—  SPECIALTY CONFERENCE  —

Genitourinary Pathology

Case 1 - Ewing's Sarcoma/PNET of the Kidney

Hema Samaratunga
Sullivan Nicolaides Pathology,
Brisbane, Australia


Click on each slide thumbnail image for an enlarged view
Case History
A 52-year- old female presented with a two week history of right flank pain and gross hematuria. Ultrasound scan revealed a 5.9 cm solid / cystic mass in the lower right kidney which on CT was a heterogeneously enhancing mass, thought to be a renal cell carcinoma. A radical nephrectomy was performed.

Gross Pathologic Features
Grossly, several masses of fleshy white focally necrotic and haemorrhagic tumor involved the lower kidney. The largest measured 6 cm and was cavitated. The tumor involved predominantly the medulla, and extended into renal vein.


Case 1 - Slide 1
Click to view with ImageScope
Click to view with a Web-Based Viewer



Case 1 - Figure 1
Low power photomicrograph illustrating broad sheets of tumor infiltrating renal parenchyma. The tumor has delicate thin walled blood vessels.
Case 1 - Figure 2
The tumor has a cystic component with many of the cysts containing serofibrinous fluid or blood.
Case 1 - Figure 3
Highly cellular tumor infiltrates renal sinus fat.

Case 1 - Figure 4
Tumor growing into proximal renal vein. Note tumor attachment to the renal vein wall.
Case 1 - Figure 5
The tumor cells are haphazardly arrayed around cysts.
Case 1 - Figure 6
Focally a vaguely nested pattern is identified.

Case 1 - Figure 7
Focal necrosis is evident.
Case 1 - Figure 8
Tumor cells are relatively monotonous with round to ovoid nuclei and a high nuclear to cytoplasmic ratio. Nuclei are vesicular with small or inconspicuous nucleoli. Mitotic figures are frequent.
Case 1 - Figure 9
In some areas, tumor cells display, a small amount of pale cytoplasm.

Case 1 - Figure 10
Focally nuclei are slightly more pleomorphic and hyperchromatic.
Case 1 - Figure 11
CAM 5.2 immunostain reveals that occasional tumor cells are positive.

Microscopic Features
A highly cellular tumor consisting of broad sheets of small cells infiltrated renal parenchyma and sinus fat. In a large part, there appeared to be little stroma with delicate blood vessels. Focally a vaguely nested pattern with more prominent stroma was identified. Cysts containing serofibrinous fluid or blood, and focal necrosis were present. Tumor cells were monotonous with ovoid vesicular nuclei, scanty pale cytoplasm and small nucleoli. Mitoses were frequent.

Immunohistochemical Features
Neoplastic cells were strongly positive in a membranous pattern for CD 99 and moderately positive for vimentin and neurone specific enolase (NSE) with rare cells positive for CAM 5.2. Tumor was negative for EMA, MNF 116, CK 7, LCA, S 100, WT-1, desmin and CD 34

FISH Analysis
PET (paraffin embedded tissue), FISH analysis revealed that the tumor was positive for Ewing's EWSR1 (22 q12) rearrangement breakapart signals.

Diagnosis
Ewing's sarcoma/PNET of kidney

Discussion
Ewing's sarcoma (ES) and primitive neuroectodermal tumor (PNET) are primitive small round cell tumors showing varying (usually minimal) neuroectodermal differentiation [1]. These show morphological overlap and have similar EWS gene rearrangements, prognosis and response to chemotherapy. Therefore these are considered closely related members of the same family known as ES/PNET or Ewing family of tumors [2].These can occur anywhere, with extra skeletal ES/PNET occurring predominantly in the deep soft tissue of the trunk. Tumors primary in the kidney are rare. Almost all ES/PNET have a translocation involving 22 q12, site of the EWSR-1 gene. About 90% have a specific t (11; 22) (q 24; q12), resulting in a chimeric FLI-1-EWSR1 protein [2, 3]. Less commonly, EWS gene rearrangement occurs from translocations involving chromosomes other than 11. Commonest of these is t (21; 22) (q 22; q12) with ERG-EWSR1 (in 5%) . Rarely, t (7; 22) (p 22; q12), with ETV1-EWSR1, t (17; 22) (q12; q12) with EIAF-EWSR1 and t (2; 22) (q 33; q12) with FEV-EWSR1 have been found [4]. A familial predisposition or association with environmental factors have not been found. ES/PNET can occur following therapy for another tumor [5].

Renal ES/PNET was first reported by Mor et al in 1994 [6]. Since then there have been many case reports and several series [7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17]. In the largest series of primary malignant neuroepithelial tumors of the kidney of 146 cases [17], 79 had features typical of ES, PNET or a similar undifferentiated neoplasm while the other 67 had features not typical of these. Since only 13 (of 45 tested) were positive with PCR for EWS/FLI1 (10 of 33 typical), it is impossible to ascertain how many were ES/PNET, even within the typical group. Considering 55 other reported cases of ES/PNET, most occurred in young adults (mean 29 years), although some occurred in children and older patients (range 4-69 years) with males more commonly affected (ratio 1.6:1). Most common presenting symptoms were abdominal/flank pain or hematuria, and rarely a palpable mass, colic, fever, night sweats, weight loss or bone pain. Tumors ranged in size from 5 to 18 cm (mean 12 cm) and were commonly gray -yellow or gray-tan lobulated soft friable masses displaying hemorrhage and necrosis. Some had cystic change.

A spectrum of histological features characterise typical ES, atypical ES (large-cell) and typical PNET. These infiltrate renal parenchyma as densely cellular sheets or lobules or vague nodules separated by fibrous septa. Confluent or "filigree" pattern of necrosis is common. Typical ES has strikingly uniform primitive small cells with round or ovoid nuclei, regular nuclear contour, powdery chromatin and inapparent nucleoli. The cytoplasm is ill-defined or vacuolated. Rosettes are rare or absent. Mitotic count can be high. Interspersed "dark" pyknotic cells, a pseudo-vascular or pseudo-alveolar pattern can be present. In some, cells show moderate nuclear enlargement, irregular nuclear contours and prominent nucleoli, consistent with atypical ES. Abundant glycogen demonstrated by PAS/ diastase is seen in many. In typical PNET, rosettes (Homer Wright, Flexner-Wintersteiner), and perivascular pseudorosettes are more obvious. Nuclei are round or oval with coarse chromatin and frequent nucleoli. Cytoplasm can be indistinct or scanty and eosinophilic [7, 18]. Some consider spindle cell morphology incompatible with ES/PNET [19]. A study of 66 genetically confirmed cases including soft tissue and visceral cases had tumors with spindle cell sarcoma-like, "adamantinoma-like" or sclerosing abundant hyalinized matrix [20]. Exceptionally rarely, glial, ganglionic and even epithelial differentiation has been reported [19]. A cytogenetically confirmed soft tissue tumor has shown osteo- cartillagenous differentiation [21]. In the series of primary malignant neuroepithelial tumors of the kidney by Parham et al [17], there were clear cell sarcoma like, malignant peripheral nerve sheath tumor like, epithelioid/rhabdoid features and paraganglioma-like cases, although only 3 cases of these were fusion positive and therefore definite for ES/PNET.

Of renal ES/PNET with immunostaining reported [7, 8, 9, 11, 16, 22], all have been strongly positive (membranous) for CD 99 and 66% for FLI-1. Cytokeratin, usually focal was found in 16% and vimentin in 77%. NSE, Chromogranin, synaptophysin, neurofilament protein (NFP) and S100 staining was found in 88%, 9%, 30%, 12%, and 25% of cases respectively. None displayed WT1, GFA or LCA. Desmin positivity has been found only in non-renal cases [7, 23].

Differential diagnosis
The differential diagnosis of renal small round cell malignancies including ES/PNET are blastema predominant Wilms' tumor (WT), neuroblastoma, monophasic synovial sarcoma (SS), embryonal rhabdomyosarcoma (RMS), desmoplastic small round cell tumor (DSRCT), clear cell sarcoma, small cell carcinoma and lymphoma. WT occurs predominantly in children with 98% < 10 years of age. Adult WT is exceptionally rare. Diagnosis of triphasic of biphasic WT with epithelial and stromal elements is straightforward, but blastema-only monophasic WT is difficult. Blastema consists of small, closely packed round/ oval cells with scanty cytoplasm, coarse nuclear chromatin and small nucleoli. Blastema can be in diffuse sheets, nodules or serpentine arrangements. Identification of the latter 2 patterns is an important clue to diagnosis. Rosettes can be present in WT [25]. Blastema is frequently vimentin positive and may show focal expression of NSE, desmin and cytokeratin. CD 99 is focally positive in upto 20% of WT. Therefore, these immunostains alone are unhelpful in differentiating from ES/PNET. WT 1 is present in about 75% of cases with no cases displaying FLI 1 immunostaining, with the converse demonstrated in ES/PNET are very useful in diagnosis [7, 24]. Cytogenetics reveal characteristic anomalies including loss of genetic material at 11p13 with the WT1 gene [25].

Neuroblastomas are derived from primordial neural crest cells populating sympathetic ganglia and adrenal medulla with tumors in these locations [26, 27]. In most renal involvement is through direct invasion from the adrenal. Neuroblastoma arising as a true intrarenal mass is rare. In contrast to ES/ PNET neuroblastoma occurs in the very young with 90% diagnosed by 5 years, with only rare adult cases. Urinary catecholamines are usually elevated. These have sheets of small round cells, which may be separated into lobules by delicate fibrous septa. Cells appear almost devoid of cytoplasm and nuclei have granular chromatin. Rosettes, neuropil and ganglionic differentiation may be found. Immunostaining reveals positivity for NSE, synaptophysin, chromogranin, S100 protein and neurofilament protein. CD 99 is negative in contrast to ES/PNET and cytokeratin is negative. In 80%, there is loss of distal 1p harbouring 2 tumour suppressor genes [26, 27].

SS has been identified in a subset of previously described cystic ES [28, 29]. This spindle cell neoplasm, the monophasic variant of which can mimic ES/PNET occurs in a similar age group (range 12-59 years). The biphasic variant shows epithelial differentiation. Tumors are highly cellular with mitotically active monomorphic plump spindled or round cells with indistinct cell borders in intersecting fascicles or sheets. Cysts from entrapped collecting ducts and calcification are common. Hypercellular with hypocellular myxoid areas with a prominent hemangiopericytoma - like pattern are clues to diagnosis. Spindle cells are positive for vimentin and BCL2, frequently positive for CD 99 and EMA, negative/ focally positive for cytokeratin and negative for desmin, actin and S100. Detection of t (X; 18) (p11.2; q11) with SYT-SSX gene fusion is diagnostic [30]. Some atypical ES without molecular confirmation of SS remain as unclassified malignant neoplasms with nests, cords and sheets of small cells within myxoid stroma. These also need to be differentiated from ES/PNET [28, 30].

DSRCT is a rare aggressive tumor usually presenting with widespread abdominal serosal involvement. It is characterised by nests, cords and sheets of undifferentiated small cells with foci of necrosis and calcification. Primary renal tumors are exceptionally rare and may not display desmoplasia [31, 32]. These may occur in children and adults. FISH result is similar to that in EWS/PNET. However, the fusion protein is different; WT1-EWSR1, due to (11; 22) (p13; q12). It is also different from EWS/PNET in displaying polyphenotypic differentiation with positivity for desmin, EMA and WT1, cytokeratins, NSE and vimentin [31, 32].

Embryonal RMS of the kidney is extremely rare and may be indistinguishable from small round cell tumours. These display immunohistochemical markers of myogenic differentiation. Clear cell sarcoma of the kidney (CCSK) is a pediatric neoplasm (mean age 36 months), which rarely occurs in adults. Most consist of a monotonous array of cells with pale cytoplasm, fine nuclear chromatin and small nucleoli. In the classic pattern, cells are arranged in sheets with a prominent branching vascular pattern. Other patterns include spindled, cystic, hyaline sclerosing and palisading. This tumor also shows EWSR 1 rearrangement, but different from EWS/PNET, t(12; 22) (q 13; q 12), with fusion of ATF1-EWSR 1. Immunoreactivity only for vimentin and BCL2 and uniform negativity for CD 99 also help differentiation from ES/PNET. CD 34, S100, desmin, cytokeratins and EMA also negative [33].

Small cell neuroendocrine carcinoma of the kidney may be metastatic from another location or primary in the kidney, and occurs in adults. These can mimic ES/PNET in having sheets, nests and trabeculae of small round to fusiform cells with hyperchromatic nuclei and inconspicuous nucleoli. Rosettes may be present. Nuclear moulding and Azzopardi phenomenon are clues to diagnosis. Dot-like cytoplasmic cytokeratin staining and variably positive neuroendocrine markers are helpful in diagnosis [34]. Renal involvement with malignant lymphoma can be secondary or primary. The most commonly encountered is in post transplant lymphoproliferative disorder. Diffuse sheets, nodules or intravascular forms are found and some can mimic ES/PNET. LCA can distinguish between these lesions. Lymphoblastic lymphoma can also be positive for CD 99 and FLI 1 [35].

Adjuvant therapies for ES/PNET have included chemotherapy, radiation and experimental therapies. Of reported cases [7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18], 23% had metastases at presentation. Follow-up data was available for 43 of whom 24 (55%) died of disease (mean time to death, 13 mo, median 10 mo). 15 patients (34%) were reported alive with no evidence of metastatic disease at last follow-up (mean duration, 22 months). Distant metastatic sites included lung, pleura, bone, mediastinum and liver. Long-term survivors are at risk (about 8%, median latency 7 years) of developing a second malignancy. These include renal medullary neuroblastoma, bone fibrosarcoma, osteosarcoma, malignant fibrous histiocytoma and leukemia. The etiology is thought to be genetic, radiation doses> 60 Gy, or alkylating agent chemotherapy [36].

Take home bullet points .
  1. Renal small cell malignancies include several distinct entities with different prognostic and therapeutic implications necessitating accurate diagnosis.

  2. These tumors have significant morphological overlap causing diagnostic difficulty.

  3. Immunohistochemistry is useful in the differential diagnosis of these tumors. However there is immunophenotypic overlap. Cytogenetic and/or molecular investigation is recommended particularly for morphologically atypical cases .

References
  1. Dehner LP. Peripheral and central primitive neuroectodermal tumors. A nosologic concept seeking a consensus. Arch Pathol Lab Med 1986; 110:997-1005

  2. Delattre O, Zucman J, Melot T, et al. The Ewing family of tumors-a subgroup of small round cell tumors defined by specific chimeric transcripts. N Engl J Med 1994; 331:294-299.

  3. Turc-Carel C, Aurias A, Mugneret F, et al. Chromosomes in Ewing's sarcoma.1. An evaluation of 85 cases and remarkable consistency of t (11; 22) (q 24; q 12). Cancer Genet Cytogenet 1988; 32:229-238.

  4. Sandbag AA, Bridge JA. Updates on cytogenetics and molecular genetics of bone and soft tissue tumors: Ewing sarcoma and peripheral primitive neuroectodermal tumors. ). Cancer Genet Cytogenet 2000; 123:1-26

  5. Spunt SL, Rodriguez-Galindo C, Fuller CE, et al. Ewing sarcoma-family tumours that arise after treatment of primary childhood cancer. Cancer 2006; 107(1): 201-206.

  6. Mor Y, Nass D, Raviv G, et al. Malignant peripheral primitive neuroectodermal tumor (PNET) of the kidney. Med Paediatr Oncol 1994; 23:437-440.

  7. Jimenez RE, Folpe AL, Lapham RL, et al. Primary Ewing's sarcoma/primitive neuroectodermal tumor of the Kidney. A clinicopathologic and immunohistochemical analysis of 11 cases. Am J Surg Pathol 2002; 26(3): 320-327.

  8. Parada D, Godoy A, Liuzzi F, et al Primary Ewing's sarcoma/primitive neuroectodermal tumor of the kidney. An infrequent finding. Arch Esp Urol 2007; 60(3): 321-325

  9. Kang SH, Perle MA, Nonaka D, et al. Primary Ewing sarcoma /PNET of the kidney: fine needle aspiration, histology, and dual colour breakapart FISH Assay. Diagn Cytopathol 2007; 35(6): 353-357.

  10. Moustafellos P, Gourgiotis S, Athanasopoulos G, et al. A spontaneously ruptured primitive neuroectodermal tumor/extraosseous Ewing's sarcoma the kidney with renal vein tumor thrombus. Int Urol Nephrol 2007; 39(2): 393-395

  11. Saxena R, Sait S, Mhawech-Fauceglia P. Ewing sarcoma/primitive neuroectodermal tumor of the kidney: a case report. Diagnosed by immunohistochemistry and molecular analysis. Ann Diagn Pathol 2006; 10 (6):363-366.

  12. Perer B, Shanberg AM, Matsunaga G, et al. Laparoscopic removal of extraosseous Ewing's sarcoma of the kidney in a pediatric patient. J Laparoendosc Adv Surg Tech A. 2006;16 (1): 74-6.

  13. Ruszat R, Casella R, Bachman A, et al. Primitive neuroectodermal tumor of the kidney with hyaline cells. Urol Int 2005; 75(2): 184-6.

  14. Dogra PN, Goel A, Kumar R, et al. Extraosseous Ewing's sarcoma of the kidney. Urol Int 2002; 69(2):150-152.

  15. Karnes RJ, Gettman MT, Anderson PM, et al. Primitive neuroectodermal tumor (extraskeletal Ewing's sarcoma) of the kidney with vena caval tumor thrombus. J Urol 2000; 164 (3 Pt 1):772

  16. Pai SA, Naresh KN, Borges AM. Primitive neuroectodermal tumors. Am J Surg Pathol 2003; 27( 8):1177.

  17. Parham DM, Roloson GJ, Feely M, et al. Primary malignant neuroepithelial tumors of the kidney: a clinicopathologic analysis of 146 adult and pediatric cases from the National Wilms' Tumor Study Group Pathology Center. Am J Surg Pathol 2001; 25(2): 133-146

  18. Enzinger FM, Weiss SW. Ewing's sarcoma/PNET tumor family and elated lesions. Soft tissue tumors. St. Louis: Mosby, 2008

  19. Fletcher CDM. Peripheral neuroectodermal tumors. Diagnostic Histopathology of Tumors. Churchill Livingstone. Third edition, 2007

  20. Folpe AL, Goldblum JR, Rubin BP, et al. Morphologic and immunophenotypic diversity in Ewing family tumours; a study of 66 genetically confirmed cases. Am J Surg Pathol 2005; 29(8): 1025-1033.

  21. Oda Y, Kinoshita Y, Tamiya S, et al. Extraskeletal primitive neuroectodermal tumor, with massive osteo-cartillagenous metaplasia. Histopathology 2000; 36(2): 188-191.

  22. Takeuchi T, Iwasaki H, Ohjimi Y, et al. Renal primitive neuroendocrine tumor: and immunohistochemical and cytogenetic analysis. Pathology International 1996; 46(4): 292-297.

  23. Parham DM, Dias P, Kelly DR et al. Desmin positivity in primitive neuroectodermal tumors of childhood. Am J Surg Pathol 1992; 16(5):483-92.

  24. Grubb GR, Yun K, Williams BR, et al. Expression of WT 1 protein in fetal kidneys and Wilms tumors. Lab Invest 1994; 71(4): 472-479.

  25. Rubin BP, Pins MR, Nielsen GP, et al. Isochromosome 7q in adult Wilms' tumors: diagnostic and pathogenetic implications. Am J Surg Pathol 2000; 24 (12): 1663-1669

  26. Lack EE. Tumors of the adrenal gland and extra adrenal paraganglia. Atlas of tumor pathology, series 3, Fascicle 19. Armed Forces Institute of Pathology 1997, Washington, DC

  27. Kozakewich HP, Perez-Atayede AR, Donovan MJ, et al Cystic neuroblastoma: emphasis on gene expression, morphology and pathogenesis. Paediatr Dev Pathol 1998; 1:17-28.

  28. Arnold MM, Beckwith JB, Faria P, et al. Embryonal sarcoma in adult and paediatric kidneys [abstract]. Mod Pathol 1995; 8 (72 A): 409.

  29. Argani P, Faria PA, Epstein JI, et al. Primary renal synovial sarcoma: molecular and morphologic delineation of an entity previously included among embryonal sarcoma the kidney. Am J Surg Pathol 2000; 24:1087-1096.

  30. Delahunt B, Beckwith JB, Eble JN, et al. Cystic embryonal sarcoma of kidney. Cancer 1998; 82: 2427-2433.

  31. Wang LL, Perlman EJ, Vujanic GM, et al. Desmoplastic small round cell tumor of the kidney in childhood. Am J Surg Pathol 2007; 31(4): 576-584

  32. Su MC, Jeng YM, Chu YC Desmoplastic small round cell tumor of the kidney. Am J Surg Pathol 2004; 28 (10): 1379-1383.

  33. Argani p, Perlman EJ, Breslow NE, et al. Clear cell sarcoma of the kidney: a review of 351 cases from the National Wilms Tumor Study Group Pathology Center Am J Surg Pathol 2000; 24(1): 4-18

  34. Lane BR, Cherry F, Jour G, et al. Renal neuroendocrine tumours: a clinicopathological study. BJU Int 2007; 100(5): 1030-1035.

  35. Folpe AL, Hill CE, Parham DM, et al Immunohistochemical detection of FLI 1 protein expression: a study of 132 round cell tumours with emphasis on CD 99-positive mimics of Ewing's sarcoma/primitive neuroectodermal tumor. Am J Surg Pathol 2000; 24 (12): 1657-1662.

  36. Smith LM, Cox RS, Donaldson SS. Second cancers in long term survivors of Ewing's sarcoma. Clin Orthop Relat Res 1992; 274:275-81.