Clinicopathological analysis of EWSR1/FUS::NFATC2 rearranged sarcoma in the left forearm: A case report

Abstract

BACKGROUND

We present a case of an EWSR1/FUS::NFATC2 rearranged sarcoma in the left forearm and analyze its clinicopathological and molecular features.

CASE SUMMARY

The patient is a 23-year-old woman. Microscopically, the tumor cells were medium-sized round cells arranged in small nests. The cytoplasm was clear, nuclei were relatively uniform, chromatin was dense, nucleoli were visible, and mitotic figures were rare. Immunohistochemically, the tumor cells were positive for Vimentin, INI-1, CD99, NKX2.2, CyclinD1, friend leukaemia virus integration 1, and NKX3.1. Next-generation sequencing revealed the presence of the EWSR1-NFATC2 fusion gene. EWSR1/FUS::NFATC2 rearranged sarcomas are rare and can easily be misdiagnosed.

CONCLUSION

Clinical imaging, immunohistochemistry, and molecular pathology should be considered to confirm the diagnosis.

Key Words: EWSR1; NFATC2; Sarcoma; Forearm; Case report

Core Tip:EWSR1/FUS::NFATC2 rearranged sarcoma is a rare and aggressive malignancy of bone or soft tissue. We report a case of EWSR1/FUS::NFATC2 rearranged sarcoma of the left forearm. Immunohistochemistry and molecular detection play an important role in distinguishing this tumor from other tumors. It is clinically insensitive to classical Ewing's sarcoma chemotherapy, therefore it is very important to recognize its presence to avoid misdiagnosis. And more cases of EWSR1/ FUS::NFATC2 rearranged sarcomas need to be collected for clinicopathological evaluation and follow-up to explore its development, prognosis, and treatment options.

INTRODUCTION

EWSR1-non-ETS fusion sarcomas involve gene partners unrelated to the ETS family. The reported incidence of EWSR1-non-ETS fusion sarcomas is 0.212 cases per million population[1]. EWSR1/FUS::NFATC2 rearranged sarcoma is the most common subtype of EWSR1-non-ETS fusion sarcoma. It is also a newly described class of solid tumors originating from bone or soft tissue with unique clinicopathological features[2-4]. EWSR1/FUS::NFATC2 rearranged sarcoma mainly occurs in the metaphysis or diaphysis of long bones and less often in soft tissues. In the long bones, the affected sites were femur, humerus, radius and tibia in descending order of frequency[5]. In soft tissues, it occurs mainly in limbs, head, neck and chest[6]. The median age of onset is approximately 32 years, and male-to-female ratio is 5:1[7-9]. Clinically, it usually presents with pain and locally destructive bone lesions that may invade the surrounding soft tissues. To date, a small number of EWSR1/FUS::NFATC2 rearranged bone tumors have been reported, many of which do not have detailed clinicopathological data[5-9]. Here, we report a case of EWSR1/FUS::NFATC2 rearranged sarcoma of the left forearm.

CASE PRESENTATION

Chief complaints

A 23-year-old female patient was admitted to the hospital 5 months after the recurrence of a tumor in her left forearm.

History of present illness

The patient had left forearm pain for 5 months, and magnetic resonance examination suggested the possibility of recurrence of malignant tumor in the left forearm.

History of past illness

The patient's left forearm tumor was diagnosed as a sclerosed epithelioid fibrosarcoma 10 years ago at a foreign hospital. Subsequently, the patient underwent tumor resection and bone grafting of the left distal radius. Chemotherapy was not administered, and no recurrence was observed in the regular review. Five months prior, pain in the dorsal left forearm recurred without an obvious trigger.

Personal and family history

No personal or family history was available.

Physical examination

A surgical scar was seen on the left forearm, which was slightly swollen. There was dorsal tenderness in the distal left forearm.

Laboratory examinations

The tumor cells were positive for Vimentin, CD99, NKX2.2, NKX3.1, INI-1, CyclinD1, and FLI1, and focally positive for epithelial membrane antigen (EMA) (Figure 1). Approximately 25% of Ki67 hotspots were positive (Figure 1C). However, cytokeratin (CK)-pan, desmin, CD45-leukocyte common antigen, S-100, mouse double minute 2, myogenic differentiation 1, myogenin, special AT-rich2, mucoprotein-4 (MUC-4), CD34, signal transducer and activator of transcription 6, MelanomA, human melanoma black 45, CD56 (neural cell adhesion molecules), neuron specific enolase, synaptophysin, ETS-related gene, transducin-like enhancer of split 1, wilms tumor protein (WT1), calretinin, Pax-8, and CD68 were negative in this case. Next-generation sequencing revealed t(20;22)(q13.2;q12.2) EWSR1-NFATC2 gene fusion (Figure 2).

Figure 1 Immunohistochemically, the tumor cells were positive for Vimentin, CD99, NKX2.2, NKX3.1, INI-1, CyclinD1 and FLI1, and focally positive for epithelial membrane antigen. A: Immunohistochemistry showed a small number of epithelial membrane antigen positives in tumor cells; B: The tumor cells were diffusely positive for Vimentin; C: About 25% of the tumor cells were positive for Ki67; D: Tumor cells were also permeated with CD99 positivity; E-I: Positive for INI1, FLI1, NKX2.2, NKX3.1, and CyclinD1, respectively. Scale bars are in the lower left corner of each image (original magnification, 630 ×).

Figure 2 Next-generation sequencing revealed t(20;22)(q13.2;q12.2) EWSR1-NFATC2 gene fusion.

Imaging examinations

Plain magnetic resonance imaging (MRI) and enhanced (3.0T) examination of the unilateral upper limb showed multiple oval and patchy abnormal signal shadows in the original surgical area and multiple abnormal signal shadows in the distal and surrounding bones, with clear boundaries; the size of the largest one was approximately 19 mm × 14 mm (Figure 3A).

Figure 3 Magnetic resonance imaging plain scan + enhancement (3.0T) indicated the location of the lesion. We could observe the tumor section in the surgical specimen, and the structure and morphology of tumor cells under the microscope. A: Multiple oval and patchy abnormal signals were found in the original surgical area of the left forearm and multiple abnormal signals were found in and around the distal bone on the plain and enhanced (3.0T) magnetic resonance imaging of the unilateral upper limb; B: The tumor involved the medullary cavity, and the texture of the cut surface was uneven and fishy; C: Tumor cells were seen in the mature bone tissue, original magnification, 50 ×; D: Tumor cells were of medium size and arranged in small nests with fibrous septa between the nests, original magnification, 200 ×; E and F: The tumor cells had clear cytoplasm, uniform nuclear morphology, dense chromatin, visible nucleoli, and rare mitotic figures. original magnification, 400 × (E), 630 × (F). Scale bars are in the lower left corner of each image.

The surgical specimen consisted of a piece of left radial tissue, approximately 9.5 cm long and 1-2.5 cm in diameter. The local surface was raised, and the cut surface was grey-white and grey-red with an uneven texture (Figure 3B). Microscopically, medium-sized round cells were arranged in small nests between mature bone tissues (Figure 3C and D). The tumor cells had a clear cytoplasm, uniform nuclear morphology, dense chromatin, visible nucleoli, and rare mitotic figures (Figure 3E and F).

FINAL DIAGNOSIS

Finally, EWSR1/FUS::NFATC2 rearranged sarcoma was diagnosed.

TREATMENT

The malignant tumor was treated with chemotherapy before surgery, and the left distal radius malignant tumor was resected and allogeneic radius was implanted under general anesthesia in September 2023.

OUTCOME AND FOLLOW-UP

Regular follow-up after surgery showed no tumor recurrence or metastasis.

DISCUSSION

This patient was a 23-year-old female who was found to have a tumor in the left forearm 10 years ago, which was diagnosed as sclerosing epithelioid fibrosarcoma in another hospital. The tumor was resected without chemotherapy, and no recurrence was found during regular review. Five months ago, back pain recurred in the left forearm, and MR examination suggested the possibility of recurrence of malignant tumor in the left forearm. Microscopically, there were medium-sized round cells arranged in small nests between the mature bone tissue. The cytoplasm of the tumor cells was clear, the nuclei were relatively uniform, the chromatin was dense, nucleoli were visible, and mitotic figures were rare. Immunohistochemical staining showed that the tumor cells were positive for Vimentin, INI-1, CD99, NKX2.2, CyclinD1, FLI1 and NKX3.1, and a few tumor cells were positive for EMA. Next generation sequencing found EWSR1 and NFATC2 gene fusion.

EWSR1/FUS::NFATC2 rearranged sarcomas are generally solid masses ranging from 4 to 18 cm with a yellowish-brown surface and hard or fleshy section[10]. Metastasis occurs mainly in the lungs and soft tissues. They mainly consist of relatively homogeneous round cells arranged in a diffuse, pseudoglandular, nested, or cord-like pattern against a fibrous or clear mucus background. The tumor nuclei are consistent in size and can have obvious pleomorphism, dense or vesicular chromatin, small or prominent nucleoli, tumor necrosis, or mitotic figures[9]. Immunohistochemically, the tumor cells were diffusely positive for CD99, PAX-7, and NKX2.2 in approximately half of the cases, and focally positive staining for cytokeratin AE1/AE3 was observed[9-12]. In addition, Aggercan and NKX3.1 are highly expressed in EWSR1/FUS::NFATC2 rearranged sarcomas, which can be used to distinguish EWSR1/FUS::NFATC2 rearranged sarcoma from Ewing's sarcoma[10,13,14]. In addition, EWSR1::NFATC2 rearranged sarcoma has a unique signal signature in FISH with the EWSR1 split probe, with frequent amplification of a single-probe (5' end) signal that is markedly different from other EWSR1-rearranged tumors. Molecular pathology showed that EWSR1-NFATC2 gene fusion is an important diagnostic marker for EWSR1::NFATC2 rearranged sarcomas. EWSR1/FUS::NFATC2 rearranged sarcoma has a potential risk of local recurrence, distant metastasis, and histological progression and is not sensitive to classical Ewing's sarcoma-specific chemotherapy[9,10,13]. Previously, EWSR1/FUS::NFATC2 rearranged sarcoma was likely to be diagnosed as Ewing sarcoma, myoepithelial tumor, ossifying fibromyxoid tumor, sclerosing epithelioid fibrosarcoma, extraskeletal myxoid chondrosarcoma, small cell osteosarcoma, etc.[3,6,15-18].

EWSR1/FUS::NFATC2 rearranged sarcoma is rare and should be distinguished from the following tumors: (1) Ewing’s sarcoma of the bone: Immunohistochemically, both sarcomas express CD99, NKX2.2, and PAX-7, which can lead to misdiagnosis, but NKX3.1, lymphoid, rhabdomyoblastic, neuroendocrine, and other markers are negative in Ewing’s sarcoma. In addition, all Ewing’s sarcomas harbor FET-ETS gene fusions[7,13,19]; (2) Myoepithelial tumors: Myoepithelial tumors of the bone are rare[20]. Histologically, the tumors were spindle-shaped with epithelioid, plasmacytoid, or clear cells. The stroma was myxochondroid, clear, or without obvious stroma. Immunohistochemically, the tumor cells were positive for myoepithelial markers such as cytokeratin, EMA, P63, alpha-smooth muscle actin (SMA), S-100, glial fibrillary acidic protein, and WT1. Some cases were positive for CD99 but negative or focally positive for NKX2.2. Approximately 50% of myoepithelial tumors harbor EWSR1-POU5F1 or PBX1 gene translocations[21,22]; (3) Ossifying fibromyxoid tumors are often well-circumscribed, lobulated, or multinodular. The tumor cells in the fibromyxoid stroma were uniform in size and bland in morphology, arranged in cords or nests, and accompanied by an intact or incomplete bone shell at the periphery. Immunohistochemically, the tumor cells were positive for S-100, Desmin, and CD99. The tumor cells were focally positive for MUC4, EMA, CK, and SMA but negative for NKX3.1 and NKX2.2. Molecular detection of the common PHF1 gene fusion, in addition to CORL1, CREBBP, and KDM2A gene fusion, has also been reported[23]; (4) Sclerosing epithelioid fibrosarcoma: Microscopically, epithelioid fibroblasts arranged in cords or nests in the sclerotic collagenous stroma, with pseudoacinar structure. The cells were uniform in size and had relatively mild morphology. Immunohistochemically, MUC4 was diffusely positive, EMA and SMA were expressed in some cases, and CK was negative. Molecular EWSR1-CREB3L1 rearrangement helps confirm the diagnosis, but additional molecular diagnostic testing is not required in cases with characteristic morphology and MUC4 expression[24,25]; (5) Extraskeletal myxoid chondrosarcoma: Primary bone lesions are rare. Histologically, the tumors were composed of ovoid, short-spindled, or rhabdoid cells arranged in cord-like or nest-like structures with myxochondroid stroma. CD99, NKX2.2 and NKX3.1 were not detected by immunohistochemistry[26]. Additionally, insulinoma-associated protein 1 is overexpressed in nearly 90% of extraskeletal myxoid chondrosarcomas. Molecular detection confirmed the presence of NR4A3 gene rearrangement[18]; and (6) Sarcomas with BCOR alterations or CIC-rearranged sarcomas: The category of undifferentiated small round cell malignant tumors is difficult to distinguish histologically, but both BCOR and CIC-rearranged sarcomas have typical molecular genetic features and lack the EWSR1 gene break and amplification signal.

CONCLUSION

EWSR1/FUS::NFATC2 rearranged sarcoma is a rare and aggressive malignancy of the bone or soft tissues. It predominantly occurs in the long bones of adult males and is clinically insensitive to chemotherapy for classical Ewing’s. EWSR1/FUS::NFATC2 rearranged sarcoma is relatively rare, and recognizing its presence is important to avoid misdiagnosis. Immunohistochemistry and molecular detection are important in distinguishing this tumor from other tumors. More cases of EWSR1/FUS::NFATC2 rearranged sarcomas should be collected for clinicopathological evaluation and follow-up to explore their development, prognosis, and treatment options.