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ORIGINAL ARTICLE  
Year : 2019  |  Volume : 62  |  Issue : 2  |  Page : 206-210
The diagnostic significance of trophoblast cell-surface antigen-2 expression in benign and malignant thyroid lesions


1 Department of Pathology, Health Sciences University, Antalya Education and Research Hospital, Antalya 07050, Turkey
2 Department of Pathology, Health Sciences University, Mehmet Akif Inan Education and Research Hospital, Sanliurfa 63300, Turkey
3 Department of General Surgery, Health Sciences University, Antalya Education and Research Hospital, Antalya 07050, Turkey

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Date of Web Publication10-Apr-2019
 

   Abstract 


Context: Thyroid cancers are the most common malignancy of the endocrine system. Over-expression of trophoblast cell-surface antigen 2 (TROP-2) in various tumors has been found to correlate with poor prognosis and aggressive tumor behavior. Aims: The aim of this study was to evaluateTROP-2 expression in thyroid neoplasms. Subjects and Methods: This study contained 152 cases, including 48 follicular nodular disease (FND), 29 follicular adenoma (FA), 57 papillary thyroid carcinoma (PTC), 12 follicular thyroid carcinoma (FTC), 3 medullary thyroid carcinoma (MTC), 2 poorly differentiated thyroid carcinoma (PDTC) and 1 undifferentiated thyroid carcinoma (UDTC). TROP-2 expression was investigated via immunohistochemistry in sections prepared from paraffin blocks of the cases. Results: The cases comprised 32 (21%) males and 120 (79%) females with a mean age of 46.8 years (range, 15-85 years). TROP-2 expression was observed in 74.6% of the malignant lesions of the thyroid except for medullary carcinoma, poorly differentiated and undifferentiated thyroid carcinoma. Immunoreactivity was 3.4% in FA, 41.7% of cases with FTC and 81.8% in PTC follicular variant (PTC fv). The difference between FA/FTC and FA/PTC follicular variant were both significant (P < 0.005, P < 0.001, respectively). There was no difference between FTC/PTC fv (P = 0.089). Conclusion: TROP-2 can be considered a useful marker for distinguishing PTC fv cases from follicular nodular disease and follicular adenoma cases because of its high sensitivity in the identification of papillary carcinomas of the thyroid.

Keywords: Follicular neoplasm, thyroid carcinoma, TROP-2

How to cite this article:
Sadullahoğlu C, Sayıner A, Süren D, Yıldırım HT, Nergiz D, Sezer C, Oruç MT. The diagnostic significance of trophoblast cell-surface antigen-2 expression in benign and malignant thyroid lesions. Indian J Pathol Microbiol 2019;62:206-10

How to cite this URL:
Sadullahoğlu C, Sayıner A, Süren D, Yıldırım HT, Nergiz D, Sezer C, Oruç MT. The diagnostic significance of trophoblast cell-surface antigen-2 expression in benign and malignant thyroid lesions. Indian J Pathol Microbiol [serial online] 2019 [cited 2019 Jun 26];62:206-10. Available from: http://www.ijpmonline.org/text.asp?2019/62/2/206/255806





   Introduction Top


Thyroid carcinoma is the most common endocrine malignancy.[1] Of all malignant cases in the thyroid, >90% are well-differentiated thyroid carcinomas (papillary thyroid carcinoma 80%, follicular thyroid carcinoma 10-15%).[1],[2] The diagnosis of benign and malignant thyroid lesions is easily made by cytomorphological and histomorphological evaluation on H and E stained slides. Diagnostic difficulties may occur in encapsulated follicular lesions with a suspicious capsule and vascular invasion and with an absence of the characteristic nuclear features of papillary carcinoma. This can lead to serious differences among pathologists in the evaluation of the same lesion.[1],[2] In morphologically overlapping cases, immunohistochemistry is necessary for the differential diagnosis. Although the most commonly used immunohistochemical (IHC) markers are Hector Battifora Mesothelial 1 (HBME-1), galectin-3 and Cytokeratin 19 (CK19), none of these markers provide 100% differential diagnosis because of expression in both neoplastic and non-neoplastic lesions.[1],[2],[3]

TROP-2, also referred to as GA733-1 (gastric antigen 733-1), MIS-1 (surface marker 1, EGP-1 (epithelial glycoprotein-1), is a transmembrane receptor glycoprotein encoded by the tumor-associated calcium signal transducer 2 (Tacstd2) gene, which is located on chromosome 1p32.[4] TROP-2 was first discovered on the surface of trophoblast cells.[5] TROP-2 over-expression is associated with diverse clinicopathological factors and poor clinical outcome in various types of epithelial cancers including breast, gastric, colorectal, endometrial and ovarian cancers.[5],[6],[7],[8],[9],[10] There are few studies in literature which have investigated TROP-2 expression in thyroid neoplasms. In some of these, the immunoreactivity of this new marker has been studied on microarrays and cytological samples. In addition, the sensitivity and specificity of TROP-2 expression have been reported to be high in papillary thyroid carcinomas.[11],[12],[13],[14],[15]

The aim of this study was to evaluate the expression of TROP-2 and, using IHC, to analyse its sensitivity in the differentiation of benign and malignant thyroid lesions.


   Subjects and Methods Top


Case selection

This retrospective study was conducted on 152 cases of thyroid neoplasm that were histopathologically diagnosed using thyroidectomy tissue at Antalya Training and Research Hospital (Turkey), between 2009 and 2016. The cases included 77 benign non-neoplastic lesions diagnosed as 48 follicular nodular disease (FND) and 29 follicular adenoma (FA) and 75 cases of malignant tumors (57 cases of PTC, 12 cases of FTC, 12 cases of MC, 2 cases of PDTC and 1 case of UDTC). The PTC cases were subdivided into papillary microcarcinoma (mPTC; n = 21), PTC classic variant (PTC cv; n = 19), PTC follicular variant (PTC fv; n = 11) and PTC oncocytic variant (PTC ov; n = 6). The study was approved by the Ethics Committee of Antalya Training and Research Hospital (Approval No: 2017-065).

Immunohistochemistry staining

All cases were reviewed by 2 pathologists to establish the histological type and characteristics of the tumours according to World Health Organization criteria (2004). All samples were routinely fixed for 24 hours in 10% buffered formaldehyde, then embedded in paraffin blocks and sections were cut of 4 μm thickness onto positively charged slides for immunohistochemical application. The tissue sections were deparaffinised in xylene and rehydrated using graded alcohol and deionised H2O. For antigen retrieval, the tissue sections were immersed in a 10% citrate buffer solution (#RE7113; Leica Microsystems, Inc., Milton Keynes, UK) and heated in the microwave at 800 W for 15 min. Endogenous peroxidase activity was blocked by incubation with 3% hydrogen peroxide for 10 min. Slides were incubated with mouse monoclonal TROP-2 (dilution, 1:400; cat. no. (B-9): sc-376746; Santa Cruz Biotechnology, Inc., Dallas, TX, USA) antibodies for 60 min at room temperature. After washing with phosphate buffered saline (PBS), the slides were incubated with biotinylated secondary antibodies (#BP-9100; undiluted; Vector Laboratories, Burlingame, CA, USA) for 20 min at 30°C, then washed with PBS for 5 min and incubated with the Peroxidase Detection system Ready to Use conjugated antibody (#RE7110K; Novocastra; Leica Microsystems, Inc.) for 20 min at room temperature. Subsequently, the sections were washed with PBS for 5 min, incubated with chromogenic 3,3'-diaminobenzidine (Leica Microsystems, Inc.) for 5 min, washed with tap water and counterstained with hematoxylin. The slides were then dehydrated in ascending grades of alcohol and stained with xylene.

Evaluation of immunohistochemical staining

Membranous staining of squamous cells of the normal skin was used as positive control. In all thyroid cases, membranous staining of the cells was accepted as positive. The proportion of positively stained cells was scored as follows; 0: negative or <5% positively stained cells, 1:6%-25% positively stained cells, 2:25%-50% positively stained cells, 3: 51%-74% positively stained cells, 4:>75% positively stained cells.[13] In the positive cases, 1+ or 2+ were accepted as focal staining pattern and diffuse staining pattern (3+ or 4+).[13]

Statistical analysis

The statistical analysis was performed using SPSS version 17 software (SPSS Inc., Chicago, U.S.A.). The Pearson Chi-square test or Fisher Exact test were used to assess the differences between groups. The sensitivity of TROP-2 was compared between the benign and malignant groups. A value of P < 0.05 was accepted as statistically significant.


   Results Top


The study cases comprised 79% (120/152) females and 21% (32/152) males with a median age of 46.8 years (range, 15-85 years). The TROP-2 expression values in the 152 thyroid lesions are shown in [Table 1]. TROP-2 expression was positive in 74.6% (56/75) of the malignant thyroid lesions and in 1.3% of benign thyroid lesions. There was a statistically significant difference (P < 0.001) between the benign and malignant lesions. Immunoreactivity was detected in only 1 of the 29 patients with FA. Hashimoto's thyroiditis was present as an additional thyroid pathological finding in 23 cases. In 5 cases of Hashimoto's thyroiditis, membranous staining was determined in oncocytic cells in and outside lymphoid aggregates [Figure 1].
Table 1: Trophoblast cell-surface antigen-2 expression in benign and malignant thyroid lesions

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Figure 1: (a and b) TROP-2 expression was not showed in follicular nodular disease (H and E, x40 and DAB, x100). (c and d) Focal membranous positivity in follicular adenoma, oncocytic variant (H and E, x40 and DAB, x400). (e and f) Hashimoto's thyroiditis was detected membranous staining in oncocytic cells in lymphoid aggregates (H and E, x40 and DAB, x400)

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TROP-2 expression was seen in 89.5% (51/57) of PTC and in 41.7% (5/12) of FC, while all MTC, PDTC and UDTC were not stained. When the PTC cases were compared according to the staining percentage of tumor cells in the follicular carcinoma cases, most of the cases showed a diffuse staining pattern [Figure 2].
Figure 2: (a and b) Follicular carcinoma showed capsule and vascular invasion (H and E, x40). (c and d) Focal positivity of TROP-2 in tumor cells (DAB, x100 and × 400)

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The TROP-2 expression distribution in the PTC cases in this study is shown in [Table 2]. All mPTC, 89.4% (17/19) of PTC cv, 81.8% (9/11) of PTC fv, and 66.6% (4/6) of PTC ov showed expression [Figure 3]. The percentage of staining of PTC fv showed more focal staining compared to other PTC.
Table 2: The expression of trophoblast cell-surface antigen-2 in subtypes of papillary thyroid carcinoma

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Figure 3: (a and b) Trop-2 expression in micropapillary (H and E, x40 and DAB, ×400), and oncocytic variant (c and d, H and E and DAB, ×400) of papillary thyroid carcinoma

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TROP-2 expression was also shown to differentiate the various follicular lesions of the thyroid [Table 3]. Immunoreactivity was detected in 3.4% (1/29) of FA, in 41.7% (5/12) of FC and in 81.8% (9/11) of PTC fv cases. The difference was found to be statistically significant between PTC fv and FA (P < 0.01), and between FA and FTC (P = 0.05). The difference between FTC and PTC fv was not statistically significant (P = 0.089).
Table 3: Trophoblast cell-surface antigen-2 expression in follicular lesions

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A review of literature and the data of the current study are presented in [Table 4].
Table 4: Sensitivity of trophoblast cell-surface antigen-2 expression in malignant thyroid neoplasm

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   Discussion Top


Thyroid carcinoma is the most common endocrine malignancy.[1] The distinction between benign and malignant lesions is very important because there are significant differences in prognostic and metastatic characteristics.[1],[16] Many studies have investigated immunohistochemistry as a complementary method in the diagnosis of thyroid tumours when the morphology is inadequate.[17],[18],[19],[20] Immunohistochemical markers such as HBME-1, Galectin 3, CK19, CİTED-1, CD44, CD56 have been found to be useful in distinguishing benign from malignant lesions.[16],[17],[18],[19],[20] However, immunoreactivity in benign follicular disease and in follicular adenoma restricts the use of the markers.[21],[22] Hürtle (oncocytic) cell is characterized by the cytoplasmatic accumulation of mitochondria, on electron microscope and has rich endogenous biotin. This cell can be seen in benign and malignant thyroid diseases, including Hashimoto's thyroiditis, oncocytic adenoma, oncocytic carcinoma or oncocytic variant of papillary thyroid carcinoma.[16],[23],[24] It may often display cytological atypia and abnormal staining with immunohistochemical markers. Cells may provide false positive results, especially when heat-induced antigen retrieval methods are used.[21] TROP-2 expression in benign thyroid lesions has been reported in only one study. However, Addati et al. indicated immunoreactivity in 2 FA and 4 non-neoplastic thyroid tissue samples.[11] In the current study, 4 of the 29 follicular adenomas had an oncocytic variant and only 1 was stained. However, Hashimoto's thyroiditis was detected as an additional pathological finding in 23 of the 152 cases and 5 of those showed membranous staining in oncocytic cells in and outside lymphoid aggregates.

In the current study, a statistically significant difference was determined between benign and malignant lesions and TROP-2 expression was most frequently observed in well-differentiated thyroid cancers of the malignant thyroid lesions. All MTC, PDTC and UDTC were negative for TROP-2. No report in literature has shown TROP-2 staining for MTC, whereas 2 PDTC and 1 UDTC were positive only in papillary areas in the study by Bychkov et al.[14] In the current study, TROP-2 expression tended to be focal staining in the FC cases compared to the PTC cases. In addition, TROP-2 expression was higher in the current study cases compared with previous findings in the literature. Bychkov et al. reported no positivity in 35 FTC cases in a study of fixed paraffin-embedded (FFPE) samples while two TMA-based studies showed positivity in 1 of 64 cases.[12],[13],[14] This difference may have been caused by the use of different clones or TMA samples.

In previous studies, the sensitivity of TROP-2 expression for PTC has been calculated to be between 50% and 90%.[11],[12],[13],[14],[15] In the current study, 89.5% sensitivity was determined for PTC. Moreover, sensitivity was 100% and 89.4% for mPTC and PTCcv cases, respectively, in accordance with the literature.[12],[13],[14],[15] In the present study, the sensitivity rate of PTC fv was high compared to the three studies in the literature which used the same antibody.[12],[13],[14] However, the study results of Adati et al. using different antibody from these studies was similar to the current study findings.[11] When TROP-2 expression was examined in terms of differentiation between benign and malignant follicular lesions, a statistically significant difference was determined between PTC fv and FA (P < 0.01), and between FA and FTC (P = 0.05), but no difference was found between FTC/PTC fv (P = 0.089).


   Conclusion Top


The results of this study demonstrated that TROP-2 has high specificity for the diagnosis of thyroid papillary carcinoma and especially for some subtypes (PTC cv, PTC fv and mPTC). TROP-2 can be considered a promising new marker that helps to differentiate between benign and malignant follicular lesions.

Financial support and sponsorship

This study was supported by specialize training council in medicine of Antalya Education and Research Hospital.

Conflicts of interest

There are no conflicts of interest.



 
   References Top

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DeLellis RA, Williams ED. Tumours of the thyroid and parathyroid. In: DeLellis RA, Lloyd RV, Heitz PU, Eng C, editors. Pathology and Genetics: Tumors of Endocrine Organs. WHO Classification of Tumors. 3rd ed. Lyon: IARC Press; 2004. p. 49-133.  Back to cited text no. 1
    
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Al Ghuzlan A, Caillou B, Schlumberger M. Galectin-3 for indeterminate thyroid cytology. Lancet Oncol 2008;9:508-10.  Back to cited text no. 2
    
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Arcolia V, Journe F, Renaud F, Leteurtre E, Gabius HJ, Remmelink M, et al. Combination of galectin-3, CK19 and HBME-1 immunostaining improves the diagnosis of thyroid cancer. Oncol Lett 2017;14:4183-9.  Back to cited text no. 3
    
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Goldenberg DM, Cardillo TM, Govindan SV, Rossi EA, Sharkey RM. Trop-2 is a novel target for solid cancer therapy with sacituzumab govitecan (IMMU-132), an antibody-drug conjugate (ADC). Oncotarget 2015;6:22496-512.  Back to cited text no. 4
    
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Lipinski M, Parks DR, Rouse RV, Herzenberg LA. Human trophoblast cell-surface antigens defined by monoclonal antibodies. Proc Natl Acad Sci U S A 1981;78:5147-50.  Back to cited text no. 5
    
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Kluger HM, Kluger Y, Gilmore-Hebert M, DiVito K, Chang JT, Rodov S, et al. CDNA microarray analysis of invasive and tumorigenic phenotypes in a breast cancer model. Lab Invest 2004;84:320-31.  Back to cited text no. 6
    
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Mühlmann G, Spizzo G, Gostner J, Zitt M, Maier H, Moser P, et al. TROP2 expression as prognostic marker for gastric carcinoma. J Clin Pathol 2009;62:152-8.  Back to cited text no. 7
    
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Fang YJ, Lu ZH, Wang GQ, Pan ZZ, Zhou ZW, Yun JP, et al. Elevated expressions of MMP7, TROP2, and survivin are associated with survival, disease recurrence, and liver metastasis of colon cancer. Int J Colorectal Dis 2009;24:875-84.  Back to cited text no. 8
    
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Bignotti E, Zanotti L, Calza S, Falchetti M, Lonardi S, Ravaggi A, et al. Trop-2 protein overexpression is an independent marker for predicting disease recurrence in endometrioid endometrial carcinoma. BMC Clin Pathol 2012;12:22.  Back to cited text no. 9
    
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Varughese J, Cocco E, Bellone S, Bellone M, Todeschini P, Carrara L, et al. High-grade, chemotherapy-resistant primary ovarian carcinoma cell lines overexpress human trophoblast cell-surface marker (Trop-2) and are highly sensitive to immunotherapy with hRS7, a humanized monoclonal anti-trop-2 antibody. Gynecol Oncol 2011;122:171-7.  Back to cited text no. 10
    
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Addati T, Achille G, Centrone M, Petroni S, Popescu O, Russo S, et al. TROP-2 expression in papillary thyroid cancer: A preliminary cyto-histological study. Cytopathology 2015;26:303-11.  Back to cited text no. 11
    
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Simms A, Jacob RP, Cohen C, Siddiqui MT. TROP-2 expression in papillary thyroid carcinoma: Potential diagnostic utility. Diagn Cytopathol 2016;44:26-31.  Back to cited text no. 12
    
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Liu H, Shi J, Lin F. The potential diagnostic utility of TROP-2 in thyroid neoplasms. Appl Immunohistochem Mol Morphol 2017;25:525-33.  Back to cited text no. 13
    
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Bychkov A, Sampatanukul P, Shuangshoti S, Keelawat S. TROP-2 immunohistochemistry: A highly accurate method in the differential diagnosis of papillary thyroid carcinoma. Pathology 2016;48:425-33.  Back to cited text no. 14
    
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Murtezaoglu AR, Gucer H. Diagnostic value of TROP-2 expression in papillary thyroid carcinoma and comparison with HBME-1, galectin-3 and cytokeratin 19. Pol J Pathol 2017;68:1-0.  Back to cited text no. 15
    
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Khan A, Nose V. Pathology of thyroid gland. In: Lloyd RV, editor. Endocrine Pathology: Differential Diagnosis and Molecular Advances. 2nd ed. New York: Springer Publishers; 2010. p. 181-235.  Back to cited text no. 16
    
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Liu Z, Yu P, Xiong Y, Zeng W, Li X, Maiaiti Y, et al. Significance of CK19, TPO, and HBME-1 expression for diagnosis of papillary thyroid carcinoma. Int J Clin Exp Med 2015;8:4369-74.  Back to cited text no. 18
    
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Palo S, Biligi DS. Differential diagnostic significance of HBME-1, CK19 and S100 in various thyroid lesions. Malays J Pathol 2017;39:55-67.  Back to cited text no. 19
    
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Dunđerović D, Lipkovski JM, Boričic I, Soldatović I, Božic V, Cvejić D, et al. Defining the value of CD56, CK19, galectin 3 and HBME-1 in diagnosis of follicular cell derived lesions of thyroid with systematic review of literature. Diagn Pathol 2015;10:196.  Back to cited text no. 20
    
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Volante M, Bozzalla-Cassione F, DePompa R, Saggiorato E, Bartolazzi A, Orlandi F, et al. Galectin-3 and HBME-1 expression in oncocytic cell tumors of the thyroid. Virchows Arch 2004;445:183-8.  Back to cited text no. 21
    
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Ma H, Yan J, Zhang C, Qin S, Qin L, Liu L, et al. Expression of papillary thyroid carcinoma-associated molecular markers and their significance in follicular epithelial dysplasia with papillary thyroid carcinoma-like nuclear alterations in Hashimoto's thyroiditis. Int J Clin Exp Pathol 2014;7:7999-8007.  Back to cited text no. 22
    
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Yang GC, Schreiner AM, Sun W. Can abundant colloid exclude oncocytic (Hürthle cell) carcinoma in thyroid fine needle aspiration? Cytohistological correlation of 127 oncocytic (Hürthle cell) lesions. Cytopathology 2013;24:185-93.  Back to cited text no. 23
    
24.
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Correspondence Address:
Canan Sadullahoğlu
Department of Pathology, Health Sciences University, Antalya Education and Research Hospital, Antalya 07050
Turkey
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/IJPM.IJPM_202_18

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