Cytological and Molecular Features of Papillary

Novel Insights from Clinical Practice Acta Cytologica DOI: 10.1159/000338395

Received: January 18, 2012 Accepted after revision: March 21, 2012 Published online: $ $ $

Cytological and Molecular Features of Papillary Thyroid Carcinoma with Prominent Hobnail Features: A Case Report Claudio Bellevicine a Immacolata Cozzolino a Umberto Malapelle a Pio Zeppa b Giancarlo Troncone a a Department of Biomorphologic and Functional Sciences, University of Naples Federico II, Naples, and b Faculty of Medicine, University of Salerno, Salerno, Italy

Established Facts • Papillary thyroid carcinoma (PTC) subtyping on fine needle aspiration (FNA) is possible and useful in guiding surgical management. • PTC with prominent hobnail features is a newly recognized histological variant with aggressive behavior. The cytological features of this variant have not yet been reported.

Novel Insights • Although the cytological features of PTC with prominent hobnail features may overlap with those of the tall cell variant, the novel ‘comet-like’ cell feature could represent the cytological counterpart of the histological hobnailing. • Thus, the presence on FNA of comet-like cells may lead to the preoperative suspicion of PTC with prominent hobnail features.

Key Words Fine needle aspiration ⴢ Thyroid ⴢ Hobnail papillary thyroid carcinoma ⴢ BRAF mutation

Abstract Background: Papillary thyroid carcinoma (PTC) with prominent hobnail features is a recently recognized PTC variant. Its histological hallmark is represented by elongated cells showing a high nuclear/cytoplasmic ratio and a hobnail appearance. Few histological studies have been performed show-

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ing aggressive clinical and pathological features. Thus, a better patient management might benefit from its early diagnosis on fine needle aspiration (FNA) samples. To date, the FNA cytology of PTC with prominent hobnail features has not been described. Case Report: We retrospectively analyzed the FNA taken from a histologically proven PTC with prominent hobnail features. Although there was a certain degree of morphological overlap with the tall cell variant,

C.B. and I.C. contributed equally to this work.

Correspondence to: Dr. Giancarlo Troncone Dipartimento di Scienze Biomorfologiche e Funzionali Università degli Studi di Napoli Federico II, via Sergio Pansini 5 IT–80131 Napoli (Italy) Tel. +39 081 746 3437, E-Mail giancarlo.troncone @ unina.it

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some peculiar cytological features were observed. In particular, the novel ‘comet-like’ cell feature could represent the cytological counterpart of the histological hobnailing. V600E BRAF mutation was observed on a matched cytohistological specimen. Conclusion: Further investigation is required to validate our diagnostic criteria, as the recognition of PTC with prominent hobnail features on FNA may prospectively enable its preoperative diagnosis, suggesting more aggressive neck surgery. Copyright © 2012 S. Karger AG, Basel

Introduction

Case Report A 57-year-old male had a single, solid nodule of the left thyroid lobe that measured 20 ! 15 mm. Ultrasound-guided FNA was performed. A cytopathologist ensured the proper smearing technique and on-site adequacy assessment. The smears were airdried and Diff-Quik stained. They were quite cellular with vari-

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Fig. 1. Low magnification of the on-site evaluated smear. Several

branching groups of epithelial thyroid cells are present. The nuclei show readily evident PTC changes. Diff-Quik. !10.

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Fine needle aspiration (FNA) is very accurate in diagnosing most papillary thyroid carcinomas (PTC). Since the clinical course of PTC is usually indolent, total thyroidectomy is the standard procedure. However, less frequently its behavior is more aggressive; in particular the tall cell, columnar, solid and diffuse sclerosing variants have a guarded clinical outcome. Thus, the correct recognition of these variants and careful PTC typing on FNA are useful for suggesting more aggressive neck surgery [1]. The term PTC with prominent hobnail features has been proposed to describe a new aggressive variant of PTC [2, 3]. Its histological hallmark is represented by elongated cells showing a high nuclear/cytoplasmic ratio and apically placed nuclei that produce a surface bulge leading to a hobnail appearance [2]. Few histological studies have been performed; thus clinical data are scarce. However, preliminary evidence seems to suggest that the rate of nodal and distant metastases in PTC with prominent hobnail features is higher than in classic PTC. From a cytological standpoint, diagnosing this tumor type preoperatively on FNA may allow for better patient management. Indeed, it is conceivable that the distinctive hobnail appearance of the neoplastic cells can also be consistently observed on cytological preparations. This point has been addressed in this study, which retrospectively analyzed an FNA taken from a PTC with prominent hobnail features.

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Fig. 2. PCR and sequencing analysis for the exon 15 BRAF gene. The electropherogram shows the presence of both mutant and wild-type BRAF alleles at the 1799 nucleotide.

ably cohesive cell groups with occasional branching; nuclear changes (enlargement, molding, crowding, grooves, and intranuclear cytoplasmic pseudoinclusions) were widespread (fig. 1). These unquestionable cytological features were conclusive for malignancy and PTC was diagnosed [4]. Moreover, the elongated and abundant cytoplasm and the prominent nuclear changes raised the possibility that the diagnosed PTC did not belong to a classic variant. The rationale behind the procedure of sample collection in our institution is to first ensure an adequate cytological diagnosis and, then, to exploit part of the diagnostic material for BRAF

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Fig. 3. Histology (HE). Whole slide microphotograph: in thyroid parenchyma, a well-demarcated neoplastic nodule is present. Noteworthy, the delicate papillary fronds imparted a cleft-like architecture to the nodule. Several distant PTC foci are also evident (arrow).

testing, whose assessment may have a prognostic value [5]. Thus, rather than preparing a cell block, the aspirate from an additional pass was directly collected in a tube containing additional nucleic preservative solution (RNAlater, Ambion) for BRAF testing. BRAF exon 15 was amplified by PCR and sequenced, as previously described [5]. The electropherogram showed the presence of both mutant and wild-type BRAF alleles (2 overlapping peaks) at the 1,799 nucleotide, diagnosing the V600E mutation (fig. 2). Histopathological Findings On gross examination, the thyroid was slightly enlarged. Serial sectioning revealed a 20-mm left lobe nodule, whose appearance was whitish, firm and infiltrating (fig. 3). Occasional distant PTC foci were also noted (fig. 3). A complex histological architecture was observed; papillae with a thin fibrovascular core were prevalent (fig. 4a). In addition, a minority of neoplastic follicles was observed. Occasionally, loss of cellular cohesiveness was evident, leading to cell detachment. The pattern of growth was infiltrative. At a higher magnification, both the papillae and neoplastic follicles were mostly lined by cuboidal to elongated cells, and their abundant cytoplasm had a well-defined border. The nuclei showed PTC features and multiple ‘soap-bubble’-like intranuclear inclusions. Their middle to apical location imparted a hobnail appearance to the cells (fig. 4b), which was also evident in the invasive front. A diagnosis of hobnail variant of PTC with prominent hobnail features was rendered according to the criteria suggested by Asioli et al. [2]. The BRAF testing performed on the matched histological sample confirmed the occurrence of exon 15 V600E mutation.

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Fig. 4. Histology (HE). a Low magnification. Thin arborizing papillae are evident. !10. b At higher power, the papillae are lined

by hobnail cells; tufts of detatching cells are also observed. !20.

Cytological Review The FNA was reviewed on the basis of the histological evidence. The original diagnosis was readily confirmed, as the nuclear features of PTC were very evident. However, on careful scrutiny additional features were noticed. Most cells were arranged in monolayered sheets; the disposition was very regular, giving a ‘tile-like’ appearance to the groups (fig. 5a). As the cells had abundant cytoplasm, neither negligible nuclear crowding nor overlapping was observed. On occasion, single cells with elongated, tapering cytoplasmic borders with a comet-like shape were noted within the groups (fig. 5b). These comet-like cells were frequently arranged in whorls (fig. 6a). Nuclear grooves and inclusions were readily seen. The former were very evident and imparted a ‘cerebriform’ shape to the nuclei (fig. 6b); the latter were multiple and small, i.e. soap-bubble-like (fig. 5a).

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Fig. 5. a Single cells with abundant cyto-

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plasm and sharp borders are aggregated in tile-like manner. No nuclear crowding and overlapping are observed. Diff-Quik. !20. b High-power view of the a neoplastic cell group. On the right, the comet-like cell is shown (arrowhead). The nucleus protrudes toward one extremity while the opposite cytoplasm is tapered. On the left, multiple soap-bubble-like nuclear inclusions are recognizable (arrow). Diff-Quik. !40.

Fig. 6. a At medium power, several comet-

like cells are arranged in whorls. DiffQuik. !20. b Multiple nuclei with cerebriform crenellated membrane (arrowheads). Diff-Quik. !20.

Discussion

PTC with prominent hobnail features has recently been described. Its clinical behavior may be aggressive, as the rate of nodal and distant metastasis is higher than classic PTC [2, 3]. Here, we show that its cytological features are also different. A sharp cell border, elongated cell shape, tile-like arrangement and soap-bubble-like intranuclear inclusions are the main features, yielding some overlap with the tall cell variant cytology [6]. Moreover, some cytological features (elongated cells and nuclear PTC changes) are also shared by hyalinizing trabecular adenoma. However, in this latter, cohesive aggregates of radially oriented elongated cells organized around DiffQuik metachromatic material are frequently reported [7, 8]. In the present case, these hyalinizing trabecular adenoma features were not observed. Conversely, an additional new feature was noted, i.e. the comet-like cells. These shapes reflected the nuclei protruding toward one extremity of the cell membrane, whereas the opposite cytoplasm was tapered (fig. 5b). The comet-like cells may represent the cytological counterpart of the hobnail features observed on histology. Further investigation is warranted to strengthen this association. To date, only the study by Hirokawa et al. [9], in a detailed description of a 4

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PTC case with septate cytoplasmic vacuole cells, describes the presence of hobnail-shaped neoplastic cells. The cerebriform nuclei (fig. 6b) represented an additional interesting feature, whose specificity may be questioned. Its infolded ‘crenellated’ nuclear membrane has already been described both in cytology and histology [10, 11]. However, while earlier evidence identified these cells only in the tall and columnar cell variant [12, 13], more recently this feature was identified even in classic PTC [10]. To date, only a few cases of PTC with prominent hobnail features have been described. Most of these had an aggressive behavior. We cannot provide pertinent data, as the follow-up of this case is very short. However, two features may suggest a potential aggressiveness: the infiltrative growth pattern and the BRAF V600E mutation. The former feature was consistently observed in the series reported by Asioli et al. [2]. Conversely, the rate of BRAF mutant was not dissimilar from that of classic PTC. More generally, the relative significance of these molecular markers is still debated. In particular, a recent paper by Eloy et al. [14] showed that on multivariate analysis an infiltrative growth pattern has a prognostic significance greater than the BRAF mutation.

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Conclusions

The hobnail variant of papillary thyroid carcinoma shows distinct cytological features, whose recognition may be useful in the diagnosis of this PTC variant. The recognition of hobnail PTC in FNA specimens may lead

to additional screening to rule out lymph node and/or distant metastasis. However, since some of the described hobnail PTC features are also shared by the tall cell variant, a prospective investigation is warranted to validate our cytological findings.

References 1 Ohori NP, Schoedel KE: Cytopathology of high-grade papillary thyroid carcinomas: tall-cell variant, diffuse sclerosing variant, and poorly differentiated papillary carcinoma. Diagn Cytopathol 1999;20:19–23. 2 Asioli S, Erickson LA, Sebo TJ, Zhang J, Jin L, Thompson GB, Lloyd RV: Papillary thyroid carcinoma with prominent hobnail features: a new aggressive variant of moderately differentiated papillary carcinoma. A clinicopathologic, immunohistochemical, and molecular study of eight cases. Am J Surg Pathol 2010; 34:44–52. 3 Motosugi U, Murata S, Nagata K, Yasuda M, Shimizu M: Thyroid papillary carcinoma with micropapillary and hobnail growth pattern: a histological variant with intermediate malignancy? Thyroid 2009;19:535–537. 4 Cibas ES, Ali SZ: The Bethesda System For Reporting Thyroid Cytopathology. Am J Clin Pathol 2009; 132:658–665. 5 Troncone G, Cozzolino I, Fedele M, Malapelle U, Palombini L: Preparation of thyroid FNA material for routine cytology and BRAF testing: a validation study. Diagn Cytopathol 2010;38:172–176.

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6 Solomon A, Gupta PK, LiVolsi VA, Baloch ZW: Distinguishing tall cell variant of papillary thyroid carcinoma from usual variant of papillary thyroid carcinoma in cytologic specimens. Diagn Cytopathol 2002; 27: 143– 148. 7 Bishop JA, Ali SZ: Hyalinizing trabecular adenoma of the thyroid gland. Diagn Cytopathol 2010;39:306–310. 8 Casey MB, Sebo TJ, Carney JA: Hyalinizing trabecular adenoma of the thyroid gland: cytologic features in 29 cases. Am J Surg Pathol 2004;28:859–867. 9 Hirokawa M, Kanahara T, Habara T, Fujimura N, Horiguchi H, Wakatsuki S, Sano T: Dilated rough endoplasmic reticulum corresponding to septate cytoplasmic vacuoles in papillary thyroid carcinoma. Diagn Cytopathol 2000;23:351–353.

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10 Bell CD, Coire C, Treger T, Volpe R, Baumal R, Fornasier VL: The ‘dark nucleus’ and disruptions of follicular architecture: possible new histological aids for the diagnosis of the follicular variant of papillary carcinoma of the thyroid. Histopathology 2001;39:33–42. 11 Mallik MK, Das DK, Mallik AA, Madda JP, Mirza K, Al-Bishi KK, George SS, Junaid TA: Dark and pale cerebriform nuclei in FNA smears of usual papillary thyroid carcinoma and its variants. Diagn Cytopathol 2004; 30: 187–192. 12 Flint A, Davenport RD, Lloyd RV: The tall cell variant of papillary carcinoma of the thyroid gland. Comparison with the common form of papillary carcinoma by DNA and morphometric analysis. Arch Pathol Lab Med 1991;115:169–171. 13 Sweeney E: Dormant cells in columnar cell carcinoma of the thyroid. Hum Pathol 1995; 26:691–693. 14 Eloy C, Santos J, Soares P, Sobrinho-Simões M: The preeminence of growth pattern and invasiveness and the limited influence of BRAF and RAS mutations in the occurrence of papillary thyroid carcinoma lymph node metastases. Virchows Arch 2011; 459: 265– 276.

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