Leiomyoma arising within a mature cystic teratoma.

Pathology (December 2013) 45(7), pp. 689–715

CORRESPONDENCE Germline SDHC mutation presenting as recurrent SDH deficient GIST and renal carcinoma

Sir, In October 2011 a 33-year-old female underwent resection of a 30 mm gastric gastrointestinal stromal tumour (GIST) and 22 mm liver metastasis with a mitotic count of 7 per 50 high power fields (HPF). Her history was remarkable for resections for three previous malignancies. In 1990 she underwent a gastric resection followed by adjuvant adriamycin and DTIC chemotherapy for a 140 mm multifocal ‘epithelioid leiomyosarcoma’ with a mitotic count of 4 per 50 HPF. In 2000 at the age of 22 she underwent a right nephrectomy for a 65 mm renal tumour for which the pathological diagnosis was ‘renal carcinoma of unclassifiable type’. In 2008 at the age of 30 she underwent thyroidectomy for papillary thyroid carcinoma. The family history was not suggestive of an inherited cancer syndrome. The patient’s sister and parents were asymptomatic and cancer free. The extended family history included a paternal aunt with colorectal carcinoma diagnosed at age 49, an additional paternal aunt with leukaemia at age 50 and two maternal aunts diagnosed with cervical cancer at ages 38 and 42. Initial genetic investigations including karyotype, p53 sequencing/MLPA and whole genome SNP array were negative. The pathology of the GIST with its accompanying liver metastasis from 2011 was reviewed. On the basis of its onset in young adulthood, gastric location, multilobulated growth pattern, absence of KIT or PDGFR mutations but diffuse strong positive staining for both KIT and DOG1 and predominance of epithelioid or plump spindled cells (Fig. 1), a diagnosis of succinate dehydrogenase (SDH) deficient GIST was suggested.1–3 The diagnosis was confirmed with immunohistochemistry for SDHB which showed negative staining of the neoplastic cells with positive staining in the non-neoplastic cells including the gastric epithelium and blood vessels (arrows in Fig. 1B,D). Additional immunohistochemistry for SDHA was positive – essentially excluding SDHA mutation.4–7 The slides from the gastric leiomyosarcoma resected in 1990 were reviewed and also showed typical features of succinate dehydrogenase deficient GIST, not an unexpected finding given that GIST (particularly in children) was only an emerging entity in the late 1980s and early 1990s and most tumours classified as leiomyosarcoma at that time or previously would now be classified as GIST. On review, the previously unclassifiable renal tumour showed identical morphology to that recently described in SDH related renal carcinoma (Fig. 2). Again immunohistochemistry for SDHB was negative with positive staining in the non-neoplastic cells including benign tubules and endothelial cells (arrows in Fig. 2B,D).8,9 The papillary carcinoma demonstrated typical morphology and showed normal positive staining for SDHB, suggesting it was more likely an incidental finding than a bona fide part of syndromic disease. A presumptive diagnosis of hereditary phaeochromocytomaparaganglioma (PGL) syndrome due to germline mutation of one Print ISSN 0031-3025/Online ISSN 1465-3931

#

of the SDH genes was made.10 After informed consent, sequential genetic testing for SDHB, SDHD and SDHC demonstrated a missense mutation in exon 5 of SDHC (c.380A>G;p.His127Arg) confirming the diagnosis. The patient is currently well with no clinical, radiological or biochemical evidence of residual or recurrent disease or phaeochromocytoma/paraganglioma. Her sister and mother have been demonstrated to be asymptomatic carriers. The SDHA, SDHB, SDHC and SDHD genes are autosomal. They encode proteins which are assembled in the mitochondria to form the mitochondrial complex II, a key respiratory enzyme which links the Krebs cycle and electron transport chain.10 Heterozygous germline mutations of these genes have been definitively linked to hereditary phaeochromocytoma/paraganglioma, SDH deficient GIST, renal carcinoma and pituitary adenoma.10,11 Because of low penetrance, and in the case of SDHD mutation parent of origin effect, the absence of a family history of hereditary neoplasia in no way excludes the possibility of germline mutation of any of the components of the mitochondrial complex 2.10 Negative immunohistochemical staining for SDHB occurs in tumours associated with double hit inactivation of any of the SDH genes, not just SDHB.1–5 In these tumours one of the two hits is almost always in the germline. Therefore, simple immunohistochemistry for SDHB performed on archived, formalin fixed, paraffin embedded tissue is a cheap and readily available screening test for germline SDHx mutation presenting with neoplasia. That is, negative immunohistochemical staining for SDHB identifies pheochromocytoma/paraganglioma, GISTs, renal carcinomas and pituitary adenomas associated with mutation of SDHA, SDHB, SDHC or SDHD. Furthermore, negative staining for SDHB also occurs in GISTs and paragangliomas associated with the Carney triad, the syndromic but non-hereditary association of GIST, paraganglioma and pulmonary chondroma.1 SDH deficient GISTs are a recently described subtype of GIST defined by negative immunohistochemical staining for SDHB.1–3 They differ from other GISTs because of their distinctive morphology, exclusive gastric location, tendency to multifocality, lymph node metastases and occurrence in childhood, absence of KIT and PDGFR mutations and a prognosis not predicted by size and mitotic rate.1,2,5 They demonstrate a unique natural history characterised by long latent periods and indolent metastases. Approximately 30% of SDH deficient GISTs are associated with germline SDHA mutation and these tumours can be identified by the occurrence of negative staining for both SDHB and SDHA.4–7 Perhaps 10–20% of SDH deficient GISTs are associated with germline mutation of SDHB, SDHC or SDHD,10 more if a paediatric population is surveyed. Other SDH deficient GISTs are associated with the Carney triad but not hereditary disease. Therefore, whenever an SDH deficient GIST is encountered it is recommended that syndromic disease (either germline mutation of SDH or the Carney triad) be considered.1,2,10 Despite this, at the present time it appears that the (slight) majority of SDH deficient GISTs are not associated with a recognisable syndrome.1,3,5 Surgical excision remains the mainstay of treatment for SDH deficient GISTs. When planning surgery, careful consideration should be given to the distinct natural history of this unique

2013 Royal College of Pathologists of Australasia

Copyright © Royal College of pathologists of Australasia. Unauthorized reproduction of this article is prohibited.

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CORRESPONDENCE

Pathology (2013), 45(7), December

Fig. 1 (A,C) Serial H&E and (B,D) SDHB stained sections of the succinate dehydrogenase deficient gastric GIST. Whilst the neoplastic cells show absent staining for SDHB, there is positive staining in non-neoplastic cells including epithelium and endothelium (arrows) which act as an internal positive control.

subtype of GIST, that is their tendency to multifocality, lymphatic spread and the relatively indolent nature of metastatic disease. Optimal medical therapy for unresectable SDH deficient GISTs is uncertain. Although their relatively slow growth may lead to confusion, there is strong evidence that they do not respond to imatinib1–3 but may respond better to second or third generation tyrosine kinase inhibitors such as sunitinib or nilotinib.1,10 Consistent overexpression of IGF1R by

these GISTs has raised hopes that IGF1R inhibition may represent a rational therapeutic target, but this is yet to be tested empirically.12 In summary, consideration of the possibility of a SDH deficient GIST or renal carcinoma based on morphology or clinical findings should lead to screening immunohistochemistry being performed on archived, formalin fixed, paraffin embedded tissue. This can then be followed up with formal

Fig. 2 (A,C) Serial H&E and (B,D) SDHB stained sections of the renal carcinoma. The neoplastic cells have clear to eosinophilic cytoplasm, a somewhat nested architecture and there are occasional intracytoplasmic vacuoles. Again the non-neoplastic cells show positive staining for SDHB (arrows) which contrasts to the negative staining in the neoplastic cells.


CORRESPONDENCE

691

genetic testing, screening for other components of syndromic disease and optimal treatment for these unique types of gastrointestinal stromal tumour and renal carcinoma.

12. Chou A, Chen J, Clarkson A, et al. Succinate dehydrogenase deficient GISTs are characterized by IGF1R overexpression. Mod Pathol 2012; 25: 1307–13.

Conflicts of interest and sources of funding: The authors state that there are no conflicts of interest to disclose.

DOI: 10.1097/PAT.0000000000000018

Anthony J. Gill*{{ Lara Liptonjj Jessica Taylorjj Diane E. Benn{§ Anne Louise Richardson{§ Mark Frydenbergô Jeremy Shapiro**{{ Roderick J. Clifton-Bligh{§ Chung W. Chow{{ Michael Bogwitzjj *Cancer Diagnosis and Pathology Group, Kolling Institute of Medical Research, Royal North Shore Hospital, St Leonards, {Department of Anatomical Pathology, Royal North Shore Hospital, St Leonards, zSydney Medical School, University of Sydney, Sydney, §Cancer Genetics, Hormones and Cancer Group, Kolling Institute of Medical Research, Royal North Shore Hospital, St Leonards, NSW, jjGenetic Medicine and Familial Cancer Centre, Royal Melbourne Hospital, Parkville, ôDepartment of Surgery, Monash Medical School, Melbourne, **Department of Medical Oncology, Cabrini Hospital, Melbourne, {{Department of Medicine, Monash Medical School, Monash University, Melbourne, and zzDepartment of Anatomical Pathology, Royal Children’s Hospital, Parkville, Vic Australia Contact Dr A. J. Gill. E-mail: [email protected] 1. Gill AJ, Chou A, Vilain R, et al. Immunohistochemistry for SDHB divides gastrointestinal stromal tumors (GISTs) into two distinct types. Am J SurgPathol 2010; 34: 636–44. 2. Miettinen M, Wang ZF, Sarlomo-Rikala M, Osuch C, Rutkowski P, Lasota J. Succinate dehydrogenase-deficient GISTs: a clinicopathologic, immunohistochemical, and molecular genetic study of 66 gastric GISTs with predilection to young age. Am J Surg Pathol 2011; 35: 1712– 21. 3. Janeway KA, Kim SY, Lodish M, et al. Defects in succinate dehydrogenase in gastrointestinal stromal tumors lacking KIT and PDGFRA mutations. Proc Natl Acad Sci USA 2011; 108: 314–8. 4. Dwight T, Benn DE, Clarkson A, et al. Loss of SDHA expression identifies SDHA mutations in succinate dehydrogenase deficient gastrointestinal stromal tumors. Am J Surg Pathol 2013; 37: 226–33. 5. Miettinen M, Killian JK, Wang ZF, et al. Immunohistochemical loss of succinate dehydrogenase subunit A (SDHA) in gastrointestinal stromal tumors (GISTs) signals SDHA germline mutation. Am J Surg Pathol 2013; 37: 234–40. 6. Oudijk L, Gaal J, Korpershoek E, et al. SDHA mutations in adult and pediatric wild-type gastrointestinal stromal tumors. Mod Pathol 2013; 26: 456–63. 7. Wagner AJ, Remillard SP, Zhang YX, Doyle LA, George S, Hornick JL. Loss of expression of SDHA predicts SDHA mutations in gastrointestinal stromal tumors. Mod Pathol 2013; 26: 289–94. 8. Gill AJ, Pachter NS, Clarkson A, et al. Renal tumors and hereditary pheochromoytoma-paraganglioma syndrome. N Engl J Med 2011; 364: 885–6. 9. Gill AJ, Pachter NS, Chou A, et al. The renal tumors associated with germline SDHB mutation show distinctive morphology. Am J Surg Pathol 2011; 35: 1578–85. 10. Gill AJ. Succinate dehydrogenase (SDH) and mitochondrial driven neoplasia. Pathology 2012; 44: 285–92. 11. Dwight T, Mann K, Benn D, et al. Familial SDHA mutation associated with pituitary adenoma and pheochromocytoma/paraganglioma. J Clin Endocrinol Metab 2013; 98: E1103–8.

Leiomyoma arising within a mature cystic teratoma

Sir, Leiomyomas of the ovary are infrequent, comprising less than 1% of ovarian neoplasms. They are typically unilateral, discovered across a wide age range of 3–102 years and may or may not co-exist with uterine leiomyomas.1,2 They have been found in association with other ipsilateral ovarian masses including endometrioma1,2 mucinous cystadenoma2 and serous cystadenofibroma.3 Leiomyoma arising within a mature cystic teratoma of the ovary has been described in a single reported case of an 18-year-old woman.4 We describe a rare case of leiomyoma of mu¨llerian origin arising within an ovarian mature cystic teratoma in a 13-year-old patient, along with a morphological and immunohistochemical characterisation of the tumour. The patient was a healthy, 13-year-old female who was found to have a pelvic mass on routine physical examination. She had reached menarche at the age of 10 years. Her periods were regular. She had no signs of virilisation or other endocrine abnormalities. An ultrasound showed a complex, predominantly cystic left ovarian mass containing a solid nodule measuring 3.1 cm. Subsequent magnetic resonance imaging (MRI) reported an ovarian mass measuring 21.5 cm in maximum dimension and confirmed an enhancing mural nodule. There was no ascites or pelvic lymphadenopathy. The operative report noted that the left ovary was occupied by a large, complex cyst of 20 cm and the right ovary had a simple cyst measuring 4 cm. Clear fluid was drained from the left cyst. A left oophorectomy was performed, as a cystectomy was deemed to be not feasible. An intra-operative frozen section of the more solid areas of the mass was reported as a dermoid cyst and a benign spindle cell lesion. The right cyst was also drained and yielded haemorrhagic fluid; thus a haemorrhagic corpus luteal cyst was favoured. The uterus, appendix and omentum appeared unremarkable, and there was no evidence of extra-ovarian disease. The left oophorectomy specimen showed an ovarian cyst with a smooth outer surface. The wall thickness ranged from 0.1 to 2.5 cm. Multiple nodules arose from the inner wall of the cyst. Some of these contained hair and sebum. The largest, measuring 3.5 cm, was solid with a tan, whorled cut surface and rubbery consistency. Sections of the largest mural nodule showed a well-circumscribed lesion composed of spindled cells arranged in short, interlacing fascicles with eosinophilic cytoplasm and bluntended nuclei (Fig. 1). There was no atypia or necrosis. Mitoses numbered up to 3/10 high power fields and no atypical mitotic figures were noted. On immunohistochemistry, the spindle cells stained diffusely positive for smooth muscle actin (SMA) and muscle specific actin and focally positive for desmin. Nuclear positivity with oestrogen receptor (ER) and progesterone receptor (PR) was also present. The tumour cells


692


CORRESPONDENCE

Fig. 1 Leiomyoma (left) arising from the mature teratoma (upper right). Inset: desmin stain.

stained negatively for S100, bcl-2 and CD34. Inhibin and calretinin highlighted scattered entrapped stromal cells only. The remaining areas of the mass showed elements typical of a mature cystic teratoma including skin with adnexal structures, mature cerebellar and glial tissue, choroid plexus, adipose tissue and cartilage. Smooth muscle was present in bundles within the cyst wall and in vessel walls, but did not appear to form a mass elsewhere in the cyst. Leiomyomas are benign smooth muscle tumours found in widespread body sites including skin, soft tissue and gastrointestinal tract. In the gynaecological organs, they arise most commonly from the myometrium of the uterus. In the ovary, possible smooth muscle origins of leiomyomas are smooth muscle metaplasia of the cortex or hilar blood vessels.5,6 In the context of a mature teratoma, smooth muscle elements intrinsic to the germ cell tumour could be the source of a leiomyoma. Morphologically, ovarian leiomyomas do not differ from extra-ovarian tumours and exhibit the same fascicular arrangement of bland spindled cells and low mitotic rates. Uterine variants including cellular, mitotically active with bizarre nuclei and myxoid leiomyomas have all been described in the ovary.1,2 On immunohistochemistry, in accordance with their smooth muscle origin, leiomyomas label with h-caldesmon, calponin, SMA, myosin and desmin. Despite the similar morphology and immunoprofile of leiomyomas in all body sites, those from the female gynaecological tract are additionally positive for ER and PR, with ER positivity reaching 100% in a series of 49 uterine leiomyomas.7 Sen et al. found the level of PR positivity distinguished between uterine and extrauterine leiomyomas, although a few leiomyomas from elsewhere in the gynaecological tract were included in the latter group.8 None of the 15 cutaneous leiomyomas showed expression of ER and PR.9 The literature on immunohistochemical labelling of ovarian leiomyomas with ER and PR is scant. In the largest series of ovarian leiomyomas by Lerwill et al., all cases which were evaluated for ER and PR were found positive; however, these constituted only four and two tumours, respectively.1 Another series of leiomyomas reported one ovarian primary positive for ER, whereas a second was negative.8 In contrast to ovarian leiomyomas, a mature cystic teratoma is a common tumour amounting to 20% of all ovarian neoplasms and occurring across all age groups.10 These tumours are composed of mature tissue elements from all three germ layers (endoderm, mesoderm and ectoderm) which can occasionally give rise to secondary neoplastic transformation. Squamous carcinoma is by far the most frequent malignancy; carcinoid and thyroid carcinomas are also commonly

encountered.11 While the malignant transformation of mature cystic teratomas is well described, the development of benign neoplasms from somatic elements of a teratoma is not reported, except for a single case report mentioned above.4 Molecular analysis, in particular loss of heterozygosity, was attempted in the current case but was unfortunately uninformative. In contrast to the observed findings, one would expect the phenotype to be non-mu¨llerian if the leiomyoma in the current case was derived from a somatic element of the teratoma. Leiomyoma arising within a mature cystic teratoma has been reported only once before, also in an adolescent female. As in our case, the tumour appeared grossly confluent with the wall of the teratoma and had the characteristic whorled, rubbery cut surface of a leiomyoma. Microscopically, neither tumour belonged to a specific subclass of leiomyomas. The smooth muscle origin of the initial case report was confirmed with muscle specific antigen immunoperoxidase staining.4 The various tissue types arising within mature cystic teratomas of the ovary have been extensively tabulated, with ectodermal derivatives such as skin, skin appendages and neural tissues being amongst the most common.11 Prostatic tissue is rarely encountered, and because of the absence of associated wolffian duct remnants, is thought to be a result of local androgen production by leutinised ovarian stromal or hilar cells.12 Interestingly, recapitulation of certain mu¨llerian tissue types such as of the uterus or fallopian tubes within ovarian teratomas is not widely described. The ER and PR positivity suggests that the leiomyoma in our case is of mu¨llerian origin, and thus more akin to leiomyoma of the uterus rather than of skin or soft tissue. It is possible that the leiomyoma is an independent tumour that co-developed with the teratoma rather than arising from elements of the latter. In conclusion, leiomyomas are among the benign neoplasms arising within mature cystic teratoma, albeit rarely. Our case presents an additional curiosity in that the leiomyoma appears to be of gyneocological origin, which is out of keeping with usual absence of mu¨llerian elements arising within mature ovarian teratomas. Conflicts of interest and sources of funding: The authors state that there are no conflicts of interest to disclose. Kiran Jakate* John Srigley{ Anna Plotkin{ *University of Toronto, {Trillium Health Partners, McMaster University, and zTrillium Health Partners, University of Toronto, Ontario, Canada Contact Dr A. Plotkin. E-mail: [email protected] 1. Lerwill MF, Sung R, Oliva E, et al. Smooth muscle tumors of the ovary: a clincopathologic study of 54 cases emphasizing prognostic criteria, histologic variants, and differential diagnosis. Am J Surg Pathol 2004; 28: 1436– 51. 2. Doss BJ, Wanek SM, Jacques SM, et al. Ovarian leiomyomas: clinicopathologic features in fifteen cases. Int J Gynecol Pathol 1999; 18: 63–8. 3. Eren F, Akpulat S, Gokaslan H, et al. Primary leiomyoma of the ovary co-existing with serous cystadenofibroma. APMIS 2005; 113: 145–7. 4. Coard KC, Frederick J. Co-existing leiomyoma and benign cystic teratoma of the ovary. West Indian Med J 1993; 42: 82–4.


CORRESPONDENCE

5. Doss BJ, Wanek SM, Jacques SM, et al. Ovarian smooth muscle metaplasia: an uncommon and possibly underrecognized entity. Int J Gynecol Pathol 1999; 18: 58–62. 6. Scully RE. Smooth-muscle differentiation in genital tract disorders. Arch Pathol Lab Med 1981; 105: 505–7. 7. Lee C, Turbin DA, Sung YV, et al. A panel of antibodies to determine site of origin and malignancy in smooth muscle tumors. Mod Pathol 2009; 22: 1519–31. 8. Sen N, Demirkan NC, Colakoglu N, et al. Are there any differences in the expression of hormonal receptors and proliferation markers between uterine and extrauterine leiomyomas? Int J Surg Pathol 2008; 16: 43–7. 9. McGinley KM, Bryant S, Kattine AA, et al. Cutaneous leiomyomas lack estrogen and progesterone receptor immunoreactivity. J Cutan Pathol 1997; 24: 241–5. 10. Talerman A, Vang R. Germ cell tumors of the ovary. In: Kurman RJ, Ellenson LH, Ronnett BM, editors. Blaustein’s Pathology of the Female Genital Tract. 6th ed. New York, NY: Springer, 2011; 873. 11. Medeiros F, Crum CP. Germ cell tumors of the ovary. In: Crum CP, Nucci MR, Lee KR, editors. Diagnostic Gynecologic and Obstetric Pathology. 2nd ed. Philadelphia, PA: Elsevier Saunders, 2011; 907. 12. McLachlin CM, Srigley JR. Prostatic tissue in mature cystic teratoma of the ovary. Am J Surg Pathol 1992; 16: 780–4.

DOI: 10.1097/PAT.0000000000000013

High grade transformation in a case of adenoid cystic carcinoma associated with Epstein–Barr virus expression Sir, We present a case of high grade transformation in a parotid adenoid cystic carcinoma associated with diffuse Epstein–Barr virus (EBV) RNA expression. The concept of transformation of a tumour to a less differentiated state is not novel and has been described in many tumours.1 Where some features of the original neoplasm has been retained, the term ‘high grade

693

transformation’ has been used.1 As this case involves expression of EBV in both the original and transformed states, it raises the question of the possible role of EBV in dedifferentiation. An 88-year-old Caucasian female presented to the otolaryngology clinic with a 2 month history of a gradually enlarging painless left pre-auricular swelling. Her medical history was significant for mild dementia and a 20 pack-year smoking history (ceased 50 years ago). A 31 21 33 mm tumour involving the left parotid was demonstrated by ultrasound and computer tomography (CT). No aggressive features, such as lymphadenopathy and facial nerve invasion, were detected clinically or radiologically. A superficial parotidectomy was performed with subsequent adjuvant radiotherapy. No metastases were found during the operation. The patient made an unremarkable recovery and was clinically free of disease after 12 months. Fine needle aspiration prior to operation was atypical and suggested features of adenoid cystic carcinoma. The excision specimen revealed an unencapsulated and lobulated tumour measuring 40 mm in size. The tumour had multiple morphologies with typical areas contiguous with solid areas and undifferentiated tumour resembling lymphoepitheliomalike carcinoma (Fig. 1). The transformed component consisted of larger and more pleomorphic cells with prominent nucleoli and areas of necrosis. In some areas the cells were elongated and spindled. A benign lymphoid infiltrate in the transformed component was noted. A transitional zone between the two morphologies was evident on haematoxylin and eosin (H&E) and was also demonstrated immunohistochemically. There was a gradual loss of myoepithelial markers CD117, SMA and calponin as the tumour became less differentiated. Cyclin-D1 expression was decreased in the transformed component while p53 was increased. No staining for HER-2 was seen. Epstein– Barr encoded RNA (EBER) in situ hybridisation (ISH) was diffusely positive in both components and negative in the normal salivary tissue (Fig. 2).

Fig. 1 (A) Typical cribriform adenoid cystic carcinoma pattern with mucin. (B) Solid adenoid cystic pattern. (C) Transformed lymphoepithelioma carcinoma-like pattern. (D) High grade tumour with necrosis.


EBER

Fig. 2 Immunohistochemistry showing gradual loss of myoepithelial cells with CD117 and increased p53 expression in the transformed tumour. Diffuse expression of EBV RNA in both the well-differentiated and transformed tumour. There is sparing of an uninvolved duct centrally.

1M (n ¼ 1); 74 years

1F (n ¼ 1); 51 years

1M (n ¼ 1); 54 years

Sato et al., 20069

Handra-Luca et al., 200910

Malhotra et al., 200911

1F (n ¼ 1); 64 years 1M (n ¼ 1); 62 years

542; parotid

Size not reported; palate

4; maxillary sinus

3; submandibular gland 2.21.5; soft palate

High-grade poorly differentiated adenocarcinoma Anaplastic cells with pleomorphic nuclei and abnormal mitoses Pleomorphic vesicular nuclei and amphophilic cytoplasm proliferating irregularly with loose solid nests or infiltratively in a small tubular and cord-like pattern Undifferentiated carcinoma with focal squamoid differentiation; staining not reported Undifferentiated carcinoma with loss of myoepithelial differentiation

Moderately dedifferentiated adenocarcinoma with frequent glandular differentiation Poorly differentiated carcinoma with scarce glandular differentiation Undifferentiated carcinoma of large cell type

Poorly differentiated adenocarcinoma or undifferentiated carcinoma Poorly differentiated adenocarcinoma Poorly differentiated cribiform adenocarcinoma or solid undifferentiated carcinoma

Dedifferentiated component

H&E

Chau et al., 2001 Ide et al., 20038

3; tongue

T2–T4; submandibular (2/7); palate (2/7); paranasal sinus; parotid, lips T2; paranasal sinus

1.7–6 (3.5); submandibular gland (2/6); maxillary sinus (2/6); nasal cavity (2/6) Size not reported; tongue, soft palate, hard palate 2–7 (4.2); Submandibular (4/11); maxillary (2/11); nasopharynx; palate

Tumour size (cm), involved glands

No local recurrence; alive at 5 m

No local recurrence; alive at 12 m

No local recurrence; metastasis 1/1; DOD 16 m

No local recurrence or metastatic disease; alive at 64 m Not reported Not reported

Local recurrence 5/6; metastatic disease 5/6; DOD 5/6 (6–69 m, mean 33.7) DOD 2/3 (within 18 mo) Local recurrence 2/11; metastatic disease 3/11; DOD 3/11 (12–15 m, mean 13); dead 2/11 (8–44 m, mean 26) Local recurrence 0/7; metastatic disease3/7; DOD 1/7 (7 m) Local recurrence; alive at 144 m

Follow-up

CORRESPONDENCE

7

Moles et al., 1999

1M (n ¼ 1); 61 years

Bonfitto et al., 20104

1F (n ¼ 1); 61 years

5F:2M (n ¼ 7); 44–65 (55) years

Cheuk et al., 1999 Seethala et al., 20071

6

2F:1M (n ¼ 3); 38,53,55 years 4F:7M (n ¼ 11); 32–72 (58) years

3

Costa et al., 2010

3M:3F (n ¼ 6); 34–70 (46.8) years

Nagao et al., 20032

5

Gender and age

CD117

Authors

Summary of reported transformations of adenoid cystic carcinoma

p53

Table 1

694 Pathology (2013), 45(7), December


CORRESPONDENCE

Transformation in salivary gland tumours is only relatively recently recognised. Previously described include transformations in acinic cell carcinoma,1 mucoepidermoid carcinoma,2 epithelial-myoepithelial carcinoma1 and adenoid cystic carcinoma.3 The study by Seethala et al. proposed criteria for diagnosis of transformation.1 These include at least three of the following: proliferation of tumour cells with at least a focal loss of myoepithelial cells surrounding tumour nests, nuclear size at least 2–3 times the size of tubular/cribiform adenoid cystic carcinoma, thickened irregular nuclear membranes, and prominent nucleoli in a majority of cells. Transformation in adenoid cystic carcinoma is relatively rare with only 36 cases described in the literature and none were associated with EBV expression.1,2,4–11 No prior cases were identified in our laboratory and to our knowledge, lymphoepithelioma-like carcinoma has not specifically been observed and EBV RNA expression has not been documented. The majority of transformed adenoid cystic carcinoma shows high grade transformation to moderate or poorly differentiated adenocarcinoma or undifferentiated carcinoma (Table 1). Transformation typically occurs in the sixth decade or later with a slight male predominance. Commonly involved sites include sinonasal seromucinous glands, palate, and submandibular glands. Parotid involvement is rare. The median survival is estimated to be approximately 12 months with a more protracted course in more differentiated tumours. Studies looking into molecular abnormalities of dedifferentiated adenoid cystic carcinomas show consistent abnormalities in p53 expression in combination with HER-2/ neu overexpression and loss of pRb expression.2,3 Cyclin-D1 overexpression is also noted in a number of cases (Table 1). EBV has been implicated in lymphoproliferative disorders (Burkitt lymphoma, Hodgkin and non-Hodgkin lymphomas), multiple sclerosis, and various epithelial neoplasms (nasopharyngeal carcinoma, gastric adenocarcinoma and lymphoepithelioma-like carcinoma) with nasopharyngeal tumours amongst the most common EBV-associated neoplasm. The relationship between EBV and adenoid cystic carcinoma is weak at best; none of the cases of adenoid cystic carcinoma transformation reported EBV expression of the tumour cells.1,2,4 – 11 In addition, tumour transformation to lymphoepithelioma-like carcinoma is also generally rare. A review of the literature shows less than 20 cases, with the most commonly reported tumour associated with lymphoepithelioma-like carcinoma transformation being intrahepatic cholangiocarcinoma.12 These tumours were shown to commonly express EBER-1 diffusely (77%, n ¼ 10) and some authors have postulated a possible role for EBV in the pathogenesis of transformed intrahepatic cholangiocarcinoma to lymphoepithelioma-like carcinoma.12 Although the exact mechanism of pathogenesis is unknown, the presence of EBV in a clonal form within many of the malignancies described suggests that the virus had entered these cells prior to their clonal expansion.12 With the limited cases studied, the current opinion is that EBV is unlikely to play a significant role in the development of

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adenoid cystic carcinoma, although EBV’s role in tumour transformation has not been explored.12 We can only postulate the sequence of EBV infection, tumourigenesis, and dedifferentiation in our case. Whether there was superinfection by EBV in a pre-existing adenoid cystic carcinoma, reactivation of latent infection, or concurrent acquisition of EBV and tumour can only be hypothesised. Conflicts of interest and sources of funding: The authors state that there are no conflicts of interest to disclose. Chanh K. Ly* Hui M. Cheng{ Tersia Vermeulen* *Department of Anatomical Pathology, PathWest Laboratory Medicine, Royal Perth Hospital, Perth, and {Royal Perth Hospital, Perth, WA, Australia Contact Dr C. K. Ly. E-mail: [email protected] 1. Seethala RR, Hunt JL, Baloch ZW, Livolsi VA, Leon Barnes E. Adenoid cystic carcinoma with high-grade transformation: a report of 11 cases and a review of the literature. Am J Surg Pathol 2007; 31: 1683–94. 2. Nagao T, Gaffey TA, Serizawa H, et al. Dedifferentiated adenoid cystic carcinoma: a clinicopathologic study of 6 cases. Mod Pathol 2003; 16: 1265– 72. 3. Cheuk W, Chan JK, Ngan RK. Dedifferentiation in adenoid cystic carcinoma of salivary gland: an uncommon complication associated with an accelerated clinical course. Am J Surg Pathol 1999; 23: 465–72. 4. Bonfitto VL, Demasi AP, Costa AF, Bonfitto JF, Araujo VC, Altemani A. High-grade transformation of adenoid cystic carcinomas: a study of the expression of GLUT1 glucose transporter and of mitochondrial antigen. J Clin Pathol 2010; 63: 615–9. 5. Costa AF, Altemani A, Vekony H, et al. Genetic profile of adenoid cystic carcinomas (ACC) with high-grade transformation versus solid type. Anal Cell Pathol 2010; 33: 217–28. 6. Moles MA, Avila IR, Archilla AR. Dedifferentiation occurring in adenoid cystic carcinoma of the tongue. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 1999; 88: 177–80. 7. Chau Y, Hongyo T, Aozasa K, Chan JK. Dedifferentiation of adenoid cystic carcinoma: report of a case implicating p53 gene mutation. Hum Pathol 2001; 32: 1403–7. 8. Ide F, Mishima K, Saito I. Small foci of high-grade carcinoma cells in adenoid cystic carcinoma represent an incipient phase of dedifferentiation. Histopathology 2003; 43: 604–6. 9. Sato K, Ueda Y, Sakurai A, et al. Adenoid cystic carcinoma of the maxillary sinus with gradual histologic transformation to high-grade adenocarcinoma: a comparative report with dedifferentiated carcinoma. Virchows Arch 2006; 448: 204–8. 10. Handra-Luca A, Planchard D, Fouret P. Docetaxel-cisplatin-radiotherapy in adenoid cystic carcinoma with high-grade transformation. Oral Oncol 2009; 45: e208–9. 11. Malhotra KP, Agrawal V, Pandey R. High grade transformation in adenoid cystic carcinoma of the parotid: report of a case with cytologic, histologic and immunohistochemical study. Head Neck Pathol 2009; 3: 310–4. 12. Henderson-Jackson E, Nasir NA, Hakam A, Nasir A, Coppola D. Primary mixed lymphoepithelioma-like carcinoma and intra-hepatic cholangiocarcinoma: a case report and review of literature. Int J Clin Exp Pathol 2010; 3: 736–41.

DOI: 10.1097/PAT.0000000000000012


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Diffuse large B-cell lymphoma/follicular lymphoma arising in a background of IgG4-related pachymeningitis

Sir, IgG4-related disease is an increasingly recognised multisystem fibro-inflammatory entity often associated with serum IgG4 elevation.1 It generally shows a relapsing remitting course and many sites of involvement including the pachymeninges have been described. Histological features include lymphoplasmacytic inflammation, a storiform pattern of fibrosis and obliterative phlebitis. A requisite number of IgG4þ cells and IgG4:IgG ratio in tissue must also be present.2 Lymphoma can develop in this context, both at sites of IgG4-related disease and uninvolved nodal and extranodal locations. We present the first known case of diffuse large B-cell lymphoma arising in the context of IgG4-related pachymeningitis, an uncommon manifestation of this multisystem disease. A 72-year-old female underwent brain imaging as part of the investigation of a 1 year history of mild confusion and cognitive impairment. Magnetic resonance imaging (MRI) found a 13 cm extensive extra-axial lobulated mass arising over the left lateral convexity (Fig. 1). Differential diagnoses included atypical meningioma and/or a dural metastasis. No additional systemic or neuroimaging was performed pre-operatively and serum tumour markers and IgG4 levels were not measured. The patient had no known history of malignancy. Left frontoparietal craniotomy was undertaken to achieve complete tumour excision. The tumour was adherent to the dura with a multinodular, exophytic growth pattern. Histology revealed two coexistent pathological processes. The dura mater was expanded by a dense lymphoplasmacytic infiltrate (Fig. 2). Scattered reactive lymphoid follicles were observed, with a paucity of other inflammatory cells. Admixed fibrocollagenous

Fig. 1 Transverse T1 weighted magnetic resonance image showing a 13 cm hyperintense extra-axial lobulated mass arising over the left lateral convexity.


bands were also present, in addition to patchy stromal sclerosis. Occasional veins showed phlebitis, sometimes associated with thrombosis. Abundant IgG4þ plasma cells were confirmed. Immunohistochemistry showed IgG4þ plasma cells up to 375 per high power field, representing 98% of all IgGþ plasma cells. Negative staining for ALK and EMA excluded inflammatory myofibroblastic tumour and lymphoplasmacyte-rich meningioma, respectively. Gomori methenamine silver (GMS), Ziehl-Neelsen for acid-fast bacilli, periodic acid-Schiff and Gram stains were all negative. Additionally, merging with the lymphoplasmacytic infiltrate was a neoplastic lymphoid population associated with a sclerotic stroma. The tumour had a biphasic appearance, with neoplastic cells showing a follicular architecture in addition to sheet-like infiltration (Fig. 3) in broadly equal proportion. Neoplastic follicles were irregularly shaped, showing accentuated mantle zones with non-polar germinal centres and inconspicuous tingible body macrophages. Tumour cells were large and pleomorphic, with round to polygonal nuclei, variably prominent nucleoli, vesicular nucleoplasm and abundant amphophilic cytoplasm. Brisk mitotic activity was observed, with Ki-67 staining 80% of tumour cell nuclei. Malignant cells stained positively for CD20, CD10 and Bcl-6, with absent staining for Mum-1, Bcl-2 and CD5. The follicular component was confirmed by preservation of CD21 positive cells in the nodular areas, with no significant centrocytic component. The morphological and immunohistochemical profile was consistent with diffuse large B-cell non-Hodgkin lymphoma, germinal centre type, with a follicular grade 3B component. This was interpreted as having arisen in a background of IgG4-related pachymeningitis. The patient had an unremarkable post-operative course. No additional sites of IgG4-related disease or lymphoma were detected and the patient completed six cycles of R-CHOP chemotherapy (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisolone). IgG4-related disease is an increasingly recognised fibroinflammatory condition which has gained prominence within the past decade. The prototype is autoimmune pancreatitis, where the first association with elevated serum IgG4 levels was noted in 2001.3 IgG4-related disease has multiple sites of potential involvement and protean manifestations.2 Disease has been described in the salivary glands, orbit, lacrimal glands, biliary tree, lung, pleura, thyroid, retroperitoneum, lymph nodes, breast, pituitary, pachymeninges, skin, liver, kidney, prostate, aorta, mediastinum and pericardium.4 IgG4-related disease typically occurs in middle-aged to elderly patients with a marked male predominance.1 Requisite histopathological features outlined in a Consensus Statement2 include a dense lymphoplasmacytic infiltrate, storiform pattern of fibrosis and obliterative phlebitis. A site specific minimum number of IgG4þ plasma cells is stipulated, in addition to the required 40% IgG4:IgG ratio. Tissue eosinophilia may be present, in addition to the tendency to form a tumefactive mass. The probability of IgG4-related disease can be reported from a histological perspective, however clinicopathological correlation is required for definitive diagnosis. IgG4-related pachymeningitis is an extremely rare entity with only 18 possible cases reported in the literature.5–13 It has been described more commonly within the cranial meninges, however intraspinal involvement has also been reported in a limited number of cases, with a thoracic


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Fig. 2 (A) Microscopic view of dura showing mixed inflammatory cell population (H&E). (B) Dense lymphoplasmacytic infiltrate higher power. (C) Phlebitis without luminal obliteration. (D) Collagenous fibrosis, lacking the storiform pattern typically reported at extra-meningeal sites. (E) Immunohistochemistry confirms abundant IgGþ plasma cells within the lymphoplasmacytic component. (F) Comparative immunohistochemistry confirms IgG4þ plasma cells constitute virtually the entire IgGþ plasma cell population.

predilection.5 A single case of IgG4-related leptomeningitis is noted,6 in addition to the possibility of cerebral parenchymal extension.7 Pre-biopsy diagnosis is confounded by the absence of symptoms or their non-specific nature. IgG4-related pachymeningitis may present with headache related to mass effect and/or cranial nerve dysfunction. Intraspinal disease is often a medical emergency with symptomatology related to spinal cord compression. The non-specific finding of a dural mass on imaging generates a broad differential list, including but not limited to meningioma, lymphoma, fibroinflammatory procesess such as inflammatory myofibroblastic tumour or idiopathic hypertrophic pachymeningitis, infection and metastasis. Increased serum IgG4 levels may be a useful adjunct to diagnosis. A normal titre may be falsely reassuring, however, as up to 40% of patients with established disease may show no serum IgG4 elevation.1 In our case, no pre-operative level was measured and testing post-initiation of chemotherapeutic regimen (including steroids) showed a normal serum IgG4 level. Histological criteria for IgG4-related pachymeningitis are not well established due to limited published data.2

Lymphoplasmacytic infiltration is consistently described, in addition to fibrosis, generally lacking the characteristic storiform pattern referred to in other sites.13 Phlebitis is an uncommon finding,5,6,10,12 although present in our case, and obliterative phlebitis has not been described in this location. An accompanying, minor eosinophilic infiltrate may be present.5,13 As a minimum, 10 IgG4þ plasma cells per high power field must be confirmed on immunohistochemical stain in meningeal disease.2 First line treatment commonly involves surgical resection, undertaken for both histological assessment and symptom control. Thereafter, steroid therapy1 is the mainstay, occasionally supplemented with other immunosuppressive medications,6 generally achieving a good radiological response and amelioration of symptoms. A case of relapsed meningeal disease is noted, 4 years after initial presentation, successfully treated with a combination of steroid and rituximab therapy.10 Patients with IgG4-related pachymeningitis require thorough investigation. Localised disease may rarely occur, however more commonly distribution reflects the systemic nature of the condition with multiple sites affected.1 Therefore, ongoing surveillance is critical to detect additional sites,

Fig. 3 (A) Enlarged and irregularly shaped follicles within sclerotic stroma (H&E). Inset: Neoplastic germinal centre with abundant centroblasts. (B) Diffuse infiltration by atypical lymphoid population.


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as well as monitoring for malignancy, with IgG4-related disease previously associated with both carcinoma and lymphoma,1 as in our case. The development of lymphoma at sites of IgG4-related disease has been broadly reported. Non-Hodgkin lymphoma is more common, with extranodal marginal zone lymphoma most likely to arise within this milieu. Mucosa-associated lymphoid tissue (MALT) lymphoma has been described within the ocular adnexae, orbit and lacrimal gland.14 A single report of high grade follicular lymphoma has been published in the context of IgG4-related dacryoadenitis. A case of composite lymphoma15 comprising extranodal marginal zone lymphoma within cervical soft tissue and classical Hodgkin lymphoma mixed cellularity subtype arising within an adjacent lymph node is described. Lymphoma has additionally been documented at uninvolved nodal and extranodal sites. Takahashi et al.16 report two cases of diffuse large B-cell lymphoma and one of B-cell lymphoma unspecified during follow-up of patients with this entity, manifesting 3–5 years after initial presentation and diagnosis of IgG4-related disease. The lymphomagenesis of IgG4-related disease is poorly understood. The composition of the lymphoplasmacytic infiltrate in IgG4-related disease is polyclonal. However, the association of lymphoma and other autoimmune diseases, such as Sjogren’s syndrome and rheumatoid arthritis is well established.17 It is postulated that chronic inflammatory stimulation plus or minus immunosuppressive therapy for autoimmune disease may be causative or contributory to malignant B-cell transformation.16 In summary, IgG4-related pachymeningitis is an uncommon entity with limited cases published. Histological criteria are outlined for this site and the probability of disease can be reported from a histological perspective, requiring clinicopathological correlation for definitive diagnosis. This is the first known association of diffuse large B-cell nonHodgkin lymphoma and IgG4-related pachymeningitis, further expanding the pool of potential sites of tumourigenesis in this entity. IgG4-related disease remains poorly understood and additional insight into pathogenesis and malignant potential are necessary in the future. Conflicts of interest and sources of funding: The authors state that there are no conflicts of interest to disclose. Nicole Lightfoot* Renate Kalnins{ Departments of Anatomical Pathology, *PeterMacCallum Cancer Centre, and {Austin Health, Melbourne, Vic, Australia Contact Dr N. Lightfoot. E-mail: [email protected] 1. Cheuk W, Chan JK. IgG4-related sclerosing disease: a critical appraisal of an evolving clinicopathologic entity. Adv Anat Pathol 2010; 17: 303–32. 2. Deshpande V, Zen Y, Chan JKC, et al. Consensus Statement on the Pathology of IgG4-related Disease. Mod Pathol 2012; 25: 1181–92. 3. Hamano H, Kawa S, Horiuchi A, et al. High Serum IgG4 concentrations in patients with sclerosing pancreatitis. N Engl J Med 2001; 344: 732–8. 4. Stone JH. IgG4-related disease: nomenclature, clinical features and treatment. Semin Diagn Pathol 2012; 29: 177–90. 5. Chan SK, Chek W, Chan KT, Chan JK. IgG4-related sclerosing pachymeningitis; a previously unrecognized form of central nervous system

6. 7. 8. 9. 10. 11. 12. 13.

14. 15.

16.

17.

involvement in IgG4-related sclerosing disease. Am J Surg Pathol 2009; 33: 1249–52. Lindstrom K, Cousar J, Lopes M. IgG4-related meningeal disease: clincopathological features and proposal for diagnostic criteria. Acta Neuropathol 2010; 120: 765–76. Kim EH, Kim SH, Cho JM, et al. Immunoglobulin G4-related hypertrophic pachymeningitis involving cerebral parenchyma. J Neurosurg 2011; 115: 1242–7. Norikane T, Yamamoto Y, Okada M, et al. Hypertrophic cranial pachymeningitis with IgG4-positive plasma cells detected by C-11 methionine PET. Clin Nucl Med 2012; 37: 108–9. Della Torre E, Bozzolo EP, Passerini G, et al. IgG4-related pachymeningitis: evidence of intrathecal IgG4 on cerebrospinal fluid analysis. Ann Intern Med 2012; 156: 401–3. Shapiro K, Bove R, Volpicelli E, et al. Relapsing course of immunoglobulin G4-related pachymeningitis. Neurology 2012; 79: 604–5. Utsuki S, Kijima C, Fujii K, et al. Investigation of IgG4-positive cell infiltration in biopsy specimens from cases of hypertrophic pachymeningitis. Clin Neuropathol 2013; 32: 84–90. Tanboon J, Felicella MM, Bilbao J, et al. Probable IgG4-related pachymeningitis: a case with transverse sinus obliteration. Clin Neuropathol 2013; 32: 291–7. Imbergamo S, Campagnolo M, Manara R, et al. Teaching neuroimages: multifocal neurologic involvement as the only manifestations of IgG4related disease. Neurology 2013; 80: e40–41. Cheuk W, Chan JKC. Lymphoproliferative lesions in IgG4-related disease. Curr Immunol Rev 2011; 7: 221–31. Cheuk W, Tam FK, Chan AN, et al. Idiopathic cervical fibrosis – a new member of IgG4-related sclerosing diseases: report of 4 cases, 1 complicated by composite lymphoma. Am J Surg Pathol 2010; 34: 1678–85. Takahashi N, Ghazale AH, Smyrk TC, et al. Possible association between IgG4-associated systemic disease with or without autoimmune pancreatitis and non-Hodgkin lymphoma. Pancreas 2009; 38: 523–6. Zintzaras E, Voulgarelis M, Moutsopoulos HM. The risk of lymphoma development in autoimmune diseases: a meta-analysis. Arch Intern Med 2005; 165: 2337–44.

DOI: 10.1097/PAT.0000000000000016

CRTC3-MAML2 fusion transcript in a bronchial mucoepidermoid carcinoma

Sir, Mucoepidermoid carcinoma (MEC) is a common salivary gland cancer and accounts for approximately 25% of all malignancies of the major and minor salivary glands.1 In contrast, primary tracheobronchial MEC is quite uncommon, accounting for less than 1% of all cases of primary tracheobronchial carcinoma.2 Approximately 50% of MEC of the salivary gland and tracheobronchial tree harbour a recurrent t(11;19)(q21;p13) translocation resulting in a CRTC1-MAML2 fusion oncogene, formed by fusion of the CREB-binding domain of CREB coactivator CRTC1 and the transactivation domain of Notch coactivator MAML2.3,4 Risks of local recurrence, metastases, and tumour-related deaths were found to be significantly lower in fusion positive patients compared to their fusion negative counterparts.4 Since tumour fusion status provides valuable diagnostic and prognostic information, molecular studies for the detection of CRTC1-MAML2 are frequently performed in MEC in the clinical setting. In 2008, Fehr et al. reported a novel fusion transcript, CRTC3-MAML2, in a case of salivary gland MEC in a 61-year-old woman.5 Since then, a few additional cases of salivary gland MEC harbouring this alternate fusion transcript had been reported.6,7 However, this fusion has never been described in tumours outside the salivary glands. Herein, we


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present for the first time a case of MEC in the tracheobronchial tree with the CRTC3-MAML2 fusion transcript. A 20-year-old man presented with non-productive cough and haemoptysis for 2 days. His medical history was significant for asthma and occasional haemoptysis (3–4 times/year) for the past 3 years. Physical examination was unremarkable. Haematological and biochemical test results were within normal limits. Electrocardiography was normal. Sputum smears and cultures were negative. Chest radiography was negative. Thoracic computed tomography (CT) scan with contrast, however, showed a 2.3 cm soft tissue mass in the left main bronchus. No mediastinal or hilar lymphadenopathy was noted. The patient underwent bronchoscopy. A large, irregular, hypervascular, polypoid mass was found in the left main bronchial orifice, obstructing the distal trachea and the left main bronchus. The mass was excised and a tracheobronchial stent was placed. The excised mass was then sent to the pathology department for examination. The fragmented specimen, measuring 2.2 1.5 0.6 cm in aggregate dimension, was fixed in 10% neutral buffered formalin, routinely processed and paraffin embedded. Tissue sections 3 mm thick were cut and stained with haematoxylin and eosin (H&E), mucicarmine, and periodic acid-Schiff with diastase. Tissue sections 4 mm thick prepared from the formalin fixed, paraffin embedded (FFPE) tissue block were deparaffinised, followed by the blockade of endogeneous peroxidases and antigen retrieval using antigenappropriate epitope retrieval. The tissue was immunohistochemically stained with antibodies to AE1/AE3, cytokeratin 7, cytokeratin 20, HMB45, p63, smooth muscle actin (Dako, Denmark), CAM 5.2 (BD Biosciences, USA), and thyroid transcription factor-1 (Novocastra, UK), per the manufacturers’ protocols. The epitopes of interest were localised using an avidin biotin immunoperoxidase method after heat-induced epitope retrieval, using 3, 3’-diaminobenzidine tetrahydrochloride as chromogen. Microscopic examination revealed a subepithelial, neoplastic cellular proliferation, organised into cysts, nests, solid sheets, and occasionally glands (Fig. 1A). The neoplasm was composed of three different cell types: mucinous, intermediate, and squamous. The mucinous cells had light eosinophilic cytoplasm. The polygonal intermediate cells had a centrally or eccentrically located bland nucleus, distinct nuclear contour, prominent nucleolus, and moderate amount of clear to light eosinophilic cytoplasm (Fig. 1B). The squamous cells demonstrated intercellular bridges. The overlying respiratory mucosa showed focal ulceration and focal squamous metaplasia.

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Nuclear pleomorphism, mitosis, necrosis, perineural or vascular invasion was not seen. Mucicarmine and periodic acid-Schiff with diastase stains highlighted the presence of intracellular and extracellular mucin. By immunohistochemistry, the tumour cells were strongly positive for cytokeratin 7, weakly positive for AE1/AE3 and CAM 5.2, and negative for cytokeratin 20, thyroid transcription factor-1, p63, smooth muscle actin, and HMB45. The combined histological and immunohistochemical findings were consistent with MEC, low to intermediate grade. Pathological grading was based on the Armed Forces Institute of Pathology (AFIP) histological classification proposed by Goode et al.8 Molecular study was then performed for the detection of CRTC3-MAML2 fusion transcript. Briefly, mRNA was extracted from FFPE tumour tissue using the Optimum FFPE RNA Isolation Kit (Ambion Diagnostics, USA), according to the manufacturer’s protocol. First strand cDNA synthesis was performed using High Capacity cDNA Reverse Transcription Kit (Applied Biosystems, USA), following the manufacturer’s instructions. The nested-polymerase chain reaction (PCR) amplification was performed in two rounds, using primers as previously described.9 The first round PCR amplification used forward (CRTCa 50 - AAGATCGCGCTGCACAATCA- 30 ) and reverse (MAML2a 50 - GGTCGCTTGCTGTTGGCAGG- 30 ) primers. The PCR reaction mixture contained 2 mL cDNA, 0.5 U AmpliTaq Gold DNA polymerase (Applied Biosystems), 2.5 mL of 10x PCR reaction buffer (with 15 mM MgCl2), 2.5 mL of 2.5 mM dNTP mixture, 0.25 mL of 10 mM primers, in a total volume of 25 mL. The PCR thermal cycling reaction was performed using the following conditions: 958C for 10 min, then 35 cycles of 958C for 45 s, 568C for 45 s, and 728C for 1 min using an Applied Biosystems 9800 Thermal Cycler. Then, 1 mL of the PCR product was used as template for the second round of PCR amplification with the forward (CRTCb 50 - GGAGGAGACGGCGGCCTTCG-30 ) and reverse (MAML2b 50 - TTGCTGTTGGCAGGAGATAG-30 ) primers using the same reaction conditions as the first round. RNA integrity and efficiency of cDNA synthesis were confirmed by RT-PCR for the housekeeping gene b-actin. PCR products were separated by electrophoresis on a 2% agarose gel and visualised by ethidium bromide staining. PCR amplicons were then purified and used as templates in cycle sequencing using BigDye Terminator Cycle Sequencing Ready Reaction Kit, version 1.1 (Applied Biosystems) in an Applied Biosystems 3130xl Genetic Analyzer. The sequences obtained were compared with those stored in GenBank databases using BLAST software (http://www.ncbi.nlm.nih.gov/blast).

B

Fig. 1 (A) The tumour mass is composed of neoplastic cells of different morphologies. Note the cystic formation and extracellular mucin (arrow) (H&E). (B) A mucicarmine stain highlights the intracellular mucin in a neoplastic mucinous cell (arrow). Note the lack of cytologic atypia (H&E).


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Patient

CRTC3

MAML2

MAML2

CRCT3

Forward

Reverse

A

B

Fig. 2 (A) Nested PCR products were resolved by electrophoresis on a 2% ethidium bromide-stained agarose gel and visualised on an UV source. A 100 bp PCR product (arrow) was detected. (B) Sequence analysis showing fusion of exon 1 of CRTC3 and exon 2 of MALM2.

Nested PCR amplified a fragment with a size of approximately 100 bp from the tumour cDNA (Fig. 2A). Sequence analysis showed that the amplified product was highly homologous to the Homo sapiens CRTC3-MAML2 fusion protein mRNA (GI:154082114), previously described by Fehr et al. in a case of salivary gland MEC.5 The product was formed by fusion of exon 1 of CRTC3 and exon 2 of MAML2 (Fig. 2B). The patient subsequently underwent a left pneumonectomy. No residual tumour was found in the bronchial stump or in the lung by macroscopic and microscopic examination. All eight peribronchial lymph nodes were also benign. Based on these clinicopathological findings, the patient received no adjuvant therapy. The post-operative course was uncomplicated. Twenty-six months following the left pneumonectomy operation, the patient was found to have tumour recurrence (2 cm) in the residual left main bronchus by bronchoscopy with biopsy. No lymphadenopathy or distant metastasis was noted on CT. The patient subsequently underwent radical radiotherapy, including two times 5 Gy endobronchial brachytherapy and 54 Gy external-beam radiotherapy (EBRT). Twelve weeks following EBRT, thoracic positron emission tomography (PET)/CT with contrast showed a single 2.59 cm soft tissue hypermetabolic mass in the bronchial stump. Bronchoscopy with biopsy confirmed the presence of persistent MEC in the medial posterior aspect of the most proximal portion of the left main bronchial stump. This persistent tumour was deemed unresectable due to its proximity to major mediastinal vessels. Thus, the patient opted for observation. At the time of writing, 5.5 years after the initial diagnosis, the patient is doing relatively well, and follow-up CT scans have so far not shown increase in tumour size post-EBRT. Shortly after the discovery by Fehr et al. of the CRTC3MAML2 fusion transcript in salivary gland MEC, Nakayama et al. screened a large series of MEC of the salivary gland and detected CRTC3-MAML2 fusion transcripts in 6% of cases. They found that CRTC3-MAML2 fusion conferred favourable clinicopathological features, including a less advanced clinical stage, negative nodal metastasis, no high grade tumour histology, no recurrence, and no tumour-related mortality after surgical resection of the tumour.6 In our patient, the CRTC3-MAML2 fusion-positive bronchial MEC also possessed no high grade histological features. While the patient did have recurrent tumour, it was felt that, based on the location of the recurrence and the absence of tumour in the pneumonectomy specimen, the persistent tumour was likely a

residual tumour, secondary to tumour seeding to the proximal left main bronchus during the pre-operative bronchoscopic procedure. Moreover, even though the tumour was not responsive to radiotherapy, it had not progressed in the past few years despite observation alone. Nonetheless, further investigations are required to elucidate the epidemiological and clinicopathological features of CRTC3-MAML2 fusion positive MEC arising in the tracheobronchial tree. Of note, the nested PCR primers used here, namely, CRTCa, MAML2a, CRTCb, and MAML2b, were published primers originally designed to amplify a CRTC1-MAML2 PCR product with a band size of 117 bp.9 However, the forward primers, CRTCa and CRTCb, were not only 100% homologous to CRTC1 but were also 100% homologous to CRTC3 (http:// www.ncbi.nlm.nih.gov/BLAST/). Additionally, these nested PCR primers can amplify a CRTC3-MAML2 PCR product of approximately 100 bp long. Therefore, direct sequencing of the breakpoint must be carried out to ascertain the identity of the fusion product and not to confuse one fusion product for another. In conclusion, we report a case of bronchial MEC harbouring the CRTC3-MAML2 fusion transcript. To our knowledge, this is the first reported case of CRTC3-MAML2 fusion positive MEC originating from the tracheobronchial tree. Conflicts of interest and sources of funding: The authors state that there are no conflicts of interest to disclose. Elaine S. Chan Joanna H. Tong Ka Fai To Department of Anatomical and Cellular Pathology, The Chinese University of Hong Kong, Hong Kong Contact Dr E. S. Chan. E-mail: [email protected] 1. Wang YL, Zhu YX, Chen TZ, et al. Clinicopathologic study of 1176 salivary gland tumors in a Chinese population: experience of one cancer center 19972007. Acta Otolaryngol 2012; 132: 879–86. 2. Leonardi HK, Jung-Legg Y, Legg MA, et al. Tracheobronchial mucoepidermoid carcinoma. Clinicopathological features and results of treatment. J Thorac Cardiovasc Surg 1978; 76: 431–8. 3. Clauditz TS, Gontarewicz A, Wang CJ, et al. 11q21 rearrangement is a frequent and highly specific genetic alteration in mucoepidermoid carcinoma. Diagn Mol Pathol 2012; 21: 134–7.


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4. Behboudi A, Enlund F, Winnes M, et al. Molecular classification of mucoepidermoid carcinomas-prognostic significance of the MECT1MAML2 fusion oncogene. Genes Chromosomes Cancer 2006; 45: 470–81. 5. Fehr A, Ro¨ser K, Heidorn K, et al. A new type of MAML2 fusion in mucoepidermoid carcinoma. Genes Chromosomes Cancer 2008; 47: 203–6. 6. Nakayama T, Miyabe S, Okabe M, et al. Clinicopathological significance of the CRTC3-MAML2 fusion transcript in mucoepidermoid carcinoma. Mod Pathol 2009; 22: 1575–81. 7. Okumura Y, Miyabe S, Nakayama T, et al. Impact of CRTC1/3-MAML2 fusions on histological classification and prognosis of mucoepidermoid carcinoma. Histopathology 2011; 59: 90–7. 8. Goode RK, El-Naggar AK. Mucoepidermoid carcinoma. In: Barnes L, Eveson J, Reichart P, Sidransky D, editors. Pathology and genetics of head and neck tumours. World Health Organization classification of tumours. Lyon: IARC Press, 2005;219–20. 9. Okabe M, Miyabe S, Nagatsuka H, et al. MECT1-MAML2 fusion transcript defines a favorable subset of mucoepidermoid carcinoma. Clin Cancer Res 2006; 12: 3902–7.

DOI: 10.1097/PAT.0000000000000014

Prominent angiotropism in a small atypical spitzoid dysplastic melanocytic naevus: what is its significance? Sir, Since the histopathological features of Spitz naevus overlap with those of melanoma, it remains a challenging diagnostic entity and a lack of consensus persists as to how to classify and manage lesions that share features of both entities.1 Angiotropism, the spread of tumour cells on the external (abluminal) surface of vessels, is a feature recently reported to be of prognostic significance in melanoma.2,3 It has also been reported to occur in atypical spitzoid tumours (AST) where its presence correlated with lymph node metastasis.4 However, it is also a virtually ubiquitous feature in congenital melanocytic naevi (CMN).2 Although the presence of

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angiotropism may correlate with clinical behaviour in bona fide AST, its significance as a histological criterion for the diagnosis and classification of melanocytic lesions remains uncertain. Herein described is a challenging melanocytic lesion from a 20-year-old male that displayed prominent angiotropism and broadens the reported spectrum of melanocytic tumours that may be associated with this phenomenon. A 20-year-old male reported a recent change in a longstanding pigmented cutaneous lesion on the neck. A shave excision was performed, the 11 8 2 mm specimen bearing a 5 mm irregular brown papule. Histologically (Fig. 1A), the lesion was a relatively circumscribed compound melanocytic lesion with accompanying mild regular acanthosis. The junctional component had a lentiginous and focally nested architecture. The melanocytes were mostly plump, often pigmented, with a variable amount of cytoplasm and variably enlarged nuclei with prominent nucleoli, focally prominent cytological atypia and junctional mitoses (Fig. 1B). There were compact junctional nests as well as vertically orientated nests of epithelioid and spindle shaped lightly pigmented melanocytes resembling those seen in Spitz naevus. Single cell pagetoid spread was noted within the epidermis, mostly centrally, but also at the periphery of the lesion (Fig. 1C). The epidermis showed surface compact and parakeratotic hyperkeratosis suggesting superficial ‘irritation’. Similar plump dermal melanocytes were present to a Breslow depth of 0.7 mm, Clark level IV. No dermal mitoses were identified. Frequent papillary and reticular dermal capillaries and small venules beneath the junctional component showed angiotropism by plump melanocytes (Fig. 2A–C), confirmed by dual melan-A and CD31 immunohistochemistry (Fig. 2D). Vessels showing angiotropism did not express D2-40 indicating they were not lymphatics. The junctional component was concerning for melanoma in situ, but in view of the overall pattern and features suggesting surface ‘irritation’ a diagnosis of ‘irritated’ spitzoid dysplastic compound naevus with angiotropism was favoured after expert review. Re-excision was

A

B

C

Fig. 1 (A) Overview scan of lesion (H&E). (B) Junctional cytological atypia with mitoses arrowed (H&E). (C) Edge of lesion with single cell pattern (H&E).


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A

C

B

D

Fig. 2 (A) Central portion of naevus with angiotropic foci arrowed (H&E). (B,C) Angiotropic melanocytes (H&E). (D) Double stain immunohistochemistry demonstrating melanocytes (Melan A, red chromagen) in a pericytic location external to the endothelial lining of small vessels (CD31, immunoperoxidase).

advised and this showed no residual lesion. Sentinel lymph node biopsy was not performed. Despite diagnostic guidelines, Spitz naevus and AST are prone to misdiagnosis as they share many histological features in common with melanoma.5 It is now well documented that it can be extremely difficult to predict the biological behaviour of AST, particularly when occurring in adult patients, and as a consequence many such tumours are regarded as being of uncertain biological potential.5 There is a lack of consensus between experts as to the diagnosis of AST, its separation from melanoma and prognosis; in fact, in one study most experts favoured a Spitz naevus for some ultimately lethal tumours.6 Molecular testing, not yet widely available, may aid diagnosis of melanoma. Melanomas harbour multiple chromosomal aberrations. Gene mutations such as of BRAF and NRAS are common in melanoma, but uncommon in typical Spitz naevi.7 Raskin et al.8 recently detected gross chromosomal aberrations in seven of 16 AST in contrast to eight Spitz naevi. The findings of this study suggest that AST might be a distinct entity with chromosomal aberrations different from melanoma and benign Spitz naevi. Nevertheless, they acknowledged that AST with chromosomal aberrations might be a low grade or indolent form of melanoma. Evidence supporting this assertion includes that such lesions tended to occur in younger patients with a more favourable outcome compared to melanoma of similar Breslow thickness and sentinel node biopsy status. Traditionally metastasis is thought to occur via an intraluminal route in lymphatics and blood vessels, but the presence of circulating melanoma cells does not predict metastasis. Metastasis can also occur via ‘extracellular lattices’. The latter includes angiotropism, in this case along external vessel lattices.9 Angiotropism has received attention in the last decade, largely due to the observations of Lugassy and Barnhill and their colleagues.2 Angiotropic melanocytes are characteristically closely opposed to the external endothelial surface

of vessels in a manner similar to pericytes and migrate in a pericytic location. The mechanism of migration along the external surface of structures has been termed ‘extravascular migratory metastasis’ (EVM).2 Histologically, Lugassy and Barnhill2 define angiotropism as unequivocal melanoma (or other tumour) cells closely opposed to the external surfaces of the endothelium of microvascular and/or lymphatic channels in linear arrays or aggregates, or in a smooth muscle location in larger vessels, in at least two or more foci at the advancing edge of the melanoma or in nearby tissue (most often within 1–2 mm) in the absence of intravascular or intralymphatic tumour aggregates. Angiotropism is reportedly far more common in melanoma than intravascular dissemination and is associated with metastasis. Indeed, it is suggested that the appearance of lymphovascular invasion may often be an artefact of vessel folding, or tissue shrinkage around clusters of melanoma.2 However, Xu et al.10 demonstrated that 35% of their melanoma cases had prognostically significant, genuine lymphatic invasion. The definition of angiotropism proposed by Barnhill and Lugassy excludes the presence of intravascular or intralymphatic tumour aggregates.2 However, the two are not mutually exclusive3 and indeed, although the presence of angiotropism has been linked to regional lymph node and distant metastasis,2 tumour growing along vessels is presumably more likely to become intravascular, which more easily explains the increased frequency of metastatic spread than simply by extravascular migratory metastasis.3 Factors associated with angiotropism include increasing Breslow thickness, dermal mitotic rate, Clark level, presence of ulceration and absence of regression. The presence of angiotropism is reported to be associated with the development of local and in-transit recurrence and is an independent predictor of reduced disease-free survival.3 Primary melanoma lymphangiogenesis determined by lymphatic vessel density (LVD) may also be positively associated


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with lymph node metastasis and may more accurately predict nodal metastasis than Breslow thickness. Furthermore, the pattern of LVD is significant and the ratio of peripheral:central LVD has been reported to be predictive of lymph node metastasis.11 However, unlike arduous LVD determination, angiotropism is a qualitative finding and easy to record in routine pathology reporting.2 EVM is strongly analogous to the migration of embryonic stem cells from the neural crest along the abluminal surfaces of vessels.2 The interaction between angiotropic melanoma cells and pericytes induces novel differential gene expression associated with cancer migration and pericytic/melanoma stem cell properties, termed ‘perivascular mimicry’.2 Gene expression profiling in angiotropic melanoma has identified 15 genes potentially involved in EVM, including those known to be associated with migration of neural crest cells (NCC). Two gene products were immunohistochemically validated in tissue sections. Overexpression of Treacle protein and under-expression of AHNAK protein was demonstrated only in angiotropic melanoma cells, strong objective evidence that EVM utilises mechanisms of NCC migration.2 Angiotropism is not exclusive to melanoma and has recently been reported in nine cases of AST, five of whom showed regional tumour spread, mostly within 2 years of diagnosis of the primary tumour. Almost all foci of angiotropism were associated with blood microvascular channels and not lymphatics.4 Documentation of angiotropism in typical Spitz naevi is lacking. Howatt and Variend12 recorded ‘lymphatic invasion’ in 14.3% of non-recurring Spitz naevi in children, although one image in that report closely resembles angiotropism. No reference was found to angiotropism in eruptive or recurrent Spitz naevi. However, angiotropism was not noted in a case of a Spitz naevus with multiple satellite lesions.13 Angiotropism has been reported once in a deep penetrating naevus.14 Angiotropism is reported to occur in 30–100% of CMN.2 Lymphatic invasion recorded in a CMN with satellites may also rather represent angiotropism.15 These findings are presumed to reflect the neural crest origin of CMN, explained by migratory phenomena and that neural crest stem cells arriving in skin (or elsewhere) via EVM may play an important role in the development of CMN. EVM in melanoma may indeed represent ‘reverse migration’, in recapitulating this pattern of embryonic migration, representing an inherent behavioural trait of melanocytes.2 Angiotropism is also described in other naevi, including a common blue naevus with satellites16 and a composite plaque-type blue naevus and neurocristic hamartoma.17 Because angiotropism is described in a broad spectrum of melanocytic lesions, the obvious question is ‘when does angiotropism become an adverse prognostic feature?’ Its significance in melanoma appears clear, and in AST one study correlated it with local recurrence and lymph node metastasis.4 However, the situation is less clear in settings such as the current case. When angiotropism is present, are those melanocytes ‘stuck’ in a perivascular location following migration into the skin, or are they in the process of exiting the skin (reverse migration)? Novel immunohistochemical markers2 may offer a new means to stratify these lesions and to guide which of them require wide excision and sentinel node biopsy.

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Conflicts of interest and sources of funding: The authors state that there are no conflicts of interest to disclose. Mark J. Wilsher* Stanley W. McCarthy{{§ Richard A. Scolyer{{§ *Douglass Hanly Moir Pathology, Macquarie Park, {Melanoma Institute Australia, zDiscipline of Pathology, Sydney Medical School, The University of Sydney, Sydney, and §Department of Tissue Pathology and Diagnostic Oncology, Royal Prince Alfred Hospital, Sydney, NSW, Australia Contact Dr M. J. Wilsher. E-mail: [email protected] 1. Dahlstrom JE, Scolyer RA, Thompson JF, Jain S. Spitz naevus: diagnostic problems and their management implications. Pathology 2004; 36: 452–7. 2. Lugassy C, Barnhill RL. Angiotropism and extravascular migratory metastasis in melanoma: from concept to gene expression. Exp Rev Dermatol 2011; 6: 303–16. 3. Van Es SL, Colman M, Thompson JF, McCarthy SW, Scolyer RA. Angiotropism is an independent predictor of local recurrence and in-transit metastasis in primary cutaneous melanoma. Am J Surg Pathol 2008; 32: 1396–403. 4. Barnhill RL, Kutzner H, Schmidt B, et al. Atypical spitzoid melanocytic neoplasms with angiotropism: a potential mechanism of locoregional involvement. Am J Dermatopathol 2011; 33: 236–43. 5. McCarthy SW, Scolyer RA. Pitfalls and important issues in the pathologic diagnosis of melanocytic tumors. Ochsner J 2010; 10: 66–74. 6. Barnhill RL, Argenyi ZB, From L, et al. Atypical Spitz nevi/tumors: lack of consensus for diagnosis, discrimination from melanoma, and prediction of outcome. Hum Pathol 1999; 30: 513–20. 7. Scolyer RA, Murali R, McCarthy SW, Thompson JF. Histologicallyambiguous (’borderline’) primary cutaneous melanocytic tumors: approaches to patient management including the roles of molecular testing and sentinel lymph node biopsy. Arch Pathol Lab Med 2010; 134: 1770–7. 8. Raskin L, Ludgate M, Iyer RK, et al. Copy number variations and clinical outcome in atypical Spitz tumors. Am J Surg Pathol 2011; 35: 243–52. 9. Zbytek B, Carlson JA, Granese J, Ross J, Mihm MC Jr, Slominski A. Current concepts of metastasis in melanoma. Expert Rev Dermatol 2008; 3: 569–85. 10. Xu X, Gimotty PA, Guerry D, et al. Lymphatic invasion revealed by multispectral imaging is common in primary melanomas and associates with prognosis. Hum Pathol 2008; 39: 901–9. 11. Shayan R, Karnezis T, Murali R, et al. Lymphatic vessel density in primary melanomas predicts sentinel lymph node status and risk of metastasis. Histopathology 2012; 61: 702–10. 12. Howatt AJ, Variend S. Lymphatic invasion in Spitz naevi. Am J Surg Pathol 1985; 9: 125–8. 13. Song JY, Kwon JA, Park CJ. Case of Spitz naevus with multiple satellite lesions. J Am Acad Dermatol 2005; 52: S48–50. 14. Hassan AS, Schulte KW, Ruzicka T, Megahed M. Linear arrangement of multiple deep penetrating nevi. Report of first case and review of literature. Arch Dermatol 2003; 139: 1608–10. 15. Sasai S, Kato T, Yoshimura T, Suetake, Tugami H. Congenital plantar melanocytic nevus with satellite lesions. Dermatology 1996; 192: 146–8. 16. Kang DS, Chung KY. Common blue naevus with satellite lesions: possible perivascular dissemination resulting in a clinical resemblance to malignant melanoma. Br J Dermatol 1999; 141: 922–5. 17. Bevona C, Tannous Z, Tsao H. Dermal melanocytic proliferation with features of a plaque-type blue nevus and neurocristic hamartoma. J Am Acad Dermatol 2003; 49: 924–9.

DOI: 10.1097/PAT.0000000000000019


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Mixed dendritic cell tumours with follicularfibroblastic dendritic cell features of lymph node

Sir, Dendritic cells are non-lymphoid, non-phagocytic, immune accessory cells that are essential for antigen presentation and they are present in both lymphoid and non-lymphoid organs. Four types of dendritic cells are present in the lymph nodes: follicular, interdigitating, Langerhans and histiocytic/fibroblastic cells.1 After the first description of a primary sarcoma in the lymph node with dendritic reticulum cell differentiation by Monda et al. in 1986,2 more than 200 cases have been reported to date. The World Health Organization (WHO) classified dendritic cell neoplasms into five groups: Langerhans cell histiocytosis (LCH), Langerhans cell sarcoma (LCS), interdigitating dendritic cell sarcoma/tumour (IDCS), follicular dendritic cell sarcoma/tumour (FDCS), and dendritic cell tumour, not otherwise specified. FDC sarcoma is an uncommon tumour that usually involves lymph nodes but may arise at a variety of extranodal sites, including the oral cavity, tonsil, gastrointestinal tract, and liver.3–7 Among reticulum cell neoplasms, fibroblastic reticular cell (FRC) tumour has been described as a new subtype, based on the expression of vimentin, SMA, and desmin.8 This tumour is an even more rare neoplasm, and few cases have been reported in English literature.8,9 Here, we report a rare case of primary lymph node dendritic cell sarcoma with mixed FDC and FRC features. An 18-year-old female patient presented with a slowgrowing mass in the left neck for 2 months. The vital signs were stable, and there was no evidence of fever or weight loss. Physical examination revealed a well-circumscribed, non-tender mass involving the left cervical areas. Computed tomography (CT) scan demonstrated extensive enlargement of the lymph nodes in the left submaxillary region. There was no radiographic evidence of lymph node enlargement in the mediastinal and abdominal areas. Laboratory examination revealed that all parameters were within normal levels. Chest X-ray also revealed no abnormality. The clinical impression was lymphadenitis or a lymphoma. An excisional biopsy of a left cervical lymph node was performed. Macroscopically, the lymph node measuring 3.5 3.5 2.8 cm was well circumscribed and had a fibrous capsule. The freshly cut surface of the mass was slightly firm, greyyellow in colour, and exhibited focal necrotic areas. Microscopically, the lymph node was almost completely effaced by a tumour with many scattered huge cells in a background of small lymphocytes and plasma cells (Fig. 1A). The tumour cells had vesicular, bizarre and irregularly folded nuclei occurring individually, sometimes with a grouping pattern but not forming tight nests. The bizarre cells had coarse chromatin and distinct nucleoli, and sometimes they contained intranuclear eosinophilic biotin inclusions (Fig. 1B). The cytoplasm was eosinophilic, but cell borders were difficult to define. The mitotic count was approximately 20–25 per 10 high power fields. Focal areas of haemorrhage and necrosis were seen, and Castleman’s disease lesion was absent. In immunohistochemical studies, the most strikingly positive stain was desmin. More than 20% of these large tumour cells were positive for desmin (Fig. 2A), and they showed dendritic

Fig. 1 (A) The tumour is comprised of many scattered huge cells in a background of small lymphocytes and plasma cells (H&E). (B) Tumour cells with vesicular and polymorphic nuclei have distinct nucleoli, and abundant ill-defined eosinophilic cytoplasm (H&E).

cell processes. A few tumour cells were positive for the FDC associated markers CD21, CD35 and D2-40 (Fig. 2B,C). There was no staining of the tumour cells for cytokeratin, SMA, S-100, CD1a, CD3, CD20, CD68, CD61 and Langerin. The proliferation index (Ki-67) of the neoplastic cells was approximately 30%. Infiltrating lymphocytes were mainly positive for CD20. In situ hybridisation for EBV-encoded RNA-1 (EBER1) was negative. The patient received post-operative radiotherapy. A total radiation dose of 60 Gray in 30 equal fractions was given for 6 weeks. Post-operative follow-up of 37 months has been uneventful. Based on the clinical and morphological results, we consider this case as mixed dendritic cell tumour with hybrid features between follicular dendritic cell and fibroblastic dendritic cell. The diagnosis of dendritic cell sarcomas can be challenging because of their morphological overlap with each other and many other neoplasms. They were previously classified as lymphomas, sarcomas, or histiocytic neoplasms. Therefore, immunohistochemical and/or ultrastructural studies are very important in their differential diagnosis. In this case, the differential diagnosis included primary tumour (dendritic cell neoplasms), metastatic carcinoma, large cell lymphoma, and acute megakaryoblastic leukaemia. In our case, histological examination revealed the lymph node was almost completely effaced by a tumour with many scattered huge cells in a


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Fig. 2 (A) Tumour cells, with a dendritic morphology, demonstrate strongly immunoreactive for desmin. (B,C) A few tumour cells are positive for the FDC associated markers (B) CD35 and (C) D2-40.

background of small lymphocytes and plasma cells. These tumour cells had vesicular nuclei with prominent nucleoli and long cytoplasmic processes that showed the dendritic morphology. Immunophenotypic studies demonstrated tumour cells were negative for cytokeratin, lymphoid cell markers (CD3 and CD20), and megakaryocyte marker (CD61), so tumours associated with these markers were excluded. In addition, the tumour cells were negative for S-100, CD1a, CD68 and Langerin, but positive for FDC associated markers CD21, CD35 and D2-40. In view of immunophenotypic

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staining and the dendritic morphology, follicular dendritic cell sarcoma had to be seriously considered, and an indeterminate cell or a Langerhans cell tumour was ruled out. On the other hand, the striking desmin staining, dendritic cytology and lack of usual morphological features of follicular dendritic cell tumour raised the alternative possibility of fibroblastic dendritic cell tumour. So, in view of the dendritic morphology and the mixed pattern of immunophenotypic staining, we consider this a mixed dendritic cell tumour with hybrid features between follicular dendritic cell and fibroblastic dendritic cell. To our knowledge, this is the second case of mixed dendritic cell tumour with hybrid features between FDC and FRC in the English literature.10 At present, the definite aetiology of FDC sarcoma and FRC tumour is poorly understood. In 10–20% of cases, FDC sarcoma is associated with Castleman’s disease, usually hyaline vascular-type Castleman’s disease, which can be concurrent with or precede the sarcoma. It has been proposed that Castleman’s disease might act as a precursor lesion to follicular dendritic cell sarcoma through a hyperplasia–dysplasia– neoplasia sequence.11 However, it is also possible that the characteristic Castleman’s disease lesion is a reaction to FDC sarcoma rather than a precursor lesion.12 EBV has been documented to be positive in a small percentage of FDCS cases, and most of these cases involve the liver or spleen and demonstrate inflammatory pseudotumour (IPT)-like morphology.4,6 In our present case, EBV was not detected within the tumour cells by in situ hybridisation for EBER1, and Castleman’s disease was absent. FRC tumour is an even more rare neoplasm than FDC sarcoma, and only few cases have been reported in the literature.8,9 Due to the limited cases, the range of features associated with FRC tumour is not yet clearly delineated. FDC sarcoma always shows a low-to-intermediate malignant potential with a tendency to local recurrence, but little risk of metastasis. It is difficult to provide treatment recommendations. Current best-available approach is to apply therapeutic guidelines similar to those used for soft tissue sarcomas of high grade. It can be effectively treated by only complete surgical resection for localised disease. In addition, adjuvant chemotherapy and radiotherapy has been reserved for patients with metastatic disease and/or after failure of primary treatment. In our present case, the patient was treated with mass resection and post-operative radiotherapy and she is alive with no evidence of disease recurrence for more than 3 years. The neoplasm in this case exhibited an indolent behaviour. This may suggest that the behaviour of these hybrid neoplasms is more akin to FDC sarcoma. In conclusion, we present a rare case of primary lymph node dendritic cell sarcoma with mixed FDC and FRC features. This tumour highlights the relationship between these dendritic cell types and might represent a new subtype or new differentiation step of dendritic cell sarcomas that is not understood at present. Acknowledgements: We would like to thank Dr John K. C. Chan of the Department of Pathology, Queen Elizabeth Hospital, Hong Kong for the consultation and some immunophenotypic staining for this case. Conflicts of interest and sources of funding: The authors state that there are no conflicts of interest to disclose.


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Xiuming Zhang Guoping Ren Hongtian Yao Zhaoming Wang Department of Pathology, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, Zhejiang Province, China Contact Dr Z. Wang. E-mail: [email protected] 1. Fonseca R, Yamakawa M, Nakamura S, et al. Follicular dendritic cell sarcoma and interdigitating reticulum cell sarcoma: a review. Am J Hematol 1998; 59: 161–7. 2. Monda L, Warnke R, Rosai J. A primary lymph node malignancy with features suggestive of dendritic reticulum cell differentiation. A report of 4 cases. Am J Pathol 1986; 122: 562–72. 3. Chan JKC, Tsang WY, Ng CS, et al. Follicular dendritic cell tumours of the oral cavity. Am J Surg Pathol 1994; 18: 148–57. 4. Shek TWH, Ho FCS, Ng IOL, et al. Follicular dendritic cell tumour of the liver: evidence for an Epstein-Barr virus related clonal proliferation of follicular dendritic cells. Am J Surg Pathol 1996; 20: 313–24. 5. Chan JKC, Fletcher CDM, Cooper K. Follicular dendritic cell sarcoma: clinicopathologic analysis of 17 cases suggesting a malignant potential higher than currently recognised. Cancer 1997; 79: 294–313. 6. Cheuk W, Chan JKC, Shek TWH, et al. Inflammatory pseudotumourlike follicular dendritic cell tumour: a distinctive low grade malignant intra-abdominal neoplasm with consistent Epstein-Barr virus association. Am J Surg Pathol 2001; 25: 721–31. 7. Dillon KM, Hill CM, Cameron CH, et al. Mediastinal mixed dendritic cell sarcoma with hybrid features. J Clin Pathol 2002; 55: 791–4. 8. Andriko JA, Kaldjian EP, Tsokos M, Abbondanzo SL, Jaffe ES. Reticulum cell neoplasms of lymph nodes: a clinicopathologic study of 11 cases with recognition of a new subtype derived from fibroblastic reticular cells. Am J Surg Pathol 1998; 22: 1048–58. 9. Sua´rez D, Izquierdo FM, Me´ndez JR, Escobar J, Cabeza A, Junco P. Tumor of fibroblastic reticular cells of lymph node coincidental with an undifferentiated endometrial stromal sarcoma. Report of a case with distinctive immunophenotype and Kikuchi-like necro-inflammatory response. APMIS 2011; 119: 216–20. 10. Jones D, Amin M, Ordonez NG, et al. Reticulum cell sarcoma of lymph node with mixed dendritic and fibroblastic features. Mod Pathol 2001; 14: 1059–67. 11. Rodrı´guez Silva H, Buchaca Faxas E, Machado Puerto I, Pe´rez Roma´n G, Pe´rez Caballero D. Castleman’s disease: Review of five cases. (Spanish.) An Med Interna 2005; 22: 24–7. 12. Chan ACL, Chan KW, Chan JKC, et al. Development of follicular dendritic cell sarcoma in hyaline-vascular Castleman’s disease of the nasopharynx: tracing its evolution by sequential biopsies. Histopathology 2001; 38: 510–8.

DOI: 10.1097/PAT.0000000000000011

The tubal fimbria: the old broom sweeps into prominence Sir, It is occasionally salutary to reflect on what are the drivers of pathologists’ observations and to concede the bidirectional crosstalk between clinical imperatives and our diagnostic capabilities. What we see is what we look at and what we look at is what is required of us. Responding to the needs of clinical colleagues (and their patients) more frequently alters our observational patterns than we might care to admit, yet the common good—shared knowledge and understanding—is an ineluctable benefit. An admirable case in point for


gynaecological pathologists is our current preoccupation with the fimbriae of the fallopian tubes, for so long simply the brooms that swept extruded oocytes on to greater things and barely expanded standard pathology textbooks by more than half a column inch. Yet the story of risk-reducing gynaecological surgery for women who carry BRCA1 or BRCA2 germline mutations and other women at high risk of developing pelvic serous cancer has changed all that with one-half of all PubMed cited references on pathology of the tubal fimbria occurring since the year 2000. Naturally enough, very many of these focus on early or occult serous cancers in the ovaries and fallopian tubes and the tubal epithelial precursors of high grade pelvic serous carcinomas, the so-called p53 signatures (Fig. 1A) and serous tubal intraepithelial carcinoma (Fig. 1B) or STIC lesions1–3 and even secretory cell outgrowths or SCOUTs4 enriching our lexicon with new epithets and acronyms. Routinely paraffin embedding all the tubal tissue is de rigueur in these high risk cases but the practice has spilled over into many routine adnexal specimens. Not surprisingly then, serendipitous ‘discoveries’ have abounded and have included such entities as transitional cell metaplasias at the fimbrial epithelial-peritoneal interface (Fig. 2A)5 and small benign fimbrial adenofibromas (Fig. 2B),6 usually by the same investigators who have reported premalignant changes in the tubal fimbria and presumably as a direct consequence of their greatly increased perusal of such tissues. The same can be said of heterotopic Leydig or hilus cells, a striking example of which we recently encountered. The patient, a 60-year-old asymptomatic woman at increased risk of developing high grade pelvic serous carcinoma due to a BRCA2 germline mutation, underwent risk-reducing bilateral salpingo-oophorectomy. She had a history of breast cancer, removed at age 32 years, with her last treatment 12 months ago and currently no evidence of active disease. The right and left adnexal structures were macroscopically completely normal with combined weights of 5.6 g and 6.1 g, respectively. All of the submitted tissue was processed and embedded according to the current laboratory protocol for such specimens.7 Histologically, neither the ovaries nor the fallopian tubes showed any epithelial neoplastic or preneoplastic changes relevant to the breast cancer or to the increased risk of adnexal carcinoma. Both ovaries, however, exhibited moderate hilus cell hyperplasia in the hilar regions and the mesovaria, the largest nodule, in the right mesovarium, measuring 2 mm across (Fig. 3A). Likewise, both fallopian tubes displayed several small, partly circumscribed nests or patternless sheets of hilus cells in fimbrial processes (Fig. 3B), some near the tips and others at the bases. These aggregates were comprised of large polyhedral cells with abundant pale eosinophilic cytoplasm, occasionally vacuolated and focally containing cytochrome pigments. No Reinke crystals were identified. These cells were strongly immunoreactive for calretinin and a-inhibin (Fig. 3C), non-reactive for CK7 and only weakly and focally reactive for pan-cytokeratin. They had rounded, uniform, pale nuclei with small eccentric nucleoli, minimal atypia and no mitoses (Fig. 3D). A background meshwork of capillary vessels was apparent. Although these aggregates showed some irregularity at their perimeters, none displayed the finely tentacular or Indian-file pattern of distribution reported by Hirschowitz and co-workers.8 Hilus cells are steroid-producing cells of uncertain histogenesis, but most likely derived from perineural or perivascular


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Fig. 1 (A) Tubal fimbrial process showing a typical p53 signature lesion, a strip of at least 12 contiguous p53 positive secretory epithelial cell nuclei. (B) Serous tubal intraepithelial carcinoma (STIC) with disordered growth of secretory type cells with high nuclear grade, but no invasion of the subjacent stroma (H&E).

Fig. 2 (A) Small discrete plaques of transitional cell metaplasia adjacent to the fimbrial epithelial-peritoneal interface. Such lesions are thought to be precursors of Walthard cell nests (H&E). (B) A minute benign fimbrial adenofibroma with stroma indistinguishable from ovarian cortical stroma and a minor epithelial component (H&E).

Fig. 3 (A) A circumscribed nodule of hyperplastic hilus cells in the right mesovarium. Two small mesonephric ductular remnants are present at centre left (H&E). (B) Sheet-like aggregates of heterotopic hilus cells within villous processes of the right tubal fimbria (H&E). (C) Same field as B immunostained for a-inhibin. (D) A patternless sheet of plump eosinophilic hilus cells with small round nuclei. Note the background meshwork of small vessels (H&E).


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fibroblasts,9 and found typically in the hilar region of the ovaries and adjacent mesovarium.10 Their presence in abnormal sites is rarely observed, in the stroma of the tubal fimbria (as here) and the walls of paratubal cysts,8,11–13 but also in the subcapsular cortex of the ovaries.8 It is arguable whether the latter position is truly heterotopic or not, inasmuch as some of the reported cases were associated with hilus cell hyperplasia and stromal luteinisation,8,11 and it may be that hilus or Leydig cells are sometimes randomly present in the subcapsular ovarian cortex and only visible under circumstances of hyperplasia or hyperstimulation. A number of cases identified by Honore and O’Hara,11 for example, were in post-partum tubal sterilisation specimens. The differential diagnosis is limited, for all practical purposes, to discriminating these small patternless sheets or aggregates of endocrine cells from the larger, well circumscribed and much more common adrenal cell nests,14 not on cytological detail but on the low power microarchitecture typical of the latter. Adrenal cell nests exhibit a peripheral capsule and defined organoid layers recapitulating those seen in the adrenal cortex. Their anatomical distribution differs from that of hilus cell heterotopias in that they are more typically seen in the mesovaria and mesosalpinges. Other unlikely possibilities might include metastatic adenocarcinoma, particularly given the clinical context and particularly if the hilus cell aggregates are poorly circumscribed or exhibit an Indian-file pattern, a problem addressed in one series,8 and militated against by the blandness and uniformity of the constituent cells and the non-epithelial immunoprofile. Hilus cell heterotopia with (as here) or without hilus cell hyperplasia appears, from all accounts, to be a pathological curiosity of no clinical significance. It serves only to remind us of the benefits of keen and targeted observation. Conflicts of interest and sources of funding: The authors state that there are no conflicts of interest to disclose. Peter Russell*{ Suzanne Hyne* Sue Valmadre{ *GynaePath, Douglass Hanly Moir Pathology, Macquarie Park, {Department of Obstetrics Gynaecology and Neonatology, University of Sydney, and zNorth Shore Private and Royal North Shore Hospitals, Sydney, NSW, Australia Contact Professor P. Russell. E-mail: [email protected] 1. Crum CP, Drapkin R, Kindelberger D, et al. Lessons from BRCA: the tubal fimbria emerges as an origin for pelvic serous cancer. Clin Med Res 2007; 5: 35–44. 2. Crum CP, McKeon FD, Xian W. BRCA, the oviduct, and the space and time continuum of pelvic serous carcinogenesis. Int J Gynecol Cancer 2012; 22 (Suppl 1): S29–34. 3. Saleemuddin A, Folkins AK, Garrett L, et al. Risk factors for a serous cancer precursor (’p53 signature’) in women with inherited BRCA mutations. Gynecol Oncol 2008; 111: 226–32. 4. Mehra K, Mehrad M, Ning G, et al. STICS, SCOUTs and p53 signatures; a new language for pelvic serous carcinogenesis. Frontiers Bioscience 2011; 3: 625–34. 5. Seidman JD, Yemelyanova A, Zaino RJ, Kurman RJ. The fallopian tubeperitoneal junction: a potential site of carcinogenesis. Int J Gynecol Pathol 2011; 30: 4–11.


6. Bossuyt V, Medeiros F, Drapkin R, et al. Adenofibroma of the fimbria: a common entity that is indistinguishable from ovarian adenofibroma. Int J Gynecol Pathol 2008; 27: 390–7. 7. Kiely BE, Friedlander ML, Milne RL, et al. Adequacy of risk-reducing gynaecologic surgery in BRCA1 or BRCA2 mutation carriers and other women at high risk of pelvic serous cancer. Famil Cancer 2011; 10: 505–14. 8. Hirschowitz L, Salmons N, Ganesan R. Ovarian hilus cell heterotopia. Int J Gynecol Pathol 2011; 30: 46–52. 9. DeFalco T, Takahashi S, Capel B. Two distinct origins for Leydig cell progenitors in the fetal testis. Dev Biol 2011; 352: 14–26. 10. Russell P, Robboy SJ. Normal ovaries, inflammatory and non-neoplastic conditions. In: Robboy SJ, Mutter GL, Pratt JH, Bentley RC, Russell P, Anderson MC, editors. Robboy’s Pathology of the Female Reproductive Tract. 2nd ed. London: Churchill Livingstone, 2009; 543–568. 11. Honore LH, O’Hara KE. Ovarian hilus cell heterotopia. Obstet Gynecol 1979; 53: 461–4. 12. Lewis JD. Hilus-cell hyperplasia of ovaries and tubes; report of a case. Obstet Gynecol 1964; 24: 728–31. 13. Palomaki JF, Blair OM. Hilus cell rest of the fallopian tube. A case report. Obstet Gynecol 1971; 37: 60–2. 14. Zheng W, Robboy SJ. Fallopian tube. In: Robboy SJ, Mutter GL, Pratt JH, Bentley RC, Russell P, Anderson MC, editors. Robboy’s Pathology of the Female Reproductive Tract. 2nd ed. London: Churchill Livingstone, 2009; 485–513.

DOI: 10.1097/PAT.0000000000000017

Hydrops fetalis, hepatic centrolobular necrosis, and hypoxic-ischaemic encephalopathy in a fetus with premature closure of foramen ovale Sir, Premature closure of foramen of ovale (PCFO) is a very rare congenital anomaly. We report a case of a stillborn fetus delivered at 20 gestational weeks. Post-mortem examination revealed PCFO, hydrops fetalis, dilatation of the right atrium, hepatic centrilobular necrosis, and hypoxic-ischaemic encephalopathy. We describe this case due to its rarity and discuss possible pathogenesis. A 37-year-old woman with no significant medical history delivered a female stillborn fetus at 20 weeks’ gestation. Screening for Down’s syndrome at 11–13 weeks of gestation was negative. A fetal autopsy was performed. The female stillborn fetus was moderately macerated and slightly small for gestational age (body weight: 259 g, expected 319 61 g; crown–heel length: 22.0 cm, expected 24.7 1.9 cm). The skin was oedematous, with particularly prominent scalp oedema (Fig. 1). No other external dysmorphic features were appreciated. Internal examination revealed scanty bilateral pleural effusions and ascites (

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