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Solitary fibrous tumor (SFT) is an anatomically ubiquitous fibroblastic neoplasm that most often affects middle-aged adults. SFT was originally described in 1931 as a pleural tumor termed ‘hemangiopericytoma’,1 but over time it has been increasingly recognized that SFT may arise at a wide range of anatomical locations. Of extrapleural tumors, ∼40% arise in subcutaneous tissue, with the remaining cases occurring in deep soft tissues, retroperitoneum, mediastinum, abdominal cavity, and meninges among other sites.

In its classic form, SFT shows variably hypocellular and hypercellular areas composed of spindled to ovoid tumor cells with scant cytoplasm and indistinct cell borders, in a ‘patternless’ architecture. Prominent branching hemangiopericytoma-like vasculature is a characteristic finding, and the stroma is usually collagenous and hyalinized, occasionally with myxoid areas.2, 3, 4, 5 Lipomatous (fat-forming) differentiation may also occur. The tumor cells of SFT are usually bland, lacking significant atypia or pleomorphism, but atypical, cellular, and malignant forms of SFT exist, reflecting the variable (although sometimes unpredictable) biological potential of this tumor.

SFT can sometimes be difficult to distinguish from some other benign mesenchymal tumors and spindle cell sarcomas, particularly in small biopsy samples. Immunohistochemistry using existing markers is of variable utility in the differential diagnosis. Traditionally, CD34 expression is the most consistent reported finding in SFT, present in up to 95% of cases. However, CD34 expression is also common in other tumors that may mimic SFT, such as soft tissue perineurioma, dermatofibrosarcoma protuberans, and spindle cell lipoma. Furthermore, SFT shows nuclear expression of β-catenin in 40% of cases, which may lead to confusion with desmoid fibromatosis.6 SFT also shows reactivity for EMA in 20–30% of cases7 and cytoplasmic staining for CD99 in ∼70% of cases.7, 8 Positivity for BCL2 is seen in virtually all SFTs,7, 9, 10 but is a nonspecific finding, as expression is also observed in a variety of other mesenchymal neoplasms.9

Until recently, very little was known about the molecular genetics of SFT. However, using whole-exome and transcriptome sequencing, three groups have recently identified NAB2–STAT6 gene fusions in the vast majority of SFTs.11, 12, 13 These genes are located close together on chromosome 12 and are transcribed in opposite directions. The fusion product results from an inversion at the 12q13 locus, which fuses NAB2 and STAT6. The resultant fusion protein, in which a repressor domain of NGFI-A binding protein 2 (EGR1 binding protein 2) (NAB2) is replaced by a transactivation domain from the carboxy terminal part of signal transducer and activator of transcription 6, interleukin-4 induced (STAT6), is believed to act as a transcriptional activator through early growth response (EGR)-mediated pathways. In a study of mesenchymal tumors of the central nervous system, Schweizer et al15 have identified the same fusion gene in meningeal SFTs and hemangiopericytomas, confirming that meningeal ‘hemangiopericytoma’ is indeed a hypercellular variant of SFT (similar to the situation at other anatomical sites);14 the authors also demonstrated that nuclear STAT6 protein overexpression by immunohistochemistry is restricted to this tumor type among meningeal tumors.

Although in most cases the diagnosis of SFT is relatively straightforward, some tumors may mimic other benign mesenchymal tumors or sarcomas, and a subset of SFTs lack expression of CD34. In this study, we investigated STAT6 protein expression by immunohistochemistry in SFT and histologic mimics to validate the utility of STAT6 as a diagnostic marker.

Materials and methods

Cases were retrieved from the surgical pathology and consult files of Brigham and Women’s Hospital, Boston, MA and Skåne University Hospital, Lund, Sweden, and the consult files of one of the authors (CDMF). Representative hematoxylin and eosin-stained slides were reviewed. Cases of SFT were classified according to the WHO Classification of Tumors as conventional (classic), malignant (>4 mitoses per 10 high-power fields, with or without hypercellularity, atypia, and infiltrative growth), and cellular or atypical in the presence of marked diffuse hypercellularity or nuclear atypia, respectively, but without increased mitotic activity.16 In total, whole-tissue sections of 231 tumors were evaluated for expression of STAT6: 60 cases of SFT (including 31 conventional (4 with prominent myxoid stroma), 9 cellular, 5 atypical, and 15 malignant variants), 20 malignant peripheral nerve sheath tumors, 10 sarcomatoid mesotheliomas, 10 cases of desmoid fibromatosis, 10 cellular schwannomas, 20 monophasic synovial sarcomas, 20 cases of dermatofibrosarcoma protuberans, 10 low-grade fibromyxoid sarcomas, 10 soft tissue perineuriomas, 10 deep fibrous histiocytomas, 10 gastrointestinal stromal tumors (spindle cell type), 21 dedifferentiated liposarcomas, 10 spindle cell lipomas, and 10 cellular angiofibromas. Six of the cases of SFT were previously shown to harbor the NAB2–STAT6 fusion gene.13

Immunohistochemistry for STAT6 was performed on 4-μm-thick formalin-fixed paraffin-embedded tissue sections following pressure cooker antigen retrieval (0.01 M citrate buffer; pH, 6.0) using a rabbit polyclonal antibody directed against the C terminus of STAT6 (1:1000; sc-621; Santa Cruz Biotechnology, Santa Cruz, CA, USA). Appropriate positive (SFT with genetically confirmed NAB2–STAT6 gene fusion) and negative controls were used throughout. It is noteworthy that during antibody optimization with low dilutions, weak nuclear staining was observed in some normal cell types (vascular endothelium, lymphocytes), whereas at higher dilutions normal tissues were completely negative, but strong staining was seen in the nuclei of a SFT confirmed to have the NAB2–STAT6 fusion gene. The extent of immunoreactivity was graded according to the percentage of positive tumor cells (0, no staining; 1+, <5%; 2+, 5–25%; 3+, 26–50%; 4+, 51–75%; and 5+, 76–100%), and the intensity of staining was graded as weak, moderate, or strong.

Results

The SFTs arose over a wide anatomic distribution, including 10 in the extremities (6 lower and 4 upper), 10 in the head and neck, 9 in the thoracic cavity (5 pleura, 3 lung, and 1 mediastinum), 7 in the abdominal cavity (including 1 each in the liver, uterus, pancreas, and urinary bladder), 7 in the pelvis, 6 in the retroperitoneum, 6 in the trunk, 4 in the inguinal region, and 1 in the meninges. The results of immunohistochemistry for STAT6 are summarized in Table 1. Fifty-nine of 60 cases of SFT (98%) showed nuclear staining for STAT6, which was usually diffuse (5+ in 41 cases; 4+ in 6 cases; 3+ in 5 cases; 2+ in 3 cases; 1+ in 4 cases) and intense (strong in 40 cases; moderate in 15 cases; and weak in 4 cases; Figure 1). In cases with less than diffuse staining (ie, 1+ or 2+) and in four cases with strong expression, heterogeneous staining in a zonal distribution pattern was often seen, with areas in the center of the tissue sections showing no (or weaker) staining, suggesting that this may be related to incomplete tissue fixation. The one SFT that was negative for STAT6 by immunohistochemistry showed histological features of malignancy, was positive for CD34, and was previously demonstrated to have a NAB2–STAT6 fusion gene. The remaining five SFTs known to harbor the NAB2–STAT6 fusion gene showed diffuse moderate-to-strong expression of STAT6. Of the 59 SFTs that were positive for STAT6, 4 were negative for CD34, 54 were positive, and in 1 case CD34 expression status was unknown.

Table 1 Summary of immunohistochemical staining for STAT6
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Figure 1
figure 1

Classic solitary fibrous tumor (SFT) composed of bland spindle cells with dense stromal collagen and branching ‘hemangiopericytoma-like’ vessels (a). Most of the tumor cells show nuclear expression of STAT6, whereas vascular endothelial cells are negative (b). Cellular SFT; the tumor cells are ovoid and there is minimal stromal collagen (c). Diffuse, strong nuclear expression of STAT6 is observed in tumor cells (d). This example of malignant SFT, which showed conspicuous mitotic activity, resembled synovial sarcoma, with monomorphic spindle cells arranged in a somewhat fascicular architecture (e). The tumor cells show nuclear expression of STAT6 (f).

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All other tumor types were negative for STAT6 (Figure 2), with the exception of 3 of 21 (14%) dedifferentiated liposarcomas, which showed predominantly weak staining (4+ weak in 1 case; 3+ weak in 1 case; 1+ weak in the well-differentiated component only of 1 case; Figure 3), and 1 of 10 deep fibrous histiocytomas, which showed 1+ weak nuclear staining.

Figure 2
figure 2

Monophasic synovial sarcoma composed of fascicles of uniform spindle cells with overlapping nuclei (a). The tumor cells are negative for STAT6 (b). Cellular spindle cell lipoma may show some overlapping features with solitary fibrous tumor (SFT) (c). STAT6 is negative in tumor cells (d).

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Figure 3
figure 3

Dedifferentiated liposarcoma with highly atypical and pleomorphic tumor cells within a fibromyxoid stroma (a). The tumor cells were positive for MDM2 and CDK4 (not shown) and showed focal weak nuclear expression of STAT6 (b).

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Eight tumors other than SFT (one low-grade fibromyxoid sarcoma, one spindle cell lipoma, one cellular angiofibroma, one cellular schwannoma, two cases of desmoid fibromatosis, and two malignant peripheral nerve sheath tumors) showed a barely perceptible ‘blush’ of staining, but lacked a distinct nuclear pattern of expression and were therefore interpreted as negative.

Discussion

SFT is a fibroblastic neoplasm with variable clinical behavior. Although most SFTs pursue a benign clinical course, recurrence or metastasis develops in 5–10% of patients. The majority of SFTs that behave aggressively show histological features of malignancy, such as >4 mitoses per 10 HPF and hypercellularity, but benign-appearing SFTs can also rarely give rise to metastases.17 Histologically, SFT may resemble some benign soft tissue tumors (eg, soft tissue perineurioma, desmoid fibromatosis, spindle cell lipoma, and cellular angiofibroma); soft tissue tumors with potential for locally aggressive behavior (eg, desmoid fibromatosis); and spindle cell sarcomas (eg, malignant peripheral nerve sheath tumor, dermatofibrosarcoma protuberans, and monophasic synovial sarcoma). The immunohistochemical profile of SFT using conventional markers is relatively nonspecific, with CD34 expression being the most consistent finding reported to date, present in 95% of cases. Other markers that are variably expressed in SFT include CD99, BCL2, nuclear β-catenin, and EMA. However, all these markers are positive in other soft tissue tumors that may mimic SFT.

Several recent studies have detected a recurrent intrachromosomal fusion between the NAB2 and STAT6 genes on chromosome 12 in SFT.11, 12, 13 This genomic inversion at the 12q13 locus results in the formation of a NAB2–STAT6 fusion gene, which can be detected by reverse transcriptase-PCR. However, as these genes are located in close proximity on 12q13, this fusion can only rarely be detected by conventional chromosomal banding or fluorescence in situ hybridization (FISH) analysis. In the study by Robinson et al,11 all 51 SFTs examined harbored the NAB2–STAT6 fusion gene. A lower frequency of this gene fusion was reported in the study by Chmielecki et al,12 being detected in just over 50% of cases, possibly reflecting diagnostic variability. Mohajeri et al13 identified NAB2–STAT6 in 90% of SFTs evaluated. The fusion gene has not been identified in other soft tissue neoplasms studied to date. These results have also been documented in meningeal SFTs (including tumors formerly known as meningeal ‘hemangiopericytoma’).15

STAT6 is a member of the STAT family of cytoplasmic transcription factors, which regulate gene expression by transmitting signals to the nucleus and binding to specific DNA promoter sequences. STAT signaling is critical for normal cellular processes such as embryonic development, innate and adaptive immune function, and regulation of cell differentiation, growth, and apoptosis.18, 19 Activation of STAT family members is a well-recognized alteration in human malignancies.19 STAT6 is composed of a DNA-binding domain, a C-terminal transcriptional activation domain, and a SH2 domain. In contrast to STAT6, NAB2 normally functions as a transcriptional repressor through its interaction with the EGR family of transcription factors and is localized to the nucleus.20 However, NAB2 gains an activation domain when fused to STAT6, and the NAB2–STAT6 fusion gene therefore acts as a transcriptional activator, inducing expression of EGR target genes, which results in increased proliferation.11 It is hypothesized that the fusion protein results in translocation to the nucleus and high levels of expression compared with normal tissues and other neoplasms, as demonstrated by immunofluorescence labeling against the C terminus of STAT611 and immunohistochemical analysis of STAT6 in SFT of the meninges, which showed diffuse nuclear expression in meningeal SFT, but not in other dural tumors.15

We found that 98% of a large cohort of SFTs (including conventional, cellular, atypical, and malignant variants) showed nuclear expression of STAT6. Staining for STAT6 was usually diffuse: 68% of cases showed reactivity for STAT6 in >75% of tumor cells. Further, the intensity of staining was strong in 67% of cases, moderate in 25%, and weak in only 7%. As discussed in the Results section, some tumors showed heterogeneity of staining, both in terms of extent and intensity, which may be because of uneven tissue fixation or loss of antigenicity in older cases for which the unstained slides were stored for extended periods of time. All other tumor types examined were negative for STAT6, except for three dedifferentiated liposarcomas and one deep fibrous histiocytoma, which showed weak staining. Schweizer et al15 reported cytoplasmic expression of STAT6 in tumors other than SFT, albeit usually weak in intensity. In our study, cytoplasmic expression was not detected, which may reflect the higher dilution of antibody used in the current study.

The finding of focal nuclear STAT6 expression in a subset of cases of dedifferentiated liposarcoma is interesting, and may suggest a role for STAT6-mediated transcriptional activity in some cases of dedifferentiated liposarcoma, given that the 12q13 locus contains several different regions that are frequently amplified in dedifferentiated liposarcoma, such as MDM2 and CDK4. In practice, dedifferentiated liposarcoma may enter the morphologic differential diagnosis with SFT, particularly for those tumors occurring in the retroperitoneum, and therefore the finding of STAT6 expression in dedifferentiated liposarcoma is a potential diagnostic pitfall. However, demonstration of positivity for MDM2 and CDK4 by immunohistochemistry21, 22 or MDM2 amplification by FISH23 can help confirm the diagnosis of dedifferentiated liposarcoma.

The discovery of a recurrent gene fusion by whole-exome and transcriptome sequencing is a novel method of detecting translocation-associated sarcomas. In the case of SFT, such methods have resulted in the identification of a useful diagnostic immunohistochemical marker, STAT6, following identification of the recurrent gene fusion product NAB2–STAT6 in this tumor type. This adds to the expanding group of markers for soft tissue tumors identified through molecular genetic methods. For example, gene expression analysis has previously identified ANO1 (anoctamin 1, calcium activated chloride channel; also known as DOG1) as a specific marker of gastrointestinal stromal tumor,24, 25, 26, 27 TLE1 (transducin-like enhancer of split 1) for synovial sarcoma,27, 28 and MUC4 (mucin 4, cell surface associated) for low-grade fibromyxoid sarcoma.29, 30 Nuclear expression of STAT6 is found in nearly all cases of SFT and is very limited in other soft tissue neoplasms. STAT6 is therefore a highly sensitive and almost perfectly specific immunohistochemical marker for SFT, and can be helpful to distinguish this tumor type from histologic mimics.