%) (40%) (60%) (34%) (5%) (1%) (47%) (53%) (42%) (11%) (28%) (15%) (1%) (3%) (61%) (16%) (10%) (13%) (40%) (23%) (22%) (14%) (1%)
EORTC, European Organisation for Research and Treatment of Cancer; MACIS, Metastases, Age, Completeness of Resection, Invasion, Size; TNM, tumor, node, metastases; UICC, Union Internationale Contre le Cancer.
‘‘low stage,’’ n = 50 vs stage II–IV ‘‘high stage,’’ n = 60), the differences between the low and high stage TNM groups in OS and RFS reached statistical significance, but not in 10-y DSS (100% vs 91%, P = .08). BRAF V600E mutation status. In comparison to the subgroup without a BRAF V600E mutation (41/116, 35%) the subgroup with mutation (75/ 116, 65%) did not show greater rates of known, aggressive, clinicopathologic characteristics such as extrathyroidal extension, lymph node metastases, higher T-categories, male sex, or a greater age. Furthermore, there was no correlation between the BRAF V600E mutation status and the clinical risk scores MACIS, EORTC, or TNM stage. V600E mutations were, however, more frequent in the
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subgroup of classic variant PTC (50/69, 73%) than in the subgroup of follicular, oxyphil, and tall cell variant PTC (25/47, 53%; P = .03). When comparing the BRAF V600E positive subgroup to the BRAF V600E-negative subgroup the 10-year OS, DSS, and RFS rates were 92% and 79%, 98% and 91%, and 98% and 88%, respectively. None of these differences were statistically significant (P > .05). Furthermore, we performed a separate analysis of BRAF mutation effects in medium and greater risk tumors based on clinical risk scores but could not find any statistical differences or trends in survival between the groups either. Cox proportional hazard regression analysis revealed significantly lesser OS rates for age greater than 45 years. Other factors such as BRAF mutation status, sex, extrathyroidal extension, lymph node metastasis, multifocality, or histologic were not associated with decreased survival. Moreover, lesser OS rates were found when comparing MACIS scores (>1 vs 1), TNM stages (stage >1 to stage 1), and EORTC scores (>2 vs 1–2). Furthermore, differences in RFS could be demonstrated for EORTC scores (>2 vs 1–2). No significant differences in RFS were observed for the other parameters (Table II). DISCUSSION A major clinical challenge in the management of patients with PTC is how to reliably distinguish those patients who need aggressive treatments to decrease potentially treatment-associated morbidity and disease mortality, especially in view of the low overall mortality rate in PTC. Both the European and the American Thyroid Associations proposed practical management guidelines in which the clinical stage was integrated by additional parameters such as the tumor histologic variant, the results of postablative whole-body scan, and serum thyroglobulin levels.22,23 Despite that, patients included in the same risk group still show heterogeneous recurrence and survival rates. In addition, risk stratification according to the guidelines is not accurate in predicting long-term patient outcome and have a very low positive predictive value.24 The increasing understanding of the molecular basis of thyroid cancer has started to be translated into clinical practice and may help to better assess the prognosis and to individualize treatment.9 The data in the literature are conflicting, and the clinical importance of molecular markers in PTC is still controversial.21 The BRAF V600E mutation is the most frequent genetic alteration in PTC.19,25 In various reports, this mutation has been associated with aggressive tumor characteristics,11-13 whereas
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Table II. Hazard ratio for OS of different risk factors Covariates T-Category >T1a N-Category N+ ETE Female sex BRAF V600E negative UICC TNM >I MACIS >1 EORTC >2
OS hazard ratio (95% CI) 2.52 1.13 2.74 0.99 0.46 5.38 4.99 14.62
(0.32–19.73) (0.34–3.48) (0.74–10.15) (0.30–3.31) (0.14–1.49) (1.76–13.19) (1.34–18.52) (2.99–71.30)
P value .3 .8 .1 .3 .1 .05* .01* .001*
*Statistically significant. EORTC, European Organisation for Research and Treatment of Cancer; ETE, extrathyroidal extension; MACIS, Metastases, Age, Completeness of Resection, Invasion, Size; OS, overall survival; TNM, tumor, node, metastases; UICC, Union Internationale Contre le Cancer.
other recent studies20,26-28 questioned its prognostic impact. The frequency of the BRAF V600E mutation is high (>50%) compared with that of the poor outcomes (around 20%).29 The aim of our study was to evaluate the relationship between BRAF V600E mutational status and clinicopathologic parameters, including three established risk scores (MACIS score, EORTC score, TNM stage) and to analyze the impact of BRAF V600E mutation on survival in comparison with established risk scores in a consecutive single institutional patient cohort with PTC in Switzerland, excluding patients with papillary microcarcinoma to avoid a selection bias. This is the first study to investigate the prevalence of BRAF V600E mutational status in PTC patients in Switzerland. In line with previous studies13,14 as well as reports from other European countries,30 the BRAF V600E mutation was detected with a prevalence of 65% in our cohort. We did not find any difference in aggressive clinicopathologic characteristics between the subgroup with the BRAF V600E mutation in comparison to the BRAF wild type subgroup. This finding is in line with a recent study by Li et al31 but in contrast to other data in the literature.11-13 Li et al found no significant associations between BRAF V600E mutation and aggressive clinicopathologic features in a consecutive patient cohort who underwent total thyroidectomy and routine lateral neck dissection. Moreover, multivariate analysis revealed that the BRAF V600E status was not an independent predictor of lymph node metastases. Similar results were demonstrated by Ito et al.20 Furthermore, Nam et al32 did not find any correlation between BRAF V600E mutation and high-risk MACIS score, lymph node metastasis, age distribution, sex, or
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cancer size; in contrast, extrathyroidal extension and multifocality were associated with the BRAF V600E mutation. We found a greater frequency of the BRAF V600E mutations in the subgroup of classic variant PTC compared to the subgroup of nonclassic variant PTC. This finding is in line with previous studies which reported an association between the BRAF mutation status and the histologic PTC subtype and was significantly related to the classic and tall cell variant PTC but not to the follicular variant.12 There is no prognostic difference between the follicular and the classic variant type,33-36 whereas the tallcell variant PTC is characterized as having a more aggressive behavior usually presenting at an older age and higher stage than their classic PTC counterparts.37-40 The subgroups of oxyphil and tall cell variants could not be analyzed separately in our study due to low number of patients. In a multi-institutional, retrospective study addressing the relationship between the BRAF V600E mutation and mortality, Xing et al30 found an association between the BRAF V600E mutation and greater cancer-related mortality. The authors suggested that the BRAF V600E mutation may promote aggressive tumor behavior and risk of PTC-related mortality by up-regulation of classic angiogenic and tumor-promoting molecules, such as vascular endothelial growth factor, matrix metalloproteinases, and nuclear transcription factors.16,30,41 Furthermore, the BRAF V600E mutation seems to influence the tumor microenvironment such as tumor adhesion, migration, invasion, and metastasis.42 Mesa et al43 demonstrated that BRAF V600E activated normal rat thyroid cells to express genes possibly promoting tumor invasion by degradation of different noncellular components of the extracellular matrix. Furthermore, BRAF V600E activation in thyroid cells appears to lead to altered function of the sodium iodide symporter and other genes metabolizing iodide, which could be responsible for the decreased ability of PTC with BRAF V600E mutations to trap radioiodine leading to treatment failure.9,44-46 In contrast, the proposed primary tumorigenic role of BRAF V600E in PTC has been questioned. BRAF V600E seems to be a secondary subclone, rather than a primary event in thyroid tumorigenesis.47 Moreover, the BRAF V600E mutation has been found in lymph node metastases, but not in the primary tumor.16 The survival estimates in our patient cohort achieved with a 10-year OS of 87% and a 10-year DSS of 96% excellent rates comparable to the literature.2,3,30 A number of formal scoring systems have been developed to estimate prognosis among
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patients diagnosed with PTC. Lang et al48 conducted a comprehensive study to determine the most predictive staging system for PTC. The relative importance of each staging system was determined by calculating the proportion of variation in survival time explained (PVE). The three highest ranked staging systems by proportion of variation in survival time explained were the MACIS, the 6th edition TNM, and EORTC. They considered the MACIS score to be the stratification system of choice for PTC. In our cohort, high clinical risk scores (MACIS, EORTC, and TNM) were associated with lesser survival rates according to Kaplan-Meier survival analysis and the importance of these clinical risk scores for OS was also confirmed by Cox proportional hazard regression. We could demonstrate that individual components of the clinical risk scores (presence of regional or distant metastasis, age at diagnosis, completeness of resection, and presence of invasion, tumor size) had no impact on survival, whereas the combination in risk score was associated with all three survival estimates (OS, DSS, RFS). In contrast BRAF V600E mutational status did not influence survival in our cohort. According to Prescott et al,49 the addition of BRAF V600E mutational status to established risk algorithms improved the discrimination of risk recurrence in patients undergoing total thyroidectomy for PTC. In their retrospective study, the 5-year, cumulative recurrence incidence among BRAF V600E patients was 20% versus 8% among BRAF wild type. Moreover, the BRAF V600E mutation was significantly associated with time to recurrence when added to different current risk classification systems (Age, Metastases, Extent, Size; MACIS; UICC-TNM). Unfortunately, the authors did not analyze the additional impact of BRAF V600E mutational status on OS or DSS. We did not find any correlation between the BRAF V600E mutational status and the clinical risk scores MACIS, EORTC, or TNM. Moreover, neither a separate analysis of the effects of the BRAF mutation status in medium and higher risk tumors nor in combination with clinical risk scores as proposed by Prescott et al49 had any influence on survival estimates. In line with our findings, no impact of the BRAF V600E mutation on survival could be demonstrated in various recent studies20,50-53 leading to a great debate concerning the prognostic role of BRAF V600E mutations in PTC. In fact, when Xing et al30 adjusted for aggressive tumor behaviors, the association with BRAF V600E was no longer statistically significant. In multivariable analyses, BRAF V600E status was not independently
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associated with mortality. It seems correct that BRAF V600E plays an important biologic role in promoting aggressive tumor behavior; however, the BRAF V600E mutation may only be a marker of aggressiveness and not a true prognostic factor. According to Sancisi et al,54 analyzing the frequency of the BRAF V600E mutation within a group of highly aggressive PTC, which had developed distant metastases and a control group BRAF V600E mutation, was not associated with the development of distant metastases or fatal outcome. In conclusion, we could not confirm a negative prognostic impact of BRAF V600E mutational status on survival. The established risk algorithms of MACIS, EORTC, or TNM offered benefit for survival estimates. Based on our results, the usefulness of BRAF V600E mutation in the management algorithm of patients of PTC remains questionable, and meanwhile, we recommend using the current staging systems for risk stratification in PTC patients.
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