The American Journal of Pathology, Vol. 184, No. 4, April 2014

TUMORIGENESIS AND NEOPLASTIC PROGRESSION

Solitary Fibrous Tumors/Hemangiopericytomas withDifferent Variants of the NAB2-STAT6 Gene Fusion AreCharacterized by Specific Histomorphology and DistinctClinicopathological FeaturesSarah Barthelmeß,* Helene Geddert,y Carsten Boltze,z Evgeny A. Moskalev,* Matthias Bieg,x Horia Sirbu,{ Benedikt Brors,x

Stefan Wiemann,k** Arndt Hartmann,* Abbas Agaimy,* and Florian Haller*

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From the Institute of Pathology* and the Department of Thoracic Surgery,{ University Hospital Erlangen, Erlangen; the Institute of Pathology,y St. Vincent’sHospital, Karlsruhe; the Institute of Pathology,z SRH-Klinikum, Gera; and the Divisions of Theoretical Bioinformaticsx and Molecular Genome Analysisk andthe Genomics and Proteomics Core Facility,** German Cancer Research Center, Heidelberg, Germany

Accepted for publication

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December 12, 2013.

Address correspondence toFlorian Haller, Dr.Med., Diag-nostic Molecular Pathology,Institute of Pathology, Univer-sity Hospital Erlangen, Kran-kenhausstrasse 8-10, D-91054Erlangen, Germany. E-mail:florian.haller@uk-erlangen.de.

opyright ª 2014 American Society for Inve

ublished by Elsevier Inc. All rights reserved

ttp://dx.doi.org/10.1016/j.ajpath.2013.12.016

Recurrent somatic fusions of the two genes, NGFI-Aebinding protein 2 (NAB2) and STAT6, locatedat chromosomal region 12q13, have been recently identified to be presumable tumor-initiatingevents in solitary fibrous tumors (SFT). Herein, we evaluated a cohort of 52 SFTs/hemangioper-icytomas (HPCs) by whole-exome sequencing (one case) and multiplex RT-PCR (all 52 cases), andidentified 12 different NAB2-STAT6 fusion variants in 48 cases (92%). All 52 cases showed strongand diffuse nuclear positivity for STAT6 by IHC. We categorized the fusion variants according totheir potential functional effects within the predicted fusion protein and found strong correlationswith relevant clinicopathological features. Tumors with the most common fusion variant, NAB2ex4-STAT6ex2/3, corresponded to classic pleuropulmonary SFTs with diffuse fibrosis and mostly benignbehavior and occurred in older patients (median age, 69 years). In contrast, tumors with thesecond most common fusion variant, NAB2ex6-STAT6ex16/17, were found in much younger patients(median age, 47 years) and represented typical HPCs from deep soft tissue with a more aggressivephenotype and clinical behavior. In summary, these molecular genetic findings support the conceptthat classic pleuropulmonary SFT and deep-seated HPC are separate entities that share commonfeatures but correlate to different clinical outcome. (Am J Pathol 2014, 184: 1209e1218; http://dx.doi.org/10.1016/j.ajpath.2013.12.016)

Supported by departmental funding from the Institute of Pathology,University Erlangen-Nuremberg.

Disclosures: None declared.

In 1942, Stout and Murray1 described a series of nine tu-mors sharing features of endothelial-lined tubes and sproutsand a pattern of well-developed branching staghorn thick-walled vessels with perivascular fibrosis, which theynamed hemangiopericytoma (HPC). In a follow-up study,Stout2 reported on 25 cases with similar features. Theconcept of HPC as a specific tumor entity was laterconsolidated in a series of 106 cases by Enzinger andSmith,3 who refined the diagnostic criteria and establishedparameters for assessment of malignancy. Also in 1942,Stout and Murray4 published a case of a solitary mesen-chymal tumor of the lung and pleura, which they namedsolitary (localized) mesothelioma of the pleura, citing a first

stigative Pathology.

.

report on a series of similar tumors by Klemperer andRabin.5 These reports were followed up later in anotherstudy establishing the characteristic features of spindle-shaped tumor cells with prominent diffuse connective tis-sue fibers within these mostly well-circumscribed tumors,later renamed as solitary fibrous tumor (SFT).6 Although, inthe initial descriptions, only tumors from the pleural cavityand lungs were included,7,8 it was not until the 1990s thatcomparable tumors were reported for other anatomical

Barthelmeß et al

locations,9,10 and the concept of extrapleural SFT wasestablished.11,12

Enzinger and Smith3 had already emphasized that themost prominent feature of HPC, the ramifying vessels, canalso be found in other mesenchymal neoplasms. An HPC-like vascular pattern has been reported for many well-defined other soft tissue sarcomas.13,14 Since then, it hasbeen debated for a long time whether HPC represents adistinct clinicopathological entity or only a non-specificvascular pattern.14e16 On the basis of histomorphologicaland clinical similarities and the lack of clear separatingdiagnostic criteria, it has been proposed that HPC and SFTrepresent different points on a broad morphological spec-trum of the same entity.16,17 Although some authors stillseparate fibrous and cellular variants of SFT, resemblingclassic SFT and HPC, respectively,16 these tumors weremerged together in the fourth edition of the World HealthOrganization classification of tumors of soft tissue.18

Furthermore, pleural and extrapleural SFTs are distin-guished according to their anatomical localization. Fea-tures suggestive of malignancy are increased mitoticcounts, nuclear atypia, tumor necrosis, and infiltrativemargins.3,7,8,19e21

Only recently, recurrent fusion events of the two genes,NGFI-Aebinding protein 2 (NAB2) and STAT6, bothlocated at chromosomal region 12q13, have been identifiedby next-generation sequencing and RT-PCR in most SFTsand likely represent an initial tumorigenic event.22e24

Different fusion variants occurring at varying frequencieshave been found in the three studies published so far, but, toour knowledge, no characteristic association with clinico-pathological features has been reported.22e24 In contrast, theoverall frequent finding of NAB2-STAT6 gene fusions andspecific nuclear expression of STAT6 in benign and ma-lignant SFTs of pleuropulmonary and different extrapleuralsites is considered as further confirmation of the unifyingconcept of SFT as a single entity.22e25

For the current study, we evaluated a cohort of 52 SFTs/HPCs for the presence of NAB2-STAT6 gene fusions. First,we performed whole-exome sequencing of a recurrentpleural SFT and identified an identical NAB2-STAT6 genefusion in two independent metachronous local recurrences.Second, we established a multiplex RT-PCR screeningassay that interrogates all possible fusion combinations ofNAB2 (seven exons) and STAT6 (23 exons) on the cDNAlevel, potentially amplifying a total of 161 different fusionvariants. Characterizing the whole cohort of 52 tumors, wewere able to identify 12 different fusion variants in 48 cases,confirming the high frequency of this recurrent geneticevent. The different fusion variants were grouped accordingto their potential functional effects among the predictedchimeric fusion proteins. Strikingly, the tumors within thetwo most frequent categories of fusion variants, NAB2ex4-STAT6ex2/3 and NAB2ex6-STAT6ex16/17, displayedspecific recurrent histomorphological characteristics anda significant association with relevant clinicopathological

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features, reminiscent of the original descriptions of SFT andHPC, respectively.1e8

Materials and Methods

Patient Cohort and STAT6 IHC

Tumors with histological and immunohistochemical (IHC)features consistent with a diagnosis of SFT or HPC wereretrieved from the surgical pathology archives of threeGerman institutions (Institute of Pathology, University Hos-pital Erlangen, Erlangen; Institute of Pathology, St Vincent’sHospital, Karlsruhe; and Institute of Pathology, SRH-Klini-kum, Gera). Primary meningeal and sinonasal tumors wereexcluded from the current series, as well as tumors displayingclassic features of myofibroma, myopericytoma, or giant-cellangiofibroma. All tumors were reclassified by an experiencedsoft tissue pathologist (A.A.) after review of the availableslides, without knowledge of the molecular findings.The IHC for STAT6 was performed as described recently

by Doyle et al,25 using a polyclonal antibody directedagainst STAT6 (1:1000 dilution, sc-621; Santa CruzBiotechnology, Santa Cruz, CA). Only unequivocal strongand diffuse nuclear staining for STAT6 was considered aspositive. Paucicellular cases with spindled tumor cells anduniform fusiform or spindled nuclei with scanty cytoplasm,arranged in a wavy and patternless pattern within a promi-nent diffuse fibrous stroma displaying typical cracking ar-tifacts, were classified as classic fibrous SFTs (Figure 1,AeC). Conventional (paucicellular) pleuropulmonary SFTswere considered the prototype of this variant. Tumors with ahigher cellularity and occasionally more rounded cells, withround to oval chromatin-rich nuclei but prominent peri-vascular fibrosis, were considered cellular SFTs (Figure 1,DeF). This cellular variant is generally of similar archi-tecture as the paucicellular fibrous variant, with the gapingstaghorn vessels being more prominent at the periphery ofthe lesion. One case (case 25) with mixed patterns of pau-cicellular and more cellular areas was included in thiscategory. In contrast, highly cellular tumors, with a well-developed net of thin-walled, dilated, or slit-like commu-nicating vessels without perivascular fibrosis/hyalinization,were designated as HPCs (Figure 1, GeI). These tumorsgenerally looked more primitive, without any fibrous stromabut occasionally with myxoid changes. Their vessels lackedany prominent connective tissue layer, and the liningendothelial cells seem to merge directly with the sur-rounding tumor cells, forming the vascular wall. Most of thelatter tumors displayed characteristic branching staghorn-like vessels that were not confined to the tumor peripherybut were frequently seen throughout the whole width. Mi-toses were counted in 10 high-power fields (HPFs). Tumorswith more than four mitoses in 10 HPFs, marked nuclearatypia, tumor necrosis, and/or infiltrative margins, weretermed histologically malignant, in contrast to histologicallybenign tumors lacking these features.3,7,8,19e21

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Figure 1 Histomorphological tumor classifica-tion. AeC: Prototype of classic pleuropulmonarySFTs. A: These tumors displayed a wavy patternlesspattern of spindled tumors cells within a promi-nent diffuse fibrous stroma. B: The tumor cells arespindled with fusiform nuclei. C: Diffuse andstrong nuclear positivity for STAT6. DeF: Pleuralcellular SFTs, second recurrence. D: Although thesetumors were more cellular, characteristic featuresof the fibrous variant are still evident. E: Peri-vascular hyalinization. F: Diffuse and strong nu-clear positivity for STAT6. GeI: Prototype oftypical deep-seated HPC. G: These tumors werehighly cellular, with a monotonous appearance andcharacteristic staghorn vessels. H: Note theabsence of perivascular hyalinization reminiscentof the original description of pericytic growthpattern. I: Diffuse and strong nuclear positivity forSTAT6. For A, B, D, E, G, and H, H&E was used.Original magnifications: �100 (A, D, and G);�400 (B, C, E, F, H, and I).

NAB2-STAT6 Fusion Variants in SFTs/HPCs

Clinical follow-up was available for 33 (63%) of 52 pa-tients, including 13 with tumor recurrences. Four patientswith recurrent tumors died of their disease 3, 5, 18, and 26years after initial diagnosis, respectively. Twenty patientshad no signs of tumor recurrence after a median of 5 years(�3 years; range, 1 to 15 years). One of these patients diedof another cause 5 years after the diagnosis. This study hasbeen approved by the local ethics committee of the Uni-versity Hospital Erlangen, Germany (number 217_12 B,19.09.2012). Signed informed consent was obtained fromall participating patients in this study.

Whole-Exome Sequencing

DNA was isolated from two fresh-frozen local recurrences ofa pleural SFT and from a matched blood sample of the samepatient serving as control. DNA was fragmented (CovarisE220; Covaris, Woburn, MA) to 150 to 200 bp, followed byenrichment of exonic regions and index tagging (AgilentSure Select, version 4; Agilent Technologies, Santa Clara,CA). Paired-end sequencing (2 � 100 bp) was performedusing a HiSeq2000 (Illumina, Inc., San Diego, CA) to a meancoverage of >120�. [Mapping and target coverage calcula-tion were done using Burrows-Wheeler Alignment (BWA)tool and the OnTarget tool (http://www.dkfz.de/gpcf/ontarget/ontarget2.tar.gz)].26,27 Breakpoints of translocations wereidentified using the clipping reveals structure (CREST) algo-rithm, which uses soft-clipped reads to discover genomic lo-cations with many partially mapped reads.28 The soft-clippedportions of the reads were assembled into a consensussequence that was afterward remapped against the reference.The genomic location of the remapped consensus sequencewas thus identified as the translocation position.

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Multiplex RT-PCR

Total RNA was isolated from formalin-fixed, paraffin-embedded (FFPE) tumor samples using the RNeasy FFPEKit and Deparaffinization Solution (Qiagen, Hilden, Ger-many). cDNA was synthesized from 700 ng of total RNAper sample with the Quantitect Reverse Transcription Kit(Qiagen) containing a combination of oligo-dT and randomprimers. A multiplex RT-PCR protocol was establishedusing forward primers located in NAB2 exons 1 to 7 andreverse primers positioned in STAT6 exons 1 to 22 (primersequences available on request) and performed by Hot-StarTaq DNA Polymerase reagents (Qiagen) with a temper-ature profile consisting of an initial step of 95�C for15 minutes, 50 cycles of 95�C for 30 seconds, 57.6�C for45 seconds, and 72�C for 1 minute, and a terminal step of72�C for 5 minutes. RT-PCR of hypoxanthine phosphor-ibosyltransferase 1 (HPRT1) was used as control for RNAintegrity. Samples with PCR products in the multiplex assaywere subsequently analyzed by omitting different forwardand reverse primers until a specific NAB2-STAT6 fusionvariant was amplified. All detected NAB2-STAT6 fusionvariants were confirmed by Sanger sequencing on a cDNAlevel.

Results

Whole-Exome Sequencing of a Recurrent Pleural SFTReveals a Genomic NAB2-STAT6 Gene Fusion

Analysis of the whole-exome sequencing data, derived fromtwo independent metachronous local recurrences of a ma-lignant pleural SFT (patient 24) (Table 1), revealed an

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Table 1 Patient and Tumor Characteristics

Case Age/sex Histological features Location Follow-upSize (cm)/mitoticcounts (10 HPFs)

Estimateddignity*

NAB2-STAT6fusion variant

STAT6IHC

Tumors with NAB2ex4-STAT6ex2/3 fusion variants1 74/M Fibrous SFT Pleura DOOC (5 years) 2.8/0 Benign 4e2 þ2 61/M Fibrous SFT Pleura NA 3.5/0 Benign 4e2 þ3 41/F Fibrous SFT Pleura AWOD (2 years) 4/0 Benign 4e2 þ4 48/F Fibrous SFT Pleura NA 4/0 Benign 4e2 þ5 69/M Fibrous SFT Pleura AWOD (4 years) 4/0 Benign 4e2 þ6 69/M Fibrous SFT Lung NA 6/0 Benign 4e2 þ7 43/F Fibrous SFT Pleura AWOD (1 year) 7/0 Benign 4e2 þ8 79/F Fibrous SFT Pleura AWOD (4 years) 8/0 Benign 4e2 þ9 67/F Fibrous SFT Lung NA 8.5/0 Benign 4e2 þ10 72/M Fibrous SFT Cervical AWOD (6 years) 8.5/0 Benign 4e2 þ11 71/F Fibrous SFT Pleura NA 9.5/0 Benign 4e2 þ12 83/M Fibrous SFT Pleura AWD (6 years) 10/0 Benign 4e2 þ13 60/M Fibrous SFT Pleura AWOD (6 years) 11/0 Benign 4e2 þ14 72/F Fibrous SFT Pleura NA 13/0 Benign 4e2 þ15 60/M Fibrous SFT Pleura NA 15.5/0 Benign 4e2 þ16 73/F Fibrous SFT Lung AWOD (2 years) 15.5/1 Benign 4e2 þ17 61/M Fibrous SFT Pleura AWOD (4 years) 18/1 Benign 4e2 þ18 83/F Fibrous SFT Pleura AWOD (1 year) 21/0 Benign 4e2 þ19 61/M Fibrous SFT Pleura NA 25/1 Benign 4e2 þ20 62/F Fibrous SFT Pleura AWOD (4 years) 26.5/0 Benign 4e2 þ21 73/M Cellular SFT Pleura NA 13/0 Benign 4e2 þ22 84/F Cellular SFT Pleura AWD (11 years) 7/7 Malignant 4e2 þ23 79/F Cellular SFT Lung NA 5/8 Malignant 4e2 þ24 65/F Cellular SFT Pleura AWD (7 years) 12/14 Malignant 4e2 þ25 78/F Cellular SFT Pleura DOT (3 years) 11.5/45 Malignant 4e2 þ26 69/F Fibrous SFT Lung AWOD (1 year) 1.6/0 Benign 4e3 þ27 58/M Cellular SFT Inguinal NA 3.5/0 Benign 4e3 þ

Tumors with NAB2ex6-STAT6ex16/17 fusion variants28 33/F Cellular SFT Paraurethral AWOD (8 years) 8/3 Benign 6e16 þ29 46/M HPC Pelvis AWOD (6 years) 2.2/3 Benign 6e16 þ30 49/F HPC Paravertebral DOT (26 years) 5/23 Malignant 6e16 þ31 y/F HPC Paravertebral AWD 3/8 Malignant 6e16 þ32 31/F HPC Parotid gland DOT (18 years) 2.7/11 Malignant 6e16 þ33 47/M HPC Paravertebral AWD (15 years) 4.5/9 Malignant 6e16 þ34 52/M HPC Paravertebral AWD 6/22 Malignant 6e16 þ35 61/F Cellular SFT Lower leg NA 1.9/0 Benign 6e17 þ36 35/M HPC Nuchal AWOD (4 years) 1/1 Benign 6e17 þ37 y/F Cellular SFT Trunk wall AWD 8.5/8 Malignant 6e17 þ38 82/M HPC Inguinal AWD (1 year) 9/53 Malignant 6e17 þ

Tumors with other NAB2-STAT6 fusion variants39 33/M Fibrous SFT Orbita NA 2.5/0 Benign 3e17 þ40 45/F Fibrous SFT Cheek AWOD (5 years) 0.6/1 Benign 3e18 þ41 65/M Fibrous SFT Trunk wall AWD (8 years) 5.5/2 Benign 3e19 þ42 54/M Cellular SFT Orbita AWOD (15 years) 1/0 Benign 3e19 þ43 39/M Cellular SFT Parotid gland NA 3.5/1 Benign 3e19 þ44 54/F Cellular SFT Lung AWOD (4 years) 3/3 Benign 4e18 þ45 67/F HPC Sublingual NA 2.9/0 Benign 5e18 þ46 49/F Cellular SFT Trunk wall AWOD (7 years) 0.9/2 Benign 7e2 þ47 67/F Cellular SFT Retroperitoneal NA 12/0 Benign 7e18 þ48 72/F Cellular SFT Retroperitoneal NA 4.2/0 Benign 7e20 þ

Tumors without detectable NAB2-STAT6 fusions49 68/F Fibrous SFT Lower leg DOT (5 years) 7/1 Benign None þ50 48/M Fibrous SFT Tongue NA 2/0 Benign None þ51 57/F Fibrous SFT Lung NA 7/0 Benign None þ52 47/F HPC Parapharyngeal AWOD (2 years) 4/0 Benign None þ*The dignity was estimated according to published criteria mitotic counts (>4/10 HPFs), marked nuclear atypia, tumor necrosis, and/or infiltrative

margins.3,7,8,19e21yBoth patients experienced recurrent disease for several years, but their age at the first tumor manifestation could not be ascertained.F, female;M,male; AWD, alivewith disease; AWOD, alivewithout disease; DOOC, diedof other cause; DOT, died of tumor;NA, not available;þ, positive nuclear staining.

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NAB2-STAT6 Fusion Variants in SFTs/HPCs

NAB2-STAT6 gene fusion in both samples, which was notpresent in the blood control (Figure 2). The genomicbreakpoints were identified using the CREST software andwere located on chr12:57486804 (NAB2 intron 4) andchr12:57508845 (upstream of STAT6). The predicted fusionprotein consisted of the N-terminal portion of NAB2encoded by exons 1 to 4, which was fused to the wholeSTAT6 protein.

NAB2-STAT6 Multiplex RT-PCR Analysis DetectsRecurrent Fusion Variants on the cDNA Level in 48 of52 SFTs/HPCs

We detected single NAB2-STAT6 fusion variants in 48(92%) of 52 cases (Table 1) using the multiplex RT-PCRapproach. The RT-PCR assay confirmed the NAB2-STAT6fusion variant identified by whole-exome sequencing in case

Figure 2 Genomic structure of the NAB2-STAT6 locus at human chromosomal rSFT. A: Wild-type organization of the affected genes [extracted from the Universbreakpoints within NAB2 intron 4 (proximal) and upstream of the STAT6 gene (distthe RepeatMaster (version open-3-2-7) track of UCSC. The distal breakpoint maps intVariant situation in consequence of the assumed inversion event in DNA extractedupstream sequence of STAT6 are inverted, so that on the mRNA level, exon 4 of NAB2Exon 1 of STAT6 corresponds to the 50-untranslated region. C: The exact mapping ofwhole-exome sequencing data. Breakpoints are indicated (arrowheads), and sequeMapping of individual reads in DNA from blood, first recurrence, and second recur

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24, as previously described. Overall, Sanger sequencing ofthe cDNA breakpoint region allowed the identification of12 different fusion variants. The three fusion variants,NAB2ex4-STAT6ex2 (n Z 25), NAB2ex6-STAT6ex16(nZ 7), and NAB2ex6-STAT6ex17 (nZ 4), were the mostfrequent events and accounted together for 75% of theobserved fusion products. All 52 tumors demonstrated astrong and diffuse nuclear positivity for STAT6 in the IHCanalysis. However, we could not find an NAB2-STAT6fusion variant by RT-PCR in four cases (8%).

The NAB2-STAT6 fusion variants were grouped accord-ing to their potential functional effects among the predictedchimeric proteins (Figure 3). A comparison with the clini-copathological patient data revealed that the different cate-gories of fusion variants were significantly correlated topatient age (Kruskal-Wallis test, P Z 0.002) (Table 2)(Figure 4A), tumor size (P Z 0.001) (Figure 4B), and

egion 12q13.13 and gene fusion breakpoints observed in a recurrent pleurality of California, Santa Cruz (UCSC) genome browser]. The locations of theal) are indicated (arrowheads). Localization of repeat structures is shown ino an MER112 element with several Alu elements upstream and downstream. B:from a recurrent pleural SFT. The genomic sequence of exon 5 of NAB2 andis spliced to exon 2 of STAT6, thus producing the observed fusion transcript.breakpoints for the proximal (NAB2) and distal (upstream STAT6) sites usingnces matching the NAB2 and STAT6 upstream regions are in black boxes. D:rence, indicating that the inversion was not present in the blood DNA.

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Figure 3 Comparison of the two most frequentcategories of NAB2-STAT6 fusion variants on thecDNA level and the predicted chimeric fusionproteins. A: Wild-type forms of the NAB2 andSTAT6 proteins with functional domains. B and C:The NAB2ex4-STAT6ex2 fusion variant encodes fora chimeric fusion protein that lacks the CID (C)terminal repressor domain of NAB2, but containsthe complete STAT6 protein (B). In contrast, theNAB2ex6-STAT6ex16 fusion variant encodes for thealmost complete NAB2 protein fused to a consid-erably truncated portion of STAT6 correspondingto the terminal TAD (C). Shown are schematic di-agrams to allow comparison of the different exonswith the respective encoded protein domains. Notethe fusion breakpoints in the Sanger chromato-grams, indicated by vertical bars.

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mitotic activity (P Z 0.001) (Figure 4C). Moreover, therewas a significant correlation with the anatomical localizationof the tumors (c2 test, P Z 7.5 � 10�8) (Figure 5A), his-tomorphological classification (P Z 2.4 � 10�7)(Figure 5B), histologically estimated dignity (P Z 0.001),and the clinical follow-up (P Z 0.016) (Figure 5C).

Most Tumors with NAB2ex4-STAT6ex2/3 FusionVariants Are Classic Fibrous Pleuropulmonary SFTs witha Benign Appearance

The most frequent fusion variant displayed the breakpointbetween NAB2 exon 4 and STAT6 exon 2 and wasobserved in 25 cases (52%). The corresponding predictedchimeric protein was composed of a truncated form ofNAB2 with loss of the C-terminal repressive domain fusedto the complete STAT6 protein (Figure 3). Two additionaltumors had a breakpoint between NAB2 exon 4 and STAT6exon 3, resulting in a probably functionally similar chimericprotein, and were thus classified within this category. Of 27tumors, 25 (93%) with an NAB2ex4-STAT6ex2/3 fusionvariant were located within the thoracic cavity, comprising20 tumors of the pleura and five tumors of the lung. Onecase was located in subcutaneous cervical tissue, and onecase was situated in the inguinal region. Overall, 25 (93%)of the 27 pleuropulmonary SFTs harbored this fusionvariant. The median age of the patients (69 years; range,41 to 84 years) was significantly higher compared withpatients with the other fusion variants. Tumors carrying the

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NAB2ex4-STAT6ex2/3 fusion variant were also signifi-cantly larger, with a mean size of 10.3 (�6.6) cm. Twenty-one of the tumors displayed the classic fibrous SFTmorphological characteristics, and six were classified ascellular SFTs. None of the 21 classic fibrous SFTs had morethan one mitotic figure in 10 HPFs, whereas four of the sixcellular SFTs showed an increased mitotic count (>4 of 10HPFs). The median mitotic count of all tumors with theNAB2ex4-STAT6ex2/3 fusion variant was 0 mitoses in 10HPFs, and the mean mitotic count was 2.9 (�9.0). Clinicalfollow-up was available for 16 of the patients. Twelve pa-tients (44%) were event free after a mean of 3 years (�2years; range, 1 to 6 years), and all had classic fibrous SFTs.Four patients (15%) developed tumor recurrences, and oneof these died of the tumor 3 years after initial diagnosis.Three of the four cases with recurrence were cellular SFTsfulfilling the histological diagnostic criteria for malignancy(> mitoses and nuclear atypia).

Most Tumors with NAB2ex6-STAT6ex16/17 FusionVariants Represent Deep-Seated Cellular SFTs and HPCsfrom the Retroperitoneum and Pelvis with CellularAppearance, Increased Mitotic Activity, and FrequentRecurrences

The two next frequent fusion variants, NAB2ex6-STAT6ex16 (n Z 7) and NAB2ex6-STAT6ex17 (n Z 4),encoded for two closely related predicted chimeric proteins,with fusion of an almost complete NAB2 protein to a largely

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Table 2 Summary of Clinicopathological Correlations among the Different Categories of NAB2-Stat6 Fusion Variants

NAB2-STAT6 fusion variant 4e2/3 6e16/17 Other P value

Relative frequencyNo. (%) 27 (56) 11 (23) 10 (21)

Sex 0.993Male 12 (44) 5 (45) 4 (40)Female 15 (56) 6 (55) 6 (60)

Age (years) 0.002Means (�SD) 67 (�11) 48 (�16) 55 (�13)Median (range) 69 (41e84) 47 (31e82) 54 (33e72)

Location 7.5 � 10�8

Thoracic 25 (92) 0 (0) 1 (10)Head/neck 1 (4) 2 (18) 5 (50)Trunk 0 (0) 1 (9) 2 (20)Extremities 1 (4) 2 (18) 0 (0)Deep soft tissue 0 (0) 6 (55) 2 (20)

Size (cm) 0.001Means (�) 10.3 (�6.6) 4.3 (�3.0) 3.7 (�3.3)Median (range) 8 (2e26) 3 (1e9) 3 (1e12)

Histological feature 2.4 � 10�7

Fibrous SFT 21 (78) 0 (0) 3 (30)Cellular SFT 6 (22) 3 (27) 6 (60)HPC 0 (0) 8 (73) 1 (10)

Mitotic counts (10 HPFs) 0.001Means (�SD) 2.9 (�9.0) 12.8 (�15.4) 1.0 (�1.1)Median (range) 0 (0e45) 8 (0e53) 1 (0e3)

Estimated dignity 0.001Benign 23 (85) 3 (27) 10 (100)Malignant 4 (15) 8 (73) 0 (0)

Clinical follow-up 0.016No recurrence 12 (44) 3 (27) 4 (40)Recurrence 4 (15) 7 (64) 1 (10)Not available 11 (41) 1 (9) 5 (50)

Data are given as number (%) unless otherwise indicated. Significant P values are indicated in bold.

NAB2-STAT6 Fusion Variants in SFTs/HPCs

truncated form of STAT6 involving only the C-terminaltransactivation domain (Figure 3). As a group, the 11 tumors(23%) with NAB2ex6-STAT6ex16/17 fusion variants weremainly deep seated, with deep paravertebral (n Z 4) andpelvic (n Z 2) localization being the most common(Figure 3A). The patients were significantly youngercompared with the patients with the NAB2ex4-STAT6ex2/3fusion variant, with a median age of 47 years (range, 31 to

Figure 4 NAB2-STAT6 fusion variants are correlated with patient age, tumorvariants occurred at a significantly older age (P Z 0.002) (A) and were significantfusion variants displayed significantly higher mitotic counts (P Z 0.001) (C). ShoOpen circles and asterisks represent mild and extreme outliers, respectively.

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82 years). Tumors with the NAB2ex6-STAT6ex16/17fusion variants were also significantly smaller, with a meansize of 4.3 (�3.0) cm. Three tumors were consideredcellular SFTs, and eight represented typical HPCs. Tumorswith the NAB2ex6-STAT6ex16/17 fusion variants hadsignificantly higher mitotic counts compared with tumorswithin the other categories (median, 8 of 10 HPFs). Sevenpatients (64%) developed tumor recurrences, and two of

size, and mitotic counts. Tumors carrying the NAB2ex4-STAT6ex2/3 fusionly larger (P Z 0.001) (B), whereas tumors with the NAB2ex6-STAT6ex16/17wn are quartile box plots with minimum, 25%, median, 75%, and maximum.

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Figure 5 NAB2-STAT6 fusion variants are correlated with anatomical localization, histological type, and clinical follow-up. Tumors with the NAB2ex4-STAT6ex2/3 fusion variants occurred almost exclusively in the lungs and pleura (P Z 7.5 � 10�8) (A) and mainly had classic fibrous SFT morphologicalcharacteristics (P Z 2.4 � 10�7) (B), whereas tumors with the NAB2ex6-STAT6ex16/17 fusion variants were frequently located in the retroperitoneum andpelvis, and corresponded to true deep-seated HPCs. C: Regarding the patients with clinical follow-up, only 4 (25%) of 16 tumors within the NAB2ex4-STAT6ex2/3 group developed a recurrence, compared with 7 (70%) of 10 tumors within the NAB2ex6-STAT6ex16/17 group (P Z 0.016).

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these patients died of their tumor. In contrast, only threepatients (27%) had no tumor recurrence. There was nofollow-up available for one patient.

Discussion

In the current study, we analyzed a cohort of 52 SFTs/HPCsregarding the presence of different variants of the recentlydescribed NAB2-STAT6 gene fusion. All 52 cases showed astrong and diffuse nuclear positivity for STAT6 in the IHCanalysis. Twelve different fusion variants were found in 48cases, correlating to a frequency of 92%. Four cases had nodetectable NAB2-STAT6 fusion variant, despite strongnuclear positivity for STAT6. Accordingly, we cannotexclude the possibility that there are other gene fusionsinvolved in a few SFTs. However, regarding the fact thatthis study was performed on RNA isolated from formalin-fixed material, it is more likely that these four cases werenegative in the RT-PCR approach, according to technicallimitations. In summary, there is a high prevalence andconcordance between STAT6 IHC and NAB2-STAT6 genefusions in tumors of the SFT/HPC spectrum.

Notably, tumors with the most common fusion variants,NAB2ex4-STAT6ex2/3 and NAB2ex6-STAT6ex16/17,were significantly correlated to patient age, anatomicallocalization, tumor size, and mitotic counts and displayeddistinct and recurrent histomorphological patterns. Mosttumors with the NAB2ex4-STAT6ex2/3 fusion variantswere located in the thoracic cavity, and these tumors weremainly paucicellular, with spindled cells arranged in acharacteristic patternless pattern and a prominent back-ground of diffuse fibrosis. Thus, they recapitulated theclassic appearance of benign pleuropulmonary SFTs in mostcases.4e8 Because these tumors were larger and occurred inolder patients (median age, 69 years), one may presume thatthese low proliferating tumors have grown slowly for a long

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time before causing symptoms. Only 4 (25%) of 16 patientswith available clinical follow-up within this group devel-oped tumor recurrence. In contrast, tumors with theNAB2ex6-STAT6ex16/17 fusion variants were mainly deepseated in the extremities and retroperitoneal or pelvic softtissues and occurred in significantly younger patients (me-dian age, 47 years). Although tumors of the latter categorywere significantly smaller, they had significantly highermitotic counts and showed more frequently aggressiveclinical behavior. Of 10 patients, 7 (70%) with follow-upwithin this group developed repeated recurrences, and twoof these patients died of their tumor. Histomorphologically,these tumors were of higher cellularity, with more plump orrounded primitive-looking cell morphological characteris-tics, lacking any fibrosis/hyalinization, and displayed char-acteristic staghorn-like branching vessels. Collectively,these data suggest that tumors of the first category, with theNAB2ex4-STAT6ex2/3 fusion variants, represent theclassic paucicellular fibrous variant of pleuropulmonarySFT,4e8 whereas tumors of the latter category, with theNAB2ex6-STAT6ex16/17 fusion variants, correspond tothe original descriptions of deep-seated HPC.1e3,13

The two genes, NAB2 and STAT6, are located close to eachother in opposing directions at the chromosomal region 12q13,with a partial overlap of 58 bp of their respective 30 ends.29

Although STAT6 is a transcription factor, NAB2 functionsas a transcriptional repressor that has noDNAbinding capacityitself. Interestingly, both proteins are involved in the regula-tion of inflammation, vessel formation, fibroblast activation,and collagen production.30,31 Moreover, these two proteinshave opposing regulatory effects on the transcription factorearly growth response 1 (EGR1),32,33 which is an importantregulator of wound healing and fibrosis.31

NAB2 has two NAB-conserved domains, NCD1 andNCD2, encoded by exons 1 to 3, and a C-terminal repressordomain encoded by exons 4 to 7.32,33 Although NCD1mediates multimerization of two to four NAB molecules

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NAB2-STAT6 Fusion Variants in SFTs/HPCs

and is required for interaction with EGR1, NCD2 and the C-terminal domain are independent and only partially redun-dant transcriptional repressor domains. The C-terminaldomain interacts with the chromodomain helicase DNA-binding protein 4 (CHD4) subunit of the nucleosome andremodeling (NuRD) complex, and was thus designated theCHD4-interacting domain (CID).34 The NuRD complexlinks multiple regulatory transcriptional processes andfunctions as a modulator of transcription.35 Although EGR1activates fibroblasts through NOX4 and induces the tran-scription of different profibrotic collagens, NAB2 is aphysiological endogenous negative regulator of fibrosis.Together, NAB2 and EGR1 orchestrate fibrotic response inwound healing.31

It is intriguing that in the tumors with the NAB2ex4-STAT6ex2/3 fusion variant, the predicted chimeric fusionprotein lacks the CID repressor domain of the NAB2 part,which implies a probably increased activity of EGR1, cor-responding to the prominent diffuse fibrosis observed inthese tumors consistently displaying the classic fibrous SFTphenotype. In contrast, in the tumors with the secondmost common fusion variant, NAB2ex6-STAT6ex16/17,comprising cellular SFTs and deep-seated HPCs, the CIDrepressor domain of the NAB2 part of the chimeric fusionprotein seems to be functionally intact, probably explainingthe much lesser extent of fibrosis in these tumors. Althoughthe STAT6 part of the predicted chimeric protein wascomplete in the first fusion variant, the second NAB2ex6-STAT6ex16/17 fusion variant encoded for a chimeric pro-tein, which lacks a large amino terminal portion of theSTAT6 protein, including the coiled-coil domain, the DNA-binding domain, and the Src-homology 2 domain. TheSrc-homology 2 domain is required for receptor binding (eg,IL-4 and IL-13) and consecutive activation of the STAT6protein, with subsequent formation of dimers that trans-locate to the nucleus.30 Only if STAT6 is present as a dimer,the DNA-binding domain allows DNA binding and fullaccomplishment of its function as a transcription factor.30

Thus, these two domains are functionally aligned and linkSTAT6 to its predominant function within the inflammatoryresponse mediated by IL-4 and IL-13.30 This function seemsto be completely abolished in the tumors with theNAB2ex6-STAT6ex16/17 fusion variant, which lacks bothdomains. In contrast, the remaining C-terminal part of theSTAT6 protein encoded by this fusion variant correlates tothe so-called transactivation domain (TAD), which isnecessary for transcriptional activation.36,37 In a directcomparison, the activity of the TAD of STAT6 was10-fold stronger compared with the corresponding TAD ofSTAT5.37 Interestingly, STAT6 TAD can be linked to RNApolymerase II and the basal transcription machinery throughthe p100 transcriptional coactivator.38 In the predictedchimeric protein encoded by the NAB2ex6-STAT6ex16/17fusion variant observed in the more proliferative cellularSFTs and HPCs, the STAT6 TAD is separated from itsphysiological role in inflammatory signaling, but may be

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located in close proximity to EGR1 target genes and theNuRD complex. Although we cannot yet show a functionaldifference of the predicted chimeric NAB2-STAT6 fusionproteins in vitro, it is likely that the fusion protein generatedby this variant has different functional effects on collagenproduction, vessel formation, and proliferation comparedwith the fusion protein encoded by the different fusionvariant observed in the fibrous SFTs.

In contrast to the current study, there were no differencesreported in the clinicopathological features found amongtumors with the different NAB2-STAT6 fusion variants inthree studies published.22e24 This might be partially relatedto the different frequencies observed for distinct fusionvariants within these studies, and also to probable hetero-geneity of the tumor cohorts among the different in-stitutions. In the study by Robinson et al,22 38 of 51 casesharbored NAB2-STAT6 fusion variants, consistent with ourNAB2ex6-STAT6ex16/17 category. Interestingly, 20 ofthese cases were located in the soft tissues of the extrem-ities, trunk, or retroperitoneum/pelvis, compared with onlythree cases with a pleuropulmonary location. In comparison,the NAB2ex4-STAT6ex2 fusion variant was present in onlythree tumors in that study, and two of these cases were ofpleuropulmonary origin. In another study published simul-taneously by Chmielecki et al,23 the most frequentlyobserved fusion found in 19 of 48 cases was the NAB2ex4-STAT6ex2 variant, and 13 of these tumors originated fromthe lungs or pleura.23 In contrast to the first study, theseauthors did not observe any fusion variants consistent withthe NAB2ex6-STAT6ex16/17 category. However, most oftheir cases (23 of 48) displayed no fusion variant, and 16 ofthese tumors were located in the deep soft tissues. Some ofthese cases may have harbored a fusion variant from theNAB2ex6-STAT6ex16/17 category, which was due totechnical issues. Regarding the third study by Mohajeriet al,24 the NAB2ex4-STAT6ex2 fusion variant was foundin 17 of 44 cases, and seven of these tumors were in apleuropulmonary location. These authors detected theNAB2ex6-STAT6ex16/17 fusion variants in 10 cases,including eight located at the extremities, trunk, or pelvis.Taken together, the published cases clearly corroborate ourcurrent findings and suggest further implications of molec-ular analysis of specific NAB2-STAT6 fusion variants intumors of the SFT/HPC spectrum for diagnostic and prog-nostic classifications. Notably, our introduced multiplex RT-PCR approach works on formalin-fixed samples and can beeasily integrated into the diagnostic workup of molecularpathology laboratories with a focus on soft tissue pathology.

Acknowledgments

We thank Simone Hebele for excellent technical assistance,Ute Ernst and Sabine Schmidt (German Cancer ResearchCenter Genomics and Proteomics Core Facility) for excel-lent support, and Christopher Previti (Genomics and

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Barthelmeß et al

Proteomics Core Facility, German Cancer Research Center,Heidelberg) for excellent help with OnTarget data analysis.

All authors have read and approved the final version ofthe manuscript.

References

1. Stout AP, Murray MR: Hemangiopericytoma: a vascular tumorfeaturing Zimmermann’s pericytes. Ann Surg 1942, 116:26e33

2. Stout AP: Hemangiopericytoma: a study of 25 cases. Cancer 1949, 2:1027e1054

3. Enzinger FM, Smith BH: Hemangiopericytoma: an analysis of 106cases. Hum Pathol 1976, 7:61e82

4. Stout AP, Murray MR: Localized pleural mesothelioma. Arch Pathol1942, 34:951e964

5. Klemperer P, Rabin CB: Primary neoplasms of the pleura: a report offive cases. Arch Pathol 1931, 11:385e412

6. Stout AP, Himadi GM: Solitary (localized) mesothelioma of the pleura.Ann Surg 1951, 133:50e64

7. England DM, Hochholzer L, McCarthy MJ: Localized benign andmalignant fibrous tumors of the pleura: a clinicopathologic review of223 cases. Am J Surg Pathol 1989, 13:640e658

8. Moran CA, Suster S, Koss MN: The spectrum of histologic growthpatterns in benign and malignant fibrous tumors of the pleura. SeminDiagn Pathol 1992, 9:169e180

9. Goodlad JR, Fletcher CD: Solitary fibrous tumour arising at unusualsites: analysis of a series. Histopathology 1991, 19:515e522

10. Suster S, Nascimento AG, Miettinen M, Sickel JZ, Moran CA: Solitaryfibrous tumors of soft tissue: a clinicopathologic and immunohisto-chemical study of 12 cases. Am J Surg Pathol 1995, 19:1257e1266

11. Chilosi M, Facchettti F, Dei Tos AP, Lestani M, Morassi ML,Martignoni G, Sorio C, Benedetti A, Morelli L, Doglioni C,Barberis M, Menestrina F, Viale G: bcl-2 Expression in pleural andextrapleural solitary fibrous tumours. J Pathol 1997, 181:362e367

12. Vallat-Decouvelaere AV, Dry SM, Fletcher CD: Atypical and malig-nant solitary fibrous tumors in extrathoracic locations: evidence of theircomparability to intra-thoracic tumors. Am J Surg Pathol 1998, 22:1501e1511

13. Tsuneyoshi M, Daimaru Y, Enjoji M: Malignant hemangiopericytomaand other sarcomas with hemangiopericytoma-like pattern. Pathol ResPract 1984, 178:446e453

14. Nappi O, Ritter JH, Pettinato G, Wick MR: Hemangiopericytoma:histopathological pattern or clinicopathologic entity? Semin DiagnPathol 1995, 12:221e232

15. Fletcher CDM: Haemangiopericytoma e a dying breed? reappraisal ofan entity and its variants: a hypothesis. CurrDiagn Pathol 1994, 1:19e23

16. Gengler C, Guillou L: Solitary fibrous tumour and haemangioper-icytoma: evolution of a concept. Histopathology 2006, 48:63e74

17. Park MS, Araujo DM: New insights into the hemangiopericytoma/solitaryfibrous tumor spectrum of tumors. Curr Opin Oncol 2009, 21:327e331

18. Fletcher CDM, Bridge JA, Lee JC: Extrapleural solitary fibrous tumour.Edited by Fletcher CDM, Bridge JA, Hogendoorn PCW, Mertens F.World Health Organisation Classification of Tumours of Soft Tissueand Bone. ed 4. Lyon, IARC Press, 2013, pp 80e82

19. Schirosi L, Lantuejoul S, Cavazza A, Murer B, Yves Brichon P,Migaldi M, Sartori G, Sgambato A, Rossi G: Pleuro-pulmonary soli-tary fibrous tumors: a clinicopathologic, immunohistochemical, andmolecular study of 88 cases confirming the prognostic value of dePerrot staging system and p53 expression, and evaluating the role of c-kit, BRAF, PDGFRs (alpha/beta), c-met, and EGFR. Am J Surg Pathol2008, 32:1627e1642

20. Cranshaw IM, Gikas PD, Fisher C, Thway K, Thomas JM, Hayes AJ:Clinical outcomes of extra-thoracic solitary fibrous tumours. Eur J SurgOncol 2009, 35:994e998

1218

21. Rao N, Colby TV, Falconieri G, Cohen H, Moran CA, Suster S:Intrapulmonary solitary fibrous tumors: clinicopathologic and immuno-histochemical study of 24 cases. Am J Surg Pathol 2013, 37:155e166

22. Robinson DR, Wu YM, Kalyana-Sundaram S, Cao X, Lonigro RJ,Sung YS, Chen CL, Zhang L, Wang R, Su F, Iyer MK,Roychowdhury S, Siddiqui J, Pienta KJ, Kunju LP, Talpaz M,Mosquera JM, Singer S, Schuetze SM, Antonescu CR, Chinnaiyan AM:Identification of recurrent NAB2-STAT6 gene fusions in solitary fibroustumor by integrative sequencing. Nat Genet 2013, 45:180e185

23. Chmielecki J, Crago AM, Rosenberg M, O’Connor R, Walker SR,Ambrogio L, Auclair D, McKenna A, Heinrich MC, Frank DA,Meyerson M: Whole-exome sequencing identifies a recurrent NAB2-STAT6 fusion in solitary fibrous tumors. Nat Genet 2013, 45:131e132

24. Mohajeri A, Tayebwa J, Collin A, Nilsson J, Magnusson L, vonSteyern FV, Brosjö O, Domanski HA, Larsson O, Sciot R, Debiec-Rychter M, Hornick JL, Mandahl N, Nord KH, Mertens F: Compre-hensive genetic analysis identifies a pathognomonic NAB2/STAT6fusion gene, nonrandom secondary genomic imbalances, and a char-acteristic gene expression profile in solitary fibrous tumor. GenesChromosomes Cancer 2013, 52:873e886

25. Doyle LA, Vivero M, Fletcher CD, Mertens F, Hornick JL: Nuclearexpression of STAT6 distinguishes solitary fibrous tumor from histo-logic mimics. Mod Pathol 2013, 27:390e395

26. Hotz-Wagenblatt A, Kats I, Scharfenberger-Schmeer M, Haldeman B,Glatting KH: OnTarget: a tool for analysing enrichment data derived fromnext generation sequencing. Presented at the 11th International Conferenceon Intelligent Systems Design and Applications (ISDA), 2011, 873e876

27. Li H, Durbin R: Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinf 2009, 25:1754e1760

28. Wang J, Mullighan CG, Easton J, Roberts S, Heatley SL, Ma J,Rusch MC, Chen K, Harris CC, Ding L, Holmfeldt L, Payne-Turner D,Fan X, Wei L, Zhao D, Obenauer JC, Naeve C, Mardis ER, Wilson RK,Downing JR,Zhang J:CRESTmaps somatic structural variation in cancergenomes with base-pair resolution. Nat Methods 2011, 8:652e654

29. Svaren J, Apel ED, Simburger KS, Jenkins NA, Gilbert DJ,Copeland NA, Milbrandt J: The Nab2 and Stat6 genes share a commontranscription termination region. Genomics 1997, 41:33e39

30. Hebenstreit D, Wirnsberger G, Horejs-Hoeck J, Duschl A: Signalingmechanisms, interaction partners, and target genes of STAT6. Cyto-kine Growth Factor Rev 2006, 17:173e188

31. Bhattacharyya S, Fang F, Tourtellotte W, Varga J: Egr-1: newconductor for the tissue repair orchestra directs harmony (regeneration)or cacophony (fibrosis). J Pathol 2013, 229:286e297

32. Svaren J, Sevetson BR, Apel ED, Zimonjic DB, Popescu NC,Milbrandt J: NAB2, a corepressor of NGFI-A (Egr-1) and Krox20, isinduced by proliferative and differentiative stimuli. Mol Cell Biol1996, 16:3545e3553

33. Ingram JL, Antao-Menezes A, Mangum JB, Lyght O, Lee PJ, Elias JA,Bonner JC: Opposing actions of Stat1 and Stat6 on IL-13-induced up-regulation of early growth response-1 and platelet-derived growth fac-tor ligands in pulmonary fibroblasts. J Immunol 2006, 177:4141e4148

34. Srinivasan R, Mager GM, Ward RM, Mayer J, Svaren J: NAB2 re-presses transcription by interacting with the CHD4 subunit of thenucleosome remodeling and deacetylase (NuRD) complex. J BiolChem 2006, 281:15129e15137

35. Denslow SA, Wade PA: The human Mi-2/NuRD complex and generegulation. Oncogene 2007, 26:5433e5438

36. Lu B, Reichel M, Fisher DA, Smith JF, Rothman P: Identification of aSTAT6 domain required for IL-4-induced activation of transcription.J Immunol 1997, 159:1255e1264

37. Moriggl R, Berchtold S, Friedrich K, Standke GJ, Kammer W,Heim M, Wissler M, Stöcklin E, Gouilleux F, Groner B: Comparisonof the transactivation domains of Stat5 and Stat6 in lymphoid cells andmammary epithelial cells. Mol Cell Biol 1997, 17:3663e3678

38. Yang J, Aittomäki S, Pesu M, Carter K, Saarinen J, Kalkkinen N, Kieff E,Silvennoinen O: Identification of p100 as a coactivator for STAT6 thatbridges STAT6 with RNA polymerase II. EMBO J 2002, 21:4950e4958

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