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Thyroid tumors of uncertain malignant potential: Morphologic and imunohistochemical analysis of 29 cases
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Original Article
Thyroid tumors of uncertain malignant potential: Morphologic and imunohistochemical analysis of 29 cases Adela Nechifor-Boila a , Angela Borda a,∗ , Geneviève Sassolas b , Zakia Hafdi-Nejjari b , Ramona C˘atan˘a c , Franc¸oise Borson-Chazot d , Nicole Berger e , Myriam Decaussin-Petrucci e a
Department of Histology, University of Medicine and Pharmacy, 38 Gheorghe Marinescu Str, Tîrgu-Mures¸ 540000, Romania Registre Rhône Alpin des cancers thyroïdiens, Centre de Médecine Nucléaire, Groupement Hospitalier Est, Hospices Civils de Lyon, 69677 Bron Cedex, France c Department of Endocrinology, University of Medicine and Pharmacy, 38 Gheorghe Marinescu Str, Tîrgu-Mures¸ 540000, Romania d Department of Endocrinology, Groupe Hospitalier Est, Hospices Civils de Lyon, Université Lyon 1, 59 boulevard Pinel, 69677 Bron Cedex, France e Department of Pathology, Centre Hospitalier Lyon Sud, Hospices Civils de Lyon, Université Lyon 1, 69495 Pierre Bénite, France b
a r t i c l e
i n f o
Article history: Received 18 May 2014 Received in revised form 7 September 2014 Accepted 5 December 2014 Keywords: Thyroid tumors of uncertain malignant potential Immunohistochemistry HBME-1 antigen CD56 antigen
a b s t r a c t Introduction: Thyroid tumors of uncertain malignant potential (TT-UMP) include follicular and welldifferentiated tumors of UMP (FT-UMP/WDT-UMP), as it refers to the presence of questionable capsular/vascular invasion or incompletely developed papillary thyroid carcinoma (PTC)-type nuclear changes. However, these tumors are difficult to diagnose in most cases. We aimed to investigate whether immunohistochemistry (HBME-1, cytokeratin-19, galectin-3, CD56 and p63) provides additional information concerning such lesions. Materials and methods: We performed an immunohistochemical analysis on 29 TT-UMP cases (22 WDTsUMP and 7 FTs-UMP) selected from the Rhone Alpes thyroid cancer registry and Departement of Pathology, Tîrgu-Mures¸ Emergency County Hospital database. The clinicopathological and follow-up data were obtained. Results: In the WDT-UMP group, HBME-1 was positive in 9/22 (40.9%) cases. CD56, a marker whose expression is reduced or absent in thyroid carcinoma, showed a “malignant” profile (no expression) in 13/22 (59.1%) cases. 7/22 (31.9%) cases were both HBME-1+ and CD56−. One case showed the coexpression of HBME-1, CD56, galectin-3 and cytokeratin-19. In the FT-UMP group, two cases were positive for HBME-1, other two for both galectin-3 and CK19 and only one case revealed a “malignant” CD56 profile. The follow-up data showed no distant metastases or persistent disease. Conclusion: Our study demonstrated very heterogeneous immunohistochemical profiles for TTs-UMP, further supporting the borderline nature of these lesions. WDTs-UMP revealed a certain tendency toward a PTC profile, suggesting a possible pathogeneticlink between these two entities. However, immunohistochemistry is still to be regarded more as a supporting diagnostic tool, while morphological criteria should always prime in the diagnostic decision. © 2014 Elsevier GmbH. All rights reserved.
Introduction Despite all past and recent efforts, conventional morphology has failed to classify some follicular-patterned thyroid lesions as benign or malignant due to the presence of some overlapping histological features [1–6]. Thus, the term “thyroid tumors of uncertain malignant potential (TT-UMP)” has generally been
∗ Corresponding author. Tel.: +40 722846985; fax: +40 265210407. E-mail address: [email protected] (A. Borda).
given to a group of borderline, encapsulated, follicular-patterned, thyroid lesions, showing architectural and/or cytological anomalies, suspicious but without fulfilling all the criteria of malignancy. However, this is a subjective area in thyroid pathology, with great inconsistency in pathological diagnosis [7–11], and an even more confusing area for the clinicians in respect of terminology. In an attempt to better classify these lesions, Williams et al. [12] has proposed in 2000 a new diagnostic terminology, that is well-differentiated and follicular tumors of UMP (WDT/FT-UMP), as it refers to the presence of incompletely developed papillary
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Please cite this article in press as: A. Nechifor-Boila, et al., Thyroid tumors of uncertain malignant potential: Morphologic and imunohistochemical analysis of 29 cases, Pathol. – Res. Pract (2015), http://dx.doi.org/10.1016/j.prp.2014.12.005
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Fig. 1. Well-differentiated thyroid tumors of uncertain malignant potential (WDTs-UMP), showing incompletely developed papillary thyroid carcinoma-type nuclear changes (HE) (a), focal and weak HBME-1 (b) and CK19 (c) immunoreactivity. Positive, membranar CD56 immunoreactivity in more that 10% of the tumor cells (considered as a negative result) in a case of WDT-UMP (d).
thyroid carcinoma (PTC)-type nuclear changes or questionable capsular/vascular invasion. The purpose of this study was to investigate whether immunohistochemistry (HBME-1, cytokeratin-19, galectin-3, CD56 and p63) provides additional information concerning such lesions.
Materials and methods Cases selection Twenty-nine cases of TT-UMP were collected from the Rhone Alpes thyroid cancer registry and Department of Pathology,
Fig. 2. A case of follicular-tumor of uncertain malignant potential (FT-UMP): an encapsulated tumor, with a dense, microfollicular architecture and minimal capsular invasion (HE) (a). Immunohistochemistry demonstrates strong and diffuse HBME-1 staining in more than 75% of the tumor cells (score 4) (b).
Please cite this article in press as: A. Nechifor-Boila, et al., Thyroid tumors of uncertain malignant potential: Morphologic and imunohistochemical analysis of 29 cases, Pathol. – Res. Pract (2015), http://dx.doi.org/10.1016/j.prp.2014.12.005
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Table 1 Antibodies used in the study. Antibody
Clone
Host species
Antigenretrieval (buffer)
Dilutiona
Source
HBME-1 Galectin-3 CK19 CD56 P63
HBME-1 9C4 RCK 108 123C3 4A4
Mouse (monoclonal) Mouse (monoclonal) Mouse (monoclonal) Mouse (monoclonal) Mouse (monoclonal)
WB (EDTA) WB (citrate, pH 6) WB (EDTA) WB (target retrieval solution, pH 6.1) WB (EDTA)
1:50 1:50 1:100 1:20 1:20
DAKO, Glostrup, Denmark Novocastra, Newcastle, UK Biogenex, Hague, the Netherlands DAKO, Glostrup, Denmark DAKO, Glostrup, Denmark
WB = water bath, 98 ◦ C. a For all antibodies used in the study the incubation time was 30 min.
Tîrgu-Mures¸ Emergency County Hospital database, between January 2002 and December 2012. The selected cases were retrieved from the surgical files of the Department of Pathology, Centre Hospitalier Lyon Sud (HCL, Lyon, France), the collaborating pathology laboratories of the Rhône Alpes region and Department of Pathology, Tîrgu-Mures¸ Emergency County Hospital. These cases consisted of TTs-UMP whose diagnosis was in accordance with the criteria proposed by Williams et al. [12]. Hematoxylin–phloxine–saffron (HPS)-stained slides from all of the cases were reviewed by a group of pathologists from the Rhone Alpes Region and Department of Pathology, Tîrgu-Mures¸ Emergency County Hospital (ANB, AB, NB and MDP) to confirm the diagnosis and classify the lesions into one of the study categories: WDT-UMP or FT-UMP (Fig 1a). The WDT-UMP and FT-UMP diagnoses were based on the criteria proposed for these tumors by Williams et al. [12]: encapsulated tumors showing incompletely developed PTC-type nuclear changes (nuclear enlargement, overlapping, irregularity of the nuclear contours, grooves, clearing or a ground glass appearance and nuclear pseudoinclusions), but no capsular invasion (WDT-UMP) or questionable capsular penetration without nuclear changes (FT-UMP) (Fig. 2a). The clinicopathological and follow-up data, when available, were obtained either from the Rhône Alpes thyroid cancer registry, either from Tîrgu-Mures¸ Emergency County Hospital database. The type of initial surgical procedure (total thyroidectomy/lobectomy), post-operative treatments and events in the period between surgery and the last clinical evaluation (follow-up period), if any, were all recorded. The ethics committee of the medical faculties (Faculté de Médecine RTH Laennec, Lyon and University of Medicine and Pharmacy, Tîrgu-Mures¸) and the state medical boards agreed to these investigations, and informed consent was obtained from all of the patients included in this study.
was considered when the tumor was entirely or almost entirely negative for this antibody (less than 10% of tumor cells stained positively). The immunostaining results for all of the cases were assessed by three pathologists (ANB, AB and MDP), and a consensus on the controversial cases was reached with the help of a triple-headed microscope.
Statistical analysis EpiInfo Software version 3.5.3 (CDC, Atlanta) was used for all statistical analysis, and a 2-tailed p-value < 0.05 was considered statistically significant.
Results Clinicopathological characteristics Twenty-two WDT-UMP cases and 7 FT-UMP cases were evaluated (Table 2). There were twenty-one females and 8 males patients included in the study, with a mean age at surgery of 30.6 ± 12.7 years. The tumors ranged in size from 0.8 to 7.5 cm, with a mean size of 3.1 ± 1.5 cm. All the tumors were encapsulated follicular neoplasms, showing either nuclear atypia that did not meet the criteria for PTC (n = 22), or incomplete capsular invasion or lack of definite vascular invasion, in the absence of nuclear atypia (n = 7). Followup information was available for 21 patients (72.4%), with a median follow-up of 4 years (minimum 1 year, maximum 9 years). None of these patients had persistent disease or had developed distant metastasis during the follow-up period.
Immunohistochemistry
Immunohistochemistry
The immunohistochemical staining was carried out on 4 m thick sections, using the labeled streptavidin–biotin–peroxidase complex system from a commercially available kit (VentanaiView DAB Detection Kit) on the VentanaNexes automated stainer (Ventana, Illkirck, France). Heat induced antigen retrieval was carried out for each antibody at 98 ◦ C using specific buffers. Sections were incubated for 30 min with the primary antibodies (Table 1). Positive and negative controls were used for each antibody. The following staining patterns were considered positive: HBME-1 membrane staining along the lateral and abluminal surfaces ± cytoplasmic, cytoplasmic and/or membrane expression of CK19 and galectin-3, and nuclear expression of p63. A positive immunoreactivity was considered when at least 10% of the tumor cells stained positively for these antibodies. The intensity of staining was further evaluated using a scoring system, as following: “1”, “2”, “3” or “4” when 10–25%, 26–50%, 51–75% or 76–100% of the tumor cells showed positive expression, respectively. For CD56, a marker whose expression is reduced or absent in thyroid carcinoma [13,14], a “malignant profile” (or positive result)
Tables 2 and 3 summarize the results of immunohistochemical staining for the five markers. In the WDT-UMP group, HBME-1 was positive in 9/22 (40.9%) cases, showing strong, diffuse staining in 4 cases (Fig. 1b). CD56 revealed a “malignant” profile (absent expression in more than 90% of the tumor cells) in 13/22 (59.1%) cases and a negative or “benign” profile (>10% of the tumor cells positive) in 9/22 (40.9%) cases (Fig. 1d). Seven out of 22 (31.9%) cases were positive for both antibodies. One case showed the co-expression of HBME-1, CD56, galectin-3 and cytokeratin-19. However, the staining for the last 2 markers was weak (less than 25% of tumor cells). None of the WDT-UMP cases stained positively for p63. In the FT-UMP group, two cases were positive for HBME-1, revealing a strong, diffuse staining in more than 50% of the tumor cells (Fig. 2b); other two cases stained positively for both galectin-3 and CK19, with variable degrees of intensity, ranging from a score of 1 to a score of 3. Only one FT-UMP case showed a “malignant” CD56 profile, with less than 10% positive tumors cells. There were 3 cases that did not stain for any of the 5 antibodies.
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Table 2 Clinicopathological data and immunohistochemical findings in 18 cases of thyroid tumors of uncertain malignant potential. No.
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29
Age/sex
22/F 34/F 16/F 32/F 31/F 24/F 30/F 29/F 29/F 21/M 34/F 18/F 33/F 11/F 36/M 44/F 41/M 38/M 28/M 45/F 41/M 38/F 35/M 19/F 25/F 31/F 16/F 67/F 47/M
Size (cm)
1.60 4.20 4.10 2.10 3.00 0.80 2.00 3.00 5.50 3.50 3.50 3.50 2.50 1.50 3.00 1.50 1.60 4.70 1.50 1.50 3.00 3.00 5.50 1.80 7.50 2.50 5.50 2.80 3.50
FT-UMP
− − − − − − − − − − − − − − − − − − − − − − + + + + + + +
WDT-UMP
+ + + + + + + + + + + + + + + + + + + + + + − − − − − − −
Follow-up (years)
9 7 1 4 1 4 2 7 8 n/a 3 1 1 2 5 n/a n/a n/a 4 7 7 n/a n/a 3 4 n/a n/a 5 5
Persistent disease ± distant metastasis
No No No No No No No No No n/a No No No No No n/a n/a n/a No No No n/a n/a No No n/a n/a No No
Score of immunohistochemistry HBME-1a
Galectin-3a
CK19a
CD56b
p63a
0 0 2 3 2 0 0 1 0 4 0 0 0 3 0 0 2 0 0 2 0 3 0 3 0 0 0 0 4
0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 2 0 0 0 0 1 0
0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 0 0 0 0 3 0
0 1 1 1 1 0 0 1 1 1 1 0 1 1 0 0 0 1 1 1 1 0 0 1 0 0 0 0 0
0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
WDT-UMP = well-differentiated tumors of uncertain malignant potential; FT-UMP = follicular tumors of uncertain malignant potential; n/a = not available. a Score for HBME-1, galectin-3, CK19 and p63 was graded as “0”, “1”, “2”, “3” or “4” when <10%, 10–25%, 26–50%, 51–75% or 76–100% of the tumor cells showed positive expression, respectively. b For CD56, a marker whose expression is reduced or absent in thyroid carcinoma, a “malignant profile” or a positive result (1) was considered when the tumor was entirely or almost entirely negative for this antibody (less than 10% of the tumor cells stained positively), while a negative result (0) was represented by the positive staining of at least 10% of the tumor cells.
Discussion In every day practice, the endocrine pathologist must provide the most appropriate diagnostic data in order that a patient with a thyroid nodule receives an adequate treatment. However, there are cases in which pathological criteria do not allow firm differentiation between benign and malignant follicular-patterned thyroid tumors, making the distinction between these two groups quite subtle and challenging. These so called “controversial” cases are generally encapsulated tumors that show no clear signs of invasion and/or have focal or questionable PTC nuclear features, also referred to as TT-UMP. In this study, we evaluated the expression and possible diagnostic role of five immunohistochemical markers (HBME-1, cytokeratin-19, galectin-3, CD56 and p63) in 29 TT-UMP cases. HBME-1, galectin-3 and cytokeratin-19 are the most frequently explored antibodies in thyroid pathology and have been shown to be sensitive markers for the diagnosis of PTC [3,4,15–19]. CD56 and p63 have been reported more recently [20,21] and CD56 in particular has been claimed to be a very sensitive and specific marker for PTC, including the follicular variant of PTC [20–22]. In our study, nine out of 22 (40.9%), 13/22 (59.1%) and 7/22 (31.8%) WDT-UMP cases revealed a “malignant” immunohistochemical profile, reflected either by a HBME-1, CD56 or both HBME-1 and CD56 positive, “malignant” expression (which means the absence of positive expression in more than 90% of the tumor cells for CD56). One WDT-UMP case showed the co-expression of four markers, with positive staining for galectin-3 and CK19 as well. Such immunohistochemical results could raise the question of whether these cases are actually PTCs, follicular variant, despite
their incomplete histological features. For encapsulated follicular variant of PTC, the diagnosis of malignancy relies on the presence of characteristic PTC nuclear features, that are nuclear enlargement, overlapping, irregularity of the nuclear contours, grooves, clearing or a ground glass appearance and nuclear pseudoinclusions [3]. In our study, the WDT-UMP diagnosis was settled based on the presence of incomplete, questionable, not “malignant enough” PTC nuclear features. All WDT-UMP cases in our study revealed nuclei that were enlarged, but not large enough, irregular, but without grooves, with only occasional clear, ground-grass appearance and no pseudoinclusions. All these nuclear criteria were evaluated by a group of thyroid pathology experts and the final diagnosis relied on a consensus. However, the subjectivity of diagnosing these Table 3 Positive staining for of HBME-1, CD56, galectin-3, CK19 and p63 in thyroid tumors of uncertain malignant potential cases in our study. Antibody
n (frequency, %) WDT-UMP (n = 22)
FT-UMP (n = 7)
9 (40.9%) 13 (59.1%) 2 (9.1%) 1 (4.5%) 0
2 (28.6%) 1 (14.3%) 2 (28.6%) 2 (28.6%) 0
Most frequent positive associations HBME-1 and CD56 7 (31.9%) 1 (4.5%) Galectin-3 and CK19
1 (14.3%) 2 (28.6%)
Single antibody HBME-1 CD56 Galectin-3 CK19 p63
WDT-UMP = well-differentiated tumors of uncertain malignant potential; FTUMP = follicular tumors of uncertain malignant potential; n/a = not available.
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difficult lesions, occurring even among expert pathologists, is well documented in the literature [8–10]. Six other WDT-UMP cases in our series had a “benign” immunohistochemical profile, with negative staining for both HBME-1 and CD56, revealing a probable benign biological behavior. This heterogeneous immunohistochemical profile of WDTUMPs has already been stated by Papotti et al. [23], who demonstrated positive HBME-1 expression in 9/13 WDT-UMP cases and some degree of positivity with either HBME-1 or galectin-3 in all of the cases except one. The authors concluded that such results reflect a possible pathogenetic link between these lesions and follicular variant of PTC and that the nuclear features of PTC are not merely artifacts. Similarly, our study, that included a larger number of TT-UMP cases, has reached the same conclusions. Regarding the FT-UMP group, 4 out of 7 (57.1%) cases had a “malignant” immunohistochemical profile, reflected either by a positive staining for both HBME-1 and CD 56 (one case), both CK19 and galectin-3 (two cases) or HBME-1 only (one case). The remaining 3 cases (43%) were negative for all of the tested antibodies. All in all, similarly to previously published studies [9,19,23], our results on TT-UMP’s imunohistochemical profile mirror their borderline nature between follicular adenoma and well-differentiated thyroid carcinoma. From a practical point of view, the most important question that arises is whether we should use immunohistochemistry as a supporting factor in our diagnostic decision in WDTs-UMP and FTs-UMP or if our diagnostic decision should be based only on morphological criteria. Because well-differentiated thyroid carcinoma generally has a “benign” outcome [3,24], long-term follow-up studies are needed to see if the biological attributes of malignancy differ among the various categories. Our results demonstrated a very heterogeneous immunohistochemical profile for these lesions and the follow-up data, which were available for most of the patients, revealed no distant metastases or persistent disease. However, a longer period of time is necessary to confirm their benign evolution. In view of these data, we consider that imunohistochemistry in WDT-UMP and FT-UMP cases should be used with caution and morphologic criteria should always be privileged. However, if imunohistochemmical findings are in conflict with the histological interpretation, it should be discussed in the pathologic report as a cautionary note to suggest the possible borderline nature of these tumors. We agree with Papotti et al. [23] that “the diagnosis of thyroid tumors of uncertain malignant potential should rest primarily on morphological criteria”. Their conclusion that “more molecular and biological information are necessary to better understand these borderline lesions” is still valid nine years after the publication of their article. Some recent studies have tried to reach the molecular insights of these tumors in order to answer this difficult question. Hofman et al. [9] have performed one of the first molecular analysis of the mutations and chromosomic rearrangements in a large series of TT-UMP cases and found that none of the WDT-UMP cases (15 cases) harbored the BRAFV600E, RET/PTC-1, RET/PTC-3 and PAX8/PPAR mutations and only few cases (3/15) were RAS mutated. Moreover, Rivera et al. [25] have shown that encapsulated follicular variant of PTCs have a molecular profile very close to follicular adenomas/carcinomas (10/28 RAS mutated and none BRAF mutated cases), while infiltrative follicular variant of PTCs have an opposite molecular profile, closer to classic PTCs (5/19 BRAF mutated and only 2/19 RAS mutated cases). Although performed by different groups of researchers, such results could highlight a similar molecular profile and probably a similar biological behavior for both WDTs-UMP and encapsulated follicular variant of PTCs, and different from infiltrative follicular variant of PTCs. The clinical consequence lining behind might be that both follicular variant of
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PTC (non-angioinvasive) and TTs-UMP (in particular WDTs-UMP) should be treated similarly. To present date, due to their unknown biologic behavior, firmly established management protocols for TTs-UMP do not exist. However, most of the authors recommend a conservative approach (lobectomy or lobectomy plus istmectomy and clinical surveillance) as treatment for these tumors, since they seem to have rather a benign outcome [23,26–28]. On the other hand, non-invasive follicular variant of PTC also behave in an indolent fashion, even after conservative treatment [29]. In summary, in our study, thyroid tumors of uncertain malignant potential demonstrated very heterogeneous immunohistochemical profiles. WDTs-UMP revealed a certain tendency toward a PTC profile (CD56− or HBME-1+ or CD56− and HBME-1+), suggesting a possible pathogenetic link between these two entities. However, immunohistochemistry is still to be regarded more as a supporting diagnostic tool, while morphological criteria should always prime in the diagnostic decision. When reaching the final diagnosis, one should always keep in mind that encapsulated, non-invasive, unifocal, follicular-patterned thyroid tumors seem to have more of an indolent biologic behavior, even after conservative therapy. Conflict of interest The authors declare that they have no conflicts of interest. Acknowledgements This paper was published under the frame of European Social Found, Human Resources Development Operational Programme 2007–2013, project no. POSDRU/159/1.5/S/136893. Also, it was made possible with the help of Région Rhône Alpes, Cancéropole Rhône Alpes, Institut National du Cancer, for which we are mostly grateful. References [1] J. Rosai, Handling of thyroid follicular patterned lesions, Endocr. Pathol. 16 (4) (2005) 279–283. [2] E. Fonseca, P. Soares, M. Cardoso-Oliveira, M. Sobrinho-Simoes, Diagnostic criteria in well-differentiated thyroid carcinomas, Endocr. Pathol. 17 (2) (2006) 109–117. [3] R.A. DeLellis, E.D. Williams, Pathology of the thyroid and parathyroid, in: R.A. DeLellis, R.V. Lloyd, P.U. Heitz (Eds.), Pathology and Genetics of Tumors of Endocrine Organs, IARC Press, Lyon, 2004, pp. 57–66. [4] V.A. LiVolsi, Papillary thyroid carcinoma: an update, Mod. Pathol. (2011) S1–S9. [5] V.A. LiVolsi, Z.W. Baloch, The many faces of follicular variant of papillary thyroid carcinoma, Pathol. Case Rev. (2009) 214–218. [6] K. Kakudo, Y. Bai, Z. Liu, Y. Li, Y. Ito, T. Ozaki, Classification of thyroid follicular cell tumors: with special reference to borderline lesions, Endocr. J. (2012) 1–12. [7] B. Franc, P. de la Salmoniere, F. Lange, C. Hoang, A. Louvel, A. de Roquancourt, F. Vilde, G. Hejblum, S. Chevret, C. Chastang, Interobserver and intraobserver reproducibility in the histopathology of follicular thyroid carcinoma, Hum. Pathol. (2003) 1092–1100. [8] M. Hirokawa, J.A. Carney, J.R. Goellner, R.A. DeLellis, C.S. Heffess, R. Katoh, M. Tsujimoto, K. Kakudo, Observer variation of encapsulated follicular lesions of the thyroid gland, Am. J. Surg. Pathol. (2002) 1508–1514. [9] V. Hofman, S. Lassalle, C. Bonnetaud, C. Butori, C. Loubatier, M. Ilie, O. Bordone, P. Brest, N. Guevara, J. Santini, B. Franc, P. Hofman, Thyroid tumours of uncertain malignant potential: frequency and diagnostic reproducibility, Virchows Arch. (2009) 21–33. [10] R.V. Lloyd, L.A. Erickson, M.B. Casey, K.Y. Lam, C.M. Lohse, S.L. Asa, J.K. Chan, R.A. DeLellis, H.R. Harach, K. Kakudo, V.A. LiVolsi, J. Rosai, T.J. Sebo, M. SobrinhoSimoes, B.M. Wenig, M.E. Lae, Observer variation in the diagnosis of follicular variant of papillary thyroid carcinoma, Am. J. Surg. Pathol. (2004) 1336–1340. [11] T.M. Elsheikh, S.L. Asa, J.K. Chan, R.A. DeLellis, C.S. Heffess, V.A. LiVolsi, B.M. Wenig, Interobserver and intraobserver variation among experts in the diagnosis of thyroid follicular lesions with borderline nuclear features of papillary carcinoma, Am. J. Clin. Pathol. (2008) 736–744. [12] E.D. Williams, Guest Editorial, Two proposals regarding the terminology of thyroid tumors, Int. J. Surg. Pathol. (2000) 181–183. [13] F. Satoh, S. Umemura, M. Yasuda, R.Y. Osamura, Neuroendocrine marker expression in thyroid epithelial tumors, Endocr. Pathol. (2001) 291–299.
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Original Article
Thyroid tumors of uncertain malignant potential: Morphologic and imunohistochemical analysis of 29 cases Adela Nechifor-Boila a , Angela Borda a,∗ , Geneviève Sassolas b , Zakia Hafdi-Nejjari b , Ramona C˘atan˘a c , Franc¸oise Borson-Chazot d , Nicole Berger e , Myriam Decaussin-Petrucci e a
Department of Histology, University of Medicine and Pharmacy, 38 Gheorghe Marinescu Str, Tîrgu-Mures¸ 540000, Romania Registre Rhône Alpin des cancers thyroïdiens, Centre de Médecine Nucléaire, Groupement Hospitalier Est, Hospices Civils de Lyon, 69677 Bron Cedex, France c Department of Endocrinology, University of Medicine and Pharmacy, 38 Gheorghe Marinescu Str, Tîrgu-Mures¸ 540000, Romania d Department of Endocrinology, Groupe Hospitalier Est, Hospices Civils de Lyon, Université Lyon 1, 59 boulevard Pinel, 69677 Bron Cedex, France e Department of Pathology, Centre Hospitalier Lyon Sud, Hospices Civils de Lyon, Université Lyon 1, 69495 Pierre Bénite, France b
a r t i c l e
i n f o
Article history: Received 18 May 2014 Received in revised form 7 September 2014 Accepted 5 December 2014 Keywords: Thyroid tumors of uncertain malignant potential Immunohistochemistry HBME-1 antigen CD56 antigen
a b s t r a c t Introduction: Thyroid tumors of uncertain malignant potential (TT-UMP) include follicular and welldifferentiated tumors of UMP (FT-UMP/WDT-UMP), as it refers to the presence of questionable capsular/vascular invasion or incompletely developed papillary thyroid carcinoma (PTC)-type nuclear changes. However, these tumors are difficult to diagnose in most cases. We aimed to investigate whether immunohistochemistry (HBME-1, cytokeratin-19, galectin-3, CD56 and p63) provides additional information concerning such lesions. Materials and methods: We performed an immunohistochemical analysis on 29 TT-UMP cases (22 WDTsUMP and 7 FTs-UMP) selected from the Rhone Alpes thyroid cancer registry and Departement of Pathology, Tîrgu-Mures¸ Emergency County Hospital database. The clinicopathological and follow-up data were obtained. Results: In the WDT-UMP group, HBME-1 was positive in 9/22 (40.9%) cases. CD56, a marker whose expression is reduced or absent in thyroid carcinoma, showed a “malignant” profile (no expression) in 13/22 (59.1%) cases. 7/22 (31.9%) cases were both HBME-1+ and CD56−. One case showed the coexpression of HBME-1, CD56, galectin-3 and cytokeratin-19. In the FT-UMP group, two cases were positive for HBME-1, other two for both galectin-3 and CK19 and only one case revealed a “malignant” CD56 profile. The follow-up data showed no distant metastases or persistent disease. Conclusion: Our study demonstrated very heterogeneous immunohistochemical profiles for TTs-UMP, further supporting the borderline nature of these lesions. WDTs-UMP revealed a certain tendency toward a PTC profile, suggesting a possible pathogeneticlink between these two entities. However, immunohistochemistry is still to be regarded more as a supporting diagnostic tool, while morphological criteria should always prime in the diagnostic decision. © 2014 Elsevier GmbH. All rights reserved.
Introduction Despite all past and recent efforts, conventional morphology has failed to classify some follicular-patterned thyroid lesions as benign or malignant due to the presence of some overlapping histological features [1–6]. Thus, the term “thyroid tumors of uncertain malignant potential (TT-UMP)” has generally been
∗ Corresponding author. Tel.: +40 722846985; fax: +40 265210407. E-mail address: [email protected] (A. Borda).
given to a group of borderline, encapsulated, follicular-patterned, thyroid lesions, showing architectural and/or cytological anomalies, suspicious but without fulfilling all the criteria of malignancy. However, this is a subjective area in thyroid pathology, with great inconsistency in pathological diagnosis [7–11], and an even more confusing area for the clinicians in respect of terminology. In an attempt to better classify these lesions, Williams et al. [12] has proposed in 2000 a new diagnostic terminology, that is well-differentiated and follicular tumors of UMP (WDT/FT-UMP), as it refers to the presence of incompletely developed papillary
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Fig. 1. Well-differentiated thyroid tumors of uncertain malignant potential (WDTs-UMP), showing incompletely developed papillary thyroid carcinoma-type nuclear changes (HE) (a), focal and weak HBME-1 (b) and CK19 (c) immunoreactivity. Positive, membranar CD56 immunoreactivity in more that 10% of the tumor cells (considered as a negative result) in a case of WDT-UMP (d).
thyroid carcinoma (PTC)-type nuclear changes or questionable capsular/vascular invasion. The purpose of this study was to investigate whether immunohistochemistry (HBME-1, cytokeratin-19, galectin-3, CD56 and p63) provides additional information concerning such lesions.
Materials and methods Cases selection Twenty-nine cases of TT-UMP were collected from the Rhone Alpes thyroid cancer registry and Department of Pathology,
Fig. 2. A case of follicular-tumor of uncertain malignant potential (FT-UMP): an encapsulated tumor, with a dense, microfollicular architecture and minimal capsular invasion (HE) (a). Immunohistochemistry demonstrates strong and diffuse HBME-1 staining in more than 75% of the tumor cells (score 4) (b).
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Table 1 Antibodies used in the study. Antibody
Clone
Host species
Antigenretrieval (buffer)
Dilutiona
Source
HBME-1 Galectin-3 CK19 CD56 P63
HBME-1 9C4 RCK 108 123C3 4A4
Mouse (monoclonal) Mouse (monoclonal) Mouse (monoclonal) Mouse (monoclonal) Mouse (monoclonal)
WB (EDTA) WB (citrate, pH 6) WB (EDTA) WB (target retrieval solution, pH 6.1) WB (EDTA)
1:50 1:50 1:100 1:20 1:20
DAKO, Glostrup, Denmark Novocastra, Newcastle, UK Biogenex, Hague, the Netherlands DAKO, Glostrup, Denmark DAKO, Glostrup, Denmark
WB = water bath, 98 ◦ C. a For all antibodies used in the study the incubation time was 30 min.
Tîrgu-Mures¸ Emergency County Hospital database, between January 2002 and December 2012. The selected cases were retrieved from the surgical files of the Department of Pathology, Centre Hospitalier Lyon Sud (HCL, Lyon, France), the collaborating pathology laboratories of the Rhône Alpes region and Department of Pathology, Tîrgu-Mures¸ Emergency County Hospital. These cases consisted of TTs-UMP whose diagnosis was in accordance with the criteria proposed by Williams et al. [12]. Hematoxylin–phloxine–saffron (HPS)-stained slides from all of the cases were reviewed by a group of pathologists from the Rhone Alpes Region and Department of Pathology, Tîrgu-Mures¸ Emergency County Hospital (ANB, AB, NB and MDP) to confirm the diagnosis and classify the lesions into one of the study categories: WDT-UMP or FT-UMP (Fig 1a). The WDT-UMP and FT-UMP diagnoses were based on the criteria proposed for these tumors by Williams et al. [12]: encapsulated tumors showing incompletely developed PTC-type nuclear changes (nuclear enlargement, overlapping, irregularity of the nuclear contours, grooves, clearing or a ground glass appearance and nuclear pseudoinclusions), but no capsular invasion (WDT-UMP) or questionable capsular penetration without nuclear changes (FT-UMP) (Fig. 2a). The clinicopathological and follow-up data, when available, were obtained either from the Rhône Alpes thyroid cancer registry, either from Tîrgu-Mures¸ Emergency County Hospital database. The type of initial surgical procedure (total thyroidectomy/lobectomy), post-operative treatments and events in the period between surgery and the last clinical evaluation (follow-up period), if any, were all recorded. The ethics committee of the medical faculties (Faculté de Médecine RTH Laennec, Lyon and University of Medicine and Pharmacy, Tîrgu-Mures¸) and the state medical boards agreed to these investigations, and informed consent was obtained from all of the patients included in this study.
was considered when the tumor was entirely or almost entirely negative for this antibody (less than 10% of tumor cells stained positively). The immunostaining results for all of the cases were assessed by three pathologists (ANB, AB and MDP), and a consensus on the controversial cases was reached with the help of a triple-headed microscope.
Statistical analysis EpiInfo Software version 3.5.3 (CDC, Atlanta) was used for all statistical analysis, and a 2-tailed p-value < 0.05 was considered statistically significant.
Results Clinicopathological characteristics Twenty-two WDT-UMP cases and 7 FT-UMP cases were evaluated (Table 2). There were twenty-one females and 8 males patients included in the study, with a mean age at surgery of 30.6 ± 12.7 years. The tumors ranged in size from 0.8 to 7.5 cm, with a mean size of 3.1 ± 1.5 cm. All the tumors were encapsulated follicular neoplasms, showing either nuclear atypia that did not meet the criteria for PTC (n = 22), or incomplete capsular invasion or lack of definite vascular invasion, in the absence of nuclear atypia (n = 7). Followup information was available for 21 patients (72.4%), with a median follow-up of 4 years (minimum 1 year, maximum 9 years). None of these patients had persistent disease or had developed distant metastasis during the follow-up period.
Immunohistochemistry
Immunohistochemistry
The immunohistochemical staining was carried out on 4 m thick sections, using the labeled streptavidin–biotin–peroxidase complex system from a commercially available kit (VentanaiView DAB Detection Kit) on the VentanaNexes automated stainer (Ventana, Illkirck, France). Heat induced antigen retrieval was carried out for each antibody at 98 ◦ C using specific buffers. Sections were incubated for 30 min with the primary antibodies (Table 1). Positive and negative controls were used for each antibody. The following staining patterns were considered positive: HBME-1 membrane staining along the lateral and abluminal surfaces ± cytoplasmic, cytoplasmic and/or membrane expression of CK19 and galectin-3, and nuclear expression of p63. A positive immunoreactivity was considered when at least 10% of the tumor cells stained positively for these antibodies. The intensity of staining was further evaluated using a scoring system, as following: “1”, “2”, “3” or “4” when 10–25%, 26–50%, 51–75% or 76–100% of the tumor cells showed positive expression, respectively. For CD56, a marker whose expression is reduced or absent in thyroid carcinoma [13,14], a “malignant profile” (or positive result)
Tables 2 and 3 summarize the results of immunohistochemical staining for the five markers. In the WDT-UMP group, HBME-1 was positive in 9/22 (40.9%) cases, showing strong, diffuse staining in 4 cases (Fig. 1b). CD56 revealed a “malignant” profile (absent expression in more than 90% of the tumor cells) in 13/22 (59.1%) cases and a negative or “benign” profile (>10% of the tumor cells positive) in 9/22 (40.9%) cases (Fig. 1d). Seven out of 22 (31.9%) cases were positive for both antibodies. One case showed the co-expression of HBME-1, CD56, galectin-3 and cytokeratin-19. However, the staining for the last 2 markers was weak (less than 25% of tumor cells). None of the WDT-UMP cases stained positively for p63. In the FT-UMP group, two cases were positive for HBME-1, revealing a strong, diffuse staining in more than 50% of the tumor cells (Fig. 2b); other two cases stained positively for both galectin-3 and CK19, with variable degrees of intensity, ranging from a score of 1 to a score of 3. Only one FT-UMP case showed a “malignant” CD56 profile, with less than 10% positive tumors cells. There were 3 cases that did not stain for any of the 5 antibodies.
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Table 2 Clinicopathological data and immunohistochemical findings in 18 cases of thyroid tumors of uncertain malignant potential. No.
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29
Age/sex
22/F 34/F 16/F 32/F 31/F 24/F 30/F 29/F 29/F 21/M 34/F 18/F 33/F 11/F 36/M 44/F 41/M 38/M 28/M 45/F 41/M 38/F 35/M 19/F 25/F 31/F 16/F 67/F 47/M
Size (cm)
1.60 4.20 4.10 2.10 3.00 0.80 2.00 3.00 5.50 3.50 3.50 3.50 2.50 1.50 3.00 1.50 1.60 4.70 1.50 1.50 3.00 3.00 5.50 1.80 7.50 2.50 5.50 2.80 3.50
FT-UMP
− − − − − − − − − − − − − − − − − − − − − − + + + + + + +
WDT-UMP
+ + + + + + + + + + + + + + + + + + + + + + − − − − − − −
Follow-up (years)
9 7 1 4 1 4 2 7 8 n/a 3 1 1 2 5 n/a n/a n/a 4 7 7 n/a n/a 3 4 n/a n/a 5 5
Persistent disease ± distant metastasis
No No No No No No No No No n/a No No No No No n/a n/a n/a No No No n/a n/a No No n/a n/a No No
Score of immunohistochemistry HBME-1a
Galectin-3a
CK19a
CD56b
p63a
0 0 2 3 2 0 0 1 0 4 0 0 0 3 0 0 2 0 0 2 0 3 0 3 0 0 0 0 4
0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 2 0 0 0 0 1 0
0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 0 0 0 0 3 0
0 1 1 1 1 0 0 1 1 1 1 0 1 1 0 0 0 1 1 1 1 0 0 1 0 0 0 0 0
0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
WDT-UMP = well-differentiated tumors of uncertain malignant potential; FT-UMP = follicular tumors of uncertain malignant potential; n/a = not available. a Score for HBME-1, galectin-3, CK19 and p63 was graded as “0”, “1”, “2”, “3” or “4” when <10%, 10–25%, 26–50%, 51–75% or 76–100% of the tumor cells showed positive expression, respectively. b For CD56, a marker whose expression is reduced or absent in thyroid carcinoma, a “malignant profile” or a positive result (1) was considered when the tumor was entirely or almost entirely negative for this antibody (less than 10% of the tumor cells stained positively), while a negative result (0) was represented by the positive staining of at least 10% of the tumor cells.
Discussion In every day practice, the endocrine pathologist must provide the most appropriate diagnostic data in order that a patient with a thyroid nodule receives an adequate treatment. However, there are cases in which pathological criteria do not allow firm differentiation between benign and malignant follicular-patterned thyroid tumors, making the distinction between these two groups quite subtle and challenging. These so called “controversial” cases are generally encapsulated tumors that show no clear signs of invasion and/or have focal or questionable PTC nuclear features, also referred to as TT-UMP. In this study, we evaluated the expression and possible diagnostic role of five immunohistochemical markers (HBME-1, cytokeratin-19, galectin-3, CD56 and p63) in 29 TT-UMP cases. HBME-1, galectin-3 and cytokeratin-19 are the most frequently explored antibodies in thyroid pathology and have been shown to be sensitive markers for the diagnosis of PTC [3,4,15–19]. CD56 and p63 have been reported more recently [20,21] and CD56 in particular has been claimed to be a very sensitive and specific marker for PTC, including the follicular variant of PTC [20–22]. In our study, nine out of 22 (40.9%), 13/22 (59.1%) and 7/22 (31.8%) WDT-UMP cases revealed a “malignant” immunohistochemical profile, reflected either by a HBME-1, CD56 or both HBME-1 and CD56 positive, “malignant” expression (which means the absence of positive expression in more than 90% of the tumor cells for CD56). One WDT-UMP case showed the co-expression of four markers, with positive staining for galectin-3 and CK19 as well. Such immunohistochemical results could raise the question of whether these cases are actually PTCs, follicular variant, despite
their incomplete histological features. For encapsulated follicular variant of PTC, the diagnosis of malignancy relies on the presence of characteristic PTC nuclear features, that are nuclear enlargement, overlapping, irregularity of the nuclear contours, grooves, clearing or a ground glass appearance and nuclear pseudoinclusions [3]. In our study, the WDT-UMP diagnosis was settled based on the presence of incomplete, questionable, not “malignant enough” PTC nuclear features. All WDT-UMP cases in our study revealed nuclei that were enlarged, but not large enough, irregular, but without grooves, with only occasional clear, ground-grass appearance and no pseudoinclusions. All these nuclear criteria were evaluated by a group of thyroid pathology experts and the final diagnosis relied on a consensus. However, the subjectivity of diagnosing these Table 3 Positive staining for of HBME-1, CD56, galectin-3, CK19 and p63 in thyroid tumors of uncertain malignant potential cases in our study. Antibody
n (frequency, %) WDT-UMP (n = 22)
FT-UMP (n = 7)
9 (40.9%) 13 (59.1%) 2 (9.1%) 1 (4.5%) 0
2 (28.6%) 1 (14.3%) 2 (28.6%) 2 (28.6%) 0
Most frequent positive associations HBME-1 and CD56 7 (31.9%) 1 (4.5%) Galectin-3 and CK19
1 (14.3%) 2 (28.6%)
Single antibody HBME-1 CD56 Galectin-3 CK19 p63
WDT-UMP = well-differentiated tumors of uncertain malignant potential; FTUMP = follicular tumors of uncertain malignant potential; n/a = not available.
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difficult lesions, occurring even among expert pathologists, is well documented in the literature [8–10]. Six other WDT-UMP cases in our series had a “benign” immunohistochemical profile, with negative staining for both HBME-1 and CD56, revealing a probable benign biological behavior. This heterogeneous immunohistochemical profile of WDTUMPs has already been stated by Papotti et al. [23], who demonstrated positive HBME-1 expression in 9/13 WDT-UMP cases and some degree of positivity with either HBME-1 or galectin-3 in all of the cases except one. The authors concluded that such results reflect a possible pathogenetic link between these lesions and follicular variant of PTC and that the nuclear features of PTC are not merely artifacts. Similarly, our study, that included a larger number of TT-UMP cases, has reached the same conclusions. Regarding the FT-UMP group, 4 out of 7 (57.1%) cases had a “malignant” immunohistochemical profile, reflected either by a positive staining for both HBME-1 and CD 56 (one case), both CK19 and galectin-3 (two cases) or HBME-1 only (one case). The remaining 3 cases (43%) were negative for all of the tested antibodies. All in all, similarly to previously published studies [9,19,23], our results on TT-UMP’s imunohistochemical profile mirror their borderline nature between follicular adenoma and well-differentiated thyroid carcinoma. From a practical point of view, the most important question that arises is whether we should use immunohistochemistry as a supporting factor in our diagnostic decision in WDTs-UMP and FTs-UMP or if our diagnostic decision should be based only on morphological criteria. Because well-differentiated thyroid carcinoma generally has a “benign” outcome [3,24], long-term follow-up studies are needed to see if the biological attributes of malignancy differ among the various categories. Our results demonstrated a very heterogeneous immunohistochemical profile for these lesions and the follow-up data, which were available for most of the patients, revealed no distant metastases or persistent disease. However, a longer period of time is necessary to confirm their benign evolution. In view of these data, we consider that imunohistochemistry in WDT-UMP and FT-UMP cases should be used with caution and morphologic criteria should always be privileged. However, if imunohistochemmical findings are in conflict with the histological interpretation, it should be discussed in the pathologic report as a cautionary note to suggest the possible borderline nature of these tumors. We agree with Papotti et al. [23] that “the diagnosis of thyroid tumors of uncertain malignant potential should rest primarily on morphological criteria”. Their conclusion that “more molecular and biological information are necessary to better understand these borderline lesions” is still valid nine years after the publication of their article. Some recent studies have tried to reach the molecular insights of these tumors in order to answer this difficult question. Hofman et al. [9] have performed one of the first molecular analysis of the mutations and chromosomic rearrangements in a large series of TT-UMP cases and found that none of the WDT-UMP cases (15 cases) harbored the BRAFV600E, RET/PTC-1, RET/PTC-3 and PAX8/PPAR mutations and only few cases (3/15) were RAS mutated. Moreover, Rivera et al. [25] have shown that encapsulated follicular variant of PTCs have a molecular profile very close to follicular adenomas/carcinomas (10/28 RAS mutated and none BRAF mutated cases), while infiltrative follicular variant of PTCs have an opposite molecular profile, closer to classic PTCs (5/19 BRAF mutated and only 2/19 RAS mutated cases). Although performed by different groups of researchers, such results could highlight a similar molecular profile and probably a similar biological behavior for both WDTs-UMP and encapsulated follicular variant of PTCs, and different from infiltrative follicular variant of PTCs. The clinical consequence lining behind might be that both follicular variant of
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PTC (non-angioinvasive) and TTs-UMP (in particular WDTs-UMP) should be treated similarly. To present date, due to their unknown biologic behavior, firmly established management protocols for TTs-UMP do not exist. However, most of the authors recommend a conservative approach (lobectomy or lobectomy plus istmectomy and clinical surveillance) as treatment for these tumors, since they seem to have rather a benign outcome [23,26–28]. On the other hand, non-invasive follicular variant of PTC also behave in an indolent fashion, even after conservative treatment [29]. In summary, in our study, thyroid tumors of uncertain malignant potential demonstrated very heterogeneous immunohistochemical profiles. WDTs-UMP revealed a certain tendency toward a PTC profile (CD56− or HBME-1+ or CD56− and HBME-1+), suggesting a possible pathogenetic link between these two entities. However, immunohistochemistry is still to be regarded more as a supporting diagnostic tool, while morphological criteria should always prime in the diagnostic decision. When reaching the final diagnosis, one should always keep in mind that encapsulated, non-invasive, unifocal, follicular-patterned thyroid tumors seem to have more of an indolent biologic behavior, even after conservative therapy. Conflict of interest The authors declare that they have no conflicts of interest. Acknowledgements This paper was published under the frame of European Social Found, Human Resources Development Operational Programme 2007–2013, project no. POSDRU/159/1.5/S/136893. Also, it was made possible with the help of Région Rhône Alpes, Cancéropole Rhône Alpes, Institut National du Cancer, for which we are mostly grateful. References [1] J. Rosai, Handling of thyroid follicular patterned lesions, Endocr. Pathol. 16 (4) (2005) 279–283. [2] E. Fonseca, P. Soares, M. Cardoso-Oliveira, M. Sobrinho-Simoes, Diagnostic criteria in well-differentiated thyroid carcinomas, Endocr. Pathol. 17 (2) (2006) 109–117. [3] R.A. DeLellis, E.D. Williams, Pathology of the thyroid and parathyroid, in: R.A. DeLellis, R.V. Lloyd, P.U. Heitz (Eds.), Pathology and Genetics of Tumors of Endocrine Organs, IARC Press, Lyon, 2004, pp. 57–66. [4] V.A. LiVolsi, Papillary thyroid carcinoma: an update, Mod. Pathol. (2011) S1–S9. [5] V.A. LiVolsi, Z.W. Baloch, The many faces of follicular variant of papillary thyroid carcinoma, Pathol. Case Rev. (2009) 214–218. [6] K. Kakudo, Y. Bai, Z. Liu, Y. Li, Y. Ito, T. Ozaki, Classification of thyroid follicular cell tumors: with special reference to borderline lesions, Endocr. J. (2012) 1–12. [7] B. Franc, P. de la Salmoniere, F. Lange, C. Hoang, A. Louvel, A. de Roquancourt, F. Vilde, G. Hejblum, S. Chevret, C. Chastang, Interobserver and intraobserver reproducibility in the histopathology of follicular thyroid carcinoma, Hum. Pathol. (2003) 1092–1100. [8] M. Hirokawa, J.A. Carney, J.R. Goellner, R.A. DeLellis, C.S. Heffess, R. Katoh, M. Tsujimoto, K. Kakudo, Observer variation of encapsulated follicular lesions of the thyroid gland, Am. J. Surg. Pathol. (2002) 1508–1514. [9] V. Hofman, S. Lassalle, C. Bonnetaud, C. Butori, C. Loubatier, M. Ilie, O. Bordone, P. Brest, N. Guevara, J. Santini, B. Franc, P. Hofman, Thyroid tumours of uncertain malignant potential: frequency and diagnostic reproducibility, Virchows Arch. (2009) 21–33. [10] R.V. Lloyd, L.A. Erickson, M.B. Casey, K.Y. Lam, C.M. Lohse, S.L. Asa, J.K. Chan, R.A. DeLellis, H.R. Harach, K. Kakudo, V.A. LiVolsi, J. Rosai, T.J. Sebo, M. SobrinhoSimoes, B.M. Wenig, M.E. Lae, Observer variation in the diagnosis of follicular variant of papillary thyroid carcinoma, Am. J. Surg. Pathol. (2004) 1336–1340. [11] T.M. Elsheikh, S.L. Asa, J.K. Chan, R.A. DeLellis, C.S. Heffess, V.A. LiVolsi, B.M. Wenig, Interobserver and intraobserver variation among experts in the diagnosis of thyroid follicular lesions with borderline nuclear features of papillary carcinoma, Am. J. Clin. Pathol. (2008) 736–744. [12] E.D. Williams, Guest Editorial, Two proposals regarding the terminology of thyroid tumors, Int. J. Surg. Pathol. (2000) 181–183. [13] F. Satoh, S. Umemura, M. Yasuda, R.Y. Osamura, Neuroendocrine marker expression in thyroid epithelial tumors, Endocr. Pathol. (2001) 291–299.
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Please cite this article in press as: A. Nechifor-Boila, et al., Thyroid tumors of uncertain malignant potential: Morphologic and imunohistochemical analysis of 29 cases, Pathol. – Res. Pract (2015), http://dx.doi.org/10.1016/j.prp.2014.12.005