Activation of the GLI Oncogene through Fusion with the β-Actin Gene (ACTB) in a Group of Distinctive Pericytic Neoplasms
2004; Elsevier BV; Volume: 164; Issue: 5 Linguagem: Inglês
10.1016/s0002-9440(10)63723-6
ISSN1525-2191
AutoresAnna Dahlén, Christopher D.�M. Fletcher, Fredrik Mertens, Jonathan A. Fletcher, Antonio R. Pérez‐Atayde, John Hicks, Maria Dębiec‐Rychter, Raf Sciot, Johan Wejde, Rikard Wedin, Nils Mandahl, Ioannis Panagopoulos,
Tópico(s)Oral and gingival health research
ResumoActivation of the GLI oncogene is an important step in the sonic hedgehog signaling pathway, and leads to, eg, tissue-specific cell proliferation during embryogenesis. GLI activity in adult tissues is restricted, but has been identified in various neoplasms, as a result of mutations in the PTCH (patched) or SMOH (smoothened) genes, encoding components of the sonic hedgehog pathway, or by amplification of GLI. Herein, we present a new mechanism of GLI activation through fusion with the β-actin gene (ACTB) in five histologically distinctive soft tissue tumors showing a t(7;12)(p21-22;q13-15) and a pericytic phenotype. Each was composed of a perivascular proliferation of monomorphic short spindle cells that stained positively for smooth muscle actin and laminin and that showed pericytic features by electron microscopy. To date, with a median follow-up of 24 months, none has behaved in an aggressive manner. Molecular genetic analysis showed that the translocation in all cases resulted in a fusion transcript including the 5′-part of ACTB and the 3′-part of GLI. The DNA-binding zinc finger domains of GLI were retained in the fusion transcripts and it is likely that the replacement of the promoter region of GLI with that of the ubiquitously expressed ACTB gene leads to deregulation of GLI expression and its downstream target genes. Activation of the GLI oncogene is an important step in the sonic hedgehog signaling pathway, and leads to, eg, tissue-specific cell proliferation during embryogenesis. GLI activity in adult tissues is restricted, but has been identified in various neoplasms, as a result of mutations in the PTCH (patched) or SMOH (smoothened) genes, encoding components of the sonic hedgehog pathway, or by amplification of GLI. Herein, we present a new mechanism of GLI activation through fusion with the β-actin gene (ACTB) in five histologically distinctive soft tissue tumors showing a t(7;12)(p21-22;q13-15) and a pericytic phenotype. Each was composed of a perivascular proliferation of monomorphic short spindle cells that stained positively for smooth muscle actin and laminin and that showed pericytic features by electron microscopy. To date, with a median follow-up of 24 months, none has behaved in an aggressive manner. Molecular genetic analysis showed that the translocation in all cases resulted in a fusion transcript including the 5′-part of ACTB and the 3′-part of GLI. The DNA-binding zinc finger domains of GLI were retained in the fusion transcripts and it is likely that the replacement of the promoter region of GLI with that of the ubiquitously expressed ACTB gene leads to deregulation of GLI expression and its downstream target genes. Soft tissue sarcomas constitute a clinically and histologically heterogeneous group of malignant neoplasms, which sometimes are difficult to distinguish from each other or from their benign counterparts.1Fletcher CDM Rydholm A Singer S Sundaram M Coindre JM Soft tissue tumours: epidemiology, clinical features, histopathological typing and grading.in: Fletcher CDM Unni KK Mertens F World Health Organization Classification of Tumours. Pathology and Genetics of Tumours of Soft Tissue and Bone. IARC Press, Lyon2002: 12-18Google Scholar The cytogenetic picture is equally variable, but the identification of recurrent tumor-specific translocations and the gene rearrangements resulting from these translocations has added significantly to diagnostic precision. Comparable reciprocal translocations, albeit apparently often less specific, have also been identified in some benign mesenchymal neoplasms, such as lipomas and leiomyomas.2Åman P Fusion genes in solid tumors.Semin Cancer Biol. 1999; 9: 303-318Crossref PubMed Scopus (83) Google Scholar, 3Ladanyi M Bridge JA Contribution of molecular genetic data to the classification of sarcomas.Hum Pathol. 2000; 31: 532-538Abstract Full Text PDF PubMed Scopus (114) Google Scholar Furthermore, the molecular genetic characterization of these recurrent cytogenetic rearrangements has also enabled a better understanding of the genetic mechanisms underlying the development of sarcomas.2Åman P Fusion genes in solid tumors.Semin Cancer Biol. 1999; 9: 303-318Crossref PubMed Scopus (83) Google Scholar, 3Ladanyi M Bridge JA Contribution of molecular genetic data to the classification of sarcomas.Hum Pathol. 2000; 31: 532-538Abstract Full Text PDF PubMed Scopus (114) Google Scholar A characteristic feature of sarcoma-associated translocations is that they result in fusion genes by the joining together of the 5′-part of one gene with the 3′-part of another gene. Typically, at least one of the two genes involved encodes a transcription factor, but also translocations leading to constitutive activation of growth factors or growth factor receptors have been identified.2Åman P Fusion genes in solid tumors.Semin Cancer Biol. 1999; 9: 303-318Crossref PubMed Scopus (83) Google Scholar In the present study, we report the finding of a novel fusion gene in five spindle cell tumors with distinctive pericytic features. Lesions of this type seem previously to be unrecognized as a discrete entity. In all of them, cytogenetic analysis revealed a t(7;12)(p21-22;q13-15), that at molecular genetic investigation was found to result in fusion of the ACTB (β-actin) and GLI (glioma-associated oncogene homologue 1) genes, neither of which has previously been implicated in fusion genes in sarcomas or other neoplasms. The five tumors included in this study were all selected initially for further molecular genetic and histopathological analyses on the basis of their cytogenetic features, ie, the presence of a translocation t(7;12)(p21-22;q13-15). One case was the subject of a previous case report.4Perez-Atayde AR Kozakewich HWP McGill T Fletcher JA Hemangiopericytoma of the tongue in a 12-year-old child: ultrastructural and cytogenetic observations.Hum Pathol. 1994; 25: 425-429Abstract Full Text PDF PubMed Scopus (18) Google Scholar Four μm hematoxylin and eosin-stained sections of all cases were examined and immunohistochemical studies were performed in all cases, using the Envision Plus detection system (DAKO, Carpinteria, CA). The antibodies, clones, dilutions, pretreatment conditions, and sources are listed in Table 1. Appropriate positive and negative controls were used throughout. Electron microscopy was performed on three tumors (cases 2, 3, and 5) in which fresh material had been suitably fixed in glutaraldehyde.Table 1Panel of Antibodies for Immunohistochemical AnalysisAntigenCloneDilutionAntigen retrievalSourceSMA1A41:20000NoneSigma, St. Louis, MOMuscle actinHHF351:500NoneDAKO, Carpinteria, CACK (AE1/AE3)AE1/AE31:20010 minute proteaseDAKOCK 8/18CAM 5.21:10010 minute proteaseBecton Dickinson, San Jose, CADesminD331:500NoneDAKOCD1056C61:1030 minute microwaveNovocastra, Newcastle, UKLamininPolyclonal1:150030 minute proteaseDAKOCOLL IVCIV 221:20020 minute proteaseDAKOS-100Polyclonal1:3000NoneDAKOCALDESh-CD1:30030 minute microwaveDAKOMelanoma (HMB45)HMB451:400NoneDAKOEMAE291:200NoneDAKOCD34Qbend 101:400NoneDAKOCD31JC/70A1:4020 minute proteaseDAKOD6D2-401:100NoneSignet Lab, Dedham, MAKITPolyclonal (A4502)1:250NoneDAKOPRPgR 6361:20030 minute microwaveDAKOAbbreviations: SMA, smooth muscle actin; CK, cytokeratin; COLL IV, collagen type IV; CALDES, caldesmon; EMA, epithelial membrane antigen; PR, progesterone receptor. Open table in a new tab Abbreviations: SMA, smooth muscle actin; CK, cytokeratin; COLL IV, collagen type IV; CALDES, caldesmon; EMA, epithelial membrane antigen; PR, progesterone receptor. Cell culturing, harvesting, and G-banding were performed as described,4Perez-Atayde AR Kozakewich HWP McGill T Fletcher JA Hemangiopericytoma of the tongue in a 12-year-old child: ultrastructural and cytogenetic observations.Hum Pathol. 1994; 25: 425-429Abstract Full Text PDF PubMed Scopus (18) Google Scholar, 5Mandahl N Methods in solid tumor cytogenetics.in: Rooney DE Human Cytogenetics: Malignancy and Acquired Abnormalities ed 3. Oxford University Press, New York2001: 165-203Google Scholar and the karyotypes were written according to the recommendations of the International System for Human Cytogenetic Nomenclature (ISCN).6Mitelman F ISCN: An International System for Human Cytogenetic Nomenclature. Karger, Basel1995Google Scholar The cytogenetic features of case 5 have been reported previously.4Perez-Atayde AR Kozakewich HWP McGill T Fletcher JA Hemangiopericytoma of the tongue in a 12-year-old child: ultrastructural and cytogenetic observations.Hum Pathol. 1994; 25: 425-429Abstract Full Text PDF PubMed Scopus (18) Google Scholar Material for metaphase and interphase FISH analysis was available for cases 1 and 5, respectively. To identify the chromosomal breakpoints in case 1, 20 bacterial artificial chromosome (BAC) probes spanning 7p21-22 or 12q13 were selected from the NCBI Map Viewer (http://www.ncbi.nlm.nih.gov/mapview), the Ensembl Genome Browser (http://www.ensembl.org) and the UCSC Human Genome Browser (http://www.genome.ucsc.edu). It should be noted that based on our FISH findings and polymerase chain reaction (PCR) results showing that it contains the ACTB locus, BAC RP11-1275H24 was substantially larger than the ∼85 kb reported at the NCBI Nucleotide Browser. Whole chromosome painting probes (Vysis, Downers Grove, IL) were also used to identify material from chromosomes 7 and 12. The BAC probes were labeled with Cy3-dCTP (Amersham, Buckinghamshire, UK), FluorX-dCTP (Amersham) or biotin-16-dUTP (Roche, Mannheim, Germany), using the Megaprime DNA labeling system (Amersham). The FISH treatments and analyses were performed as described.7Dahlén A Debiec-Rychter M Pedeutour F Domanski HA Höglund M Bauer HCF Rydholm A Sciot R Mandahl N Mertens F Clustering of deletions on chromosome 13 in benign and low-malignant lipomatous tumors.Int J Cancer. 2003; 103: 616-623Crossref PubMed Scopus (86) Google Scholar Total RNA was extracted from frozen tissue (cases 1 to 4) or cell cultures (case 5), using the Trizol-reagent according to the manufacturer's recommendations (Gibco BRL, Täby, Sweden). For the cDNA synthesis, 5 μg of total RNA were reverse-transcribed in a 20-μl reaction, containing 50 mmol/L Tris-HCl, pH 8.3 (at 25°C), 75 mmol/L KCl, 3 mmol/L MgCl2, 10 mmol/L dithiothreitol, 1 mmol/L of each dNTP, 0.5 pmol/L random hexamers, 28 U RNase inhibitor (RNA guard, Amersham), and 400 U M-MLV reverse transcriptase (Invitrogen, Stockholm, Sweden). The reaction was incubated for 1 hour at 37°C, followed by 5 minutes at 65°C. As an internal quality control, 1 μl of cDNA was amplified by PCR using ABL1-specific primers. Reverse transcriptase (RT) PCR was used for the detection of the ACTB-GLI and the reciprocal GLI-ACTB fusion transcripts. The nucleotide sequences for all primers used for PCR amplification and sequence analyses are presented in Table 2. One μl of cDNA was used as template for PCR, and all PCRs described below were performed under the same conditions, ie, the 50-μl reaction contained 20 mmol/L Tris-HCl, 50 mmol/L KCl, 1.25 mmol/L MgCl2, 0.8 mmol/L dNTPs, 0.5 μmol/L of each primer, and 1 U Platinum TaqDNA Polymerase (Invitrogen). After an initial denaturation for 5 minutes at 95°C, 30 cycles of 1 minute at 94°C, 1 minute at 60°C, and 1 minute at 72°C were run using a PCT-200 DNA Engine (MJ Research, Waltham, MA), followed by a final extension for 10 minutes at 74°C. For the nested RT-PCR, 1 μl of the first PCR products was used as template.Table 2Primers for RT-PCR and Direct SequencingPrimer*F, forward primer; R, reverse primer.SequencePositionAccession no.GLI336F5′-ACCTCTGTCGGATGCCAGCCTGG336–358NM_005269GLI412F5′-TCGCGATGCACATCTCCAGGAGG412–434NM_005269GLI520F5′-TCCTTTGGGGTCCAGCCTTGTGG520–542NM_005269GLI577R5′-AAGGGTCCCCGGGACTGAGGATG599–577NM_005269GLI628R5′-CGGCACTTGCCAACCAGCATGTC650–628NM_005269GLI720R5′-AGGTCCTCCCGCCCATCCAGC740–720NM_005269GLI868R5′-GTGGCACACGAACTCCTTCCGCTC891–868NM_005269GLI938R5′-TCTGCGCATGTGAACCACCAGCA960–938NM_005269GLI1246R5′-GCCGTTTGGTCACATGGGCGTC1267–1246NM_005269GLI1389R5′-CCCCAGGGCTTGGCTGTGGC1408–1389NM_005269GLI1477R5′-TGCCCCCTGCATTGCCAGTCAT1498–1477NM_005269ACT18F5′-CACAGAGCCTCGCCTTTGCCGA18–39NM_001101ACT61F5′-CCGCCAGCTCACCATGGATGATG61–83NM_001101ACT80F5′-GATGATATCGCCGCGCTCGTCG80–101NM_001101ACT106F5′-CAACGGCTCCGGCATGTGCAA106–126NM_001101ACT288F5′-AGCACGGCATCGTCACCAACTGG288–310NM_001101ACT351F5′-AGCTGCGTGTGGCTCCCGAGG351–371NM_001101ACT520R5′-CACCGGAGTCCATCACGATGCCA542–520NM_001101ACT594R5′-AGCCAGGTCCAGACGCAGGATGG616–594NM_001101* F, forward primer; R, reverse primer. Open table in a new tab A total of 8 ACTB-specific, and 11 GLI-specific primers were selected, to allow a precise mapping of the fusion transcripts (Figure 1a and Table 2). In cases 1 to 4, the ACTB-GLI fusion transcripts were amplified with primer pairs ACT61F-GLI868R, ACT18F-GLI1246R, ACT61F-GLI1477R, and ACT61F-GLI938R, respectively. A reciprocal GLI-ACTB fusion was detected by nested PCR in case 1, using the primer pair GLI412F-ACT594R, in the first round of PCR, and GLI520F-ACT520R for the second. In case 5, nested PCR was required for the detection of all fusion sequences. The ACTB-GLI and GLI-ACTB fusion transcripts were amplified in a first round of RT-PCR with primer pairs ACT80F-GLI1246 and GLI336-ACT594R, respectively. For nested PCR, the corresponding primer pairs ACT106F-GLI868R and GLI412F-ACT520R were used. All PCR products (15 μl) were analyzed on 1.5% agarose gels stained with ethidium bromide. For sequence analysis, the band corresponding to the expected PCR product was excised, purified using the QIAquick gel extraction kit (Qiagen, Hilden, Germany), and sequenced in a 20-μl reaction with various primers (Table 2), using the dideoxy procedure with an ABI Prism BigDye terminator cycle sequencing ready reaction kit on the Applied Biosystems (Foster City, CA) model 310 DNA sequencing system. The BLAST software (http://www.ncbi.nlm.nih.gov/blast) was used for the analysis of ACTB [accession numbers: NM_001101 (mRNA), M10277 (complete cds)], and GLI [accession number: NM_005269 (mRNA)] sequence data. Clinical data are summarized in Table 3. All tumors were primary lesions, three being located in the tongue and one each in the stomach and calf. Three patients were female and two were male, ranging in age at presentation from 11 to 65 years (median, 27 years). Tumor size ranged from 0.8 to 5.5 cm (median, 2.4 cm). Two of the tongue lesions (cases 2 and 3) were treated with preoperative chemotherapy with no clear evidence of response. All were locally resected, with tumor-positive resection margins in cases 3 and 5. Follow-up so far, ranging from 22 to 120 months (median, 24 months), has revealed no evidence of recurrence or metastasis.Table 3Clinical Data and Cytogenetic FindingsCaseSex/ageLocalizationSize*Diameter in cm.Follow-up†Follow-up in months; NED, no evidence of disease.Karyotype1M/61Calf2NED 2445,XY,t(7;12)(p22;q13),inv(10)(p11q21)c,der(15;16)(q10;p10)[25]2F/27Tongue0.8NED 6046,XX,t(7;12)(p22;q13)[20]3M/11Tongue5NED 2245,XY,der(1)t(1;?7)(p36;p?22),add(2)(p25),add(5)(p15),add(6)(p?21.3),der(7)t(7;12)(p11.2;p11.2),der(7)t(7;12)(p21;q?15),−12[19]4F/65Stomach5.5NED 2446,XX,t(7;12)(?p22;?q15)[17]5F/12Tongue2.4NED 12046,XX,t(1;13)(p22;q21),t(7;12)(p22;q13)[20]* Diameter in cm.† Follow-up in months; NED, no evidence of disease. Open table in a new tab Histologically each tumor showed remarkably similar morphology, seemingly somewhat distinct from any currently well-defined entity. The tumors had a multilobulated, infiltrative growth pattern and were each composed of uniform spindle-shaped cells with small amounts of pale eosinophilic cytoplasm and ovoid-to-tapered nuclei with vesicular chromatin and often a single small nucleolus (Figure 2, A and B). These spindle cells were consistently arranged around numerous small, thin-walled arborizing vessels, which were readily highlighted by immunopositivity for CD34 (Figure 2C). There was no significant cytological atypia or pleomorphism. Mitoses in all cases numbered less than 1 per 10 high-power fields. Three of the cases showed a focally myxoid stroma. In two cases there was prominent and multifocal subendothelial protrusion of tumor cells into vascular lumina (Figure 2D), in a manner reminiscent of myopericytic neoplasms. Two of the tongue lesions showed focal surface ulceration and the gastric lesion showed areas of stromal hemorrhage and hyalinization but no true tumor necrosis was seen in any case. Immunohistochemical analysis (Table 4) revealed that tumor cells in all cases showed focal to extensive positivity for smooth muscle actin (Figure 2E), as well as multifocal pericellular positivity for laminin (Figure 2F), consistent with the presence of an external lamina. Three cases each also showed positivity for collagen type IV and for CD10. Staining for keratins, desmin, S-100 protein, and CD34 was consistently negative. One tumor each showed cytoplasmic positivity for D2-40 (case 4) and focal epithelial membrane antigen positivity (case 1) of uncertain significance.Table 4Immunohistochemical Findings*Additional results not listed in the table above include negativity for progesterone receptor (cases 3 to 5), for CD31 (cases 1, 4), and for KIT (cases 4, 5).CaseSMAHHF-35CKDesminCD10LamininCOLL IVS100CALDESHMB45EMACD34D2-401+NP−−NP+ (focal)NPNP−NP+ (focal)NPNP2+NP−−NP+ (focal)+ (focal)−NP−−−−3+ (focal)NP−−++ (focal)+ (focal)−NP−−−−4+ (focal)−−−+++−−−−−+5+ (focal)−−−++ (focal)−−NP−NP−−Abbreviations: NP, not performed; SMA, smooth muscle actin; CK, keratin; COLL IV, collagen type IV; CALDES, caldesmon; EMA, epithelial membrane antigen.* Additional results not listed in the table above include negativity for progesterone receptor (cases 3 to 5), for CD31 (cases 1, 4), and for KIT (cases 4, 5). Open table in a new tab Abbreviations: NP, not performed; SMA, smooth muscle actin; CK, keratin; COLL IV, collagen type IV; CALDES, caldesmon; EMA, epithelial membrane antigen. Three tumors examined ultrastructurally (cases 2, 3, and 5) each showed similar features (Figure 3). The tumor cells were closely arranged around capillary size vessels and exhibited short segments of external lamina, occasional intermediate-type junctions, cytoplasmic pools of free glycogen, subplasmalemmal thickenings, and bundles of thin filaments with focal densities at the cytoplasmic periphery. These findings indicate partial smooth muscle differentiation, as seen in modified smooth muscle cells including pericytes. All cases were cytogenetically analyzed after short-term culturing, and a t(7;12)(p21-22;q13-q15) was found in all cases, being the sole abnormality in two of them (Table 3). In case 1, metaphase FISH analysis with the 7p-specific probes RP11-1275H24 and RP11-93G19, and the 12q-specific probes RP11-181L23 and RP11-772E1 resulted in split signals in the t(7;12)-carrying cells (Figure 4a). On the corresponding normal homologues, intact signals were seen. In case 5, only interphase nuclei from cell cultures in passage 6 were available for FISH analysis. Hybridization with probes RP11-1275H24 and RP11-772E1 revealed split signals in ∼1% (8 and 11 nuclei, respectively) of the nuclei (Figure 4a). Based on these FISH results, ACTB and GLI were considered potential target genes in 7p22 and 12q13, respectively (Figure 4, b and c). RT-PCR with different combinations of ACTB forward and GLI reverse primers amplified cDNA fragments, strongly suggesting the presence of an ACTB-GLI fusion gene (Figure 5). Amplified products were analyzed by direct sequencing for an exact characterization of the corresponding fusion points. The molecular genetic findings are summarized in Figure 1b. Analyses regarding expression of the FLJ11467 gene or a potential FLJ11467-GLI fusion gene were consistently negative (data not shown). In case 1, the primer pair ACT61F-GLI868R amplified a 270-bp product in which ACTB exon 2 (nucleotide 196, NM_001101) was fused to nucleotide 758 within exon 7 of GLI (NM_005269). Insertion of a guanine at the breakpoint retained the open reading frame (Figure 6). In case 2, the primer pair ACT18F-GLI1246R amplified a 754-bp product corresponding to an ACTB-GLI fusion transcript containing ACTB intron 1 sequences. In this transcript, ACTB exon 1 and nucleotides 313 to 319 of ACTB intron 1 (M10277) were joined to nucleotides 1017 to 1043 of ACTB intron 1 (M10277), which in turn was fused to GLI exon 6 (nucleotide 613, NM_005269) (Figure 6). In case 3, the ACT61F-GLI1477 primer combination amplified an 1172-bp product corresponding to a fusion of ACTB exon 3 (nucleotide 436, NM_001101) to GLI exon 7 (nucleotide 703, NM_005269) (Figure 6). In case 4, a 484-bp product was amplified with primer pair ACT61F-GLI938R, corresponding to a fusion of ACTB exon 2 (nucleotide 196, NM_001101) to GLI exon 6 (nucleotide 613, NM_005269) (Figure 6). Nested PCR was required for the amplification of ACTB-GLI fusion transcripts in case 5. Amplification with primer pair ACT80F-GLI1246R yielded no visible band at gel electrophoresis. Nested PCR with primer pair ACT106F-GLI868R resulted in the amplification of several bands (Figure 5), two of which were analyzed by direct sequencing. The first band (610 bp) corresponded to an in-frame fusion of ACTB exon 3 (nucleotide 436, NM_001101) to GLI exon 6 (nucleotide 613, NM_005216). Sequencing of the second band (712 bp) revealed an alternative transcript containing ACTB intron 3 sequences. Thus, in this transcript, ACTB exon 3 (nucleotide 1587, M10277), and nucleotides 1588 to 1592 of ACTB intron 3, were fused to nucleotides 1706 to 1604 of ACTB intron 3 (M10277), and joined to GLI exon 6 (nucleotide 613, NM_005269) (Figure 6). Results from genomic PCR were in good agreement with the RT-PCR findings in all five cases (data not shown). Reciprocal transcripts were detected by nested PCR in cases 1 and 5. Amplification with primer pairs GLI412F-ACT594R and GLI336F-ACT594R, in cases 1 and 5, respectively, did not yield any visible product at gel electrophoresis, but nested PCR with the corresponding primer pairs GLI520F-ACT520R (case 1) and GLI412F-ACT520R (case 5), amplified fragments that were analyzed by direct sequencing. In case 1, the amplified 289-bp fragment corresponded to a fusion of GLI exon 6 (nucleotide 702, NM_00592) to ACTB exon 4 (nucleotide 434, NM_001101) (Figure 6). In case 5, the amplified 162-bp fragment corresponded to a fusion of GLI exon 4 (nucleotide 467, NM_005269) with ACTB exon 4 (nucleotide 437, NM_001101) (Figure 6). We have identified a discrete group of previously uncharacterized neoplasms that are remarkably homogeneous at the morphological, cytogenetic, and molecular genetic levels. These lesions that, to date, seem benign (albeit with quite limited follow-up) have cytoarchitectural, immunohistochemical, and ultrastructural features highly suggestive of pericytic differentiation and it seems most likely that these lesions fall within the recently recognized spectrum of myopericytic neoplasms.8McMenamin ME Myopericytoma.in: Fletcher CDM Unni KK Mertens F World Health Organization Classification of Tumours. Pathology and Genetics of Tumours of Soft Tissue and Bone. IARC Press, Lyon2002: 138-139Google Scholar Arguably they could be regarded as the true hemangiopericytomas, but, given the loosely used manner in which the latter term has been used in the past 30 years and in view of the resulting confusion, then this terminology seems undesirable at least in the short term. In fact, in the new World Health Organization classification of soft tissue tumors, hemangiopericytoma has been discarded as a discrete entity and is regarded as synonymous with cellular examples of solitary fibrous tumor.9Guillou L Fletcher JA Fletcher CDM Mandahl N Extrapleural solitary fibrous tumour and haemangiopericytoma.in: Fletcher CDM Unni KK Mertens F World health Organization Classification of Tumours. Pathology and Genetics of Tumours of Soft Tissue and Bone. IARC Press, Lyon2002: 86-90Google Scholar In these circumstances (and in line with the established trend in hematolymphoid neoplasia), we suggest the term “pericytoma with t(7;12)” for these lesions, thereby reflecting both their morphological and cytogenetic characteristics. Notably, this group of lesions, which was first identified through their shared karyotypic features, harbors not only a novel, seemingly tumor-specific chromosomal translocation but also a previously unrecognized mechanism of GLI activation. Tumors that might enter the morphological differential diagnosis include cellular examples of solitary fibrous tumor (often termed “hemangiopericytoma” in the past), which have more patternless architecture, more prominent stromal collagen, larger branching vessels, and show consistent CD34 positivity; myofibroma(tosis), which generally has a biphasic appearance with fascicular myoid areas and is most frequent in young children; monophasic synovial sarcoma, which is more fascicular and shows immunopositivity for epithelial membrane antigen and/or keratin; and mesenchymal chondrosarcoma that shows more round cell morphology, greater nuclear atypia, and has foci of cartilaginous differentiation. An additional consideration could be metastatic endometrial stromal sarcoma, in which the vessels are more rounded (spiral arteriole-like) and which often shows focal keratin and desmin immunopositivity as well as consistent positivity for progesterone receptor. The list of identified fusion genes in soft tissue neoplasms, mostly sarcomas, is steadily increasing and it has become clear that most of them involve at least one transcription factor gene.2Åman P Fusion genes in solid tumors.Semin Cancer Biol. 1999; 9: 303-318Crossref PubMed Scopus (83) Google Scholar, 3Ladanyi M Bridge JA Contribution of molecular genetic data to the classification of sarcomas.Hum Pathol. 2000; 31: 532-538Abstract Full Text PDF PubMed Scopus (114) Google Scholar At the molecular genetic level, the fusion of genes causes either overexpression of normally silent genes when placed under the influence of a strong promoter, or else expression of abnormal, chimeric, gene products.2Åman P Fusion genes in solid tumors.Semin Cancer Biol. 1999; 9: 303-318Crossref PubMed Scopus (83) Google Scholar The ACTB-GLI fusion gene described herein fits well with this pattern. The ACTB gene, encoding an important cytoskeletal component, is under the control of a conserved, strong, and complex promoter that assures a high level of expression in nonmuscle cells.10Ng S-Y Gunning P Eddy R Ponte P Leavitt J Shows T Kedes L Evolution of the functional human β-actin gene and its multi-pseudogene family: conservation of noncoding regions and chromosomal dispersion of pseudogenes.Mol Cell Biol. 1985; 5: 2720-2732Crossref PubMed Google Scholar, 11Ng S-Y Gunning P Liu S-H Leavitt J Kedes L Regulation of the human β-actin promoter by upstream and intron domains.Nucleic Acids Res. 1989; 17: 601-615Crossref PubMed Scopus (58) Google Scholar In contrast to the ubiquitous expression of ACTB, GLI expression seems to be restricted to a few tissue types, including the fallopian tube, testis, and myometrium. GLI is the archetype for the human Krüppel gene family, characterized by five DNA-binding zinc finger domains linked by highly conserved histidine-cysteine motifs.12Kinzler KW Ruppert JM Bigner SH Vogelstein B The GLI gene is a member of the Kruppel family of zink finger proteins.Nature. 1988; 332: 371-374Crossref PubMed Scopus (316) Google Scholar, 13Kinzler KW Vogelstein B The GLI gene encodes a nuclear protein which binds specific sequences in the human genome.Mol Cell Biol. 1990; 10: 634-642Crossref PubMed Scopus (409) Google Scholar Activation of GLI genes, also including GLI2 and GLI3, constitute the last known step in the sonic hedgehog (SHH) signaling pathway, and they thus function as direct effectors of the mediated signal. The SHH gene is expressed during embryogenesis and directs tissue-specific cell proliferation. Binding of SHH to the transmembrane receptor PTCH (patched) releases the repression of SMOH (smoothened) exerted by PTCH, which results in transduction of the signal.14Villavicencio EH Walterhouse DO Iannaccone PM The sonic hedgehog-patched-gli pathway in human development and disease.Am J Hum Genet. 2000; 67: 1047-1054Abstract Full Text Full Text PDF PubMed Scopus (332) Google Scholar The resulting activation of GLI induces an up- and down-regulation of multiple target genes involved in, eg, cell-cycle regulation, cell adhesion, apoptosis, signal transduction, and cell proliferation.15Yoon JW Kita Y Frank DJ Majewski RR Konicek BA Nobrega MA Jacob H Walterhouse D Iannaccone P Gene expression profiling leads to identification of GLI1 binding elements in target genes and a role for multiple downstream pathways in GLI1-induced cell transformation.J Biol Chem. 2002; 7: 5548-5555Crossref Scopus (267) Google Scholar Lack of SHH signaling during embryogenesis is associated with human developmental birth defects; mutations in SHH cause holoprosencephaly,16Roessler E Belloni E Gaudenz K Jay P Berta P Scherer SW Tsui LC Muenke M Mutations in the human Sonic Hedgehog gene cause holoprosencephaly.Nat Genet. 1996; 14: 357-360Crossref PubMed Scopus (933) Google Scholar and PTCH mutations are found in patients with the Gorlin syndrome.17Hahn H Wicking C Zaphiropoulos PG Gailani MR Shanley S Chidambaram A Vorechovsky I Holmberg E Unden AB Gillies S Negus K Smyth I Pressman C Leffell DJ Gerrard B Goldstein AM Dean M Toftgard R Chenevix-Trench G Wainwright B Bale AE Mutations in the human homolog of drosophila patched in the nevoid basal cell carcinoma syn
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