Role of p40 and Cytokeratin 5/6 in the Differential Diagnosis of Sinonasal Undifferentiated Carcinoma Lavleen Singh M.D, Richa Ranjan M.D, Sudheer Arava M.D, M.K. Singh M.D PII: DOI: Reference:
S1092-9134(14)00004-5 doi: 10.1016/j.anndiagpath.2014.01.003 YADPA 50911
To appear in:
Annals of Diagnostic Pathology
Received date: Revised date: Accepted date:
18 August 2013 22 November 2013 5 January 2014
Please cite this article as: Singh Lavleen, Ranjan Richa, Arava Sudheer, Singh MK, Role of p40 and Cytokeratin 5/6 in the Differential Diagnosis of Sinonasal Undifferentiated Carcinoma, Annals of Diagnostic Pathology (2014), doi: 10.1016/j.anndiagpath.2014.01.003
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT Role of p40 and Cytokeratin 5/6 in the Differential Diagnosis of Sinonasal Undifferentiated Carcinoma
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Lavleen Singh M.D, Richa Ranjan M.D, Sudheer Arava M.D, M K Singh M.D Department of Pathology, All India Institute of Medical Sciences, New Delhi-110029 Abstract
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Sinonasal undifferentiated carcinoma is an epithelial neoplasm of sinonasal region which does not exhibit a squamous or glandular differentiation. The challenge in diagnosis of this entity is the rarity of the disease, the varying morphology of the tumor which leads to gamut of differential diagnosis and the paucity of consistent immunohistochemical markers except pancytokeratin. Forty one cases of sinonasal epithelial neoplasm consisting of 11 cases of SNUC and 10 cases each of high grade (grade 3 &4) esthesioneuroblastoma , undifferentiated nasopharyngeal carcinoma and poorly differentiated squamous cell carcinoma of the sinonasal region were analysed for morphology and immunoexpression of CK5/6 and p40.. It was found that SNUC did not exhibit immunohistochemical expression of p40 and CK 5/6, suggesting that these could be useful negative immune markers for diagnosis of SNUC. Key Words: p40, CK5/6, SNUC, Epithelial sinonasal malignancies
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Introduction
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Sinonasal undifferentiated carcinoma (SNUC) continues to be an enigma for histopathologists and clinicians, after nearly 3 decades of its first description by Frierson et al . It is believed to originate from schneiderian epithelium or from the nasal ectoderm of the paranasal sinuses.[1] World Health Organization defines SNUC as a rare, highly aggressive carcinoma that typically presents with locally extensive disease.[2] It is difficult to differentiate this tumor from other small blue round cell tumor of the sinonasal tract purely on morphological grounds. The differential diagnosis of SNUC is wide and includes poorly differentiated squamous cell carcinoma(PDSCC), undifferentiated nasopharyngeal carcinoma(NPC), high grade olfactory neuroblastoma (ONB), small cell undifferentiated neuroendocrine carcinoma (SCNEC), NUT- Midline carcinoma(NMC), poorly differentiated adenocarcinoma of salivary and non salivary type, mucosal malignant melanoma, nasal-type natural killer (NK)/T-cell lymphoma and rhabdomyosarcoma. In the present series we studied the morphology of prototype cases of SNUC. We also analyzed the utility of p40 and CK 5/6 as a negative markers of SNUC to differentiate it from the other epithelial tumors of the region. There has been no study about immunoexpression of p40 in Sino nasal undifferentiated carcinomas, till date. Material and Method Eleven consecutive cases of sinonasal undifferentiated carcinoma (SNUC)diagnosed over a period of 4 years (2008-12) were retrieved from histopathology archives of Department of Pathology, of our institute. The cases were reviewed and confirmed by 2 pathologist independently (LS & RR). Ten cases each of high grade (grade 3 &4) ONB , undifferentiated NPC and PDSCC of the sinonasal region were
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taken as controls. The above diagnoses were based on light microscopy and immunohistochemistry for pancytokeratin, CK 19, EMA, synaptophysin, chromogranin, CD 56, S-100 and EBV- LMP-1. A representative block containing the largest amount of viable tumor was selected in each case. Unstained sections from each of these blocks were submitted for determination of p-40 and CK 5/6 expression byimmunohistochemical analysis. Sections were incubated with the antibodies p40 (∆Np63) (polyclonal, dilution 1:400; Scy Tek laboratories, Lohan, Utah) and CK5/6 (monoclonal, dilution 1:100, Bio SB, Santa Barbara, CA) for 60 minutes at room temperature. The primary antibody was then detected by a secondary biotinylated antibody and a streptavidin-peroxidase conjugate. Immunostaining results for p-40 and CK5/6 were evaluated semiquantitatively according to the percentage of positive tumor cells, ie, less than 1%(score 0) 1% to 10%(score 1), 11% to 50%( score 2), and more than 50%( score 3) and whether immunoreactive tumor cells showed mild(score 1), moderate(score 2) or intense(score 3) immunostaining. The percentage positivity and intensity scores were added to get a total score which ranged from 0 to 6. To exclude equivocal reactions a total score of 3 or more was considered as a diagnostically relevant positive reaction. The positive immunostain was localized in cytoplasm and nucleus for anti-CK5/6 and p40 antibody respectively. Results
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In total we studied 41 cases of epithelial tumor with overlapping histological and immunohistochemical features. There were a total of 11 cases of SNUC with an age range of 14 to 72 years and a mean age of 46 years. There were 9 males and 2 females (M: F=4.5:1). Two of the cases in our study were localized to sphenoid sinus and nasopharynx, the rest were extensive and involved sinonasal tract. On light microscopy most of the cases showed presence of tumor cells in varying patterns including trabeculae, sheets,lobules and nests. The individual tumor cells were medium to large, round to oval with hyperchromatic nucleus, inconspicuous or prominent nucleoli and a varying amount of eosinophilic cytoplasm lacking syncytial quality.[Fig. 1a] In addition 1 of case of SNUC showed nuclear molding, absent nucleoli and focal spindling simulating a small cell neuroendocrine carcinoma morphologically.[Fig. 1b]None of the cases exhibited any evidence of keratinisation, gland formation, neurofibrillary matrix and neural rosettes. On immunohistochemistry the cases of SNUC showed positivity for pancytokeratin and variable positivity for NSE, CK19 and EMA. SNUC were consistently negative for synaptophysin, chromogranin, CD 56, S-100 and EBV- LMP-1. The immune expression of p40 and CK5/6 were observed in none of the SNUC or ONBs. all 10 cases of poorly differentiated squamous cell carcinomas of the sinonasal tract and 9 out of the 10 cases of nasopharyngeal carcinoma showed immunopositivity for CK5/6 and p40.[Fig.2 a, b, c, d] In the present study the staining was strong and diffuse in nasopharyngeal carcinoma cases and poorly differentiated squamous cell carcinomas of the sinonasal tract. We observed that none of the cases of ONB showed even focal positivity for p-40 or CK5/6 in compared to SNUC in which we did find a single case of focal weak expression of CK5/6 and p40, which was regarded negative(total score 2). No discordance was seen in results of CK5/6 and p40. (Table 1) Discussion
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SNUC is a high grade tumor with probable origin from schenederian epithelium or the nasal ectoderm which by definition lacks any glandular or squamous differentiation. [1] Evolution of symptoms is typically of short duration (weeks to months) with evidence of local invasion and destruction of orbital and/or cranial bones.[3] Only about 100 cases of SNUC has been reported in literature.[2] The largest series consisting of 36 cases was reported by Jeng et al. [4] This tumor shows a male preponderance with a wide age range.[2] The male female ratio was 4.5:1 and age range was 14 to 72 years in the present study. Thus SNUC should be considered in the differential of a malignant tumor in almost any age group. Morphology is the most precious tool for diagnosis(Table 2) but is insufficient alone to differentiate SNUC from other small blue round cell tumor occurring in sinonasal area especially where the material is limited and tumor is high grade. Microscopically, SNUC consist of sheets or large nests of medium-sized cells with high nucleo cytoplasmic ratio and often prominent nucleoli . The large cell nests often exhibit central, comedo like necrosis. Occasionally, SNUC grow in wide trabeculae or ribbons and may have a vaguely organoid pattern. Mitotic rate is usually high and vascular invasion is often noted. Focal tumor areas might show nuclear spindling and hyperchromatic nuclei with absent nucleoli ,similar to small cell neuroendocrine carcinoma.
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Hyam's grading system classifies ONB into 4 prognostically significant grades based on architecture,pleomorphism, neurofibrillary background, rosettes, mitosis, necrosis, gland formation and calcification.[5] In cases of grade 1 and 2 ONB where the prototype morphology in the form of fibrillary cell processes, vascular stroma, Homer Wright and Flexner wintersteiner rossetes and presence of ganglion cells are seen, diagnosis is straight forward. In grade 3 and 4 ONB differentiation from SNUC is difficult since the characteristic features of ONB are focal or absent. [Fig 3 a]]. Another problem is the occurrence of ONB with foci of adenocarcinoma, squamous cell carcinoma and even undifferentiated carcinoma. Thus one has to specifically look for the above mentioned components in each case of ONB and accordingly diagnose the case as ONB or collision tumor. [6][ Fig 3 b]
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The subtle microscopic clues for the diagnosis of poorly differentiated squamous carcinoma, may be identification of an in-situ carcinoma component and/or contiguity with the overlying surface epithelium. In addition presence of broad interconnecting or ramifying cords of neoplastic polygonal cells with distinct cell boundaries are features more commonly identified in SCC. [7] However the presence of overlying dysplasia does not excludes the diagnosis of SNUC as it has been documented in few cases of SNUC as well. [2]When the characteristic age group, clinical presentation, site, morphology EBV immunopositivity are present, the diagnosis of nasopharyngeal carcinoma is simple. The undifferentiated nasopharyngeal carcinoma is arranged in syncytium, trabeculae or irregular islands with variable amount of mature lymphoid component. Individual tumor cells have large prominent nucleoli. [8] Since SNUC might show prominent nucleoli and mild lymphoplasmacytic inflammatory infiltrate around the tumor islands, the differentiation is difficult purely on morphological grounds. Further complicating this matter are reports of EBV positivity in stray cases of SNUC especially in Asians. [9] Other tumors which need mention as a close morphological differential are NUT-midline carcinoma , small cell neuroendocrine carcinoma, high-grade of salivary and non salivary type. The NMC is a relatively new entity morphologically simulating a poorly differentiated carcinoma. The focal squamous differentiation is reported in around 80% of NMC. [10] The definitive diagnosis of NMC
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requires demonstration of NUT rearrangement by dual color, split-apart FISH, or identification of a BRD4-NUT fusion transcript in RT-PCR.[11] A NUT antibody has become recently available which is reported to have a sensitivity of 87%, and a specificity of 100%. [12] A useful practical guideline is to consider NMC in the differential diagnosis of any midline poorly differentiated carcinoma in a nonsmoker, in the absence of EBV or HPV immunopositivity.[1]
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Poorly differentiated subtype of sinonasal intestinal type adenocarcinoma(Barnes solid type, PTCC III) [13,14] which occur in solid sheets and are cytologically high grade can be a close differential of SNUC. The histomorphological clues like focal tubule formation and mucinous background are pivotal in clinching the diagnosis. A positive staining for Ber-EP4, B72.3 and Leu M-1 may help in difficult cases. [15]
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Adenoid cystic carcinoma is the commonest salivary gland type adenocarcinoma occuring in sinonasal region though the whole gamut of salivary gland tumors can occur in this region. As the dedifferentiated variants of salivary type adenocarcinoma may be a close differential of SNUC, a meticulous search for areas with classical morphology may provide answer in challenging cases.
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Small cell neuroendocrine carcinoma(SCNEC) are composed of small to intermediate sized cells with oval or round hyperchromatic nuclei and absent nucleoli. The nuclei show molding. The tumor cells are arranged in sheets, nests or trabeculae and have extensive necrosis. Crush artifact and a high mitotic rate are common. Morphological similarities exist between SCNET and other round cell neoplasms of the sinonasal tract. The Immunohistochemical profile separates out the SCNET from the rest of the group but differentiating from ONB is challenging. In this regards calcitonin and neurofilament could be of some use. [16]
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Sinonasal undifferentiated carcinoma cannot be distinguished from other tumors of this region (with the possible exception of melanoma) on the basis of imaging features. [17]The problem is compounded in patients who present late with large lesions.
The importance of meticulous light microscopic examination cannot be understated however the use of immunohistochemistry is mandatory in arriving at the accurate diagnosis. The specific lineage markers like LCA, CD3, CD 20,desmin, myogenin, myo D1, HMB-45 and Melan A help differentiate SNUC from lymphoma , rhabdomyosarcoma, and melanoma .As for sinonasal epithelial tumors the problem is compounded by overlapping morphological and immunohistochemical features.The most consistent immunohistochemical marker in SNUC is pancytokeratin, however, one third of ONB show focal immunopositivity for pancytokeratin. [18] About 50% cases of SNUC show positivity for EMA, NSE and p53. [19] Studies have been done on p16 and HPV in sinonasal tumors and HPV DNA have been quoted to be present in a minority of cases of SNUC while p16 have been found to be expressed more frequently. [20, 21] As there is paucity of positive markers we studied the utility of p40 and CK5/6 as negative markers to differentiate SNUC from other epithelial tumors of the sinonasal area. Cytokeratin 5/6 is a commercially available monoclonal antibody which recognizes basal cell–type high-molecular weight cytokeratins and is a sensitive, although not entirely specific, marker associated with a squamous differentiation in carcinomas.
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[22]This immnuhistochemical marker has beenstudied in epithelial tumors of the sinonasal region by some authors. Franchi et al differentiated keratinizing SCC, nonkeratizing SCC and SNUC on basis of differential cytokeratin staining patterns . In this study SNUC, was characterized by the expression of cytokeratins of simple epithelia, such as CK8, CK7, and CK19 in contrast both keratinizing and nonkeratinizing forms of SCC showed preferential expression of high molecular weight cytokeratin including CK5/6. [23] In this study none of their cases of SNUC showed positivity for CK5/6. We found similar results in our study where all the cases of SNUC and esthesioneuroblastoma were negative for CK5/6. The cases of poorly differentiated squamous cell carcinoma and nasopharyngeal carcinoma were consistently positive for CK5/6 except a single case of nasopharyngeal carcinoma.The antibody p40 recognizes ∆Np63 exclusively, which is an isoform of p63 and is more specific than p63 in the diagnosis of squamous tumors.[24] Bourne et al studied the expression of p63 in various tumors of the sinonasal tract. A positive immunostaining for p63 was identified in 20% and 36 % of sinonasal undifferentiated carcinoma and esthesioneuroblastoma respectively. The p63 positivity of both the above tumors was weak and focal as compared to nasopharyngeal carcinoma where it was strong and diffusely positive. [25] No study has been done till date to depict p40 as a negative marker in SNUC. In the present study we immunostained SNUC along with cases of high grade esthesioneuroblastoma , undiffferentiated nasopharyngeal carcinoma and poorly differentiated squamous cell carcinoma of the sinonasal region. As brought out in our results none of the SNUC was positive for either CK 5/6 orp40. In comparison the undifferentiated nasopharyngeal carcinoma and poorlydifferentiated squamous cell carcinoma both exhibited for CK 5/6 an p40 in 90% an 100% cases respectively. None of the ONB was positive for either of studied markers. Our results were in concordance with that of Bourne et al who studied p63 and with Franchi et al who studied CK 5/6.
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Conclusion
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SNUC is an epithelial tumor of the sinonasal region which is not supposed to show a squamous or glandular differention and thus is a diagnosis of exclusion.Hence we need to substantiate our diagnosis of SNUC by an extensive immunohistochemical panel of lineage specific and lineage associated markers . We found that p40 and CK5/6 are robust and dependable negative markers to establish the diagnosis of SNUC with reasonable accuracy.
References: 1. Frierson HF Jr, Mills SE, Fechner RE, Taxy JB, Levine PA. Sinonasal undifferentiated carcinoma. An aggressive neoplasm derived from schneiderian epithelium and distinct from olfactory neuroblastoma. Am J Surg Pathol. 1986 Nov;10(11):771-9. 2. H.F. Frierson, Jr. Sinonasal undifferentiated carcinoma. In : Leon Barnes,John W. Eveson, Peter Reichart David Sidransky, editors. Pathology and Genetics of Head and Neck Tumours. IARCPress Lyon, 2005;19.
ACCEPTED MANUSCRIPT 3. Ejaz A, Wenig BM. Sinonasal undifferentiated carcinoma. Clinical and pathologic features and a discussion on classification, cellular differentiation, and differential diagnosis. Adv Anat Pathol. 2005;12:134-143.
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4. Jeng YM, Sung MT, Fang CL, Huang HY, Mao TL, Cheng W, Hsiao CH. Sinonasalundifferentiated carcinoma and nasopharyngeal-type undifferentiated carcinoma:two clinically, biologically, and histopathologically distinct entities. Am J Surg Pathol. 2002 Mar;26(3):371-6.
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5. Hyams V J 1982 olfactory neuroblastoma(case 6).In: Batsakis J G, Hyam VJ, Morales A R, editors. Special tumors of head and neck. ACSP Press, Chicago, p 24-29
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6. Miller DC, Goodman ML, Pilch BZ, Shi SR, Dickersin GR, Halpern H, Norris CM Jr. Mixed olfactory neuroblastoma and carcinoma. A report of two cases. Cancer. 1984 Nov 1;54(9):2019-28.
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7. Bridge JA, Bowen JM, Smith RB. The small round blue cell tumors of the sinonasal area. Head Neck Pathol. 2010 Mar;4(1):84-93. 8. Iezzoni JC, Mills SE. "Undifferentiated" small round cell tumors of the sinonasal tract: differential diagnosis update. Am J Clin Pathol. 2005 Dec;124 Suppl:S110-21.
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9. Gallo O, Di Lollo S, Graziani P, Gallina E, Baroni G. Detection of Epstein-Barr virus genome in sinonasal undifferentiated carcinoma by use of in situ hybridization. Otolaryngol Head Neck Surg. 1995 Jun;112(6):659-64.
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10 French CA, Kutok JL, Faquin WC, Toretsky JA, Antonescu CR, Griffin CA, Nose V, Vargas SO, Moschovi M, Tzortzatou-Stathopoulou F, Miyoshi I, Perez-Atayde AR, Aster JC, Fletcher JA. Midline carcinoma of children and young adults with NUT rearrangement. J Clin Oncol. 2004 Oct;22(20):4135-9.
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11. Engleson J, Soller M, Panagopoulos I, Dahlen A, Dictor M, Jerkeman M. Midline carcinoma with t(15;19) and BRD4-NUT fusion oncogene in a 30-year-old female with response to docetaxel and radiotherapy. BMC cancer 2006;6:69. 12. Haack H, Johnson LA, Fry CJ, Crosby K, Polakiewicz RD, Stelow EB, Hong SM, Schwartz BE, Cameron MJ, Rubin MA, Chang MC, Aster JC, French CA. Diagnosis of NUT midline carcinoma using a NUT-specific monoclonal antibody. Am J Surg Pathol.2009 Jul;33(7):984-91.13. 13. Barnes L. Intestinal-type adenocarcinoma of the nasal cavity and paranasal sinuses. Am J Surg Pathol. 1986 Mar;10(3):192-202. 14. Kleinsasser O, Schroeder HG. Adenocarcinomas of the inner nose after exposure to wood dust. Morphological findings and relationships between histopathology and clinical behavior in 79 cases. Arch Otorhinolaryngol. 1988;245(1):1-15. 15. McKinney CD, Mills SE, Franquemont DW. Sinonasal intestinal-type adenocarcinoma: immunohistochemical profile and comparison with colonic adenocarcinoma. Mod Pathol. 1995 May;8(4):421-6.
ACCEPTED MANUSCRIPT 16. Bayardo Perez-Ordonez, Salvatore M Caruana, Andrew G Huvos. Small cell neuroendocrine carcinoma of the nasal cavity and paranasal sinuses. Human Pathology Volume 29, Issue 8, August 1998, 826–832
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17. Mohammed A. Gomaa, Moustafa S. Hammad, Abobakr Abdelmoghny, Ashraf M. Elsherif, Heba M. Tawfik. Magnetic Resonance Imaging Versus Computed Tomography and Different Imaging Modalities in Evaluation of Sinonasal Neoplasms Diagnosed by Histopathology Clin Med Insights Ear Nose Throat. 2013; 6: 9–15.
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18. Frierson HF Jr, Ross GW, Mills SE, Frankfurter A. Olfactory neuroblastoma. Additional immunohistochemical characterization. Am J Clin Pathol. 1990 Nov;94(5):547-53.
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19. Cerilli LA, Holst VA, Brandwein MS, Stoler MH, Mills SE. Sinonasal undifferentiated carcinoma: immunohistochemical profile and lack of EBV association. Am J Surg Pathol. 2001 Feb;25(2):156-63.
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20. Beth Wadsworth, Jeffery M. Bumpous, Alvin W. Martin, Michael R. Nowacki, Alfred B. Jenson, Hanan Farghaly Expression of p16 in Sinonasal Undifferentiated Carcinoma (SNUC) Without Associated Human Papillomavirus (HPV) Head Neck Pathol. 2011 December; 5(4): 349–354.
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21. El-Mofty SK, Lu DW. Prevalence of high-risk human papillomavirus DNA in nonkeratinizing (cylindrical cell) carcinoma of the sinonasal tract: a distinct clinicopathologic and molecular disease entity. Am J Surg Pathol. 2005 Oct;29(10):1367-72.
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22. Moll R, Franke WW, Schiller DL, Geiger B, Krepler R. The catalog of human cytokeratins: patterns of expression in normal epithelia, tumors and cultured cells. Cell. 1982 Nov;31(1):11-24.
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23. Franchi A, Moroni M, Massi D, Paglierani M, Santucci M. Sinonasal undifferentiated carcinoma, nasopharyngeal-type undifferentiated carcinoma, and keratinizing and nonkeratinizing squamous cell carcinoma express different cytokeratin patterns. Am J Surg Pathol. 2002 Dec;26(12):1597-604. 24. Hibi K, Trink B, Patturajan M, Westra WH, Caballero OL, Hill DE, Ratovitski EA, Jen J, Sidransky D. AIS is an oncogene amplified in squamous cell carcinoma.Proc Natl Acad Sci U S A. 2000 May 9;97(10):54627. 25. Bourne TD, Bellizzi AM, Stelow EB, Loy AH, Levine PA, Wick MR, Mills SE. p63 Expression in olfactory neuroblastoma and other small cell tumors of the sinonasal tract. Am J Clin Pathol. 2008 Aug;130(2):213-8.
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Figure 1
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Figure 2
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Figure 3
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CK5/6
- (focal weak staining in 1 case) - (focal weak staining in 1 case)
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+(9/10)
+
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p40
PDSCC
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Undifferentiated NPC +(9/10)
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NMC
Nests
Islands, broad interconnectin g cords
Cellular characteristics
Medium sized, round cells
Nucleus
Small round cells
Medium sized cells with indistinct cell membrane
Polygonal cells with distinct boundary
Round, vesicular
Salt and pepper chromatin
Round, vesicular
Nucleoli
Prominent
Large and prominent
Mitosis
Frequent
Absent to conspicuous (depending on grade) Variable
Necrosis
Prominent
Variable
Variable
Inflammation
Absent to Scant
Absent
Other Features
---
IHC
PanCK,p63 +/-, NSE+/-
Fibrillary stroma, rossetes, vascular stroma, sustentacula r cells NSE, CD56, S100*, Syn+/-, CG+/-
Prominent mature lymphoplasmacyti c infiltrate ---
Specific genetic alteration
poorly differentiated carcinoma, squamous differentiation+ /Monomorphic
Sheets, occasional gland
Sheets, nests or trabeculae
variable
Round, vesicular
*
Round
Conspicuous
*
Prominent
Small round cells with moulding Round , artifectual spindling Absent
Frequent Frequent
Absent
* * *
Overlying epithelial dysplasia / in situ component
*
Glands, tubules, Azzopardy mucin effect
CK, p63,EBER,
Pan CK, p16+/-
P63, anti-NUT
EMA,B72.3,Be PanCK,NSE, r-EP4, CDX-2*, CG,SYN,CD5 6
Frequent
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Sheets, Lobules
SNEC
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PDSCC
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Architecture
Poorly differentiated adenocarcinoma
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Undifferentiated NPC Syncytium, irregular islands
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ONB
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SNUC
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Characteristics
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Table 2
Frequent Frequent
NUT-BRD rearrangement
Brisk Frequent Variable
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Figure 1a. SNUC showing large nests of medium-sized cells with high nucleo cytoplasmic ratio and often prominent nucleoli.
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Figure 1b. Tumor showing nuclear molding,hyperchromatic nuclei with absent nucleoli and focal spindling simulating a small cell neuroendocrine carcinoma.
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Figure 2a&b Positive immunostaining for p40 and CK5/6 in Poorly differentiated squamous cell carcinoma. Figure 2 c&d Positive immunostaining for p40 and CK5/6 in nasopharyngeal carcinoma.
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Figure 3a. Olfactory neuroblastoma grade 3, the tumor lacks neurofibrillary matrix and includes malignant round cells with increased mitotic activity. Figure 3b. Olfactory neuroblastoma with areas of squamous differentiation
S1092-9134(14)00004-5 doi: 10.1016/j.anndiagpath.2014.01.003 YADPA 50911
To appear in:
Annals of Diagnostic Pathology
Received date: Revised date: Accepted date:
18 August 2013 22 November 2013 5 January 2014
Please cite this article as: Singh Lavleen, Ranjan Richa, Arava Sudheer, Singh MK, Role of p40 and Cytokeratin 5/6 in the Differential Diagnosis of Sinonasal Undifferentiated Carcinoma, Annals of Diagnostic Pathology (2014), doi: 10.1016/j.anndiagpath.2014.01.003
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT Role of p40 and Cytokeratin 5/6 in the Differential Diagnosis of Sinonasal Undifferentiated Carcinoma
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Lavleen Singh M.D, Richa Ranjan M.D, Sudheer Arava M.D, M K Singh M.D Department of Pathology, All India Institute of Medical Sciences, New Delhi-110029 Abstract
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Sinonasal undifferentiated carcinoma is an epithelial neoplasm of sinonasal region which does not exhibit a squamous or glandular differentiation. The challenge in diagnosis of this entity is the rarity of the disease, the varying morphology of the tumor which leads to gamut of differential diagnosis and the paucity of consistent immunohistochemical markers except pancytokeratin. Forty one cases of sinonasal epithelial neoplasm consisting of 11 cases of SNUC and 10 cases each of high grade (grade 3 &4) esthesioneuroblastoma , undifferentiated nasopharyngeal carcinoma and poorly differentiated squamous cell carcinoma of the sinonasal region were analysed for morphology and immunoexpression of CK5/6 and p40.. It was found that SNUC did not exhibit immunohistochemical expression of p40 and CK 5/6, suggesting that these could be useful negative immune markers for diagnosis of SNUC. Key Words: p40, CK5/6, SNUC, Epithelial sinonasal malignancies
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Introduction
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Sinonasal undifferentiated carcinoma (SNUC) continues to be an enigma for histopathologists and clinicians, after nearly 3 decades of its first description by Frierson et al . It is believed to originate from schneiderian epithelium or from the nasal ectoderm of the paranasal sinuses.[1] World Health Organization defines SNUC as a rare, highly aggressive carcinoma that typically presents with locally extensive disease.[2] It is difficult to differentiate this tumor from other small blue round cell tumor of the sinonasal tract purely on morphological grounds. The differential diagnosis of SNUC is wide and includes poorly differentiated squamous cell carcinoma(PDSCC), undifferentiated nasopharyngeal carcinoma(NPC), high grade olfactory neuroblastoma (ONB), small cell undifferentiated neuroendocrine carcinoma (SCNEC), NUT- Midline carcinoma(NMC), poorly differentiated adenocarcinoma of salivary and non salivary type, mucosal malignant melanoma, nasal-type natural killer (NK)/T-cell lymphoma and rhabdomyosarcoma. In the present series we studied the morphology of prototype cases of SNUC. We also analyzed the utility of p40 and CK 5/6 as a negative markers of SNUC to differentiate it from the other epithelial tumors of the region. There has been no study about immunoexpression of p40 in Sino nasal undifferentiated carcinomas, till date. Material and Method Eleven consecutive cases of sinonasal undifferentiated carcinoma (SNUC)diagnosed over a period of 4 years (2008-12) were retrieved from histopathology archives of Department of Pathology, of our institute. The cases were reviewed and confirmed by 2 pathologist independently (LS & RR). Ten cases each of high grade (grade 3 &4) ONB , undifferentiated NPC and PDSCC of the sinonasal region were
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taken as controls. The above diagnoses were based on light microscopy and immunohistochemistry for pancytokeratin, CK 19, EMA, synaptophysin, chromogranin, CD 56, S-100 and EBV- LMP-1. A representative block containing the largest amount of viable tumor was selected in each case. Unstained sections from each of these blocks were submitted for determination of p-40 and CK 5/6 expression byimmunohistochemical analysis. Sections were incubated with the antibodies p40 (∆Np63) (polyclonal, dilution 1:400; Scy Tek laboratories, Lohan, Utah) and CK5/6 (monoclonal, dilution 1:100, Bio SB, Santa Barbara, CA) for 60 minutes at room temperature. The primary antibody was then detected by a secondary biotinylated antibody and a streptavidin-peroxidase conjugate. Immunostaining results for p-40 and CK5/6 were evaluated semiquantitatively according to the percentage of positive tumor cells, ie, less than 1%(score 0) 1% to 10%(score 1), 11% to 50%( score 2), and more than 50%( score 3) and whether immunoreactive tumor cells showed mild(score 1), moderate(score 2) or intense(score 3) immunostaining. The percentage positivity and intensity scores were added to get a total score which ranged from 0 to 6. To exclude equivocal reactions a total score of 3 or more was considered as a diagnostically relevant positive reaction. The positive immunostain was localized in cytoplasm and nucleus for anti-CK5/6 and p40 antibody respectively. Results
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In total we studied 41 cases of epithelial tumor with overlapping histological and immunohistochemical features. There were a total of 11 cases of SNUC with an age range of 14 to 72 years and a mean age of 46 years. There were 9 males and 2 females (M: F=4.5:1). Two of the cases in our study were localized to sphenoid sinus and nasopharynx, the rest were extensive and involved sinonasal tract. On light microscopy most of the cases showed presence of tumor cells in varying patterns including trabeculae, sheets,lobules and nests. The individual tumor cells were medium to large, round to oval with hyperchromatic nucleus, inconspicuous or prominent nucleoli and a varying amount of eosinophilic cytoplasm lacking syncytial quality.[Fig. 1a] In addition 1 of case of SNUC showed nuclear molding, absent nucleoli and focal spindling simulating a small cell neuroendocrine carcinoma morphologically.[Fig. 1b]None of the cases exhibited any evidence of keratinisation, gland formation, neurofibrillary matrix and neural rosettes. On immunohistochemistry the cases of SNUC showed positivity for pancytokeratin and variable positivity for NSE, CK19 and EMA. SNUC were consistently negative for synaptophysin, chromogranin, CD 56, S-100 and EBV- LMP-1. The immune expression of p40 and CK5/6 were observed in none of the SNUC or ONBs. all 10 cases of poorly differentiated squamous cell carcinomas of the sinonasal tract and 9 out of the 10 cases of nasopharyngeal carcinoma showed immunopositivity for CK5/6 and p40.[Fig.2 a, b, c, d] In the present study the staining was strong and diffuse in nasopharyngeal carcinoma cases and poorly differentiated squamous cell carcinomas of the sinonasal tract. We observed that none of the cases of ONB showed even focal positivity for p-40 or CK5/6 in compared to SNUC in which we did find a single case of focal weak expression of CK5/6 and p40, which was regarded negative(total score 2). No discordance was seen in results of CK5/6 and p40. (Table 1) Discussion
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SNUC is a high grade tumor with probable origin from schenederian epithelium or the nasal ectoderm which by definition lacks any glandular or squamous differentiation. [1] Evolution of symptoms is typically of short duration (weeks to months) with evidence of local invasion and destruction of orbital and/or cranial bones.[3] Only about 100 cases of SNUC has been reported in literature.[2] The largest series consisting of 36 cases was reported by Jeng et al. [4] This tumor shows a male preponderance with a wide age range.[2] The male female ratio was 4.5:1 and age range was 14 to 72 years in the present study. Thus SNUC should be considered in the differential of a malignant tumor in almost any age group. Morphology is the most precious tool for diagnosis(Table 2) but is insufficient alone to differentiate SNUC from other small blue round cell tumor occurring in sinonasal area especially where the material is limited and tumor is high grade. Microscopically, SNUC consist of sheets or large nests of medium-sized cells with high nucleo cytoplasmic ratio and often prominent nucleoli . The large cell nests often exhibit central, comedo like necrosis. Occasionally, SNUC grow in wide trabeculae or ribbons and may have a vaguely organoid pattern. Mitotic rate is usually high and vascular invasion is often noted. Focal tumor areas might show nuclear spindling and hyperchromatic nuclei with absent nucleoli ,similar to small cell neuroendocrine carcinoma.
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Hyam's grading system classifies ONB into 4 prognostically significant grades based on architecture,pleomorphism, neurofibrillary background, rosettes, mitosis, necrosis, gland formation and calcification.[5] In cases of grade 1 and 2 ONB where the prototype morphology in the form of fibrillary cell processes, vascular stroma, Homer Wright and Flexner wintersteiner rossetes and presence of ganglion cells are seen, diagnosis is straight forward. In grade 3 and 4 ONB differentiation from SNUC is difficult since the characteristic features of ONB are focal or absent. [Fig 3 a]]. Another problem is the occurrence of ONB with foci of adenocarcinoma, squamous cell carcinoma and even undifferentiated carcinoma. Thus one has to specifically look for the above mentioned components in each case of ONB and accordingly diagnose the case as ONB or collision tumor. [6][ Fig 3 b]
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The subtle microscopic clues for the diagnosis of poorly differentiated squamous carcinoma, may be identification of an in-situ carcinoma component and/or contiguity with the overlying surface epithelium. In addition presence of broad interconnecting or ramifying cords of neoplastic polygonal cells with distinct cell boundaries are features more commonly identified in SCC. [7] However the presence of overlying dysplasia does not excludes the diagnosis of SNUC as it has been documented in few cases of SNUC as well. [2]When the characteristic age group, clinical presentation, site, morphology EBV immunopositivity are present, the diagnosis of nasopharyngeal carcinoma is simple. The undifferentiated nasopharyngeal carcinoma is arranged in syncytium, trabeculae or irregular islands with variable amount of mature lymphoid component. Individual tumor cells have large prominent nucleoli. [8] Since SNUC might show prominent nucleoli and mild lymphoplasmacytic inflammatory infiltrate around the tumor islands, the differentiation is difficult purely on morphological grounds. Further complicating this matter are reports of EBV positivity in stray cases of SNUC especially in Asians. [9] Other tumors which need mention as a close morphological differential are NUT-midline carcinoma , small cell neuroendocrine carcinoma, high-grade of salivary and non salivary type. The NMC is a relatively new entity morphologically simulating a poorly differentiated carcinoma. The focal squamous differentiation is reported in around 80% of NMC. [10] The definitive diagnosis of NMC
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requires demonstration of NUT rearrangement by dual color, split-apart FISH, or identification of a BRD4-NUT fusion transcript in RT-PCR.[11] A NUT antibody has become recently available which is reported to have a sensitivity of 87%, and a specificity of 100%. [12] A useful practical guideline is to consider NMC in the differential diagnosis of any midline poorly differentiated carcinoma in a nonsmoker, in the absence of EBV or HPV immunopositivity.[1]
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Poorly differentiated subtype of sinonasal intestinal type adenocarcinoma(Barnes solid type, PTCC III) [13,14] which occur in solid sheets and are cytologically high grade can be a close differential of SNUC. The histomorphological clues like focal tubule formation and mucinous background are pivotal in clinching the diagnosis. A positive staining for Ber-EP4, B72.3 and Leu M-1 may help in difficult cases. [15]
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Adenoid cystic carcinoma is the commonest salivary gland type adenocarcinoma occuring in sinonasal region though the whole gamut of salivary gland tumors can occur in this region. As the dedifferentiated variants of salivary type adenocarcinoma may be a close differential of SNUC, a meticulous search for areas with classical morphology may provide answer in challenging cases.
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Small cell neuroendocrine carcinoma(SCNEC) are composed of small to intermediate sized cells with oval or round hyperchromatic nuclei and absent nucleoli. The nuclei show molding. The tumor cells are arranged in sheets, nests or trabeculae and have extensive necrosis. Crush artifact and a high mitotic rate are common. Morphological similarities exist between SCNET and other round cell neoplasms of the sinonasal tract. The Immunohistochemical profile separates out the SCNET from the rest of the group but differentiating from ONB is challenging. In this regards calcitonin and neurofilament could be of some use. [16]
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Sinonasal undifferentiated carcinoma cannot be distinguished from other tumors of this region (with the possible exception of melanoma) on the basis of imaging features. [17]The problem is compounded in patients who present late with large lesions.
The importance of meticulous light microscopic examination cannot be understated however the use of immunohistochemistry is mandatory in arriving at the accurate diagnosis. The specific lineage markers like LCA, CD3, CD 20,desmin, myogenin, myo D1, HMB-45 and Melan A help differentiate SNUC from lymphoma , rhabdomyosarcoma, and melanoma .As for sinonasal epithelial tumors the problem is compounded by overlapping morphological and immunohistochemical features.The most consistent immunohistochemical marker in SNUC is pancytokeratin, however, one third of ONB show focal immunopositivity for pancytokeratin. [18] About 50% cases of SNUC show positivity for EMA, NSE and p53. [19] Studies have been done on p16 and HPV in sinonasal tumors and HPV DNA have been quoted to be present in a minority of cases of SNUC while p16 have been found to be expressed more frequently. [20, 21] As there is paucity of positive markers we studied the utility of p40 and CK5/6 as negative markers to differentiate SNUC from other epithelial tumors of the sinonasal area. Cytokeratin 5/6 is a commercially available monoclonal antibody which recognizes basal cell–type high-molecular weight cytokeratins and is a sensitive, although not entirely specific, marker associated with a squamous differentiation in carcinomas.
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[22]This immnuhistochemical marker has beenstudied in epithelial tumors of the sinonasal region by some authors. Franchi et al differentiated keratinizing SCC, nonkeratizing SCC and SNUC on basis of differential cytokeratin staining patterns . In this study SNUC, was characterized by the expression of cytokeratins of simple epithelia, such as CK8, CK7, and CK19 in contrast both keratinizing and nonkeratinizing forms of SCC showed preferential expression of high molecular weight cytokeratin including CK5/6. [23] In this study none of their cases of SNUC showed positivity for CK5/6. We found similar results in our study where all the cases of SNUC and esthesioneuroblastoma were negative for CK5/6. The cases of poorly differentiated squamous cell carcinoma and nasopharyngeal carcinoma were consistently positive for CK5/6 except a single case of nasopharyngeal carcinoma.The antibody p40 recognizes ∆Np63 exclusively, which is an isoform of p63 and is more specific than p63 in the diagnosis of squamous tumors.[24] Bourne et al studied the expression of p63 in various tumors of the sinonasal tract. A positive immunostaining for p63 was identified in 20% and 36 % of sinonasal undifferentiated carcinoma and esthesioneuroblastoma respectively. The p63 positivity of both the above tumors was weak and focal as compared to nasopharyngeal carcinoma where it was strong and diffusely positive. [25] No study has been done till date to depict p40 as a negative marker in SNUC. In the present study we immunostained SNUC along with cases of high grade esthesioneuroblastoma , undiffferentiated nasopharyngeal carcinoma and poorly differentiated squamous cell carcinoma of the sinonasal region. As brought out in our results none of the SNUC was positive for either CK 5/6 orp40. In comparison the undifferentiated nasopharyngeal carcinoma and poorlydifferentiated squamous cell carcinoma both exhibited for CK 5/6 an p40 in 90% an 100% cases respectively. None of the ONB was positive for either of studied markers. Our results were in concordance with that of Bourne et al who studied p63 and with Franchi et al who studied CK 5/6.
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SNUC is an epithelial tumor of the sinonasal region which is not supposed to show a squamous or glandular differention and thus is a diagnosis of exclusion.Hence we need to substantiate our diagnosis of SNUC by an extensive immunohistochemical panel of lineage specific and lineage associated markers . We found that p40 and CK5/6 are robust and dependable negative markers to establish the diagnosis of SNUC with reasonable accuracy.
References: 1. Frierson HF Jr, Mills SE, Fechner RE, Taxy JB, Levine PA. Sinonasal undifferentiated carcinoma. An aggressive neoplasm derived from schneiderian epithelium and distinct from olfactory neuroblastoma. Am J Surg Pathol. 1986 Nov;10(11):771-9. 2. H.F. Frierson, Jr. Sinonasal undifferentiated carcinoma. In : Leon Barnes,John W. Eveson, Peter Reichart David Sidransky, editors. Pathology and Genetics of Head and Neck Tumours. IARCPress Lyon, 2005;19.
ACCEPTED MANUSCRIPT 3. Ejaz A, Wenig BM. Sinonasal undifferentiated carcinoma. Clinical and pathologic features and a discussion on classification, cellular differentiation, and differential diagnosis. Adv Anat Pathol. 2005;12:134-143.
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4. Jeng YM, Sung MT, Fang CL, Huang HY, Mao TL, Cheng W, Hsiao CH. Sinonasalundifferentiated carcinoma and nasopharyngeal-type undifferentiated carcinoma:two clinically, biologically, and histopathologically distinct entities. Am J Surg Pathol. 2002 Mar;26(3):371-6.
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5. Hyams V J 1982 olfactory neuroblastoma(case 6).In: Batsakis J G, Hyam VJ, Morales A R, editors. Special tumors of head and neck. ACSP Press, Chicago, p 24-29
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6. Miller DC, Goodman ML, Pilch BZ, Shi SR, Dickersin GR, Halpern H, Norris CM Jr. Mixed olfactory neuroblastoma and carcinoma. A report of two cases. Cancer. 1984 Nov 1;54(9):2019-28.
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7. Bridge JA, Bowen JM, Smith RB. The small round blue cell tumors of the sinonasal area. Head Neck Pathol. 2010 Mar;4(1):84-93. 8. Iezzoni JC, Mills SE. "Undifferentiated" small round cell tumors of the sinonasal tract: differential diagnosis update. Am J Clin Pathol. 2005 Dec;124 Suppl:S110-21.
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9. Gallo O, Di Lollo S, Graziani P, Gallina E, Baroni G. Detection of Epstein-Barr virus genome in sinonasal undifferentiated carcinoma by use of in situ hybridization. Otolaryngol Head Neck Surg. 1995 Jun;112(6):659-64.
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10 French CA, Kutok JL, Faquin WC, Toretsky JA, Antonescu CR, Griffin CA, Nose V, Vargas SO, Moschovi M, Tzortzatou-Stathopoulou F, Miyoshi I, Perez-Atayde AR, Aster JC, Fletcher JA. Midline carcinoma of children and young adults with NUT rearrangement. J Clin Oncol. 2004 Oct;22(20):4135-9.
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11. Engleson J, Soller M, Panagopoulos I, Dahlen A, Dictor M, Jerkeman M. Midline carcinoma with t(15;19) and BRD4-NUT fusion oncogene in a 30-year-old female with response to docetaxel and radiotherapy. BMC cancer 2006;6:69. 12. Haack H, Johnson LA, Fry CJ, Crosby K, Polakiewicz RD, Stelow EB, Hong SM, Schwartz BE, Cameron MJ, Rubin MA, Chang MC, Aster JC, French CA. Diagnosis of NUT midline carcinoma using a NUT-specific monoclonal antibody. Am J Surg Pathol.2009 Jul;33(7):984-91.13. 13. Barnes L. Intestinal-type adenocarcinoma of the nasal cavity and paranasal sinuses. Am J Surg Pathol. 1986 Mar;10(3):192-202. 14. Kleinsasser O, Schroeder HG. Adenocarcinomas of the inner nose after exposure to wood dust. Morphological findings and relationships between histopathology and clinical behavior in 79 cases. Arch Otorhinolaryngol. 1988;245(1):1-15. 15. McKinney CD, Mills SE, Franquemont DW. Sinonasal intestinal-type adenocarcinoma: immunohistochemical profile and comparison with colonic adenocarcinoma. Mod Pathol. 1995 May;8(4):421-6.
ACCEPTED MANUSCRIPT 16. Bayardo Perez-Ordonez, Salvatore M Caruana, Andrew G Huvos. Small cell neuroendocrine carcinoma of the nasal cavity and paranasal sinuses. Human Pathology Volume 29, Issue 8, August 1998, 826–832
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17. Mohammed A. Gomaa, Moustafa S. Hammad, Abobakr Abdelmoghny, Ashraf M. Elsherif, Heba M. Tawfik. Magnetic Resonance Imaging Versus Computed Tomography and Different Imaging Modalities in Evaluation of Sinonasal Neoplasms Diagnosed by Histopathology Clin Med Insights Ear Nose Throat. 2013; 6: 9–15.
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18. Frierson HF Jr, Ross GW, Mills SE, Frankfurter A. Olfactory neuroblastoma. Additional immunohistochemical characterization. Am J Clin Pathol. 1990 Nov;94(5):547-53.
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19. Cerilli LA, Holst VA, Brandwein MS, Stoler MH, Mills SE. Sinonasal undifferentiated carcinoma: immunohistochemical profile and lack of EBV association. Am J Surg Pathol. 2001 Feb;25(2):156-63.
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20. Beth Wadsworth, Jeffery M. Bumpous, Alvin W. Martin, Michael R. Nowacki, Alfred B. Jenson, Hanan Farghaly Expression of p16 in Sinonasal Undifferentiated Carcinoma (SNUC) Without Associated Human Papillomavirus (HPV) Head Neck Pathol. 2011 December; 5(4): 349–354.
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21. El-Mofty SK, Lu DW. Prevalence of high-risk human papillomavirus DNA in nonkeratinizing (cylindrical cell) carcinoma of the sinonasal tract: a distinct clinicopathologic and molecular disease entity. Am J Surg Pathol. 2005 Oct;29(10):1367-72.
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22. Moll R, Franke WW, Schiller DL, Geiger B, Krepler R. The catalog of human cytokeratins: patterns of expression in normal epithelia, tumors and cultured cells. Cell. 1982 Nov;31(1):11-24.
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23. Franchi A, Moroni M, Massi D, Paglierani M, Santucci M. Sinonasal undifferentiated carcinoma, nasopharyngeal-type undifferentiated carcinoma, and keratinizing and nonkeratinizing squamous cell carcinoma express different cytokeratin patterns. Am J Surg Pathol. 2002 Dec;26(12):1597-604. 24. Hibi K, Trink B, Patturajan M, Westra WH, Caballero OL, Hill DE, Ratovitski EA, Jen J, Sidransky D. AIS is an oncogene amplified in squamous cell carcinoma.Proc Natl Acad Sci U S A. 2000 May 9;97(10):54627. 25. Bourne TD, Bellizzi AM, Stelow EB, Loy AH, Levine PA, Wick MR, Mills SE. p63 Expression in olfactory neuroblastoma and other small cell tumors of the sinonasal tract. Am J Clin Pathol. 2008 Aug;130(2):213-8.
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Figure 1
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Figure 2
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Figure 3
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CK5/6
- (focal weak staining in 1 case) - (focal weak staining in 1 case)
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p40
PDSCC
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Undifferentiated NPC +(9/10)
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NMC
Nests
Islands, broad interconnectin g cords
Cellular characteristics
Medium sized, round cells
Nucleus
Small round cells
Medium sized cells with indistinct cell membrane
Polygonal cells with distinct boundary
Round, vesicular
Salt and pepper chromatin
Round, vesicular
Nucleoli
Prominent
Large and prominent
Mitosis
Frequent
Absent to conspicuous (depending on grade) Variable
Necrosis
Prominent
Variable
Variable
Inflammation
Absent to Scant
Absent
Other Features
---
IHC
PanCK,p63 +/-, NSE+/-
Fibrillary stroma, rossetes, vascular stroma, sustentacula r cells NSE, CD56, S100*, Syn+/-, CG+/-
Prominent mature lymphoplasmacyti c infiltrate ---
Specific genetic alteration
poorly differentiated carcinoma, squamous differentiation+ /Monomorphic
Sheets, occasional gland
Sheets, nests or trabeculae
variable
Round, vesicular
*
Round
Conspicuous
*
Prominent
Small round cells with moulding Round , artifectual spindling Absent
Frequent Frequent
Absent
* * *
Overlying epithelial dysplasia / in situ component
*
Glands, tubules, Azzopardy mucin effect
CK, p63,EBER,
Pan CK, p16+/-
P63, anti-NUT
EMA,B72.3,Be PanCK,NSE, r-EP4, CDX-2*, CG,SYN,CD5 6
Frequent
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SNEC
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Architecture
Poorly differentiated adenocarcinoma
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Undifferentiated NPC Syncytium, irregular islands
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Characteristics
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Table 2
Frequent Frequent
NUT-BRD rearrangement
Brisk Frequent Variable
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Figure 1a. SNUC showing large nests of medium-sized cells with high nucleo cytoplasmic ratio and often prominent nucleoli.
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Figure 1b. Tumor showing nuclear molding,hyperchromatic nuclei with absent nucleoli and focal spindling simulating a small cell neuroendocrine carcinoma.
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Figure 2a&b Positive immunostaining for p40 and CK5/6 in Poorly differentiated squamous cell carcinoma. Figure 2 c&d Positive immunostaining for p40 and CK5/6 in nasopharyngeal carcinoma.
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Figure 3a. Olfactory neuroblastoma grade 3, the tumor lacks neurofibrillary matrix and includes malignant round cells with increased mitotic activity. Figure 3b. Olfactory neuroblastoma with areas of squamous differentiation
