Pulmonary adenocarcinoma with signet ring cell features: a comprehensive study from 3 distinct patient cohorts.

Comprehensive biological characteristics of pulmonary adenocarcinomas with signet ring cell features (SRC⁺) are not well known. Herein, we systematically evaluated clinical and molecular features of SRC⁺ cases with particular attention to smoking status. Surgically treated lung adenocarcinomas (n=763) with follow-up ≥5 years in 3 cohorts were reviewed: all patients in 2006 to 2007 ("all-comers," n=222; 168 ever-smokers), a never-smoker cohort (n=266), and a cohort of ever-smokers (n=275). SRC⁺ tumors had ≥10% of SRCs agreed by 2 pathologists. SRC⁺ cases were tested for rearrangement of ALK and ROS1, as well as 187 known mutations in 10 oncogenes including EGFR, KRAS, BRAF, ERBB2, JAK2, AKT1, AKT2, KIT, MET, and PIK3CA. Overall, 53 of 763 cases (7%) were SRC⁺. In the 2006 to 2007 "all comer" cohort, 9% were SRC⁺. In the never-smoker cohort, 9% were SRC⁺. In the smoker cohort, 3% were SRC⁺. Univariable analysis showed that SRC⁺ never-smokers had shorter overall and disease-free survival (P=0.006 and 0.0004, respectively), but the significance faded in the multivariable analysis. For the other 2 cohorts, crude 5-year survival was decreased by 6% to 27% in SRC⁺ cases without reaching statistical significance. In SRC⁺ tumors, KRAS mutation was most common (29%), followed by ALK (26%), EGFR (18%), ROS1 (6%), BRAF (6%), and PIK3CA (3%). In summary, SRC⁺ tumors in never-smokers had a worse survival by univariable analysis only. SRC⁺ cases seemed enriched for ALK⁺ and ROS1⁺, and other mutations were generally in keeping with the patient's smoking status.

Analysis of MAML2 rearrangement in mucoepidermoid carcinoma of the thymus.

Primary thymic mucoepidermoid carcinoma (TMEC) is rare. High-grade TMEC can be difficult to distinguish from poorly differentiated squamous cell carcinoma and adenosquamous carcinoma. A strong association between mucoepidermoid carcinoma (MEC) and t(11;19)(q21;p13) has been observed in other anatomical sites. Although this translocation is largely considered a disease-defining event for MEC, its incidence in TMEC has not been explored. In this study, we evaluate the value of identifying MAML2 rearrangement by fluorescence in situ hybridization (FISH) to distinguish TMEC from poorly differentiated squamous cell carcinoma and adenosquamous carcinoma. Cases of TMEC, moderate to poorly differentiated squamous cell carcinoma, and adenosquamous carcinoma were re-reviewed by 3 surgical pathologists and classified according to the current World Health Organization classification of thymic tumors (2004). Cases of TMEC were histologically graded using the Brandwein system. FISH was used to detect MAML2 rearrangements using a break-apart probe. FISH for MAML2 rearrangement was performed on cases of TMEC (n = 2), thymic squamous cell carcinoma (n = 5), and thymic adenosquamous carcinoma (n = 3). The 2 cases of TMEC showed MAML2 rearrangement. All other tested cases did not show rearrangement of MAML2. In conclusion, using FISH to identify MAML2 rearrangement is a valuable diagnostic tool in the evaluation of thymic malignancies, specifically, distinguishing TMEC from squamous cell carcinoma and adenosquamous carcinoma. These findings also suggest that TMEC has both histomorphologic and cytogenetic similarities to cases of MEC arising from other anatomical sites.

Endobronchial lipomatous tumors: clinicopathologic analysis of 12 cases with molecular cytogenetic evidence supporting classification as "lipoma".

Lipomatous lesions rarely involve the bronchial tree, and detailed morphologic and molecular cytogenetic analysis of these tumors is lacking. The clinicopathologic features of 12 endobronchial lipomatous neoplasms were studied, with ancillary fluorescence in situ hybridization performed in subsets of cases for CPM, which is amplified in atypical lipomatous tumors/well-differentiated liposarcomas (ALT/WDL), and HMGA1 and HMGA2, which are often rearranged in lipomas. The cases occurred predominately in older men (91%) (age range 44 to 80 y, mean 65 y). Most patients (80%) had a former or current history of heavy smoking (20 to 100 pack-years). Three patients had concurrent pulmonary squamous cell carcinoma, and 1 had a history of multiple lung cancers. Most lesions were small (<2.5 cm) and discovered incidentally. A subset of tumors showed atypical morphologic features that would be suggestive of ALT/WDL in soft tissue sites, including regions of fibrosis and scattered hyperchromatic stromal cells. However, all cases with atypia were CPM negative and behaved in a clinically benign manner. Seven cases were tested for HMGA1 and HMGA2 rearrangement; 4 showed HMGA2 rearrangement, and 1 showed HMGA1 rearrangement, consistent with lipomas. Two cases were negative for HMGA1/2 rearrangements. We conclude that endobronchial lipomatous neoplasms represent lipomas, even in the presence of morphologic features suggestive of ALT/WDL. Ancillary fluorescence in situ hybridization testing may be very valuable in the analysis of these rare tumors, as true ALT/WDL seem to be very rare or nonexistent at this anatomic site.

MET and EGFR mutations identified in ALK-rearranged pulmonary adenocarcinoma: molecular analysis of 25 ALK-positive cases.

INTRODUCTION: Oncogenic ALK kinase activity associated with ALK gene rearrangement is the target of crizotinib, an ALK inhibitor recently approved by the Food and Drug Administration for the treatment of ALK-rearranged (ALK+) non-small cell lung cancers. ALK+ status is generally thought to be mutually exclusive of epidermal growth factor receptor (EGFR) and KRAS mutations. However, the mutation status of other genes is not widely known in ALK+ tumors. The aim of this study is to survey for mutations involving other genes in 25 ALK+ cases confirmed by fluorescent in situ hybridization. METHODS: Using the DNA extracted from formalin-fixed paraffin-embedded tumor samples, a MassArray-based Lung Cancer Mutations Screening Panel was performed to test for 179 individual mutations in 10 genes, including EGFR, KRAS, BRAF, ERBB2, JAK2, AKT1, AKT2, KIT, MET and PIK3CA, which have been implicated in lung carcinogenesis and/or considered as potential therapeutic targets. RESULTS: Five of 25 ALK+ cases showed additional genetic abnormalities, which were verified by gene sequencing. One patient had EGFR del L747-S752. The remaining four mutations were in the MET gene: MET N375S (n = 2) and MET R988C (n = 2). No MET amplification was found by fluorescent in situ hybridization in the four cases with MET mutation. No mutations were detected in the other genes tested. CONCLUSIONS: In summary, additional mutations were found in 20% of ALK+ cases involving two of the 10 genes tested. Our study highlights that EGFR mutation can be present in ALK+ tumors, though uncommon. Clinical implication of MET mutation in our cases is uncertain and further study is needed.

Fusion of dynactin 1 to anaplastic lymphoma kinase in inflammatory myofibroblastic tumor.

Inflammatory myofibroblastic tumor is an uncommon neoplasm that occurs more often in younger patients. Approximately 50% of inflammatory myofibroblastic tumors are characterized by anaplastic lymphoma kinase fusion genes, more commonly TPM3-anaplastic lymphoma kinase and TPM4-anaplastic lymphoma kinase. Herein, we report a novel fusion of dynactin 1 to anaplastic lymphoma kinase in a neck inflammatory myofibroblastic tumor diagnosed in a 7-year-old girl. Histologic evaluation showed a perineurioma-like bland spindle cell neoplasm with positive immunohistochemical staining for anaplastic lymphoma kinase, S-100, and CD34 but negative for epithelial membrane antigen. Standard cytogenetic analysis showed a der(2)t(2;12)(p23;q11). Fluorescence in situ hybridization demonstrated separation of the anaplastic lymphoma kinase locus. 5'-rapid amplification of complementary DNA ends polymerase chain reaction identified an in-frame fusion of dynactin 1 exon 16 on chromosome 2 to anaplastic lymphoma kinase exon 20. Reverse transcription-polymerase chain reaction with specific primers and direct sequencing confirmed the fusion. The structure of the fusion protein retains the cytoskeleton-associated protein-glycine domain and coiled coil domain of dynactin 1 and the receptor tyrosine kinase domain of anaplastic lymphoma kinase. This novel fusion gene is structurally similar to other previously described anaplastic lymphoma kinase fusion genes and may be associated with the unusual morphology and immunophenotype of this tumor.

HMGA gene rearrangement is a recurrent somatic alteration in polypoid endometriosis.

The pathogenesis of endometriosis is unclear, and several genetic, endocrine, immune, and environmental agents have been evaluated with no putative causative factors identified. Here, we show somatic genetic alterations involving HMGA1 (6p21) and HMGA2 (12q15) in 3 cases of polypoid endometriosis. The lesions involved the small bowel mesentery and perirectal soft tissue in 1 case and the posterior vaginal fornix and sigmoid colon serosa in 2 other cases, respectively. All had a polypoid configuration with cystically dilated irregular glands and fibrotic stroma, containing thick-walled vessels. Conventional cytogenetic analysis of 1 case showed 46,XX,t(5;12)(q13;q15) in all metaphases. Fluorescence in situ hybridization studies confirmed the balanced rearrangement of HMGA2. HMGA1 rearrangements were present in 2 additional cases. Rearrangements were exclusively found in the stromal component but not in the glandular component. These findings suggest that HMGA rearrangements likely contribute to the pathogenesis of endometriosis. However, additional studies are needed to better define the biologic role of this genetic alteration.

Worse disease-free survival in never-smokers with ALK+ lung adenocarcinoma.

INTRODUCTION: The EML4-anaplastic lymphoma kinase (ALK) translocation is a recognized oncogenic driver in non-small cell lung cancer. We investigated immunohistochemistry (IHC) screening with fluorescence in situ hybridization (FISH) confirmation for ALK detection and estimated the prevalence of ALK positivity in our patient cohort of never-smokers, together with differences in clinical outcomes and prognostic factors for patients with ALK-positive and ALK-negative tumors. METHODS: We designed a three-phase study (training, validation, and testing) in 300 never-smokers with lung adenocarcinoma from the observational Mayo Clinic Lung Cancer Cohort. Tumor samples were tested using IHC and FISH, and concordance between the methods was assessed. Clinical outcomes were assessed via 5-year progression- or recurrence-free survival from diagnosis. Prognostic factors for ALK-positive tumors and metastases were also investigated. RESULTS: ALK-positive patients were significantly (p < 0.05) younger and had higher grade tumors than ALK-negative patients. ALK positivity was 12.2% by IHC and confirmed at 8.2% of tumors by FISH, with complete concordance between IHC 3+/0 and FISH+/- assessments, respectively. Five-year risk of progression or recurrence was doubled for patients with ALK-positive compared with ALK-negative tumors; ALK-positive tumors also appeared to be associated with a higher risk of brain and liver metastases. CONCLUSIONS: Our findings suggest that ALK positivity is associated with a significantly poor outcome in nonsmoking-related adenocarcinoma and that ALK-positive tumors may be associated with an increased risk of brain and liver metastases compared with ALK-negative disease. Consequently, an unmet medical need exists in ALK-positive lung cancer patients, and effective ALK-specific therapies are needed.

Identification of a disease-defining gene fusion in epithelioid hemangioendothelioma.

Integrating transcriptomic sequencing with conventional cytogenetics, we identified WWTR1 (WW domain-containing transcription regulator 1) (3q25) and CAMTA1 (calmodulin-binding transcription activator 1) (1p36) as the two genes involved in the t(1;3)(p36;q25) chromosomal translocation that is characteristic of epithelioid hemangioendothelioma (EHE), a vascular sarcoma. This WWTR1/CAMTA1 gene fusion is under the transcriptional control of the WWTR1 promoter and encodes a putative chimeric transcription factor that joins the amino terminus of WWTR1, a protein that is highly expressed in endothelial cells, in-frame to the carboxyl terminus of CAMTA1, a protein that is normally expressed only in brain. Thus, CAMTA1 expression is activated inappropriately through a promoter-switch mechanism. The gene fusion is present in virtually all EHEs tested but is absent from all other vascular neoplasms, demonstrating it to be a disease-defining genetic alteration. A sensitive and specific break-apart fluorescence in situ hybridization assay was also developed to detect the translocation and will assist in the evaluation of this diagnostically challenging neoplasm. The chimeric WWTR1/CAMTA1 transcription factor may represent a therapeutic target for EHE and offers the opportunity to shed light on the functions of two poorly characterized proteins.

Nodular fasciitis: a novel model of transient neoplasia induced by MYH9-USP6 gene fusion.

Nodular fasciitis (NF) is a relatively common mass-forming and self-limited subcutaneous pseudosarcomatous myofibroblastic proliferation of unknown pathogenesis. Due to its rapid growth and high mitotic activity, NF is often misdiagnosed as a sarcoma. While studying the USP6 biology in aneurysmal bone cyst and other mesenchymal tumors, we identified high expression levels of USP6 mRNA in two examples of NF. This finding led us to further examine the mechanisms underlying USP6 overexpression in these lesions. Upon subsequent investigation, genomic rearrangements of the USP6 locus were found in 92% (44 of 48) of NF. Rapid amplification of 5'-cDNA ends identified MYH9 as the translocation partner. RT-PCR and direct sequencing revealed the fusion of the MYH9 promoter region to the entire coding region of USP6. Control tumors and tissues were negative for this fusion. Xenografts of cells overexpressing USP6 in nude mice exhibited clinical and histological features similar to human NF. The identification of a sensitive and specific abnormality in NF holds the potential to be used diagnostically. Considering the self-limited nature of the lesion, NF may represent a model of 'transient neoplasia', as it is, to our knowledge, the first example of a self-limited human disease characterized by a recurrent somatic gene fusion event.

High-resolution genomic mapping reveals consistent amplification of the fibroblast growth factor receptor substrate 2 gene in well-differentiated and dedifferentiated liposarcoma.

Well-differentiated liposarcoma (WDLS) is one of the most common malignant mesenchymal tumors and dedifferentiated liposarcoma (DDLS) is a malignant tumor consisting of both WDLS and a transformed nonlipogenic sarcomatous component. Cytogenetically, WDLS is characterized by the presence of ring or giant rod chromosomes containing several amplified genes, including MDM2, TSPAN31, CDK4, and others mainly derived from chromosome bands 12q13-15. However, the 12q13-15 amplicon is large and discontinuous. The focus of this study was to identify novel critical genes that are consistently amplified in primary (nonrecurrent) WDLS and with potential relevance for future targeted therapy. Using a high-resolution (5.0 kb) "single nucleotide polymorphism"/copy number variation microarray to screen the whole genome in a series of primary WDLS, two consistently amplified areas were found on chromosome 12: one region containing the MDM2 and CPM genes, and another region containing the FRS2 gene. Based on these findings, we further validated FRS2 amplification in both WDLS and DDLS. Fluorescence in situ hybridization confirmed FRS2 amplification in all WDLS and DDLS tested (n = 57). Real time PCR showed FRS2 mRNA transcriptional upregulation in WDLS (n = 19) and DDLS (n = 13) but not in lipoma (n = 5) and normal fat (n = 9). Immunoblotting revealed high expression levels of phospho-FRS2 at Y436 and slightly overexpression of total FRS2 protein in liposarcoma but not in normal fat or preadipocytes. Considering the critical role of FRS2 in mediating fibroblast growth factor receptor signaling, our findings indicate that FRS2 signaling should be further investigated as a potential therapeutic target for liposarcoma.

HMGA2 expression analysis in cytological and paraffin-embedded tissue specimens of thyroid tumors by relative quantitative RT-PCR.

The distinction between benign and malignant thyroid tumors in some cytological and histological specimens remains challenging. The aim of this study was to evaluate the use of High Mobility Group A2 (HMGA2) mRNA expression to distinguish benign from malignant thyroid tumors in cytological and histological specimens. RNA samples from 170 thyroid formalin-fixed paraffin-embedded (FFPE) tissues and 226 fine needle aspiration (FNA) specimens were analyzed by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The FFPE tissues included 34 follicular adenomas, 10 Hürthle cell adenomas (HA), 6 hyperplastic nodules, 4 atypical adenomas, 44 classic papillary thyroid carcinomas (PTC), 29 follicular variant of PTC, 23 follicular thyroid carcinomas, 17 Hürthle cell carcinomas (HC), and 3 anaplastic thyroid carcinomas. The FNA specimens included 55 follicular adenomas, 34 HA, 20 hyperplastic nodules, 8 Hashimoto thyroiditis, 32 PTC, 24 follicular variant of PTC, 30 follicular thyroid carcinomas, 21 HC, and 2 anaplastic thyroid carcinomas. HMGA2 mRNA levels were expressed as relative fold change after normalizing with a calibrator. HMGA2 expression in thyroid carcinomas (16.8-fold for FFPE and 18.2-fold for FNA) was significantly higher than in benign lesions (0.8-fold for FFPE and 0.8-fold for FNA). HMGA2 expression in HC was relatively low (1.8-fold for FFPE and 8.5-fold for FNA) compared with the other types of carcinomas. HMGA2 expression values of 4.5-fold and 5.9-fold were used as cutoff points for FFPE and FNA (excluding HA and HC), respectively, to separate benign and malignant thyroid tumors, with 97.5% clinical specificity and 79.8% sensitivity for FFPE, and 95.2% clinical specificity and 88.6% sensitivity for the FNA specimens. Conventional RT-PCR supported the qRT-PCR results. Detection of HMGA2 mRNA expression by qRT-PCR may be a useful tool to assist in the diagnosis of well-differentiated thyroid carcinomas. The 1-step qRT-PCR method is a sensitive, accurate, and reliable technique for gene expression analysis of thyroid tumors.

Reticular angiomatoid "malignant" fibrous histiocytoma--a case report with cytogenetics and molecular genetic analyses.

Angiomatoid "malignant" fibrous histiocytoma is a rare sarcoma of low malignant potential that occurs most commonly in the extremities of children and young adults. Herein, we present a case of angiomatoid malignant fibrous histiocytoma with unusual histologic features arising in the mediastinum of an 80-year-old man. The tumor exhibited a reticular growth pattern and myxoid stroma. The tumor cells expressed epithelial membrane antigen and desmin. Cytogenetic analysis revealed the translocation t(2;22)(q33;q12). Molecular genetic analysis confirmed the rearrangement of the EWSR1 locus and the presence of the EWSR1/CREB1 fusion. This report expands the clinicopathologic spectrum of angiomatoid malignant fibrous histiocytoma and underscores the value of integrating morphologic, immunophenotypic, and molecular findings in the identification of its unusual morphologic variants.

Correlation of IHC and FISH for ALK gene rearrangement in non-small cell lung carcinoma: IHC score algorithm for FISH.

INTRODUCTION: Accurate, cost-effective methods for testing anaplastic lymphoma kinase gene rearrangement (ALK+) are needed to select patients with non-small cell lung carcinoma for ALK-inhibitor therapy. Fluorescent in situ hybridization (FISH) is used to detect ALK+, but it is expensive and not routinely available. We explored the potential of an immunohistochemistry (IHC) scoring system as an affordable, accessible approach. METHODS: One hundred one samples were obtained from an enriched cohort of never-smokers with adenocarcinoma from the Mayo Clinic Lung Cancer Cohort. IHC was performed using the ALK1 monoclonal antibody with ADVANCE detection system (Dako) and FISH with dual-color, break-apart probe (Abbott Molecular) on formalin-fixed, paraffin-embedded tissue. RESULTS: Cases were assessed as IHC score 0 (no staining; n = 69), 1+ (faint cytoplasmic staining, n = 21), 2+ (moderate, smooth cytoplasmic staining; n = 3), or 3+ (intense, granular cytoplasmic staining in ≥10% of tumor cells; n = 8). All IHC 3+ cases were FISH+, whereas 1 of 3 IHC 2+ and 1 of 21 IHC 1+ cases were FISH+. All 69 IHC 0 cases were FISH-. Considering FISH a gold-standard reference in this study, sensitivity and specificity of IHC were 90 and 97.8%, respectively, when 2+ and 3+ were regarded as IHC positive and 0 and 1+ as IHC negative. CONCLUSIONS: IHC scoring correlates with FISH and may be a useful algorithm in testing ALK+ by FISH in non-small cell lung carcinoma, similar to human epidermal growth factor-2 testing in breast cancer. Further study is needed to validate this approach.

Fusion of HMGA2 to COG5 in uterine leiomyoma.

Uterine leiomyomas are smooth muscle tumors most commonly seen in middle-aged women. Approximately 10% of these tumors contain rearrangements of the chromatin-remodeling gene HMGA2 at the chromosome band 12q14.3. Herein, we report on a uterine leiomyoma with a novel HMGA2 fusion gene. A 44-year-old woman presented with a 20-cm mass uterine leiomyoma. From a histological standpoint, the tumor exhibited extensive hyalinization, very low mitotic activity (<1/10 HPH), and no cytologic atypia. Smooth muscle differentiation was confirmed by the expression of smooth muscle actin and desmin. Standard cytogenetic analysis showed the reciprocal translocation t(7;12)(q31.2;q14.3). Fluorescence in situ hybridization analysis confirmed a balanced rearrangement of the HMGA2 locus in 80% of the cells. 3'RACE reverse-transcription polymerase chain reaction identified the fusion of HMGA2 exon 4 to the COG5 locus on 7q31 (component of oligomeric golgi complex 5 isoform). The fusion sequence is predicted to encode a 96-amino acid chimeric protein that retains all three DNA-binding domains (AT hooks) of HMGA2, but that is shorter than the original HMGA2 protein. Since the general structure of the fusion gene is similar to other previously described HMGA2 fusions, its biologic activity is predicted to be likely similar.

Diagnostic utility of IMP3 expression in thyroid neoplasms: a quantitative RT-PCR study.

The capability of molecular markers to differentiate between benign and malignant well-differentiated thyroid tumors remains unclear. The aim of this study was to evaluate the use of insulin-like growth factor II mRNA binding protein-3 (IMP3) mRNA expression to distinguish benign from malignant thyroid tumors. RNA samples from 80 formalin-fixed, paraffin-embedded thyroid tissues, including 22 usual papillary thyroid carcinomas (PTCs), 18 follicular variants of PTC, 5 follicular thyroid carcinomas, 33 follicular adenomas, and 2 hyperplastic nodules, were used for quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis. IMP3 mRNA expression levels in thyroid tumors were expressed as relative fold change (fold) after normalization with normal thyroid RNA. The results showed that thyroid carcinomas including PTC, follicular variants of PTC, and follicular thyroid carcinomas have significantly higher IMP3 expression levels with 48.3, 35.3, and 43.8 fold, respectively, compared with benign thyroid lesions (2.8 fold). Using the IMP3 expression value of 5 fold as a cutoff point to separate benign and malignant thyroid tumors, IMP3 qRT-PCR analysis had a 91.4% clinical specificity and 86.7% clinical sensitivity for the diagnosis of well-differentiated thyroid carcinomas. Conventional RT-PCR and immunohistochemical analysis for IMP3 in a subset of cases supported the qRT-PCR results. These results indicate that detection of IMP3 mRNA expression levels by qRT-PCR may be a useful molecular marker to assist in the diagnosis of well-differentiated thyroid carcinomas.

HMGA1 and HMGA2 rearrangements in mass-forming endometriosis.

Endometriosis is a common gynecologic disorder characterized by ectopic endometrium associated with pelvic pain and infertility. The pathogenesis of endometriosis is unclear, and several genetic, endocrine, immune, and environmental agents have been studied as putative causative factors. However, consistent somatic genetic alterations have not been identified. Rarely, endometriosis presents as a mass lesion with an infiltrative pattern reminiscent of malignancy. We describe cytogenetic and molecular cytogenetic findings of mass-forming endometriosis. The index case of pulmonary endometriosis underwent conventional and molecular cytogenetics analysis. In addition, 16 cases of mass-forming endometriosis, 11 cases of usual endometriosis, and six endometriomas were investigated by fluorescence in situ hybridization (FISH) for HMGA1 and HMGA2 loci, performed on paraffin-embedded thin tissue sections with custom-designed probes. The index patient had an endometriotic lung nodule, with a 46,XX, t(5;6)(q13;p21) karyotype and HMGA1 rearrangement by FISH. A second patient had decidualized endometriosis forming a large abdominal mass and HMGA1 rearrangement by FISH. Of the 15 other cases of mass-forming endometriosis, one had HMGA1 rearrangement and two had HMGA2 rearrangement. The rearrangements were found in the stromal component exclusively. None of the usual endometriosis cases or endometriomas had HMGA1 or HMGA2 rearrangements. In conclusion, mass-forming endometriosis is an uncommon subset of endometriosis that harbors HMGA1 or HMGA2 rearrangements in up to 29% of cases. The present findings support the concept that endometriosis is clonal and that rearrangement of HMGA genes likely contributes to its pathogenesis.

Carboxypeptidase M: a biomarker for the discrimination of well-differentiated liposarcoma from lipoma.

The discrimination between well-differentiated liposarcomas/atypical lipomatous tumors and lipomas can be diagnostically challenging at the histological level. However, cytogenetic identification of ring and giant rod chromosomes supports the diagnosis of well-differentiated liposarcoma/atypical lipomatous tumor. These abnormal chromosomes are mainly composed of amplified genomic sequences derived from chromosome 12q13-15, and contain several genes, including MDM2, CDK4 (SAS), TSPAN31, HMGA2, and others. MDM2 is consistently amplified in well-differentiated liposarcomas/atypical lipomatous tumors, and up to 25% in other sarcomas. As part of a large genomic study of lipomatous neoplasms, we initially found CPM to be consistently amplified in well-differentiated liposarcomas/atypical lipomatous tumors. To further explore this initial finding, we investigated the copy number status of MDM2 and CPM by fluorescent in situ hybridization (FISH) on a series of 138 tumors and 17 normal tissues, including 32 well-differentiated liposarcoma/atypical lipomatous tumors, 63 lipomas, 11 pleomorphic lipomas, 2 lipoblastomas, 30 other tumors and 17 normal fat samples. All 32 well-differentiated liposarcoma/atypical lipomatous tumors showed amplification of MDM2 and CPM, usually >20 copies per cell. The other tumors lacked MDM2 and/or CPM amplification. Chromogenic in situ hybridization confirmed the above results on a subset of these tumors (n=27). These findings suggest that identification of CPM amplification could be used as an alternative diagnostic tool for the diagnosis of well-differentiated liposarcoma/atypical lipomatous tumors.

Pseudosarcomatous fibroblastic/myofibroblastic proliferation in perinephric adipose tissue adjacent to renal cell carcinoma: a lesion mimicking well-differentiated liposarcoma.

Cytologically atypical stromal cells were found in the perinephric adipose tissue, mimicking well-differentiated liposarcoma in 12 of 59 (20%) consecutive nephrectomy specimens that were resected for renal cell carcinoma. Morphologically, the atypical cells included enlarged, hyperchromatic spindle cells and floret-type multinucleate cells. Of 59, 10 (17%) renal cell carcinomas invaded through the renal capsule into the perinephric adipose tissue. Of these cases, three (30%) contained the aforementioned atypical cells. In contrast, 9 of 49 cases without extrarenal invasion (18%) contained the atypical stromal cells. Of the 12 cases with atypical stromal cells, 3 (25%) were associated with extrarenal involvement. The atypical spindle cells exhibited focal to variable positivity for smooth muscle actin and desmin in 3 of the 14 cases (including two cases from our consultation files) each. Cytokeratin AE1/AE3, cytokeratin Cam 5.2, cytokeratin 7, epithelial membrane antigen, and S-100 were negative in all cases. Amplification of MDM2 gene region, which is commonly observed in well-differentiated liposarcoma, was absent by fluorescence in situ hybridization (FISH) in the atypical stromal cells. Immunohistochemistry and FISH suggest that the atypical cells are most consistent with reactive fibroblasts/myofibroblasts. Recognition of these atypical fibroblasts/myofibroblasts may help in avoiding the potential pitfall of misdiagnosing them as well-differentiated liposarcoma.

Identification of novel HMGA2 fusion sequences in lipoma: evidence that deletion of let-7 miRNA consensus binding site 1 in the HMGA2 3' UTR is not critical for HMGA2 transcriptional upregulation.

Lipoma is a benign tumor composed of mature adipocytes and one of the most common mesenchymal tumors seen in adults. Rearrangement of HMGA2 in chromosome band 12q15 has been found in approximately 60-70% of ordinary lipomas with cytogenetic abnormalities. Herein, we report two novel HMGA2 fusion sequences in lipomas with chromosome 12 rearrangements. Cytogenetic studies showed 12q abnormalities in both cases, and fluorescence in situ hybridization (FISH) confirmed the involvement of HMGA2 in each instance. Rapid amplification of cDNA ends (RACE) PCR experiments revealed that one lipoma contained a fusion of the HMGA2 3' untranslated region (UTR) to a genomic area downstream of the DYRK2 locus on 12q15; the second lipoma showed a fusion of the HMGA2 3' UTR to a genomic sequence upstream of the DCN locus on 12q21. In both instances the breakpoint on HMGA2 occurred downstream to let-7 miRNA (microRNA) consensus binding site (CBS) 1. These two and several other previously reported tumors containing HMGA2 3' UTR rearrangements show breakpoints after let-7 miRNA CBS 1, which suggests that the elimination of this miRNA binding site is not critical for driving HMGA2 transcriptional upregulation.