Primitive Neuroectodermal Tumor of the Gallbladder

Ewing sarcoma/primitive neuroectodermal tumor (ES/PNET), also referred to previously as peripheral neuroepithelioma or peripheral neuroblastoma, usually occurs in the deep soft tissue of the extremities or paravertebral areas. Primitive neuroectodermal tumor shows the same chromosomal translocations (t[11;22][q24;q12] and t[21;22][q22;q12]) as extraskeletal ES, and these tumors are widely regarded as the opposite ends of a spectrum of the same disease entity (ES/PNET family). Extraskeletal ES usually occurs in patients younger than 30 years, whereas PNET can occur in patients older than 40 years. Ewing sarcoma also lacks neuronal differentiation, whereas PNET shows neuronal differentiation with a formation of Homer-Wright rosettes and positive immunoreactivity for neuroendocrine markers. Although tumors of the ES/ PNET family have been reported in unusual sites, including the kidney,1–3 urinary bladder,4 testis,5 uterus,6 ovary,7 lung,8 myocardium,9 parotid gland,10 esophagus,11 pancreas,12 small intestine,13 and hepatic duct,13 to our knowledge no case has yet been reported in the gallbladder. We present a case of primary PNET occurring in the gallbladder of a 53-year-old woman and report the light microscopic, immunohistochemical, ultrastructural, and molecular findings for this tumor.

A 53-year-old woman presented with 1-month history of epigastric pain, anorexia, and nausea. Her previous medical history was unremarkable. Physical examination revealed a nontender palpable gallbladder with positive Courvoisier sign. The preoperative laboratory studies showed mild anemia and normal ranges of serum tumor markers, including α-fetoprotein, carcinoembryonic antigen, and CA 19-9. Abdominal and pelvic computed tomographic scans and ultrasonography showed an intraluminal polypoid mass with heterogenous enhancement in the gallbladder (Figure 1). The patient subsequently underwent cholecystectomy with partial segmentectomy of the right hepatic lobe with the diagnosis of gallbladder cancer. The patient refused adjuvant treatment, and the last follow-up examination at 3 months after surgery showed no evidence of recurrence.

Grossly, a polypoid mass was present in the fundus of the gallbladder and measured 7 × 4 × 3.5 cm. The cut surface of the mass was whitish tan, friable, and mostly necrotic. A separately received fragment of liver tissue was unremarkable. Microscopically, the tumor was composed of diffuse solid sheets of monotonous, small round cells with an intervening fibrovascular stroma and necrotic foci. Numerous Homer-Wright rosettes with a central fibrillary core were identified (Figure 2). The tumor cells showed hyperchromatic nuclei containing coarse or fine powdery chromatin, indistinct pale scanty cytoplasm, and inconspicuous nucleoli (Figure 3). Mitotic figures were frequent (>20/10 high-power fields). The tumor extended to the muscle and the perimuscular connective tissue layer without involvement of the serosa. The 4 pericholedochal lymph nodes were free of tumor. Immunohistochemical staining of the tumor cells showed intense positive membranous immunoreactivity for MIC2/CD99 protein (1:25; Dako Corporation, Carpinteria, Calif) (Figure 4, A) and positive intracytoplasmic immunoreactivity for neuron-specific enolase (1:1000; Dako) and synaptophysin (1:100; DiNonA, Seoul, Korea) (Figure 4, B). The tumor cells showed negative immunoreactivity for leukocyte common antigen (1:200; Dako), desmin (1:200; Dako), and S100 (1:200; Zymed Laboratories, Inc, South San Francisco, Calif). Ultrastructurally, the tumor cells contained a few intracytoplasmic organelles without evidence of neurosecretory granules or neurofilaments. The characteristic chromosomal translocations, such as t(11;22) (q24;q12) (EWSR1/FLI1) and t(21;22)(q22;q21) (EWSR1/ERG) were not detected when the reverse transcriptase–polymerase chain reaction was performed on 2 different paraffin blocks.

Ewing sarcoma/PNET is the second most common sarcoma among children and young adults, and PNET occurs in a broader age range than extraskeletal ES.3,6,9,10,14 Primitive neuroectodermal tumor occurs outside the brain, spinal cord, and sympathetic nervous system and is mostly found within the deep soft tissue of the extremities and the paravertebral areas. The primary visceral PNETs are extremely rare1–10,12,13 and have been reported most commonly in the kidney.1–3 Primitive neuroectodermal tumor is considered to be a more differentiated form of the ES/ PNET family, and PNET with poor rosette formation overlaps with ES and a malignant small round cell tumor of the thoracopulmonary region (ie, Askin tumor). All these tumors share the same chromosomal translocation (t[11; 22][q24;q12]) and expression of p30/32 cell surface glycoprotein antigen. For the diagnosis of PNET over extraskeletal ES, positive immunoreactivity for at least 2 neuronal markers, light microscopic evidence of Homer- Wright rosettes, or ultrastructural evidence of neuronal differentiation, such as interdigitating cell processes and neurosecretory granules, are required. The current case showed positive immunoreactivity for neuron-specific enolase and synaptophysin, as well as Homer-Wright rosette formation, favoring PNET over extraskeletal ES.

The MIC2 gene on the short arms of the X and Y chromosomes encodes the cell surface glycoprotein p30/32. Immunohistochemical staining for MIC2/CD99 is known to be highly sensitive for diagnosis of the ES/PNET family, but other small round cell tumors, such as T-lymphoblastic lymphoma, poorly differentiated synovial sarcoma, small cell osteosarcoma, rhabdomyosarcoma, desmoplastic small round cell tumor, small cell carcinoma, and Merkel cell carcinoma, can also show positive immunoreactivity for MIC2/CD99.14 Thus, the MIC2/CD99 marker should be used as part of a panel of immunohistochemical stainings because of the lack of the specificity. The current case could exclude the possibility of other small round cell tumors, such as lymphoma, rhabdomyosarcoma, and melanoma, by negative immunoreactivity for leukocyte common antigen, desmin, and S100, respectively.

Ninety percent to 95% of tumors in the ES/PNET family reveal several characteristic chromosomal translocations involving the ES gene (EWS) on chromosome 22q12, and members of the ETS family of oncogenes show characteristic translocations such as fli-1 on chromosome 11q24, ERG on chromosome 21q22, etv-1 on chromosome 7q22, E1AF on chromosome 17q12, and FEV on chromosome 2q33.14 After the formation of various fusion gene products, the ETS oncogenes encoding DNA-binding transcription factors are subject to regulation from the EWS promoter.14 The most common chromosomal translocation, t(11;22)(q24;q12), can be detected by reverse transcriptase–polymerase chain reaction or fluorescence in situ hybridization in more than 90% of members of the ES/PNET family with chromosomal translocations.15 In the current case, the chromosomal translocations, including t(11; 22)(q24;q12) and t(21;22)(q22;q12), were not demonstrated; however, the possibility of translocations in other ETS family oncogenes cannot be entirely excluded. Since the chromosomal translocation t(11;22)(q24;q12) has also been reported in other tumors, such as olfactory neuroblastoma, small cell osteosarcoma, and mesenchymal chondrosarcoma,14 a careful histologic examination as well as a proper use of immunohistochemical markers, ultrastructural studies, and molecular or cytogenetic analyses are needed for correct diagnosis of the ES/PNET family.

The clinical significance of differentiation between PNET and ES has not yet been determined because of the small number of cases and limited follow-up periods. Some authors have reported that patients with PNET had more aggressive clinical courses than those with ES3,7,8; however, more cases should be collected for further analysis. To the best of our knowledge, this is the first reported case of PNET of the gallbladder.