EWSR1-POU5F1 fusion in soft tissue myoepithelial tumors. A molecular analysis of sixty-six cases, including soft tissue, bone, and visceral lesions, showing common involvement of the EWSR1 gene

Abstract

The diagnosis of myoepithelial (ME) tumors outside salivary glands remains challenging, especially in unusual clinical presentations, such as bone or visceral locations. A few reports have indicated EWSR1 gene rearrangement in soft tissue ME tumors, and, in one case each, the fusion partner was identified as either PBX1 or ZNF444. However, larger studies to investigate whether these genetic abnormalities are recurrent or restricted to tumors in soft tissue locations are lacking. Sixty-six ME tumors mainly from soft tissue (71%), but also from skin, bone, and visceral locations, characterized by classic morphological features and supporting immunoprofile were studied. Gene rearrangements in EWSR1, FUS, PBX1, and ZNF444 were investigated by fluorescence in situ hybridization. EWSR1 gene rearrangement was detected in 45% of the cases. A EWSR1-POU5F1 fusion was identified in a pediatric soft tissue tumor by 3'Rapid Amplification of cDNA Euds (RACE) and subsequently confirmed in four additional soft tissue tumors in children and young adults. An EWSR1-PBX1 fusion was seen in five cases, whereas EWSR1-ZNF444 and FUS gene rearrangement was noted in one pulmonary tumor each. In conclusion, EWSR1 gene rearrangement is a common event in ME tumors arising outside salivary glands, irrespective of anatomical location. EWSR1-negative tumors were more often benign, superficially located, and showed ductal differentiation, suggesting the possibility of genetically distinct groups. A subset of soft tissue ME tumors with clear cell morphology harbor an EWSR1-POU5F1 fusion, which can be used as a molecular diagnostic test in difficult cases. These findings do not support a pathogenetic relationship between soft tissue ME tumors and their salivary gland counterparts.

Fig. 1

BAC probe sets used to determine breakpoints by FISH for EWSR1, FUS, POU5F1, PBX1, and ZNF444.

Fig. 2

Morphologic spectrum of soft tissue ME tumors showing an EWSR1 gene rearrangement, composed of: (A) undifferentiated small blue cells with ill-defined cell borders (100x; 7 year-old female, peri-orbital mass); (B) rhabdoid morphology with eosinophilic cytoplasm (400x; 2 year-old female, mediastinum); (C) epithelioid cells with clear cytoplasm arranged in nests in a sclerotic background (200x, 20year-old male, subcutaneous foot); (D) bland spindle cell proliferation in a sclerotic background (100x, fibula, 16-year old); (E) fibrotic lesion with hypocellular epithelioid cells (200x; 32 year-old male, subcutaneous ankle); (F) EWSR1 gene rearrangement by FISH showing break-apart signal; (G) nested epithelioid morphology of an intra-osseous ME tumor (40x, humerus, 23 year-old male), showing diffuse and strong S100 protein (H) and by FISH in addition to EWSR1 gene rearrangement, amplification of the 3’end of the EWSR1 gene (I).

Fig. 3

Pathologic and molecular findings in a deep-seated arm ME tumor from a 9-year old male, showing (A) epithelioid cells with abundant clear cytoplasm, arranged in a nested growth pattern, being immunoreactive for (B) S100 protein and (C) EMA, and by (D) 3’RACE showing fusion of exon 7 of EWSR1 with the last portion of POU5F1 intron 1; (E) this fusion transcript structure is similar with the one previously reported in eccrine hidradenoma/ mucoepidermoid carcinoma (Moller et al., 2008); the gene fusion results were further confirmed by (F) RT-PCR: showing an amplified product in lane 1; lane 2 is the negative control; M, size marker and by (G) FISH, with a break-apart signal for POU5F1 probe.

Fig. 4

ME tumors characterized by EWSR1-PBX1 fusion: (A) 37-year old male with a left hip lesion, showing a bland spindle cell proliferation embedded in a fibrotic stroma (200x); (B) 6-year old female with a locally recurrent forearm mass showing bone invasion, composed of uniform epithelioid cells with scant clear cell cytoplasm (200x); (C) partial karyotype showing a t(1;22)(q23;q12), confirmed by FISH to have a rearrangement of (D) PBX1 and (E) EWSR1.

Fig. 5

Pulmonary ME tumor from a 64-year old female showing (A) epithelioid cells with clear cytoplasm separated by a prominent collagenous stroma (200x), which had a (B) ZNF444 rearrangement by FISH.