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Variable expression of keratins and nearly uniform lack of thyroid transcription factor 1 in thyroid anaplastic carcinoma
Variable Expression of Keratins and Nearly Uniform Lack of Thyroid Transcription Factor 1 in Thyroid Anaplastic Carcinoma MARKKU MIETTINEN, MD, AND KAARLE O. FRANSSILA, MD Thyroid anaplastic (undifferentiated) carcinomas (TACs) comprise a morphologically heterogeneous group of tumors, which can arise in the background of differentiated papillary or follicular carcinoma. The thyroid epithelial differentiation varies in these tumors and has not been completely characterized. In this study, we immunohistochemically analyzed different variants TACs from 35 patients by using antibodies specific to 9 different keratin polypeptides, epithelial membrane antigen, thyroid transcription factor I (TTF-1), and thyroglobulin. These tumors were histologically divided into 3 categories: squamoid-cohesive (SC, 13 tumors), spindle cell sarcomatous (SS, 8 cases) and intermediate group, including tumors with giant cells and solid epithelioid components (GC, 18 tumors); 4 tumors had 2 components. The patients ages ranged from 40 to 89 years, with a mean age in all groups of 70 years. TTF-1 was present in only 2 of 9 of the SC tumors, and absent in all other TACs, but was present in entrapped differentiated components. Thyroglobulin was absent in all but 1 case. A complex keratin (K) pattern of stratified epithelia was typically seen in the SC tumors with extensive K7, K8, K17, K18, and K19, and variable K13 and K14 expression; EMA was also present.
K16 was limited to squamous pearls in 1 tumor, and K10 was absent. The GC carcinomas typically had K8 and K18, whereas the expression of K7 was variable and that of K14, K17, and K19 sporadic; EMA was variably present in half of the cases. The keratins in spindle cell sarcomatous tumors were usually limited to K7, K8, and K18, often in limited numbers of cells. EMA was present in 1 case only. These results indicate a complex pattern of keratins in squamoid and giant cell TACs, similar to papillary carcinoma and suggesting the possibility of relationship. There was a progressive loss of epithelial differentiation and keratins in sarcomatoid TACs. Loss of TTF-1 is a nearly uniform feature of TAC and disallows the use of this marker to pinpoint a thyroid origin of these tumors. HUM PATHOL 31:1139-1145. Copyright © 2000 by W.B. Saunders Company Key words: thyroid transcription factor, keratins, EMA immunohistochemistry. Abbreviations: TAC, thyroid anaplastic (undifferentiated) carcinoma; PC, papillary carcinoma; FC, follicular carcinoma; TTF-1, thyroid transcription factor 1; SC, squamoid cohesive; GC, giant cell.
Thyroid anaplastic (undifferentiated) carcinoma (TAC) is a designation for a group of thyroid carcinomas that do not show papillary, follicular, or neuroendocrine differentiation. These tumors constitute only approximately 5% of all thyroid carcinomas. They are typically aggressively invasive, often metastasizing, and are usually associated with a short patient survival. Histologically, TACs may show squamoid, giant cell, and sarcomatoid spindle cell patterns.1-7 Previous studies have shown that TACs are generally positive for keratins, but occasional tumors have been keratin-negative.8-13 However, 1 study showed 13 of 29 TACs to be keratin-negative, even if 9 different antibodies were used.14 Papillary (PC) and follicular carcinomas (FC) have been extensively characterized for their composition in of keratin polypeptides of the Moll catalog.9,14-17 They both express simple epithelial keratins K7, K8, K18, and commonly K19. Complex epithelial keratins, including K5/K14 and K17, have
been found in PC but generally not in FC.16-18 However, the keratin composition of TACs has not been similarly characterized with a similar panel and could give a clue of the nature and interelationships of these tumors. Thyroid differentiation has been previously extensively analyzed by immunohistochemical demonstration thyroglobulin, a thyroid-specific thyroid hormone– binding colloid protein generally not found in TACs.2 Thyroid transcription factor I (TTF-1) is a new marker for thyroid differentiation. TTF-1 is a DNA-binding transcriptional regulator protein localized in the nuclei of thyroid follicular cells, subsets of respiratory and alveolar epithelium and diencephalon. It also has been shown to be present in the corresponding tumors, including thyroid carcinomas and subsets of pulmonary carcinomas, especially adenocarcinomas and small cell carcinomas.19-23 Although the differentiated thyroid carcinomas have been found to be TTF-1 positive, results of only 3 thyroid anaplastic carcinomas have been published; all were negative.20 In this study, we analyzed the epithelial and thyroid differentiation in TACs from 35 patients by using a panel of antibodies that recognize 9 individual keratin polypeptides in the Moll catalog, including keratins of both simple and stratified epithelia. TTF was generally absent. Although the squamoid TACs show a complex keratin pattern similar to that of papillary carcinoma, the sarcomatoid TACs showed lessened keratin expression. These findings suggest some similarities between squamoid TACs and papillary carcinomas and show progressive loss of thyroid and epithelial differentiation in the sarcomatoid variants.
From the Department of Soft Tissue Pathology, Armed Forces Institute of Pathology, Washington, DC; and the Department of Pathology, Helsinki University Central Hospital, Helsinki, Finland. Accepted for publication June 13, 2000. The opinions and assertions contained herein are the expressed views of the authors and are not to be construed as official or reflecting the views of the Departments of the Army or Defense. Address correspondence and reprint requests to Markku Miettinen, MD, Department of Soft Tissue Pathology, Armed Forces Institute of Pathology, 14th Street and Alaska Ave, NW, Washington, DC 20306-6000. Copyright © 2000 by W.B. Saunders Company 0046-8177/00/3109-0023$10.00/0 doi: 10.1053/hupa.2000.16667
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MATERIALS AND METHODS Tumors
Squamoid Cohesive TACs
Thyroid anaplastic (undifferentiated) carcinomas (TACs) of 35 patients were obtained from the files of the Helsinki University Hospital. The clinical features of these patients have been previously published.24 All patients included in this study died of disease. Hematoxylin and eosin–stained slides of all tumors were histologically examined, including careful evaluation for components of preexisting differentiated papillary or follicular carcinoma. All preexisting differentiated tumors with ground-glass type nuclei were considered papillary irrespective of the architectural pattern. Ten papillary and 5 follicular carcinomas were studied for TTF-1 for comparative purposes. All tumors were negative for synaptophysin and chromogranin in immunohistochemical evaluation before being entered in the study, and none of the patients had evidence for medullary carcinoma or multiple endocrine neoplasia. The diagnosis of lymphoma was excluded in all cases (tumors negative for CD20 and CD45 [leukocyte common antigen]). All tumors were primary in the thyroid, and there was no clinical evidence of other primary tumors.
Immunohistochemistry The antibodies and their specificities, dilutions, and sources are listed in Table 1. All antibodies have been previously characterized.25-33 The immunostains were performed by using avidin-biotin detection system with diaminobenzidine as the chromogen. Avidin-biotin block (Dako, Carpinteria, CA) was used before the application of primary antibody. Negative and positive controls were used in each run.
RESULTS Anaplastic (undifferentiated) thyroid carcinomas from 35 patients were studied. There were 25 women and 10 men of median age 70 years (range, 40 to 89; mean, 70 years). These tumors were histologically classified into 3 categories: squamoid cohesive (SC), giant cell (GC), and spindle cell sarcomatoid. Four patients had both SC and GC components, which were scored separately for immunoreactivity. The immunohistochemical results have been summarized in Table 2.
TABLE 1. Polypeptide
There were 9 pure squamoid carcinomas and additional squamoid components in four tumors, all of which had a major large giant cell component. These tumors occurred in 7 women and 6 men with a median age of 70 years (mean, 71 years; range, 55 to 89 years). One of these tumors had elements of preexisting follicular carcinoma. Histologically, these tumors showed cohesive clusters of large, polygonal epithelial cells with ample, variably eosinophilic cytoplasm with various degrees of squamoid features. In some cases, the tumor cells were separated by fibrous septa, whereas in others they formed solid sheets with minimal stroma (Fig 1A, C). Extensive keratin expression was detected in most cases. Nearly all of them showed K7, K8, K18, and K19 in most tumor cells. Epithelial membrane antigen was detected in all but 1 case, although it often was limited to small foci of tumor cells. Keratins of complex epithelia were expressed variably. K14 was detected in all cases, although the proportion of positive cells varied between 1% and 100% (median, 15%) (Fig 1B). Most cases showed at least focal reactivity for K17, although this was only rarely extensive (Fig 1D). Focal positivity for K13 was common (Fig 1E), and reactivity for K16 was seen in 1 case strictly limited to keratin pearls (Fig 1F). K10 was not detected. All but 2 tumors were negative for TTF-1, whereas positive normal or differentiated carcinomatous elements were seen in all but 2 cases. One positive tumor showed distinct nuclear staining in most of the squamoid epithelial cells (Fig 1G, H), and another showed a 10% TTF-1–positive population. Only the latter case among this group was positive for thyroglobulin; all other positive elements in other tumors represented either entrapped normal thyroid or preexisting differentiated elements. In contrast, all 10 papillary and 5 follicular carcinomas had nuclear positivity for TTF-1.
Monoclonal Antibodies, Their Sources and Dilutions, and the Pretreatment Modalities Used in This Study Clone
Pretreatment
Antibody Dilution
Source
Reference
Keratin 7 Keratin 8
OV-TL 12/30 Cam5.2
PrVIII* Pepsin†
1:50 1:40
33 28
Keratin Keratin Keratin Keratin Keratin Keratin Keratin TTF-1 EMA
LHP1 KS-1A3 LL002 LL025 E3 DC-10 RCK 108 8G7G3/1 E29
MW‡ MW‡ MW‡ MW‡ MW‡ MW‡ MW‡ PrVIII1 MW‡
1:50 1:100 1:100 1:40 1:40 1:40 1:50 1:50 1:50
Dako Beckton-Dickinson, Mountain view, CA Novocastra Novocastra Novocastra Novocastra Novocastra Novocastra Dako, Carpinteria, CA Biocare, Walnut Creek, CA Dako
10 13 14 16 17 18 19
29 32 31 30 26 25 27
* PrVIII ⫽ 0.05% Sigma protease type VIII, 3 minutes in 0.1 mol/L phosphate buffer, pH 7.8 at 37°C. † Pepsin ⫽ crude pepsin (0.05% in HCl, pH 2.0) for 30 min at 37°C. ‡ MW ⫽ microwawe heating adjusted to near to boiling in EDTA-buffer 20 minutes at pH 6.0, followed by a 20-minute cooling period.
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TABLE 2. Summary of the Expression of Seven Keratin Polypeptides and Epithelial Membrane Antigens in Variants of Thyroid Anaplastic Carcinomas Squamoid (n ⫽ 13) Keratin 7 ⬎10 of tumor cells ⬍10% of tumor cells Negative Keratin 8 ⬎30 of tumor cells ⬍30% of tumor cells Negative Keratin 18 ⬎30 of tumor cells ⬍30% of tumor cells Negative Keratin 19 ⬎30 of tumor cells ⬍30% of tumor cells Negative Keratin 17 ⬎30 of tumor cells ⬍30% of tumor cells Negative Keratin 13 ⬎30 of tumor cells ⬍30% of tumor cells Negative Keratin 14 ⬎30 of tumor cells ⬍30% of tumor cells Negative Epithelial membrane antigen ⬎30 of tumor cells ⬍30% of tumor cells Negative
Giant Cell/Solid Spindle Cell Epithelioid Sarcomatoid (n ⫽ 16) (n ⫽ 7)
12 1 0
9 4 5
1 4 3
10 3 0
10 2 6
1 5 2
12 0 1
12 2 4
3 1 4
10 2 1
4 3 10
0 2 5
4 5 4
1 3 15
0 2 6
4 4 2
0 3 15
0 0 7
6 5 0
1 6 10
0 1 6
6 5 1
3 3 10
0 1 6
cells in only 3 cases and focally in 4 additional cases. Expression of complex epithelial keratins was sporadic. K14, K17, and K13 were seen in 7, 4, and 3 cases, respectively. Only 1 of these had more than focal positivity for K14 and K17 (30% to 50% of tumor cells); all others had only scattered positive cells. K10 and K16 were not detected. All tumors were negative for TTF-1 and thyroglobulin; 7 cases had TTF-1–positive differentiated components in which the nuclei were brightly positive (Fig 2F). TAC With Spindle Cell Sarcomatoid Features All 8 tumors occurred in women of median age of 63 years (range, 40 to 86 years). No elements of differentiated thyroid carcinoma were found in these tumors. This group of tumors were composed of undifferentiated spindle cells in a dense, sclerosing, collagenrich background (Fig 3A), except for 1 tumor that had a matrix-poor, noncollagenous background. Keratin-positive cells were detected most often with the antibody to K8 (6 of 8), whereas K7-, K18-, and K19-positive cells were detected in 5, 4, and 2 cases, respectively. Although many cases showed limited numbers of keratin-positive cells, in 3 of 8 cases most tumor cells were positive for K18, but only 1 of these cases showed significant numbers of K7- and K8-positive cells (Fig 3B). None of these tumors showed K19-positivity in more than 5% of tumor cells. One tumor was negative with all keratin antibodies. The complex epithelial keratins were seen only occasionally. Two tumors both scattered K14 and K17 positive cells. K10, K13, and K16 were not detected. All cases were negative for TTF-1 and thyroglobulin. EMA-positive cells were present in 1 case only (focally).
TACs With Giant Cell/Solid Epithelioid Patterns These 18 tumors, of which 4 tumors also had small squamoid cohesive foci, occurred in 12 women and 6 men of median age 70 years (mean, 71 years; range, 53 to 89 years). The keratins of the squamoid foci were evaluated with the squamoid tumors (see above). Six of these tumors arose in the background of papillary carcinoma, and 1 with a follicular carcinoma. The tumors showed features intermediate between squamoid epithelial and spindle cell sarcomatous patterns. They were composed of epithelioid cells with variable cellular cohesion ranging from dyscohesive pattern to solid sheets of tumor cells. Large cytoplasmic giant cells containing usually 1 irregular, sometimes lobated, nucleus were often present (Fig 2A, C). The amount of extracellular matrix was sparse. All but 2 cases (16 of 18) showed positive cells at least with 1 of the keratin antibodies. Most cases showed extensive reactivity for K7, K8, and K18 (Fig 2B). K18 was the most abundant keratin and was detected in 12 of 18 cases in over 20% of tumor cells, and focally in 2 additional cases (Fig 2D). K19 was usually absent or scant (Fig 2E); it was seen in more than 20% of tumor
DISCUSSION Thyroid anaplastic (undifferentiated) carcinomas (TACs) constitute a morphologically heterogeneous group, of which three different patterns are generally recognized: squamoid, giant cell, and sarcomatoid.2-4 In this study, we examined TACs of different types from 35 patients for epithelial and thyroid differentiation by using a panel of antibodies specific to individual keratins, epithelial membrane antigen, thyroglobulin, and thyroid transcription factor I. We divided the TACs in 3 groups, to better examine the spectrum of differentiation in these tumors. Thyroid transcription factor 1 (TTF-1) is a DNAbinding transcriptional activator protein that regulates the expression of certain key thyroid specific genes, for example, that of thyroglobulin.19 TTF-1 also is expressed in bronchial epithelia and has been shown to be present in differentiated but not anaplastic thyroid carcinoma, subsets of pulmonary adenocarcinomas, and pulmonary small cell carcinomas.22-24 We found
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FIGURE 1. Patterns of keratins and TTF-1 in squamoid anaplastic thyroid carcinomas. (A) This anaplastic carcinoma has cohesive sheets of squamoid cells in a collageneous stroma. (B) The tumor cells are strongly and uniformly positive for keratin 14. (C) This anaplastic carcinoma is composed of solid sheets of squamoid epithelial cells. (D) The tumor cells are extensively positive for keratin 17. (E) Focal keratin 13 in a squamoid anaplastic carcinoma with dyscohesive tumor cells. (F) Only a squamous epithelial keratin pearl is positive in this anaplastic carcinoma. (G) A squamoid anaplastic carcinoma with necrosis shows nuclear TTF-1 positivity (H).
consistent nuclear TTF-1 expression in the normal thyroid, in the differentiated carcinoma components, and in papillary and follicular carcinomas studied as controls. However, with few exceptions, TTF-1 was absent in TAC cells. Therefore, TTF-1 cannot be used as a marker for thyroid origin of TACs. The 2 positive tumors were both squamoid variants of TAC, neither of them showing differentiated carcinoma components. TTF-1 therefore may have a limited usefulness in the identification of thyroid origin of the squamoid variant of TAC. The general absence of thyroglobulin in the anaplastic components paralleled the absence of TTF-1, consistent with the concept that TTF-1 regulates thyro-
globulin expression.19 However, thyroglobulin was present in the preexisting differentiated elements and in entrapped thyroid. Previous series have shown thyroglobulin expression in tumors cells to be exceptional in TACs.3,12 There are 20 different epithelial (cyto)keratins (excluding the hair and nail keratins) belonging to 2 isoelectric groups according to the Moll catalog15,34: I (A, acidic comprising K9 to K20) and II (B, basic comprising K1 to K8). Two of them (K3 of group II and K12 of group I) are specific for cornea, and the other keratins can be roughly divided into 2 functional groups: keratins of simple, nonstratified epithelia and keratins of complex epithelia. The
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FIGURE 2. Patterns of keratins and TTF-1 in giant cell/solid epithelioid anaplastic carcinoma. (A) An anaplastic giant cell/solid epithelioid carcinoma with pleomorphic epithelioid cells. The tumor cells extensively positive for keratin 7 (B). (C) An anaplastic giant cell/solid epithelioid carcinoma with islands of preexisting papillary carcinoma with uniform presence of keratin 18 (D), complete loss of keratin 19 (E) and TTF-1 (F); the latter 2 markers are present only in the papillary carcinoma components.
former, K7, K8, K18, and K19, including the lowest molecular weight keratins in groups I and II, are generally typical of glandular and other simple, nonstratified epithelia. Of the complex epithelial keratins, the high-molecular-weight keratins K1, K2, K9, K10, and K11 are generally restricted to keratinizing squamous epithelia, such as epidermis, whereas K4 and K13 are present in internal squamous epithelia and urothelia. K5 and K14 and K17, in turn, are present in many basal cells such as myoepithelia and some basal cells of squamous epithelia and skin adnexa.15,35-38
The squamoid TACs showed extensive keratin expression with a complex pattern somewhat paralleling the complex pattern of keratins in squamous epithelia. These tumors also prominently expressed all simple epithelial keratins, of which K7, K8, and K18 are often absent in well-differentiated squamous cell carcinomas, although they become expressed in poorly differentiated squamous cell carcinomas. In their keratin pattern, SCs resemble papillary thyroid carcinomas by often having similar complexity in the keratin profile with coexpression of simple epithelial keratins with K5/14 and K17.17,18 This finding is consistent with the fact that
FIGURE 3. Sarcomatoid anaplastic carcinoma shows spindled tumor cells in a collagenous stroma (A). The tumor cells are positive for keratin 8.
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some of these tumors may have originated from papillary carcinoma. Relative, partial loss of simple epithelial keratins and nearly total loss of complex epithelial keratins K13, K14, and K17 was typically observed in the TACs that were composed of sheets or poorly cohesive poorly differentiated epithelioid cells or giant cells; these tumors represented the largest group of TACs and were intermediate between the squamoid and spindle cell sarcomatous TACs. Most tumors of this group contained simple epithelial keratin–positive cells and therefore could be verified as epithelial. However, there was a remarkable loss of K19 in this group of tumors, compared with the possible squamoid component or preexisting papillary or follicular elements. Nevertheless, presence of complex epithelial keratins, although to lesser degree, could suggest relationship with papillary carcinoma, which, in fact, was verified in several cases. Loss of keratins in carcinomas is rare but has been described in vitro with hepatocellular carcinoma cells that lost their keratin expression on dedifferentiation in culture.39 Posttranslational downregulation also has been shown as a possible mechanism that lessens the keratin expression in TACs.40 It has been shown in oral squamous carcinomas that downregulation of K19 is associated with an increased invasive potential.41 Also, the expression of stratified epithelial keratins has been shown to reflect increased aggressiveness among ductal carcinomas of the breast.42 Both of these phenomenona may occur in TACs that are typically highly aggressive tumors. The sarcomatoid TACs that are composed of spindle cells in a sclerotic, collagenous background typically show extensive loss of keratins. In these tumors, keratins are often limited to simple epithelial keratins, typically in only scattered cells. However, the keratin expression K7, K8, K18, and rarely K19 still supports epithelial origin of these tumors. We believe that by overall similarity, the totally keratinnegative tumors are also carcinomas rather than sarcomas, although no epithelial markers can be demonstrated. In summary, we have evaluated 35 anaplastic thyroid carcinomas for epithelial markers and TTF-1 to better understand the cell differentiation in these tumors. Our results show nearly complete absence of TTF-1 in TACs, indicating that this marker is not generally useful in establishing thyroid origin for these tumor. The complex keratin patterns seen in squamoid, and to lesser degree giant cell epithelioid tumors, is somewhat similar to the keratin patterns seen in papillary carcinoma, suggesting a possible histogenetic relationship. Keratin reactivity (especially for simple epithelial keratins) can be shown at least focally in most tumors, supporting their epithelial origin. REFERENCES 1. Franssila K: Value of histologic classification of thyroid cancer. Acta Pathol Microbiol Scand 225:5-76, 1971 (suppl)
2. Nishiyama R, Dunn EL, Thompson NW: Anaplastic spindlecell and giant-cell tumors of the thyroid. Cancer 30:113-127, 1972 3. Carangiu ML, Steeper T, Zampi G, et al: Anaplastic thyroid carcinoma: A study of 70 cases. Am J Clin Pathol 83:135-158, 1985 4. Rosai J, Saxe´n EA, Woolner L: Undifferentiated and poorly differentiated carcinoma. Semin Diagn Pathol 2:123-136, 1985 5. Shvero J, Gal R, Avidor I, et al: Anaplastic thyroid carcinoma: A clinical, histologic, and immunohistochemical study. Cancer 62: 319-325, 1988 6. Venkatesh YSS, Ordonez NG, Schultz PN, et al: Anaplastic carcinoma of the thyroid: A clinicopathologic study of 121 cases. Cancer 66:321-330, 1990 7. Lampertico P: Anaplastic (sarcomatoid) carcinoma of the thyroid gland. Semin Diagn Pathol 10:159-168, 1993 8. Miettinen M, Franssila K, Lehto VP, et al: Expression of intermediate filament proteins in thyroid gland and in thyroid tumors. Lab Invest 50:262-270, 1984 9. Dockhorn-Dworniczak B, Franke WW, Czernobilsky B, et al: Patterns of expression of cytoskeletal proteins in human thyroid gland and thyroid carcinomas. Differentiation 35:53-71, 1987 10. Henzen-Longmans SC, Mullink H, Ramaekers FC, et al: Expression of cytokeratins and vimentin in epithelial cells of normal and pathologic thyroid tissue. Virchows Arch A Pathol Anat Histopathol 410:347-354, 1987 11. Hurlimann J, Gardiol D, Scazziga B: Immunohistology of anaplastic thyroid carcinoma: A study of 43 cases. Histopathology 11:567-580, 1987 12. Livolsi VA, Brooks JJ, Arendash-Durand B: Anaplastic thyroid tumors: Immunohistology. Am J Clin Pathol 87:434-442, 1987 13. Ordonez NG, El-Naggar AK, Hickey RC, et al: Anaplastic thyroid carcinoma: Immunocytochemical study of 32 cases. Am J Clin Pathol 96:15-24, 1991 14. Schro¨der S, Wodzynski A, Padberg B: Zytokeratinexpression benigner und maligner epithelialer Schilddrusentumoren. Eine immunohistologische Unterschung an 154 Neoplasien inter Einsatz von8 verschiedene monoklonalen Zytokeratinantikorpern. Pathologe 17:425-432, 1996 15. Moll R, Franke WW, Schiller DL, et al: The catalog of human cytokeratins: Patterns of expression in normal epithelia, tumors and cultured cells. Cell 31:11-24, 1982 16. Fonseca E, Nesland JM, Ho¨ie J, et al: Patterns of expression of intermediate cytokeratin filaments in the thyroid gland: An immunohistochemical study of simple and stratified epithelial-type cytokeratins. Virchows Arch 430:239-246, 1997 17. Miettinen M, Kovatich AJ, Ka¨rkka¨inen P: Patterns of keratin polypeptides in papillary and follicular thyroid lesions. Virchows Arch 431:407-413, 1997 18. Baloch ZW, Abraham S, Roberts S, et al: Differential expression of cytokeratins in follicular variant of papillary carcinoma: an immunohistochemical study and its diagnostic utility. HUM PATHOL 30:1166-1171, 1999 19. Bingle CD: Thyroid transcription factor-1. Int J Biochem Cell Biol 29:1471-1473, 1997 20. Fabbro G, Di Loreto C, Beltrami CA, et al: Expression of thyroid-specific transcription factors TTF-1 and PAX-8 in human thyroid neoplasms. Cancer Res 54:4744-4749, 1994 21. Holzinger A, Dingle S, Bejarano PA, et al: Monoclonal antibody to thyroid transcription factor-1: Production, characterization, and usefulness in tumor diagnosis. Hybridoma 15:49-53, 1996 22. Bejarano PA, Baughman RP, Biddinger PW, et al: Surfactant proteins and thyroid transcription factor-1 in pulmonary and breast cazrcinomas. Mod Pathol 9:445-452, 1996 23. Folpe AL, Gown AM, Lamps LW, et al: Thyroid transcription fator-1: Immunohistochemical evaluation in pulmonary neuroendocrine tumors. Mod Pathol 12:5-8, 1999 24. Voutilainen PE, Multanen M, Haapiainen RK, et al: Anaplastic thyroid carcinoma survival. World J Surg 23:975-978, 1999 25. Bartek J, Vojtesek B, Staskova Z, et al: A series of 14 new monoclonal monoclonal antibodies to keratins: Characterizations and value in diagnostic histopathology. J Pathol 164:215-224, 1991 26. Guelstein VI, Tchypysheva TA, Ermilova VD, et al: Monoclonal antibody mapping of keratins 8 and 17 and of vimentin in normal human mammary gland, benign tumors, dysplasias and breast cancer. Int J Cancer 42:147-153, 1988
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THYROID UNDIFFERENTIATED CARCINOMA (Miettinen & Franssila) 27. Klianieko J, el-Naggar A, DeBraud F: Keratins 6, 13 and 19 (1993) Differential expression in squamous cell carcinoma of the head and neck. Anal Quant Cytol Histol 15:335-340, 1993 28. Kuruc N, Franke WW: Transient coexpression of desmin and cytokeratins 8 and 18 in developing myocardial cells of some vertebrate species. Differentiation 38:177-193, 1986 29. Leigh IM, Purkis PE, Whitehead P, et al: Monospecific monoclonal antibodies to keratin 1 carboxy terminal (synthetic polypeptide) and to keratin 10 a markers of epidermal differentiation. Br J Dermatol 129:110-119, 1993 30. Markey AC, Lane EB, Churchill LJ, et al: Expression of simple epithelial keratins 8 and 18 in epidermal neoplasia. J Invest Dermatol 97:763-770, 1991 31. Purkis PE, Steel JB, Mackenzie IC, et al: Antibody markers of basal cells in complex epithelia. J Cell Sci 97:39-50, 1990 32. van Muijen GNP, Ruiter DJ, Franke WW: Cell-type heterogeneity of cytokeratin expression in complex epithelia and carcinomas as demonstrated by monoclonal antibodies specific for cytokeratins 4 and 13. Exp Cell Res 162:97-113, 1986 33. van Niekerk CC, Jap PHK, Ramaekers FCS, et al: Immunohistochemical demonstration of keratin 7 in routinely fixed paraffinembedded human tissues. J Pathol 165:145-152, 1991 34. Sun T-T, Eichner R, Shcermer A, et al: Classification, expression and possible mechanisms of evolution of mammalian epithelial keratins: A unifying model, in Levine AJ, van de Voude
GF, Topp WC, et al (eds): Cancer Cell I/The Transformed Phenotype. Cold Spring Harbor, NY, Cold Spring Harbor Laboratory, 1985, pp 169-176 35. Nagle RB: Intermediate filaments: A review of the basic biology. Am J Surg Pathol 12:4-16, 1988 (suppl 1) 36. Miettinen M: Keratin immunohistochemistry: Update on applications and pitfalls. Pathol Annu 24:133-145, 1993 37. Moll R: Cytokeratins in the histological diagnosis of malignant tumors. Int J Biol Markers 9:63-69, 1994 38. Moll R, Krepler R, Franke WW: Complex cytokeratin polypeptide patterns observed in certain human carcinomas. Differentiation 23:256-269, 1983 39. Venetianer A, Schiller DL, Magin T, et al: Cessation of cytokeratin expression in a rat hepatoma cell line lacking differentiated functions. Nature 305:730-733, 1983 40. Paine ML, Gibbins JR, Chew KE, et al: Loss of keratin expression in anaplastic carcinoma cells due to posttranscriptional downregulation acting in trans. Cancer Res 52:6603-6611, 1992 41. Crowe DL, Milo GE, Shuler CF: Keratin 19 downregulation by oral squamous cell carcinoma lines increases invasive potential. J Dent Res 78:1256-1263, 1999 42. Malzahn K, Mitze M, Thoenes M, et al: Biological and prognostic significance of stratified epithelial cytokeratins in infiltrating ductal breast carcinomas. Virchows Arch 433:119-129, 1998
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K16 was limited to squamous pearls in 1 tumor, and K10 was absent. The GC carcinomas typically had K8 and K18, whereas the expression of K7 was variable and that of K14, K17, and K19 sporadic; EMA was variably present in half of the cases. The keratins in spindle cell sarcomatous tumors were usually limited to K7, K8, and K18, often in limited numbers of cells. EMA was present in 1 case only. These results indicate a complex pattern of keratins in squamoid and giant cell TACs, similar to papillary carcinoma and suggesting the possibility of relationship. There was a progressive loss of epithelial differentiation and keratins in sarcomatoid TACs. Loss of TTF-1 is a nearly uniform feature of TAC and disallows the use of this marker to pinpoint a thyroid origin of these tumors. HUM PATHOL 31:1139-1145. Copyright © 2000 by W.B. Saunders Company Key words: thyroid transcription factor, keratins, EMA immunohistochemistry. Abbreviations: TAC, thyroid anaplastic (undifferentiated) carcinoma; PC, papillary carcinoma; FC, follicular carcinoma; TTF-1, thyroid transcription factor 1; SC, squamoid cohesive; GC, giant cell.
Thyroid anaplastic (undifferentiated) carcinoma (TAC) is a designation for a group of thyroid carcinomas that do not show papillary, follicular, or neuroendocrine differentiation. These tumors constitute only approximately 5% of all thyroid carcinomas. They are typically aggressively invasive, often metastasizing, and are usually associated with a short patient survival. Histologically, TACs may show squamoid, giant cell, and sarcomatoid spindle cell patterns.1-7 Previous studies have shown that TACs are generally positive for keratins, but occasional tumors have been keratin-negative.8-13 However, 1 study showed 13 of 29 TACs to be keratin-negative, even if 9 different antibodies were used.14 Papillary (PC) and follicular carcinomas (FC) have been extensively characterized for their composition in of keratin polypeptides of the Moll catalog.9,14-17 They both express simple epithelial keratins K7, K8, K18, and commonly K19. Complex epithelial keratins, including K5/K14 and K17, have
been found in PC but generally not in FC.16-18 However, the keratin composition of TACs has not been similarly characterized with a similar panel and could give a clue of the nature and interelationships of these tumors. Thyroid differentiation has been previously extensively analyzed by immunohistochemical demonstration thyroglobulin, a thyroid-specific thyroid hormone– binding colloid protein generally not found in TACs.2 Thyroid transcription factor I (TTF-1) is a new marker for thyroid differentiation. TTF-1 is a DNA-binding transcriptional regulator protein localized in the nuclei of thyroid follicular cells, subsets of respiratory and alveolar epithelium and diencephalon. It also has been shown to be present in the corresponding tumors, including thyroid carcinomas and subsets of pulmonary carcinomas, especially adenocarcinomas and small cell carcinomas.19-23 Although the differentiated thyroid carcinomas have been found to be TTF-1 positive, results of only 3 thyroid anaplastic carcinomas have been published; all were negative.20 In this study, we analyzed the epithelial and thyroid differentiation in TACs from 35 patients by using a panel of antibodies that recognize 9 individual keratin polypeptides in the Moll catalog, including keratins of both simple and stratified epithelia. TTF was generally absent. Although the squamoid TACs show a complex keratin pattern similar to that of papillary carcinoma, the sarcomatoid TACs showed lessened keratin expression. These findings suggest some similarities between squamoid TACs and papillary carcinomas and show progressive loss of thyroid and epithelial differentiation in the sarcomatoid variants.
From the Department of Soft Tissue Pathology, Armed Forces Institute of Pathology, Washington, DC; and the Department of Pathology, Helsinki University Central Hospital, Helsinki, Finland. Accepted for publication June 13, 2000. The opinions and assertions contained herein are the expressed views of the authors and are not to be construed as official or reflecting the views of the Departments of the Army or Defense. Address correspondence and reprint requests to Markku Miettinen, MD, Department of Soft Tissue Pathology, Armed Forces Institute of Pathology, 14th Street and Alaska Ave, NW, Washington, DC 20306-6000. Copyright © 2000 by W.B. Saunders Company 0046-8177/00/3109-0023$10.00/0 doi: 10.1053/hupa.2000.16667
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MATERIALS AND METHODS Tumors
Squamoid Cohesive TACs
Thyroid anaplastic (undifferentiated) carcinomas (TACs) of 35 patients were obtained from the files of the Helsinki University Hospital. The clinical features of these patients have been previously published.24 All patients included in this study died of disease. Hematoxylin and eosin–stained slides of all tumors were histologically examined, including careful evaluation for components of preexisting differentiated papillary or follicular carcinoma. All preexisting differentiated tumors with ground-glass type nuclei were considered papillary irrespective of the architectural pattern. Ten papillary and 5 follicular carcinomas were studied for TTF-1 for comparative purposes. All tumors were negative for synaptophysin and chromogranin in immunohistochemical evaluation before being entered in the study, and none of the patients had evidence for medullary carcinoma or multiple endocrine neoplasia. The diagnosis of lymphoma was excluded in all cases (tumors negative for CD20 and CD45 [leukocyte common antigen]). All tumors were primary in the thyroid, and there was no clinical evidence of other primary tumors.
Immunohistochemistry The antibodies and their specificities, dilutions, and sources are listed in Table 1. All antibodies have been previously characterized.25-33 The immunostains were performed by using avidin-biotin detection system with diaminobenzidine as the chromogen. Avidin-biotin block (Dako, Carpinteria, CA) was used before the application of primary antibody. Negative and positive controls were used in each run.
RESULTS Anaplastic (undifferentiated) thyroid carcinomas from 35 patients were studied. There were 25 women and 10 men of median age 70 years (range, 40 to 89; mean, 70 years). These tumors were histologically classified into 3 categories: squamoid cohesive (SC), giant cell (GC), and spindle cell sarcomatoid. Four patients had both SC and GC components, which were scored separately for immunoreactivity. The immunohistochemical results have been summarized in Table 2.
TABLE 1. Polypeptide
There were 9 pure squamoid carcinomas and additional squamoid components in four tumors, all of which had a major large giant cell component. These tumors occurred in 7 women and 6 men with a median age of 70 years (mean, 71 years; range, 55 to 89 years). One of these tumors had elements of preexisting follicular carcinoma. Histologically, these tumors showed cohesive clusters of large, polygonal epithelial cells with ample, variably eosinophilic cytoplasm with various degrees of squamoid features. In some cases, the tumor cells were separated by fibrous septa, whereas in others they formed solid sheets with minimal stroma (Fig 1A, C). Extensive keratin expression was detected in most cases. Nearly all of them showed K7, K8, K18, and K19 in most tumor cells. Epithelial membrane antigen was detected in all but 1 case, although it often was limited to small foci of tumor cells. Keratins of complex epithelia were expressed variably. K14 was detected in all cases, although the proportion of positive cells varied between 1% and 100% (median, 15%) (Fig 1B). Most cases showed at least focal reactivity for K17, although this was only rarely extensive (Fig 1D). Focal positivity for K13 was common (Fig 1E), and reactivity for K16 was seen in 1 case strictly limited to keratin pearls (Fig 1F). K10 was not detected. All but 2 tumors were negative for TTF-1, whereas positive normal or differentiated carcinomatous elements were seen in all but 2 cases. One positive tumor showed distinct nuclear staining in most of the squamoid epithelial cells (Fig 1G, H), and another showed a 10% TTF-1–positive population. Only the latter case among this group was positive for thyroglobulin; all other positive elements in other tumors represented either entrapped normal thyroid or preexisting differentiated elements. In contrast, all 10 papillary and 5 follicular carcinomas had nuclear positivity for TTF-1.
Monoclonal Antibodies, Their Sources and Dilutions, and the Pretreatment Modalities Used in This Study Clone
Pretreatment
Antibody Dilution
Source
Reference
Keratin 7 Keratin 8
OV-TL 12/30 Cam5.2
PrVIII* Pepsin†
1:50 1:40
33 28
Keratin Keratin Keratin Keratin Keratin Keratin Keratin TTF-1 EMA
LHP1 KS-1A3 LL002 LL025 E3 DC-10 RCK 108 8G7G3/1 E29
MW‡ MW‡ MW‡ MW‡ MW‡ MW‡ MW‡ PrVIII1 MW‡
1:50 1:100 1:100 1:40 1:40 1:40 1:50 1:50 1:50
Dako Beckton-Dickinson, Mountain view, CA Novocastra Novocastra Novocastra Novocastra Novocastra Novocastra Dako, Carpinteria, CA Biocare, Walnut Creek, CA Dako
10 13 14 16 17 18 19
29 32 31 30 26 25 27
* PrVIII ⫽ 0.05% Sigma protease type VIII, 3 minutes in 0.1 mol/L phosphate buffer, pH 7.8 at 37°C. † Pepsin ⫽ crude pepsin (0.05% in HCl, pH 2.0) for 30 min at 37°C. ‡ MW ⫽ microwawe heating adjusted to near to boiling in EDTA-buffer 20 minutes at pH 6.0, followed by a 20-minute cooling period.
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TABLE 2. Summary of the Expression of Seven Keratin Polypeptides and Epithelial Membrane Antigens in Variants of Thyroid Anaplastic Carcinomas Squamoid (n ⫽ 13) Keratin 7 ⬎10 of tumor cells ⬍10% of tumor cells Negative Keratin 8 ⬎30 of tumor cells ⬍30% of tumor cells Negative Keratin 18 ⬎30 of tumor cells ⬍30% of tumor cells Negative Keratin 19 ⬎30 of tumor cells ⬍30% of tumor cells Negative Keratin 17 ⬎30 of tumor cells ⬍30% of tumor cells Negative Keratin 13 ⬎30 of tumor cells ⬍30% of tumor cells Negative Keratin 14 ⬎30 of tumor cells ⬍30% of tumor cells Negative Epithelial membrane antigen ⬎30 of tumor cells ⬍30% of tumor cells Negative
Giant Cell/Solid Spindle Cell Epithelioid Sarcomatoid (n ⫽ 16) (n ⫽ 7)
12 1 0
9 4 5
1 4 3
10 3 0
10 2 6
1 5 2
12 0 1
12 2 4
3 1 4
10 2 1
4 3 10
0 2 5
4 5 4
1 3 15
0 2 6
4 4 2
0 3 15
0 0 7
6 5 0
1 6 10
0 1 6
6 5 1
3 3 10
0 1 6
cells in only 3 cases and focally in 4 additional cases. Expression of complex epithelial keratins was sporadic. K14, K17, and K13 were seen in 7, 4, and 3 cases, respectively. Only 1 of these had more than focal positivity for K14 and K17 (30% to 50% of tumor cells); all others had only scattered positive cells. K10 and K16 were not detected. All tumors were negative for TTF-1 and thyroglobulin; 7 cases had TTF-1–positive differentiated components in which the nuclei were brightly positive (Fig 2F). TAC With Spindle Cell Sarcomatoid Features All 8 tumors occurred in women of median age of 63 years (range, 40 to 86 years). No elements of differentiated thyroid carcinoma were found in these tumors. This group of tumors were composed of undifferentiated spindle cells in a dense, sclerosing, collagenrich background (Fig 3A), except for 1 tumor that had a matrix-poor, noncollagenous background. Keratin-positive cells were detected most often with the antibody to K8 (6 of 8), whereas K7-, K18-, and K19-positive cells were detected in 5, 4, and 2 cases, respectively. Although many cases showed limited numbers of keratin-positive cells, in 3 of 8 cases most tumor cells were positive for K18, but only 1 of these cases showed significant numbers of K7- and K8-positive cells (Fig 3B). None of these tumors showed K19-positivity in more than 5% of tumor cells. One tumor was negative with all keratin antibodies. The complex epithelial keratins were seen only occasionally. Two tumors both scattered K14 and K17 positive cells. K10, K13, and K16 were not detected. All cases were negative for TTF-1 and thyroglobulin. EMA-positive cells were present in 1 case only (focally).
TACs With Giant Cell/Solid Epithelioid Patterns These 18 tumors, of which 4 tumors also had small squamoid cohesive foci, occurred in 12 women and 6 men of median age 70 years (mean, 71 years; range, 53 to 89 years). The keratins of the squamoid foci were evaluated with the squamoid tumors (see above). Six of these tumors arose in the background of papillary carcinoma, and 1 with a follicular carcinoma. The tumors showed features intermediate between squamoid epithelial and spindle cell sarcomatous patterns. They were composed of epithelioid cells with variable cellular cohesion ranging from dyscohesive pattern to solid sheets of tumor cells. Large cytoplasmic giant cells containing usually 1 irregular, sometimes lobated, nucleus were often present (Fig 2A, C). The amount of extracellular matrix was sparse. All but 2 cases (16 of 18) showed positive cells at least with 1 of the keratin antibodies. Most cases showed extensive reactivity for K7, K8, and K18 (Fig 2B). K18 was the most abundant keratin and was detected in 12 of 18 cases in over 20% of tumor cells, and focally in 2 additional cases (Fig 2D). K19 was usually absent or scant (Fig 2E); it was seen in more than 20% of tumor
DISCUSSION Thyroid anaplastic (undifferentiated) carcinomas (TACs) constitute a morphologically heterogeneous group, of which three different patterns are generally recognized: squamoid, giant cell, and sarcomatoid.2-4 In this study, we examined TACs of different types from 35 patients for epithelial and thyroid differentiation by using a panel of antibodies specific to individual keratins, epithelial membrane antigen, thyroglobulin, and thyroid transcription factor I. We divided the TACs in 3 groups, to better examine the spectrum of differentiation in these tumors. Thyroid transcription factor 1 (TTF-1) is a DNAbinding transcriptional activator protein that regulates the expression of certain key thyroid specific genes, for example, that of thyroglobulin.19 TTF-1 also is expressed in bronchial epithelia and has been shown to be present in differentiated but not anaplastic thyroid carcinoma, subsets of pulmonary adenocarcinomas, and pulmonary small cell carcinomas.22-24 We found
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FIGURE 1. Patterns of keratins and TTF-1 in squamoid anaplastic thyroid carcinomas. (A) This anaplastic carcinoma has cohesive sheets of squamoid cells in a collageneous stroma. (B) The tumor cells are strongly and uniformly positive for keratin 14. (C) This anaplastic carcinoma is composed of solid sheets of squamoid epithelial cells. (D) The tumor cells are extensively positive for keratin 17. (E) Focal keratin 13 in a squamoid anaplastic carcinoma with dyscohesive tumor cells. (F) Only a squamous epithelial keratin pearl is positive in this anaplastic carcinoma. (G) A squamoid anaplastic carcinoma with necrosis shows nuclear TTF-1 positivity (H).
consistent nuclear TTF-1 expression in the normal thyroid, in the differentiated carcinoma components, and in papillary and follicular carcinomas studied as controls. However, with few exceptions, TTF-1 was absent in TAC cells. Therefore, TTF-1 cannot be used as a marker for thyroid origin of TACs. The 2 positive tumors were both squamoid variants of TAC, neither of them showing differentiated carcinoma components. TTF-1 therefore may have a limited usefulness in the identification of thyroid origin of the squamoid variant of TAC. The general absence of thyroglobulin in the anaplastic components paralleled the absence of TTF-1, consistent with the concept that TTF-1 regulates thyro-
globulin expression.19 However, thyroglobulin was present in the preexisting differentiated elements and in entrapped thyroid. Previous series have shown thyroglobulin expression in tumors cells to be exceptional in TACs.3,12 There are 20 different epithelial (cyto)keratins (excluding the hair and nail keratins) belonging to 2 isoelectric groups according to the Moll catalog15,34: I (A, acidic comprising K9 to K20) and II (B, basic comprising K1 to K8). Two of them (K3 of group II and K12 of group I) are specific for cornea, and the other keratins can be roughly divided into 2 functional groups: keratins of simple, nonstratified epithelia and keratins of complex epithelia. The
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FIGURE 2. Patterns of keratins and TTF-1 in giant cell/solid epithelioid anaplastic carcinoma. (A) An anaplastic giant cell/solid epithelioid carcinoma with pleomorphic epithelioid cells. The tumor cells extensively positive for keratin 7 (B). (C) An anaplastic giant cell/solid epithelioid carcinoma with islands of preexisting papillary carcinoma with uniform presence of keratin 18 (D), complete loss of keratin 19 (E) and TTF-1 (F); the latter 2 markers are present only in the papillary carcinoma components.
former, K7, K8, K18, and K19, including the lowest molecular weight keratins in groups I and II, are generally typical of glandular and other simple, nonstratified epithelia. Of the complex epithelial keratins, the high-molecular-weight keratins K1, K2, K9, K10, and K11 are generally restricted to keratinizing squamous epithelia, such as epidermis, whereas K4 and K13 are present in internal squamous epithelia and urothelia. K5 and K14 and K17, in turn, are present in many basal cells such as myoepithelia and some basal cells of squamous epithelia and skin adnexa.15,35-38
The squamoid TACs showed extensive keratin expression with a complex pattern somewhat paralleling the complex pattern of keratins in squamous epithelia. These tumors also prominently expressed all simple epithelial keratins, of which K7, K8, and K18 are often absent in well-differentiated squamous cell carcinomas, although they become expressed in poorly differentiated squamous cell carcinomas. In their keratin pattern, SCs resemble papillary thyroid carcinomas by often having similar complexity in the keratin profile with coexpression of simple epithelial keratins with K5/14 and K17.17,18 This finding is consistent with the fact that
FIGURE 3. Sarcomatoid anaplastic carcinoma shows spindled tumor cells in a collagenous stroma (A). The tumor cells are positive for keratin 8.
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some of these tumors may have originated from papillary carcinoma. Relative, partial loss of simple epithelial keratins and nearly total loss of complex epithelial keratins K13, K14, and K17 was typically observed in the TACs that were composed of sheets or poorly cohesive poorly differentiated epithelioid cells or giant cells; these tumors represented the largest group of TACs and were intermediate between the squamoid and spindle cell sarcomatous TACs. Most tumors of this group contained simple epithelial keratin–positive cells and therefore could be verified as epithelial. However, there was a remarkable loss of K19 in this group of tumors, compared with the possible squamoid component or preexisting papillary or follicular elements. Nevertheless, presence of complex epithelial keratins, although to lesser degree, could suggest relationship with papillary carcinoma, which, in fact, was verified in several cases. Loss of keratins in carcinomas is rare but has been described in vitro with hepatocellular carcinoma cells that lost their keratin expression on dedifferentiation in culture.39 Posttranslational downregulation also has been shown as a possible mechanism that lessens the keratin expression in TACs.40 It has been shown in oral squamous carcinomas that downregulation of K19 is associated with an increased invasive potential.41 Also, the expression of stratified epithelial keratins has been shown to reflect increased aggressiveness among ductal carcinomas of the breast.42 Both of these phenomenona may occur in TACs that are typically highly aggressive tumors. The sarcomatoid TACs that are composed of spindle cells in a sclerotic, collagenous background typically show extensive loss of keratins. In these tumors, keratins are often limited to simple epithelial keratins, typically in only scattered cells. However, the keratin expression K7, K8, K18, and rarely K19 still supports epithelial origin of these tumors. We believe that by overall similarity, the totally keratinnegative tumors are also carcinomas rather than sarcomas, although no epithelial markers can be demonstrated. In summary, we have evaluated 35 anaplastic thyroid carcinomas for epithelial markers and TTF-1 to better understand the cell differentiation in these tumors. Our results show nearly complete absence of TTF-1 in TACs, indicating that this marker is not generally useful in establishing thyroid origin for these tumor. The complex keratin patterns seen in squamoid, and to lesser degree giant cell epithelioid tumors, is somewhat similar to the keratin patterns seen in papillary carcinoma, suggesting a possible histogenetic relationship. Keratin reactivity (especially for simple epithelial keratins) can be shown at least focally in most tumors, supporting their epithelial origin. REFERENCES 1. Franssila K: Value of histologic classification of thyroid cancer. Acta Pathol Microbiol Scand 225:5-76, 1971 (suppl)
2. Nishiyama R, Dunn EL, Thompson NW: Anaplastic spindlecell and giant-cell tumors of the thyroid. Cancer 30:113-127, 1972 3. Carangiu ML, Steeper T, Zampi G, et al: Anaplastic thyroid carcinoma: A study of 70 cases. Am J Clin Pathol 83:135-158, 1985 4. Rosai J, Saxe´n EA, Woolner L: Undifferentiated and poorly differentiated carcinoma. Semin Diagn Pathol 2:123-136, 1985 5. Shvero J, Gal R, Avidor I, et al: Anaplastic thyroid carcinoma: A clinical, histologic, and immunohistochemical study. Cancer 62: 319-325, 1988 6. Venkatesh YSS, Ordonez NG, Schultz PN, et al: Anaplastic carcinoma of the thyroid: A clinicopathologic study of 121 cases. Cancer 66:321-330, 1990 7. Lampertico P: Anaplastic (sarcomatoid) carcinoma of the thyroid gland. Semin Diagn Pathol 10:159-168, 1993 8. Miettinen M, Franssila K, Lehto VP, et al: Expression of intermediate filament proteins in thyroid gland and in thyroid tumors. Lab Invest 50:262-270, 1984 9. Dockhorn-Dworniczak B, Franke WW, Czernobilsky B, et al: Patterns of expression of cytoskeletal proteins in human thyroid gland and thyroid carcinomas. Differentiation 35:53-71, 1987 10. Henzen-Longmans SC, Mullink H, Ramaekers FC, et al: Expression of cytokeratins and vimentin in epithelial cells of normal and pathologic thyroid tissue. Virchows Arch A Pathol Anat Histopathol 410:347-354, 1987 11. Hurlimann J, Gardiol D, Scazziga B: Immunohistology of anaplastic thyroid carcinoma: A study of 43 cases. Histopathology 11:567-580, 1987 12. Livolsi VA, Brooks JJ, Arendash-Durand B: Anaplastic thyroid tumors: Immunohistology. Am J Clin Pathol 87:434-442, 1987 13. Ordonez NG, El-Naggar AK, Hickey RC, et al: Anaplastic thyroid carcinoma: Immunocytochemical study of 32 cases. Am J Clin Pathol 96:15-24, 1991 14. Schro¨der S, Wodzynski A, Padberg B: Zytokeratinexpression benigner und maligner epithelialer Schilddrusentumoren. Eine immunohistologische Unterschung an 154 Neoplasien inter Einsatz von8 verschiedene monoklonalen Zytokeratinantikorpern. Pathologe 17:425-432, 1996 15. Moll R, Franke WW, Schiller DL, et al: The catalog of human cytokeratins: Patterns of expression in normal epithelia, tumors and cultured cells. Cell 31:11-24, 1982 16. Fonseca E, Nesland JM, Ho¨ie J, et al: Patterns of expression of intermediate cytokeratin filaments in the thyroid gland: An immunohistochemical study of simple and stratified epithelial-type cytokeratins. Virchows Arch 430:239-246, 1997 17. Miettinen M, Kovatich AJ, Ka¨rkka¨inen P: Patterns of keratin polypeptides in papillary and follicular thyroid lesions. Virchows Arch 431:407-413, 1997 18. Baloch ZW, Abraham S, Roberts S, et al: Differential expression of cytokeratins in follicular variant of papillary carcinoma: an immunohistochemical study and its diagnostic utility. HUM PATHOL 30:1166-1171, 1999 19. Bingle CD: Thyroid transcription factor-1. Int J Biochem Cell Biol 29:1471-1473, 1997 20. Fabbro G, Di Loreto C, Beltrami CA, et al: Expression of thyroid-specific transcription factors TTF-1 and PAX-8 in human thyroid neoplasms. Cancer Res 54:4744-4749, 1994 21. Holzinger A, Dingle S, Bejarano PA, et al: Monoclonal antibody to thyroid transcription factor-1: Production, characterization, and usefulness in tumor diagnosis. Hybridoma 15:49-53, 1996 22. Bejarano PA, Baughman RP, Biddinger PW, et al: Surfactant proteins and thyroid transcription factor-1 in pulmonary and breast cazrcinomas. Mod Pathol 9:445-452, 1996 23. Folpe AL, Gown AM, Lamps LW, et al: Thyroid transcription fator-1: Immunohistochemical evaluation in pulmonary neuroendocrine tumors. Mod Pathol 12:5-8, 1999 24. Voutilainen PE, Multanen M, Haapiainen RK, et al: Anaplastic thyroid carcinoma survival. World J Surg 23:975-978, 1999 25. Bartek J, Vojtesek B, Staskova Z, et al: A series of 14 new monoclonal monoclonal antibodies to keratins: Characterizations and value in diagnostic histopathology. J Pathol 164:215-224, 1991 26. Guelstein VI, Tchypysheva TA, Ermilova VD, et al: Monoclonal antibody mapping of keratins 8 and 17 and of vimentin in normal human mammary gland, benign tumors, dysplasias and breast cancer. Int J Cancer 42:147-153, 1988
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GF, Topp WC, et al (eds): Cancer Cell I/The Transformed Phenotype. Cold Spring Harbor, NY, Cold Spring Harbor Laboratory, 1985, pp 169-176 35. Nagle RB: Intermediate filaments: A review of the basic biology. Am J Surg Pathol 12:4-16, 1988 (suppl 1) 36. Miettinen M: Keratin immunohistochemistry: Update on applications and pitfalls. Pathol Annu 24:133-145, 1993 37. Moll R: Cytokeratins in the histological diagnosis of malignant tumors. Int J Biol Markers 9:63-69, 1994 38. Moll R, Krepler R, Franke WW: Complex cytokeratin polypeptide patterns observed in certain human carcinomas. Differentiation 23:256-269, 1983 39. Venetianer A, Schiller DL, Magin T, et al: Cessation of cytokeratin expression in a rat hepatoma cell line lacking differentiated functions. Nature 305:730-733, 1983 40. Paine ML, Gibbins JR, Chew KE, et al: Loss of keratin expression in anaplastic carcinoma cells due to posttranscriptional downregulation acting in trans. Cancer Res 52:6603-6611, 1992 41. Crowe DL, Milo GE, Shuler CF: Keratin 19 downregulation by oral squamous cell carcinoma lines increases invasive potential. J Dent Res 78:1256-1263, 1999 42. Malzahn K, Mitze M, Thoenes M, et al: Biological and prognostic significance of stratified epithelial cytokeratins in infiltrating ductal breast carcinomas. Virchows Arch 433:119-129, 1998
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