Assessment of MDM2 Gene Locus Amplification by Fluorescence In-Situ Hybridization in Juvenile Ossifying Fibroma.

Juvenile ossifying fibroma (JOF) is an uncommon benign fibro-osseous lesion (BFOL) of the maxillofacial bones with a locally aggressive nature and a high recurrence rate. Murine Double Minute 2 (MDM2) is an oncogene located at chromosome 12 (12q13-15) that inhibits the tumor suppressor gene TP53. The presence of MDM2 gene locus amplification is a useful molecular adjunct in the evaluation of some sarcomas, including low-grade intramedullary osteosarcoma (LGIOS). JOF and LGIOS have some overlapping clinical and histopathological features. The aim of this study is to evaluate a series of JOF for the presence of MDM2 gene locus amplification using fluorescence in-situ hybridization (FISH). MATERIALS AND METHODS: With IRB approval, a search of the institutional files of the archives of the Oral Pathology and Surgical Pathology biopsy services at the University of Florida Health was performed. The cases were re-evaluated by an oral pathology resident, an oral and maxillofacial pathologist, and a bone and soft tissue pathologist. Cases with consensus in diagnosis were selected (n = 9) for MDM2 testing. Testing by FISH for MDM2 gene locus amplification was applied to all retrieved cases. RESULTS: The examined cases were all negative for MDM2 gene locus amplification via FISH testing. CONCLUSION: In our small series, JOF did not demonstrate MDM2 gene locus abnormality, a characteristic of LGIOS. This finding suggests that JOF has a distinct underlying pathogenesis. If confirmed in a larger series, these findings may be useful in distinguishing these two entities in cases with overlapping features or when minimal biopsy material is available.

Histomorphometric and molecular characterization of stromal and mineralized components in fibro-osseous lesions.

BACKGROUND: Indistinct and analogous histopathological features of various fibro-osseous lesions make establishing a definitive diagnosis a challenge. There is a need for additional molecular and histochemical tools to support and differentiate these lesions in order to establish a concrete diagnosis. MATERIALS AND METHODS: A retrospective analysis of biopsied lesions in formalin-fixed paraffin-embedded sections (10 cases each of fibrous dysplasia, ossifying fibroma, and cement-osseous dysplasia) retrieved from the archives was studied for immunoexpression of osteocalcin (quantitative analysis in osteocytes), collagen characterization using Azan, Picrosirus, and Toluidine blue stain for evaluating intensity and localization of collagen fibers, and morphometric analysis of vasculature (for evaluating mean vessel density as square microns). RESULTS: Positive immunostaining of osteocalcin suggested mutations of the GNAS-1 gene found in fibrous dysplasia indirectly, as it is a negative regulator of bone formation. Osteocalcin immunopositivity was quantitatively measured in the fibro-osseous lesions, with fibrous dysplasia measuring 14.47 ± 3.628 as compared to ossifying fibroma measuring 5.23 ± 1.33, followed by cemento-osseous dysplasia measuring 2.30 ± 1.409. Toluidine blue suggests the presence of oxytalan fibers (resistant to acid hydrolysis) in ossifying fibroma and cemento-osseous dysplasia, pointing toward the pathogenesis of the lesion. Azan stain and Picrosirus (under a polarizing microscope) helped in distinguishing hard tissue characteristics (70% of cases of fibrous dysplasia showed only a magenta component followed by intermixed magenta with a blue component in 20% of cases and only 10% of cases showed magenta with blue borders whereas for ossifying fibroma, 40% of cases depicted magenta with blue borders along with the other 40% with intermixed magenta with blue component). The mean vessel density was also highest in fibrous dysplasia measuring 7.90 ± 1.079 (in Sq. micron area), followed by ossifying fibroma and cemento-osseous dysplasia. CONCLUSION: The diagnosis of fibro-osseous lesions by hematoxylin and eosin alone is confusing and thus should be supported by relatively simple histomorphometric analysis for better treatment outcomes. At the diagnostic stage of fibro-osseous lesions, evaluation of intralesional vessel size, reliable molecular marker, and histochemical nature can aid in differentiating fibrous dysplasia from central ossifying fibroma and cemento-osseous dysplasia alongside, other clinical, radiographic and pathological criteria. These parameters help in the diagnostic decision-making of fibro-osseous lesions.

Comprehensive Genomic Analysis of Cemento-Ossifying Fibroma.

Cemento-ossifying fibroma (COF) of the jaws is currently classified as a benign mesenchymal odontogenic tumor, and only targeted approaches have been used to assess its genetic alterations. A minimal proportion of COFs harbor CDC73 somatic mutations, and copy number alterations (CNAs) involving chromosomes 7 and 12 have recently been reported in a small proportion of cases. However, the genetic background of COFs remains obscure. We used a combination of whole-exome sequencing and RNA sequencing to assess somatic mutations, fusion transcripts, and CNAs in a cohort of 12 freshly collected COFs. No recurrent fusions have been identified among the 5 cases successfully analyzed by RNA sequencing, with in-frame fusions being detected in 2 cases (MARS1::GOLT1B and PARG::BMS1 in one case and NCLN::FZR1 and NFIC::SAMD1 in the other case) and no candidate fusions identified for the remaining 3 cases. No recurrent pathogenic mutations were detected in the 11 cases that had undergone whole-exome sequencing. A KRAS p.L19F missense variant was detected in one case, and 2 CDC73 deletions were detected in another case. The other variants were of uncertain significance and included variants in PC, ACTB, DOK6, HACE1, and COL1A2 and previously unreported variants in PTPN14, ATP5F1C, APOBEC1, HDAC5, ATF7IP, PARP2, and ACTR3B. The affected genes do not clearly converge on any signaling pathway. CNAs were detected in 5/11 cases (45%), with copy gains involving chromosome 12 occurring in 3/11 cases (27%). In conclusion, no recurrent fusions or pathogenic variants have been detected in the present COF cohort, with copy gains involving chromosome 12 occurring in 27% of cases.

When molecular outsmarts morphology: Malignant ossifying fibromyxoid tumors masquerading as osteosarcomas, including a novel CREBZF::PHF1 fusion.

We present two cases of malignant ossifying fibromyxoid tumor (OFMT) which eluded diagnosis due to compelling clinicopathologic mimicry, compounded by similarly elusive underlying molecular drivers. The first is of a clavicle mass in a 69 year-old female, which histologically showed an infiltrative nested and trabeculated proliferation of monomorphic cells giving rise to scattered spicules of immature woven bone. Excepting SATB2 positivity, the lesion showed an inconclusive immunoprofile which along with negative PHF1 FISH led to an initial diagnosis of high-grade osteosarcoma. Next generation sequencing (NGS) revealed a particularly rare CREBBP::BCORL1 fusion. The second illustrates the peculiar presentation of a dural-based mass in a 52 year-old female who presented with neurologic dyscrasias. Sections showed a sheeted monotonous proliferation of ovoid to spindle cells, but in contrast to Case #1, the tumor contained an exuberance of reticular osteoid and woven bone deposition mimicking malignant osteogenic differentiation. NGS showed a novel CREBZF::PHF1 fusion. Both tumors recurred locally less than 1 year post-operatively. As such we reiterate that careful morphologic examination is axiomatic to any diagnosis in our discipline, but this paradigm must shift to recognize that molecular diagnostics can provide closure where traditional tools have notable limitations.

Ossifying Fibromyxoid Tumor of the Genitourinary Tract: Report of 4 Molecularly Confirmed Cases of a Diagnostic Pitfall.

Ossifying fibromyxoid tumors (OFMTs) are rare mesenchymal neoplasms which typically present in the superficial subcutaneous tissues and have not been reported to arise in visceral organs. We now report 4 molecularly confirmed cases of OFMT involving the genitourinary tract. All patients were males, ranging in age from 20 to 66 years (mean: 43 y). One case each arose in the kidney, ureter, perirenal soft tissue, and penis. All neoplasms demonstrated bland epithelioid to spindled cells set in a variably fibrous to fibromyxoid stroma, and only 1 had a peripheral shell of lamellar bone. All cases appeared well-circumscribed on gross/radiologic examination, though the primary renal neoplasm permeated between native renal tubules. By immunohistochemistry, S100 protein was negative in all 4 cases, while desmin was positive in 2 cases. In 2 cases, the Illumina TruSight RNA Fusion Panel demonstrated a PHF1::TFE3 and EP400::PHF1 fusion, respectively. In the remaining 2 cases, PHF1 gene rearrangement was confirmed by fluorescence in situ hybridization analysis. Due to unusual clinical presentation, lack of S100 positivity, and only occasional bone formation, the correct diagnosis was challenging in the absence of molecular testing. In summary, OFMT may rarely present primarily in the genitourinary tract. Given their nonspecific morphology and immunophenotype, molecular analysis is crucial to establish the correct diagnosis.

Genomic Profiling of the Craniofacial Ossifying Fibroma by Next-Generation Sequencing.

BACKGROUND: Ossifying fibroma (OF) of the craniofacial skeleton is a fibro-osseous lesion characterized by various patterns of bone formation in a cellular fibroblastic stroma. The molecular landscape of OF remains mostly unknown. There are a few known pathogenic abnormalities in OF, including HRPT2 mutations in conventional OF and SATB2 translocations in juvenile psammomatoid OF. On the other hand, conflicting reports exist regarding MDM2 gene amplification and chromosomal copy number alterations (CNA) in OF. METHODS: Surgically removed biopsies and curettage specimens from OF patients were obtained. Clinical, radiographic, and pathologic features of tumors were reviewed. Genomic DNA was extracted from formalin-fixed, paraffin-embedded blocks of tumor tissue. Capture-based DNA next-generation sequencing targeting the coding regions 529 cancer genes and select introns was performed. RESULTS: We identified 17 OF cases from 8 male and 8 female patients with mean age of 22 years (range 1-58 years). Nine case occurred in the gnathic bones and 8 in the extragnathic craniofacial bones. These cases included 3 juvenile psammomatoid OF, 6 conventional OF and 8 juvenile trabecular OF. Large-scale CNAs were present in 6 of 17 cases. Seven cases (41%) had focal amplifications including FOSB (n = 2, 11%), FOS (n = 4, 23%), COL1A1 (n = 4, 23%) and TBX3 (n = 5, 29%). Three cases (17%) had pathogenic CDC73 mutations. No cases showed focal MDM2 amplification. CONCLUSIONS: Here, we provided a comprehensive molecular characterization of OF that reveals a heterogeneous genetic profile with occasional large-scale CNAs (n = 6, 35%). FOS, FOSB, and TBX3 genes that regulate AP-1 transcriptional complex are frequently altered in OF (n = 7, 41%), chiefly in juvenile trabecular OF. These genes encode transcription factors that act as downstream effectors of the MAP kinase signaling pathway. MDM2 amplification is an exceedingly rare event in OF, if present at all, so identification of this event should continue to raise concern for low-grade gnathic osteosarcoma. In summary, our findings suggest that OF represents a heterogeneous group of tumors at the genetic level but dysregulation of the AP-1 pathway may play a role in pathogenesis of juvenile trabecular OF.

Psammomatoid Ossifying Fibroma Is Defined by SATB2 Rearrangement.

Psammomatoid ossifying fibroma (PsOF), also known as juvenile PsOF, is a benign fibro-osseous neoplasm predominantly affecting the extragnathic bones, particularly the frontal and ethmoid bones, with a preference for adolescents and young adults. The clinical and morphologic features of PsOF may overlap with those of other fibro-osseous lesions, and additional molecular markers would help increase diagnostic accuracy. Because identical chromosomal breakpoints at bands Xq26 and 2q33 have been described in 3 cases of PsOF located in the orbita, we aimed to identify the exact genes involved in these chromosomal breakpoints and determine their frequency in PsOF using transcriptome sequencing and fluorescence in situ hybridization (FISH). We performed whole RNA transcriptome sequencing on frozen tissue in 2 PsOF index cases and identified a fusion transcript involving SATB2, located on chromosome 2q33.1, and AL513487.1, located on chromosome Xq26, in one of the cases. The fusion was validated using reverse transcription (RT)-PCR and SATB2 FISH. The fusion lead to a truncated protein product losing most of the functional domains. Subsequently, we analyzed an additional 24 juvenile PsOFs, 8 juvenile trabecular ossifying fibromas (JTOFs), and 11 cemento-ossifying fibromas (COFs) for SATB2 using FISH and found evidence of SATB2 gene rearrangements in 58% (7 of 12) of the evaluable PsOF cases but not in any of the evaluable JTOF (n = 7) and COF (n = 7) cases. A combination of SATB2 immunofluorescence and a 2-color SATB2 FISH in our index case revealed that most tumor cells harboring the rearrangement lacked SATB2 expression. Using immunohistochemistry, 65% of PsOF, 100% of JTOF, and 100% of COF cases showed moderate or strong staining for SATB2. In these cases, we observed a mosaic pattern of expression with >25% of the spindle cells in between the bone matrix, with osteoblasts and osteocytes being positive for SATB2. Interestingly, 35% (8 of 23) of PsOFs, in contrast to JTOFs and COFs, showed SATB2 expression in <5% of cells. To our knowledge, this is the first report that shows the involvement of SATB2 in the development of a neoplastic lesion. In this study, we have showed that SATB2 rearrangement is a recurrent molecular alteration that appears to be highly specific for PsOF. Our findings support that PsOF is not only morphologically and clinically but also genetically distinct from JTOF and COF.

First presentation of a frameshift mutation in the SETD2 gene of a juvenile psammomatoid ossifying fibroma (JPOF) associated with an aneurysmal bone cyst.

BACKGROUND: The rarity of juvenile psammomatoid ossifying fibroma (JPOF) and lack of cytogenetic studies prompted us to report a novel SETD2 gene mutation in a benign odontogenic tumour. CASE PRESENTATION: A 21-year-old man presented with a hard, expanded mandibular cortex. Computed tomography revealed multilocular radiopacity in the mandible; this was reconstructed via segmental mandibulectomy using a vascularised iliac crest flap. Based on the clinical and histological findings, we diagnosed JPOF associated with an aneurysmal bone cyst. Microscopically, the solid area was characterised by many rounded or angular ossicles in a cellular fibrous stroma. The stromal cells were spindle-like or stellate. Next-generation sequencing detected a frame shift mutation of the SETD2 gene, while the copy number was normal. CONCLUSIONS: Our findings suggest further genetic studies should be performed to assess whether this mutation is related to tumour genesis. .

A PHF1-TFE3 fusion atypical ossifying fibromyxoid tumor with prominent collagenous rosettes: Case report with a brief review.

Ossifying fibromyxoid tumor (OFMT) is a rare mesenchymal neoplasm of uncertain line of differentiation that can be subdivided into typical, atypical, and malignant tumors. Cytogenetically, OFMT is characterized by recurrent gene rearrangement involving PHF1 in up to 85% of cases. The most common PHF1 fusion partner is EP400, present in approximately half of cases. Most recently, a novel fusion of PHF1-TFE3 was identified in about 10% of PHF1-rearranged OFMTs. Herein, we report a unique case of PHF1-TFE3 fusion atypical OFMT with prominent collagenous rosettes. A 50-year-old male patient presented with a slowly growing, painless mass in the right foot for 4 years. Gross examination showed a 3.5-cm, subcutaneous well-circumscribed, lobulated mass. Microscopic examination revealed a well-demarcated but un-encapsulated tumor without a peripheral bony shell. The neoplasm was composed of mildly atypical spindle to ovoid cells with increased mitosis (2 mitoses per 10 high-power fields) arranged in a multinodular manner within a fibromyxoid stroma, which contained numerous small, irregular collagenous rosettes surrounded by radiating growth of tumor cells. The neoplastic cells were diffusely positive for TFE3 and CD10. RNA sequencing revealed an in-frame fusion between PHF1 exon 12 and TFE3 exon 7. Subsequent Fluorescence in-situ hybridization analyses demonstrated positive for rearrangements of both the PHF1 and TFE3 loci. The patient was free of disease at 63 months' follow-up. Our case exhibits atypical features and prominent collagenous rosettes, expanding the morphological spectrum of OFMT with PHF1-TFE3 fusion.

Round cell tumor with a myxoid matrix harboring a PHF1-TFE3 fusion: Myoepithelial neoplasm or ossifying fibromyxoid tumor?

Myoepithelial tumors arising in soft tissue are uncommon and mostly manifest a benign clinical course, although a malignant form does exist. An EWSR1 gene rearrangement is a common event in these tumors. Ossifying fibromyxoid tumor, a rare soft tissue neoplasm of uncertain differentiation, may have overlapping histologic and immunophenotypic features with myoepithelial tumors, but frequently harbors a PHF1 gene rearrangement. Interestingly, a PHF1-TFE3 fusion has been recently reported in both entities. Here we report a case of a malignant soft tissue tumor demonstrating myoepithelial differentiation and harboring a PHF1-TFE3 fusion. Despite being slow-growing and lacking significant cytologic atypia at initial presentation, the patient deteriorated rapidly with local recurrence and distant metastases. A discussion of the potential clinicopathologic implications of a PHF1-TFE3 fusion in these entities is also developed.

Copy number alteration profiling facilitates differential diagnosis between ossifying fibroma and fibrous dysplasia of the jaws.

Ossifying fibroma (OF) and fibrous dysplasia (FD) are two fibro-osseous lesions with overlapping clinicopathological features, making diagnosis challenging. In this study, we applied a whole-genome shallow sequencing approach to facilitate differential diagnosis via precise profiling of copy number alterations (CNAs) using minute amounts of DNA extracted from morphologically correlated microdissected tissue samples. Freshly frozen tissue specimens from OF (n = 29) and FD (n = 28) patients were obtained for analysis. Lesion fibrous tissues and surrounding normal tissues were obtained by laser capture microdissection (LCM), with ~30-50 cells (5 000-10 000 µm2) per sample. We found that the rate of recurrent CNAs in OF cases was much higher (44.8%, 13 of 29) than that in FD cases (3.6%, 1 of 28). Sixty-nine percent (9 of 13) of the CNA-containing OF cases involved segmental amplifications and deletions on Chrs 7 and 12. We also identified eight CNA-associated genes (HILPDA, CALD1, C1GALT1, MICALL2, PHF14, AIMP2, MDM2, and CDK4) with amplified expression, which was consistent with the copy number changes. We further confirmed a jaw lesion with a previous uncertain diagnosis due to its ambiguous morphological features and the absence of GNAS mutation as OF based on the typical Chr 12 amplification pattern in its CNA profile. Moreover, analysis of a set of longitudinal samples collected from an individual with a cellular lesion in suspicion of OF at the first surgery, recurrence and the latest malignant transformation revealed identical CNA patterns at the three time points, suggesting that copy number profiling can be used as an important tool to identify borderline lesions or lesions with malignant potential. Overall, CNA profiling of fibro-osseous lesions can greatly improve differential diagnosis between OF and FD and help predict disease progression.

Copy number alteration profiling facilitates differential diagnosis between ossifying fibroma and fibrous dysplasia of the jaws / 国际口腔科学杂志·英文版

Ossifying fibroma (OF) and fibrous dysplasia (FD) are two fibro-osseous lesions with overlapping clinicopathological features, making diagnosis challenging. In this study, we applied a whole-genome shallow sequencing approach to facilitate differential diagnosis via precise profiling of copy number alterations (CNAs) using minute amounts of DNA extracted from morphologically correlated microdissected tissue samples. Freshly frozen tissue specimens from OF (n = 29) and FD (n = 28) patients were obtained for analysis. Lesion fibrous tissues and surrounding normal tissues were obtained by laser capture microdissection (LCM), with ~30-50 cells (5 000-10 000 µm

Primary malignant ossifying fibromyxoid tumour of the bone. A clinicopathologic and molecular report of two cases.

OBJECTIVE: To report the exceptional occurrence of ossifying fibromyxoid tumour (OFMT) as a primary bone lesion. OFMT is a rare soft tissue tumour of uncertain differentiation and variable malignant potential, that occurs in adults with a slight male predominance. It is typically located in the subcutis or in the skeletal muscles of the extremities, followed by trunk or head and neck. METHODS: Two cases of OFMT proven to arise from bone are presented. The first is a 65-year old female with a history of rib "osteosarcoma", presenting with an inferior lobe left lung mass. The second is a man with a lytic lesion of the 5th cervical vertebra that recurred shortly after resection. Following H&E and immunohistochemical examination, tumour samples were analysed by NGS and by break-apart FISH to detect rearrangement of the PHF1 and TFE3 genes. RESULTS: PHF1 gene-rearrangement was identified by FISH on both the primary and the metastatic lesion of first patient. NGS identified a PHF1(intron1) and EPC1 (exon 10) fusion transcript later confirmed by positive PHF1 rearrangement on FISH in the second case. CONCLUSIONS: The demonstration of PHF1 gene rearrangements represents a fundamental ancillary diagnostic test when presented with challenging examples of OFMT.

PHF1 fusions cause distinct gene expression and chromatin accessibility profiles in ossifying fibromyxoid tumors and mesenchymal cells.

Ossifying fibromyxoid tumor (OFMT) is a soft tissue tumor frequently displaying gene fusions, most of which affect the PHF1 gene. PHF1 encodes plant homeodomain finger protein 1, which is involved in various processes regulating gene transcription, including those orchestrated by the polycomb repressor complex 2. Here, a series of 37 OFMTs, including 18 typical, 9 atypical, and 10 malignant variants, was analyzed with regard to transcriptomic features, gene fusion and copy number status, and/or single-nucleotide variants. The effects on gene expression and chromatin accessibility of three detected fusions (EP400-PHF1, MEAF6-PHF1, and PHF1-TFE3) were further evaluated in fibroblasts. Genomic imbalances showed a progression-related pattern, with more extensive copy number changes among atypical/malignant lesions than among typical OFMTs; loss of the RB1 gene was restricted to atypical/malignant OFMTs, occurring in one-third of the cases. RNA sequencing identified fusion transcripts in >80% of the cases analyzed, including a novel CSMD1-MEAF6. The gene-expression profile of OFMT was distinct from that of other soft tissue tumors, with extensive transcriptional upregulation of genes in OFMT. These findings were largely recapitulated in gene fusion-expressing fibroblast lines, suggesting that genes involved in, e.g., Wnt signaling and/or being regulated through trimethylation of lysine 27 in histone 3 (H3K27me3) are pivotal for OFMT development. The genes showing differentially higher expression in fusion-expressing cells paralleled increased chromatin accessibility, as revealed by ATAC sequencing. Thus, the present study suggests that OFMT develops through gene fusions that have extensive epigenetic consequences.

Novel recurrent PHF1-TFE3 fusions in ossifying fibromyxoid tumors.

Ossifying fibromyxoid tumor (OFMT) is an uncommon mesenchymal neoplasm of uncertain differentiation and intermediate malignant potential. Recurrent PHF1 gene rearrangements are detected in up to 80% of OFMTs. We describe the clinicopathologic features of five OFMTs harboring a novel PHF1-TFE3 fusion. In two cases, RNA sequencing identified a fusion transcript composed of PHF1 exon 11 fused to TFE3 exon 3, whereas in a third case PHF1 exon 12 was fused to TFE3 exon 7. A FISH break-apart assay revealed rearrangements in both PHF1 and TFE3 genes in all cases. The cohort included three males and two females with a median age of 64 years. One OFMT originated in the scapula, while four occurred in the deep soft tissues. Two OFMTs had typical features, whereas three were classified as malignant. Despite uniform cytologic features and fibromyxoid stroma compatible with an OFMT diagnosis, none showed a peripheral shell of lamellar bone. S100 expression was focally present in only one case, while desmin was positive in three cases. All tumors showed strong nuclear immunopositivity for TFE3. All three malignant OFMTs developed metastases, either regionally or to the lung. One patient died of disease 1 year after diagnosis, while the remaining two are alive with disease. In summary, we report novel recurrent PHF1-TFE3 fusions in a subset of OFMTs with aggressive clinical behavior. The PHF1-TFE3 fusions resulted in consistent protein TFE3 overexpression which can be used as a reliable screening tool, adding OFMT as another tumor driven by TFE3 oncogenic activation pathway.

Deletion of Menin in craniofacial osteogenic cells in mice elicits development of mandibular ossifying fibroma.

Ossifying fibroma (OF) is a rare benign tumor of the craniofacial bones that can reach considerable and disfiguring dimensions if left untreated. Although the clinicopathological characteristics of OF are well established, the underlying etiology has remained largely unknown. Our work indicates that Men1-a tumor suppressor gene responsible of Multiple endocrine neoplasia type 1-is critical for OF formation and shows that mice with targeted disruption of Men1 in osteoblasts (Men1Runx2Cre) develop multifocal OF in the mandible with a 100% penetrance. Using lineage-tracing analysis, we demonstrate that loss of Men1 arrests stromal osteoprogenitors in OF at the osterix-positive pre-osteoblastic differentiation stage. Analysis of Men1-lacking stromal spindle cells isolated from OF (OF-derived MSCs (OFMSCs)) revealed a downregulation of the cyclin-dependent kinase (CDK) inhibitor Cdkn1a, consistent with an increased proliferation rate. Intriguingly, the re-expression of Men1 in Men1-deficient OFMSCs restored Cdkn1a expression and abrogated cellular proliferation supporting the tumor-suppressive role of Men1 in OF. Although our work presents the first evidence of Men1 in OF development, it further provides the first genetic mouse model of OF that can be used to better understand the molecular pathogenesis of these benign tumors and to potentially develop novel treatment strategies.

MicroRNA profiling reveals dysregulated microRNAs and their target gene regulatory networks in cemento-ossifying fibroma.

BACKGROUND: Cemento-ossifying fibroma (COF) is a benign fibro-osseous neoplasm of uncertain pathogenesis, and its treatment results in morbidity. MicroRNAs (miRNA) are small non-coding RNAs that regulate gene expression and may represent therapeutic targets. The purpose of the study was to generate a comprehensive miRNA profile of COF compared to normal bone. Additionally, the most relevant pathways and target genes of differentially expressed miRNA were investigated by in silico analysis. METHODS: Nine COF and ten normal bone samples were included in the study. miRNA profiling was carried out by using TaqMan® OpenArray® Human microRNA panel containing 754 validated human miRNAs. We identified the most relevant miRNAs target genes through the leader gene approach, using STRING and Cytoscape software. Pathways enrichment analysis was performed using DIANA-miRPath. RESULTS: Eleven miRNAs were downregulated (hsa-miR-95-3p, hsa-miR-141-3p, hsa-miR-205-5p, hsa-miR-223-3p, hsa-miR-31-5p, hsa-miR-944, hsa-miR-200b-3p, hsa-miR-135b-5p, hsa-miR-31-3p, hsa-miR-223-5p and hsa-miR-200c-3p), and five were upregulated (hsa-miR-181a-5p, hsa-miR-181c-5p, hsa-miR-149-5p, hsa-miR-138-5p and hsa-miR-199a-3p) in COF compared to normal bone. Eighteen common target genes were predicted, and the leader genes approach identified the following genes involved in human COF: EZH2, XIAP, MET and TGFBR1. According to the biology of bone and COF, the most relevant KEGG pathways revealed by enrichment analysis were proteoglycans in cancer, miRNAs in cancer, pathways in cancer, p53-, PI3K-Akt-, FoxO- and TGF-beta signalling pathways, which were previously found to be differentially regulated in bone neoplasms, odontogenic tumours and osteogenesis. CONCLUSION: miRNA dysregulation occurs in COF, and EZH2, XIAP, MET and TGFBR1 are potential targets for functional analysis validation.

The Wnt/ß-catenin pathway is deregulated in cemento-ossifying fibromas.

OBJECTIVE: The molecular pathogenesis of cemento ossifying fibroma (COF) is unclear. The purpose of this study was to investigate mutations in 50 oncogenes and tumor suppressor genes, including APC and CTNNB1, in which mutations in COF have been previously reported. In addition, we assessed the transcriptional levels of the Wnt/ß-catenin pathway genes in COF. STUDY DESIGN: We used a quantitative polymerase chain reaction array to evaluate the transcriptional levels of 44 Wnt/ß-catenin pathway genes in 6 COF samples, in comparison with 6 samples of healthy jaws. By using next-generation sequencing (NGS) in 7 COF samples, we investigated approximately 2800 mutations in 50 genes. RESULTS: The expression assay revealed 12 differentially expressed Wnt/ß-catenin pathway genes in COF, including the upregulation of CTNNB1, TCF7, NKD1, and WNT5 A, and downregulation of CTNNBIP1, FRZB, FZD6, RHOU, SFRP4, WNT10 A, WNT3 A, and WNT4, suggesting activation of the Wnt/ß-catenin signaling pathway. NGS revealed 5 single nucleotide variants: TP53 (rs1042522), PIK3 CA (rs2230461), MET (rs33917957), KIT (rs3822214), and APC (rs33974176), but none of them was pathogenic. CONCLUSIONS: Although NGS detected no oncogenic mutation, deregulation of key Wnt/ß-catenin signaling pathway genes appears to be relevant to the molecular pathogenesis of COF.

Unusual skull tumors with psammomatoid bodies: a diagnostic challenge.

AIM: We describe a series of three diagnostically challenging, histologically similar fibro-osseous skull masses. METHODS: The cases were identified in our archives among 50,000 neuropathology specimens. A comprehensive review of the histological, immunohistochemical, ultrastructural, and imaging features as well as the clinical outcome was performed. RESULTS: The routine histology was similar in all 3 cases and showed spindle cell proliferations with frequent calcospheres or psammomatoid bodies. There was no evidence of an underlying subdural component. Immunohistochemistry for the meningioma markers EMA and SSTR2A raised the possibility of intraosseous meningioma, as all 3 lesions were convincingly positive for epithelial membrane antigen (EMA) and 1 lesion was convincingly positive for the somatostatin receptor subtype 2A (SSTR2A); weak, questionable positivity for SSTR2 was present in the remaining 2 cases. In addition, electron microscopy was available in 1 case and showed features consistent with meningioma. CONCLUSIONS: Overall, the findings were most consistent with intraosseous meningioma. Primary intraosseous meningiomas are rare lesions that may present a diagnostic challenge. It is important to consider meningiomas in the differential diagnosis, as extradural meningiomas are associated with an increased risk of recurrence and may occasionally undergo malignant transformation.
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