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Nasopharyngeal hyalinizing clear cell carcinoma with EWSR1 rearrangements diagnosed by fluorescence in situ hybridization
Auris Nasus Larynx 42 (2015) 412–415
Contents lists available at ScienceDirect
Auris Nasus Larynx journal homepage: www.elsevier.com/locate/anl
Nasopharyngeal hyalinizing clear cell carcinoma with EWSR1 rearrangements diagnosed by fluorescence in situ hybridization Atsushi Fukuda a,*, Yohei Tagami b, Akira Takasawa b, Shintaro Sugita c, Rinnosuke Kuramoto a, Suguru Imai a, Tadashi Hasegawa c, Keiji Iizuka a a b c
Department of Otolaryngology, Kushiro City General Hospital, 1-12, Syunkodai, Kushiro-shi, Hokkaido 085-0822, Japan Department of Pathology, Sapporo Medical University School of Medicine, Minami 1, Nishi 17, Chuou-ku, Sapporo-shi, Hokkaido 060-8556, Japan Department of Surgical Pathology, Sapporo Medical University School of Medicine, Minami 1, Nishi 16, Chuou-ku, Sapporo-shi, Hokkaido 060-8543, Japan
A R T I C L E I N F O
A B S T R A C T
Article history: Received 1 January 2015 Accepted 19 February 2015 Available online 21 March 2015
Hyalinizing clear cell carcinoma (HCCC) is a rare, low-grade salivary gland neoplasm with a predilection for the palate and tongue. A 63-year-old woman presented a 14 14 17-mm mass at the roof of the nasopharynx. Endoscopic resection was performed via a transnasal approach. Histopathological findings of the salivary gland tumor indicated hyalinization of the stroma and neoplastic cells with clear cytoplasm without mucin. Fluorescence in situ hybridization analysis revealed that the tumor cells were positive for EWSR1 rearrangement. We finally diagnosed this case as HCCC of the nasopharynx. EWSR1 rearrangements are non-existent in other salivary gland tumors with clear cell change; thus, the identification of this rearrangement was very useful in accurately diagnosing HCCC. ß 2015 Elsevier Ireland Ltd. All rights reserved.
Keywords: Hyalinizing clear cell carcinoma Nasopharynx EWSR1 rearrangement FISH Salivary gland tumor
1. Introduction Hyalinizing clear cell carcinoma (HCCC) was first described by Milchgrub et al. as a rare carcinoma of the minor salivary glands [1], representing less than 1% of all salivary tumors [2]. The tumor is characterized by low-grade morphology with only occasional metastatic spread that can achieve good overall treatment outcome [3]. HCCC exclusively comprises epithelial cells with optically clear cytoplasm that are positive for cytokeratin and negative for S-100 protein and smooth muscle actin [2]. HCCC has a wide differential diagnosis that includes other clear cell-containing tumors, such as epithelial–myoepithelial carcinoma, mucoepidermoid carcinoma, and myoepithelial carcinoma [3]. Hence, it is sometimes very challenging to distinguish HCCC from other clear cell mimics based on a small biopsy sample. Antonescu et al. [4] recently described the consistent expression of the Ewing sarcoma breakpoint region 1-activating transcription factor 1 (EWSR1-ATF1) fusion protein in HCCC. Furthermore, Shah et al. [5] recently found that EWSR1 rearrangements were not a
* Corresponding author at: Department of Otolaryngology, Kushiro City General Hospital, Kushiro-city, Hokkaido 085-0822, Japan. Tel.: +81 0154 41 6121; fax: +81 0154 41 4080. E-mail address: [email protected] (A. Fukuda). http://dx.doi.org/10.1016/j.anl.2015.02.015 0385-8146/ß 2015 Elsevier Ireland Ltd. All rights reserved.
feature in other salivary gland tumors, suggesting that this EWSR1 rearrangement is specific to HCCC. The occurrence of HCCC in the nasophyarynx is very rare. To the best of our knowledge, there has been only one other report of nasopharyngeal HCCC identified by EWSR1 rearrangement [6]. Here, we report a case of nasopharyngeal HCCC in a 63-year-old Japanese woman and comment on the usefulness of the identification of EWSR1 rearrangement by fluorescence in situ hybridization (FISH) for the confirmation of the diagnosis. 2. Case report A 63-year-old woman was referred to our department with a 5year history of an asymptomatic, slow-growing, nasopharyngeal mass annually monitored by brain magnetic resonance imaging (MRI). Before presentation, she underwent a comprehensive cranial examination, including fluorine-18 fluorodeoxyglucose positron emission tomography/computed tomography (FDG-PET/ CT). MRI revealed a 14 14 17-mm, expansile mass at the roof of the nasopharynx with low intensity on a T1-weighted image (WI) and slightly high intensity on T2WI. Contrast-enhanced T1weighted MRI revealed a virtual homogenous enhancement of the mass. Further, FDG-PET revealed an uptake of 18F-FDG by the nasopharyngeal mass with a maximum standardized uptake value (max) of 5.8 (Fig. 1). There was no evidence of lymph node or distant metastases. Nasopharyngoscopy revealed a protruded
A. Fukuda et al. / Auris Nasus Larynx 42 (2015) 412–415
413
Fig. 1. MRI showed a 14 14 17-mm, expansile mass at the roof of the nasopharynx with low intensity on a T1-weighted image (WI) (A). Contrast-enhanced T1-weighted MRI revealed virtual homogenous enhancement of the mass (B, C). FDG-PET revealed 18F-FDG uptake by the nasopharyngeal mass with a maximum standardized uptake value (max) of 5.8 (D).
lesion with a smooth surface, approximately 20 mm in diameter, located on the roof of the nasopharynx (Fig. 2). Biopsy of the mass was performed and a tumor derived from a salivary gland was suspected; therefore, an incisional biopsy was performed using an endoscopic transnasal approach under general anesthesia. The tumor was totally excised macroscopically. The pathological
Fig. 2. Nasopharyngoscopy revealed a protruded lesion with a smooth surface, approximately 20 mm in diameter located on the roof of the nasopharynx.
findings were consistent with hyalinizing clear cell carcinoma, as described below. There have been no indications of recurrence or metastasis for 1 year without adjuvant therapy. 3. Pathological findings The tumor was 15 15 10 mm in size. The lesion was microscopically covered by protrusions of pseudostratified ciliated epithelium with minor salivary glands observed in the marginal regions. The tumor was formed of sheets, nests, and cords of tumor cells with fibrosis and/or hyalinization of the stroma (Fig. 3A), and slight myxomatous changes were observed. High-power magnification revealed that the sheets and nests were composed of welldelineated cells, most with clear cytoplasm and some with eosinophilic cytoplasm (Fig. 3B). The nuclei were not oval but polygonal. The number of mitotic figures was less than one per 10 high-power fields. Moreover, the cytoplasm of the clear cells was reactive with periodic-acid Schiff stain only before diastase digestion, suggesting the presence of glycogen. Very few cells were found positive by mucicarmine staining. Immunohistochemical studies demonstrated that the tumor cells were positive for cytokeratin AE1/AE3 (Fig. 3C), p40 (Fig. 3D), p63, and epithelial membrane antigen, whereas negative for glial fibrillary acidic protein, cluster of differentiation 10, high-molecular-weight kininogen, S100-protein, smooth muscle actin, and focally positive for vimentin. FISH analysis was performed on formalin-fixed paraffinembedded tissue sections of the tumor using a commercially
414
A. Fukuda et al. / Auris Nasus Larynx 42 (2015) 412–415
Fig. 3. Histopathological features of the tumor. (A) Low power view of the tumor. The tumor exhibited typical characteristics of hyalinizing clear cell carcinoma (HCCC), including the formation of nests and cords composed of polygonal to round tumor cells and hyaline bands in the stroma. (B) High power view of the tumor. Tumor cells with clear cytoplasm were embedded in abundant hyalinous stroma. (C) Tumor cells were positive for cytokeratin AE1/AE3. (D) Most tumor cells were positive for p40.
available EWSR1 (22q12) dual color, break apart rearrangement probe (Abbot Molecular Inc., Des Plaines, IL, USA). FISH analysis was performed according to the manufacturer’s instructions, as described in a previous study [7]. An interphase FISH analysis then revealed split signals of EWSR1 rearrangement in 92% of the tumor cells (Fig. 4). We finally diagnosed this case as HCCC of the minor salivary glands and surgical margins were positive.
4. Discussion HCCC was first described as a separate entity by Milchgrub et al. [1] as a low-grade, rare salivary gland neoplasm made up of clear cells that form cords and nests in hyalinized stroma. A majority of HCCC cases are presumed to arise from minor salivary glands [1], while the most common sites of origin are the
Fig. 4. Fluorescence in situ hybridization (FISH) showed split signals, indicating EWSR1 rearrangement (arrow heads).
palate (27.5%) and tongue (25.5%), followed by the floor of the mouth (8.1%) and other locations with minor salivary glands. HCCC exhibits a slight prediction for middle aged-females (60.2%) [6] and occurrence in the nasopharynx is exceptionally rare. Including the present case, till date, there have been only 10 cases of HCCC occurring in the nasopharynx [6,8]. This tumor is reportedly confused with monomorphic variants of epithelial–myoepithelial carcinoma, mucin-depleted mucoepidermoid carcinoma, and myoepithelial carcinoma. However, HCCC has been since separated from these entities because of its lack of apparent squamous, mucinous, and myoepithelial differentiation [1]. Furthermore, differential diagnosis is extensive and includes a number of neoplasms with clear cell changes. The usual suspects and other rare tumors considered in the differential diagnosis include clear cell variants of squamous cell carcinoma, myoepithelial carcinoma, mucoepidermoid carcinoma, clear cell odontogenic carcinoma, calcifying epithelial odontogenic tumors (in bone), epithelial–myoepithelial carcinoma, acinic cell carcinoma, oncocytoma, myoepithelioma, and metastatic renal cell carcinoma [3]. Many soft tissue tumors, including sarcomas, are associated with EWSR1 translocations. EWSR1 rearrangement has been recently described by Antonescu et al. [4] in 18 of 22 (82%) HCCC cases with ATF1 as the partner gene in 13 of 14 (93%) cases. In addition, Shah et al. [5] reported that 13 of 15 (87%) HCCC cases demonstrated the EWSR1 rearrangement and EWSR1 rearrangement was not a feature of the vast majority of salivary gland tumors. If these reports are correct, the sensitivity and specificity of EWSR1 rearrangement by FISH for a diagnosis of HCCC are 82–87% and 100%, suggesting that EWSR1 rearrangement is a hallmark of HCCC and can be used to distinguish this lesion from other salivary gland neoplasms. In the present case, although we considered a differential diagnosis of mucoepidermoid carcinoma based on morphologic features and immunohistochemical studies, we finally diagnosed the case as HCCC of the nasopharynx as a result of identifying EWSR1 rearrangement by FISH.
A. Fukuda et al. / Auris Nasus Larynx 42 (2015) 412–415
HCCC is currently classified as a ‘‘clear cell carcinoma, not otherwise specified (NOS)’’ by the World Health Organization (WHO) ‘‘blue book’’ on head and neck tumors [9]. It is considered by the WHO to be a diagnosis of exclusion. However, the EWSR1ATF1 fusion is specific for HCCC within the context of salivary neoplasms. Brandwein and Wei [10] argued that ‘‘NOS’’ is not applicable for a diagnosis of HCCC. Fulciniti et al. [11] diagnosed a recurrent, poorly differentiated HCCC of the parotid gland by identifying EWSR1 translocation by FISH analysis of fine needle cytology (FNC) sampling of three separate soft tissue masses in the pre-masseterine area. Jin et al. [12] reported EWSR1 rearrangement in a case of recurrent HCCC at the base of the tongue with high-grade transformation by FISH, which is a reliable and relatively accessible method to diagnose HCCC in samples acquired by FNC as well as small biopsies. Further, many molecular biology labs are already equipped to identify EWSR1 rearrangement by FISH. Last, this diagnostic method is useful for diagnosing HCCC cases with poor differentiation or highgrade transformation, which are very difficult to correctly diagnose by only morphologic features and immunohistochemical patterns. HCCC is a relatively rare disease and there is currently no consensus on a standard treatment. The recommended treatment for HCCC is wide surgical resection as benefits from adjuvant treatment have not been confirmed and the effect of chemotherapy remains unknown. Post-operative radiotherapy is recommended when a total surgical removal of the primary tumor is not attainable [13]. However, Gon et al. [2] reported a case of locoregional recurrence after 6 years, despite all surgical margins being free of neoplasms and administration of three cycles of adjuvant radiotherapy, suggesting that the therapeutic value of radiotherapy for HCCC may be limited. Although tumors in HCCC have a propensity for locoregional recurrence in the order of 12–17% and occasional lymph node or distant metastases have been noted [3], the prognosis is excellent. Su et al. [14] reported two patients who underwent wide local resection for HCCC and remained disease-free at the primary site but were diagnosed with cervical lymph node metastases 10 and 14 years later. Further, because HCCC is associated with late recurrence, even years later, patients should be continuously followed up. While rare, cases of dedifferentiated HCCC have been reported [12,15]; therefore, it is important to consider such a diagnosis.
5. Conclusion In conclusion, we reported a case of nasopharyngeal HCCC with EWSR1 rearrangements detected by FISH. EWSR1 rearrangements have not been observed in any other salivary gland tumors with clear cell change; therefore, the identification of these characteristics was very useful to accurately diagnosis HCCC. It is possible that the EWSR1-ATF1 fusion oncogene may
415
be a future therapeutic target with diagnostic and prognostic implications in HCCC. Conflict of interest The authors declare no conflict of interest associated with this manuscript. Acknowledgement The authors would like to thank Dr Tomoyuki Oouchi (Department of Pathology, Keiyukai Sapporo Hospital) for his expert advice. References [1] Milchgrub S, Gnepp DR, Vuitch F, Delgado R, Albores-Saavedra J. Hyalinizing clear cell carcinoma of salivary gland. Am J Surg Pathol 1994;18:74–82. [2] Gon S, Bhattacharyya A, Majumdar B, Das TK. Post-radiotherapy locoregional recurrence of hyalinizing clear cell carcinoma of palate. J Cancer Res Ther 2013;9:281–3. [3] Weinreb I. Hyalinizing clear cell carcinoma of salivary gland: a review and update. Head Neck Pathol 2013;7:S20–9. [4] Antonescu CR, Katabi N, Zhang L, Sung YS, Seethala RR, Jordan RC, et al. EWSR1ATF1 fusion is a novel and consistent finding in hyalinizing clear-cell carcinoma of salivary gland. Genes Chromosomes Cancer 2011;50:559–70. [5] Shah AA, LeGallo RD, van Zante A, Frierson HF, Mills SE, Berean KW, et al. EWSR1 genetic rearrangements in salivary gland tumors: a specific and very common feature of hyalinizing clear cell carcinoma. Am J Surg Pathol 2013;37:571–8. [6] Ceballos Sa´enz C, Argyris PP, Manivel JC, Urias Barreras CM, Koutlas IG. Nasopharyngeal hyalinizing clear cell carcinoma: report of the histopathologic features of a case showing EWSR1 rearrangements by FISH and literature review. Int J Surg Pathol 2014;22:667–72. [7] Yamaguchi U, Hasegawa T, Morimoto Y, Tateishi U, Endo M, Nakatani F, et al. A practical approach to the clinical diagnosis of Ewing’s sarcoma/primitive neuroectodermal tumour and other small round cell tumours sharing EWS rearrangement using new fluorescence in situ hybridization probes for EWSR1 on formalin-fixed, paraffin wax-embedded tissue. J Clin Pathol 2005;58:1051–6. [8] Nakano T, Yamamoto H, Nishijima T, Tamiya S, Shiratsuchi H, Nakashima T, et al. Hyalinizing clear cell carcinoma with EWSR1-ATF1 fusion gene: report of three cases with molecular analyses. Virchows Arch 2015;466:37–43. [9] Barnes L, Eveson JW, Reichart P, Sidransky D, editors. World Health Organization Classification of Tumours. Pathology and genetics of head and neck tumours. Lyon: IARC Press; 2005. [10] Brandwein-Gensler M, Wei S. Envisioning the next WHO head and neck classification. Head Neck Pathol 2014;8:1–15. [11] Fulciniti F, Pia Curcio M, Liguori G, Aquino G, Botti G, Campanile AC, et al. Hyalinizing clear cell carcinoma of the parotid gland: report of a recurrent case with aggressive cytomorphology and behavior diagnosed on fine-needle cytology sample. Diagn Cytopathol 2014;42:63–8. [12] Jin R, Craddock KJ, Irish JC, Perez-Ordonez B, Weinreb I. Recurrent hyalinizing clear cell carcinoma of the base of tongue with high-grade transformation and EWSR1 gene rearrangement by FISH. Head Neck Pathol 2012;6:389–94. [13] Pujary K, Rangarajan S, Nayak DR, Balakrishnan R, Ramakrishnan V. Hyalinizing clear cell carcinoma of the base of tongue. Int J Oral Maxillofac Surg 2008;37:93–6. [14] Su HK, Wang BY, Mannan AASR, Dewey EH, Alpert EH, Reis LL, et al. Very delayed cervical lymph node metastases from hyalinizing clear cell carcinoma: report of 2 cases. Head Neck 2015;37:E19–21. [15] O’Regan E, Shandilya M, Gnepp DR, Timon C, Toner M. Hyalinizing clear cell carcinoma of salivary gland: an aggressive variant. Oral Oncol 2004;40:348–52.
Contents lists available at ScienceDirect
Auris Nasus Larynx journal homepage: www.elsevier.com/locate/anl
Nasopharyngeal hyalinizing clear cell carcinoma with EWSR1 rearrangements diagnosed by fluorescence in situ hybridization Atsushi Fukuda a,*, Yohei Tagami b, Akira Takasawa b, Shintaro Sugita c, Rinnosuke Kuramoto a, Suguru Imai a, Tadashi Hasegawa c, Keiji Iizuka a a b c
Department of Otolaryngology, Kushiro City General Hospital, 1-12, Syunkodai, Kushiro-shi, Hokkaido 085-0822, Japan Department of Pathology, Sapporo Medical University School of Medicine, Minami 1, Nishi 17, Chuou-ku, Sapporo-shi, Hokkaido 060-8556, Japan Department of Surgical Pathology, Sapporo Medical University School of Medicine, Minami 1, Nishi 16, Chuou-ku, Sapporo-shi, Hokkaido 060-8543, Japan
A R T I C L E I N F O
A B S T R A C T
Article history: Received 1 January 2015 Accepted 19 February 2015 Available online 21 March 2015
Hyalinizing clear cell carcinoma (HCCC) is a rare, low-grade salivary gland neoplasm with a predilection for the palate and tongue. A 63-year-old woman presented a 14 14 17-mm mass at the roof of the nasopharynx. Endoscopic resection was performed via a transnasal approach. Histopathological findings of the salivary gland tumor indicated hyalinization of the stroma and neoplastic cells with clear cytoplasm without mucin. Fluorescence in situ hybridization analysis revealed that the tumor cells were positive for EWSR1 rearrangement. We finally diagnosed this case as HCCC of the nasopharynx. EWSR1 rearrangements are non-existent in other salivary gland tumors with clear cell change; thus, the identification of this rearrangement was very useful in accurately diagnosing HCCC. ß 2015 Elsevier Ireland Ltd. All rights reserved.
Keywords: Hyalinizing clear cell carcinoma Nasopharynx EWSR1 rearrangement FISH Salivary gland tumor
1. Introduction Hyalinizing clear cell carcinoma (HCCC) was first described by Milchgrub et al. as a rare carcinoma of the minor salivary glands [1], representing less than 1% of all salivary tumors [2]. The tumor is characterized by low-grade morphology with only occasional metastatic spread that can achieve good overall treatment outcome [3]. HCCC exclusively comprises epithelial cells with optically clear cytoplasm that are positive for cytokeratin and negative for S-100 protein and smooth muscle actin [2]. HCCC has a wide differential diagnosis that includes other clear cell-containing tumors, such as epithelial–myoepithelial carcinoma, mucoepidermoid carcinoma, and myoepithelial carcinoma [3]. Hence, it is sometimes very challenging to distinguish HCCC from other clear cell mimics based on a small biopsy sample. Antonescu et al. [4] recently described the consistent expression of the Ewing sarcoma breakpoint region 1-activating transcription factor 1 (EWSR1-ATF1) fusion protein in HCCC. Furthermore, Shah et al. [5] recently found that EWSR1 rearrangements were not a
* Corresponding author at: Department of Otolaryngology, Kushiro City General Hospital, Kushiro-city, Hokkaido 085-0822, Japan. Tel.: +81 0154 41 6121; fax: +81 0154 41 4080. E-mail address: [email protected] (A. Fukuda). http://dx.doi.org/10.1016/j.anl.2015.02.015 0385-8146/ß 2015 Elsevier Ireland Ltd. All rights reserved.
feature in other salivary gland tumors, suggesting that this EWSR1 rearrangement is specific to HCCC. The occurrence of HCCC in the nasophyarynx is very rare. To the best of our knowledge, there has been only one other report of nasopharyngeal HCCC identified by EWSR1 rearrangement [6]. Here, we report a case of nasopharyngeal HCCC in a 63-year-old Japanese woman and comment on the usefulness of the identification of EWSR1 rearrangement by fluorescence in situ hybridization (FISH) for the confirmation of the diagnosis. 2. Case report A 63-year-old woman was referred to our department with a 5year history of an asymptomatic, slow-growing, nasopharyngeal mass annually monitored by brain magnetic resonance imaging (MRI). Before presentation, she underwent a comprehensive cranial examination, including fluorine-18 fluorodeoxyglucose positron emission tomography/computed tomography (FDG-PET/ CT). MRI revealed a 14 14 17-mm, expansile mass at the roof of the nasopharynx with low intensity on a T1-weighted image (WI) and slightly high intensity on T2WI. Contrast-enhanced T1weighted MRI revealed a virtual homogenous enhancement of the mass. Further, FDG-PET revealed an uptake of 18F-FDG by the nasopharyngeal mass with a maximum standardized uptake value (max) of 5.8 (Fig. 1). There was no evidence of lymph node or distant metastases. Nasopharyngoscopy revealed a protruded
A. Fukuda et al. / Auris Nasus Larynx 42 (2015) 412–415
413
Fig. 1. MRI showed a 14 14 17-mm, expansile mass at the roof of the nasopharynx with low intensity on a T1-weighted image (WI) (A). Contrast-enhanced T1-weighted MRI revealed virtual homogenous enhancement of the mass (B, C). FDG-PET revealed 18F-FDG uptake by the nasopharyngeal mass with a maximum standardized uptake value (max) of 5.8 (D).
lesion with a smooth surface, approximately 20 mm in diameter, located on the roof of the nasopharynx (Fig. 2). Biopsy of the mass was performed and a tumor derived from a salivary gland was suspected; therefore, an incisional biopsy was performed using an endoscopic transnasal approach under general anesthesia. The tumor was totally excised macroscopically. The pathological
Fig. 2. Nasopharyngoscopy revealed a protruded lesion with a smooth surface, approximately 20 mm in diameter located on the roof of the nasopharynx.
findings were consistent with hyalinizing clear cell carcinoma, as described below. There have been no indications of recurrence or metastasis for 1 year without adjuvant therapy. 3. Pathological findings The tumor was 15 15 10 mm in size. The lesion was microscopically covered by protrusions of pseudostratified ciliated epithelium with minor salivary glands observed in the marginal regions. The tumor was formed of sheets, nests, and cords of tumor cells with fibrosis and/or hyalinization of the stroma (Fig. 3A), and slight myxomatous changes were observed. High-power magnification revealed that the sheets and nests were composed of welldelineated cells, most with clear cytoplasm and some with eosinophilic cytoplasm (Fig. 3B). The nuclei were not oval but polygonal. The number of mitotic figures was less than one per 10 high-power fields. Moreover, the cytoplasm of the clear cells was reactive with periodic-acid Schiff stain only before diastase digestion, suggesting the presence of glycogen. Very few cells were found positive by mucicarmine staining. Immunohistochemical studies demonstrated that the tumor cells were positive for cytokeratin AE1/AE3 (Fig. 3C), p40 (Fig. 3D), p63, and epithelial membrane antigen, whereas negative for glial fibrillary acidic protein, cluster of differentiation 10, high-molecular-weight kininogen, S100-protein, smooth muscle actin, and focally positive for vimentin. FISH analysis was performed on formalin-fixed paraffinembedded tissue sections of the tumor using a commercially
414
A. Fukuda et al. / Auris Nasus Larynx 42 (2015) 412–415
Fig. 3. Histopathological features of the tumor. (A) Low power view of the tumor. The tumor exhibited typical characteristics of hyalinizing clear cell carcinoma (HCCC), including the formation of nests and cords composed of polygonal to round tumor cells and hyaline bands in the stroma. (B) High power view of the tumor. Tumor cells with clear cytoplasm were embedded in abundant hyalinous stroma. (C) Tumor cells were positive for cytokeratin AE1/AE3. (D) Most tumor cells were positive for p40.
available EWSR1 (22q12) dual color, break apart rearrangement probe (Abbot Molecular Inc., Des Plaines, IL, USA). FISH analysis was performed according to the manufacturer’s instructions, as described in a previous study [7]. An interphase FISH analysis then revealed split signals of EWSR1 rearrangement in 92% of the tumor cells (Fig. 4). We finally diagnosed this case as HCCC of the minor salivary glands and surgical margins were positive.
4. Discussion HCCC was first described as a separate entity by Milchgrub et al. [1] as a low-grade, rare salivary gland neoplasm made up of clear cells that form cords and nests in hyalinized stroma. A majority of HCCC cases are presumed to arise from minor salivary glands [1], while the most common sites of origin are the
Fig. 4. Fluorescence in situ hybridization (FISH) showed split signals, indicating EWSR1 rearrangement (arrow heads).
palate (27.5%) and tongue (25.5%), followed by the floor of the mouth (8.1%) and other locations with minor salivary glands. HCCC exhibits a slight prediction for middle aged-females (60.2%) [6] and occurrence in the nasopharynx is exceptionally rare. Including the present case, till date, there have been only 10 cases of HCCC occurring in the nasopharynx [6,8]. This tumor is reportedly confused with monomorphic variants of epithelial–myoepithelial carcinoma, mucin-depleted mucoepidermoid carcinoma, and myoepithelial carcinoma. However, HCCC has been since separated from these entities because of its lack of apparent squamous, mucinous, and myoepithelial differentiation [1]. Furthermore, differential diagnosis is extensive and includes a number of neoplasms with clear cell changes. The usual suspects and other rare tumors considered in the differential diagnosis include clear cell variants of squamous cell carcinoma, myoepithelial carcinoma, mucoepidermoid carcinoma, clear cell odontogenic carcinoma, calcifying epithelial odontogenic tumors (in bone), epithelial–myoepithelial carcinoma, acinic cell carcinoma, oncocytoma, myoepithelioma, and metastatic renal cell carcinoma [3]. Many soft tissue tumors, including sarcomas, are associated with EWSR1 translocations. EWSR1 rearrangement has been recently described by Antonescu et al. [4] in 18 of 22 (82%) HCCC cases with ATF1 as the partner gene in 13 of 14 (93%) cases. In addition, Shah et al. [5] reported that 13 of 15 (87%) HCCC cases demonstrated the EWSR1 rearrangement and EWSR1 rearrangement was not a feature of the vast majority of salivary gland tumors. If these reports are correct, the sensitivity and specificity of EWSR1 rearrangement by FISH for a diagnosis of HCCC are 82–87% and 100%, suggesting that EWSR1 rearrangement is a hallmark of HCCC and can be used to distinguish this lesion from other salivary gland neoplasms. In the present case, although we considered a differential diagnosis of mucoepidermoid carcinoma based on morphologic features and immunohistochemical studies, we finally diagnosed the case as HCCC of the nasopharynx as a result of identifying EWSR1 rearrangement by FISH.
A. Fukuda et al. / Auris Nasus Larynx 42 (2015) 412–415
HCCC is currently classified as a ‘‘clear cell carcinoma, not otherwise specified (NOS)’’ by the World Health Organization (WHO) ‘‘blue book’’ on head and neck tumors [9]. It is considered by the WHO to be a diagnosis of exclusion. However, the EWSR1ATF1 fusion is specific for HCCC within the context of salivary neoplasms. Brandwein and Wei [10] argued that ‘‘NOS’’ is not applicable for a diagnosis of HCCC. Fulciniti et al. [11] diagnosed a recurrent, poorly differentiated HCCC of the parotid gland by identifying EWSR1 translocation by FISH analysis of fine needle cytology (FNC) sampling of three separate soft tissue masses in the pre-masseterine area. Jin et al. [12] reported EWSR1 rearrangement in a case of recurrent HCCC at the base of the tongue with high-grade transformation by FISH, which is a reliable and relatively accessible method to diagnose HCCC in samples acquired by FNC as well as small biopsies. Further, many molecular biology labs are already equipped to identify EWSR1 rearrangement by FISH. Last, this diagnostic method is useful for diagnosing HCCC cases with poor differentiation or highgrade transformation, which are very difficult to correctly diagnose by only morphologic features and immunohistochemical patterns. HCCC is a relatively rare disease and there is currently no consensus on a standard treatment. The recommended treatment for HCCC is wide surgical resection as benefits from adjuvant treatment have not been confirmed and the effect of chemotherapy remains unknown. Post-operative radiotherapy is recommended when a total surgical removal of the primary tumor is not attainable [13]. However, Gon et al. [2] reported a case of locoregional recurrence after 6 years, despite all surgical margins being free of neoplasms and administration of three cycles of adjuvant radiotherapy, suggesting that the therapeutic value of radiotherapy for HCCC may be limited. Although tumors in HCCC have a propensity for locoregional recurrence in the order of 12–17% and occasional lymph node or distant metastases have been noted [3], the prognosis is excellent. Su et al. [14] reported two patients who underwent wide local resection for HCCC and remained disease-free at the primary site but were diagnosed with cervical lymph node metastases 10 and 14 years later. Further, because HCCC is associated with late recurrence, even years later, patients should be continuously followed up. While rare, cases of dedifferentiated HCCC have been reported [12,15]; therefore, it is important to consider such a diagnosis.
5. Conclusion In conclusion, we reported a case of nasopharyngeal HCCC with EWSR1 rearrangements detected by FISH. EWSR1 rearrangements have not been observed in any other salivary gland tumors with clear cell change; therefore, the identification of these characteristics was very useful to accurately diagnosis HCCC. It is possible that the EWSR1-ATF1 fusion oncogene may
415
be a future therapeutic target with diagnostic and prognostic implications in HCCC. Conflict of interest The authors declare no conflict of interest associated with this manuscript. Acknowledgement The authors would like to thank Dr Tomoyuki Oouchi (Department of Pathology, Keiyukai Sapporo Hospital) for his expert advice. References [1] Milchgrub S, Gnepp DR, Vuitch F, Delgado R, Albores-Saavedra J. Hyalinizing clear cell carcinoma of salivary gland. Am J Surg Pathol 1994;18:74–82. [2] Gon S, Bhattacharyya A, Majumdar B, Das TK. Post-radiotherapy locoregional recurrence of hyalinizing clear cell carcinoma of palate. J Cancer Res Ther 2013;9:281–3. [3] Weinreb I. Hyalinizing clear cell carcinoma of salivary gland: a review and update. Head Neck Pathol 2013;7:S20–9. [4] Antonescu CR, Katabi N, Zhang L, Sung YS, Seethala RR, Jordan RC, et al. EWSR1ATF1 fusion is a novel and consistent finding in hyalinizing clear-cell carcinoma of salivary gland. Genes Chromosomes Cancer 2011;50:559–70. [5] Shah AA, LeGallo RD, van Zante A, Frierson HF, Mills SE, Berean KW, et al. EWSR1 genetic rearrangements in salivary gland tumors: a specific and very common feature of hyalinizing clear cell carcinoma. Am J Surg Pathol 2013;37:571–8. [6] Ceballos Sa´enz C, Argyris PP, Manivel JC, Urias Barreras CM, Koutlas IG. Nasopharyngeal hyalinizing clear cell carcinoma: report of the histopathologic features of a case showing EWSR1 rearrangements by FISH and literature review. Int J Surg Pathol 2014;22:667–72. [7] Yamaguchi U, Hasegawa T, Morimoto Y, Tateishi U, Endo M, Nakatani F, et al. A practical approach to the clinical diagnosis of Ewing’s sarcoma/primitive neuroectodermal tumour and other small round cell tumours sharing EWS rearrangement using new fluorescence in situ hybridization probes for EWSR1 on formalin-fixed, paraffin wax-embedded tissue. J Clin Pathol 2005;58:1051–6. [8] Nakano T, Yamamoto H, Nishijima T, Tamiya S, Shiratsuchi H, Nakashima T, et al. Hyalinizing clear cell carcinoma with EWSR1-ATF1 fusion gene: report of three cases with molecular analyses. Virchows Arch 2015;466:37–43. [9] Barnes L, Eveson JW, Reichart P, Sidransky D, editors. World Health Organization Classification of Tumours. Pathology and genetics of head and neck tumours. Lyon: IARC Press; 2005. [10] Brandwein-Gensler M, Wei S. Envisioning the next WHO head and neck classification. Head Neck Pathol 2014;8:1–15. [11] Fulciniti F, Pia Curcio M, Liguori G, Aquino G, Botti G, Campanile AC, et al. Hyalinizing clear cell carcinoma of the parotid gland: report of a recurrent case with aggressive cytomorphology and behavior diagnosed on fine-needle cytology sample. Diagn Cytopathol 2014;42:63–8. [12] Jin R, Craddock KJ, Irish JC, Perez-Ordonez B, Weinreb I. Recurrent hyalinizing clear cell carcinoma of the base of tongue with high-grade transformation and EWSR1 gene rearrangement by FISH. Head Neck Pathol 2012;6:389–94. [13] Pujary K, Rangarajan S, Nayak DR, Balakrishnan R, Ramakrishnan V. Hyalinizing clear cell carcinoma of the base of tongue. Int J Oral Maxillofac Surg 2008;37:93–6. [14] Su HK, Wang BY, Mannan AASR, Dewey EH, Alpert EH, Reis LL, et al. Very delayed cervical lymph node metastases from hyalinizing clear cell carcinoma: report of 2 cases. Head Neck 2015;37:E19–21. [15] O’Regan E, Shandilya M, Gnepp DR, Timon C, Toner M. Hyalinizing clear cell carcinoma of salivary gland: an aggressive variant. Oral Oncol 2004;40:348–52.