Pamela A. Althof

Cytogenetic and molecular cytogenetic findings in 43 aneurysmal bone cysts: aberrations of 17p mapped to 17p13.2 by fluorescence in situ hybridization

Aneurysmal bone cyst is a benign, cystic lesion of bone composed of blood-filled spaces separated by fibrous septa. Relatively few cases of aneurysmal bone cyst have been cytogenetically characterized, yet abnormalities of the short arm of chromosome 17 appear to be recurrent. In this study, conventional cytogenetic analysis of 43 aneurysmal bone cyst specimens from 38 patients over a 12-year period revealed clonal chromosomal abnormalities in 12 specimens. Karyotypic anomalies of 17p, including a complex translocation and inversion, were identified in eight of these 12 specimens. In an effort to further define the aberrant 17p breakpoint, fluorescence in situ hybridization (FISH) analyses were performed using a series of probe combinations spanning a 5.1 Mb region between the TP53 (17p13.1) and Miller–Dieker lissencephaly syndrome (17p13.3) gene loci. These studies revealed the critical breakpoint locus at 17p13.2, flanked proximally by an RP11-46I8, RP11-333E1, and RP11-457I18 bacterial artificial chromosome (BAC) probe cocktail and distally by an RP11-198F11 and RP11-115H24 BAC and RP5-1050D4 P1 artificial chromosome (PAC) probe cocktail. Overall, abnormalities of the 17p13.2 locus were identified by metaphase and/or interphase cell FISH analysis in 22 of 35 (63%) aneurysmal bone cyst specimens examined including 26 karyotypically normal specimens. These cytogenetic and molecular cytogenetic findings expand our knowledge of chromosomal alterations in aneurysmal bone cyst, further localize the critically involved 17p breakpoint, and provide an alternative approach (ie FISH) for detecting 17p abnormalities in nondividing cells of aneurysmal bone cysts. The latter could potentially be utilized as an adjunct in diagnostically challenging cases.

Genome wide copy number analysis of paediatric Burkitt lymphoma using formalin-fixed tissues reveals a subset with gain of chromosome 13q and corresponding miRNA over expression.

The majority of paediatric Burkitt lymphoma (pBL) patients that relapse will die of disease, but markers for this high‐risk subset are unknown. MYC translocations characterize pBL, but additional genetic changes may relate to prognosis and serve as potential biomarkers. We utilized a molecular inversion probe single nucleotide polymorphism assay to perform high resolution, genome‐wide copy number analysis on archival formalin‐fixed, paraffin‐embedded pBL and germline tissues. We identified copy number abnormalities (CNAs) in 18/28 patients (64%) with a total of 62 CNAs that included 32 gains and 30 copy number losses. We identified seven recurrent CNAs including 1q gain (7/28, 25%), 13q gain (3/28, 11%), and 17p loss (4/28, 14%). The minimum common amplified region on 13q was at 13q31 and included the MIR17HG (MIR17‐92) locus. Samples with this gain had higher levels of MIR17 RNA and showed a tendency for early relapse. Tumour‐specific uniparental disomy was identified in 32% of cases and usually was recurrent. These results demonstrate that high‐resolution copy number analysis can be performed on archival lymphoma tissue specimens, which has significance for the study of rare diseases.

B‐cell lymphoma, unclassifiable, with features intermediate between diffuse large B‐cell lymphoma and burkitt lymphoma: study of 39 cases

B‐cell lymphoma, unclassifiable (B‐UCL), with features intermediate between diffuse large B‐cell lymphoma and Burkitt lymphoma, is a poorly characterized entity. Therefore, we investigated cases of B‐UCL treated by the Nebraska Lymphoma Study Group (NLSG). We searched the NLSG registry for years 1985–2010 for cases of B‐UCL. Immunohistochemical stains and fluorescence in situ hybridization studies for MYC, BCL2 and BCL6 gene rearrangements were performed. Among the 39 cases studied, 54% were male and 46% were female, with a median age of 69 years. The majority of patients presented with advanced‐stage disease (62%) and had high (3–5) International Prognostic Index (IPI) scores (54%). The median overall survival (OS) was only 9 months and the 5‐year OS was 30%. Patients with low IPI scores (0–2) had a better survival than those with high scores (3–5). The cases were genetically heterogeneous and included 11 ‘double‐hit’ lymphomas with rearrangements of both MYC and BCL2 or BCL6. None of the immunohistochemical or genetic features was predictive of survival. This B‐cell lymphoma is a morphologically‐recognizable entity with a spectrum of genetic abnormalities. New and better treatments are needed for this aggressive lymphoma.

Mammary analog secretory carcinoma, low-grade salivary duct carcinoma, and mimickers: a comparative study.

Mammary analog secretory carcinoma (MASC) is a recently recognized low-grade salivary carcinoma characterized by a specific ETV6 rearrangement. We describe 14 new MASCs and examine their immunophenotypic and genetic profiles in the context of look-alikes, namely, low-and high-grade salivary duct carcinoma and acinic cell carcinoma. ETV6 rearrangement, and robust expression of mammaglobin and S100, were demonstrated in 11/11, 14/14, and 12/14 MASCs, respectively. All low-grade salivary duct carcinomas coexpressed S100/mammaglobin (6/6); none harbored ETV6 rearrangements (0/5). Given that S100/mammaglobin coexpression and absence of zymogen granules are features of both MASC and low-grade salivary duct carcinoma, these two are best distinguished histologically. The former is predominantly an extraductal neoplasm with bubbly pink cytoplasm, whereas the latter is a distinct intraductal micropapillary and cribriform process. Querying ETV6 gene status may be necessary for difficult cases. No acinic cell carcinoma expressed mammaglobin (0/13) or harbored an ETV6 rearrangement (0/7); only 1/13 acinic cell carcinomas weakly expressed S100. DOG1 expression was limited or absent among all tumor types, except acinic cell carcinoma which expressed DOG1 diffusely in a canalicular pattern. Therefore, histology and immunohistochemistry (mammaglobin, S100, DOG1) suffices in distinguishing acinic cell carcinoma from both MASC and low-grade salivary duct carcinoma. HER2 (ERBB2) amplification was detected in only 1/10 acinic cell carcinomas, but none of the MASCs or low-grade salivary duct carcinomas tested. High-grade salivary duct carcinomas frequently expressed mammaglobin (11/18) and harbored HER2 amplifications (13/15); none harbored ETV6 rearrangements (0/12). High-grade salivary duct carcinomas can easily be distinguished from these other entities by histology and HER2 amplification.

Aberrations of 6q13 mapped to the COL12A1 locus in chondromyxoid fibroma

Chondromyxoid fibroma, a rare benign bone tumor, may be mistaken for chondrosarcoma. Although cytogenetic studies of chondromyxoid fibroma are few, rearrangements of the long arm of chromosome 6, frequently expressed as an inv(6)(p25q13), are prominent. In this study, conventional cytogenetic analysis of 16 chondromyxoid fibroma samples from 14 patients revealed rearrangements of chromosome 6 in 10 of 11 clonally abnormal specimens. In addition to 6q13 rearrangements, recurrent 6p25 and 6q25 anomalies were detected. Notably, an identical t(6;9)(q25;q22) translocation was identified in two cases, suggesting that it represents a distinct translocation of chondromyxoid fibroma. In an effort to further define the aberrant 6q13 breakpoint and identify the molecular consequences, a fluorescence in situ hybridization (FISH)-based positional cloning strategy on chondromyxoid fibroma abnormal metaphase and interphase cells using a series of bacterial and plasmid artificial chromosome (BAC/PAC) probe combinations spanning a 6.1 Mb region was employed. The breakpoint on 6q13 was located within the COL12A1 gene, a collagen gene purportedly involved in another benign bone tumor, subungual exostosis. The findings of this study expand our knowledge of chromosomal alterations in chondromyxoid fibroma, identify COL12A1 as the likely gene candidate within the recurrent 6q13 breakpoint, and provide an alternative approach for detecting 6q13 anomalies in nondividing cells of chondromyxoid fibroma. The latter could potentially be utilized as an adjunct in diagnostically challenging cases.

Immunohistochemical and molecular cytogenetic evaluation of potential targets for tyrosine kinase inhibitors in Langerhans cell histiocytosis.

Langerhans cell histiocytosis is a rare disorder of Langerhans cells, a component of the dendritic cell system, with an unknown pathogenesis. Conventional therapy for patients with Langerhans cell histiocytosis is usually effective, but some patients are refractory to treatment or develop toxicity. Thus, there is a need for innovative therapies. Recently, some cases of Langerhans cell histiocytosis were reported to express platelet-derived growth factor receptors α and β or c-KIT by immunohistochemistry, and some of these patients had a clinical response to imatinib mesylate. Other hematologic disorders with PDGFRα or PDGFRβ gene rearrangements also have responded to imatinib mesylate. The aim of this study was to evaluate immunohistochemical and molecular markers in Langerhans cell histiocytosis that would identify cases for possible treatment with tyrosine kinase inhibitors. We investigated formalin-fixed, paraffin-embedded tissue sections from 14 cases of Langerhans cell histiocytosis. As controls, we included cases of inflammatory dermatitis (n = 5) and dermatopathic lymphadenitis (n = 7). We performed immunohistochemistry for S100, CD1a, c-KIT, and platelet-derived growth factor receptors α and β. Fluorescence in situ hybridization analysis to detect rearrangements of the PDGFRα or PDGFRβ genes was also performed. Four (28.5%) of 14 cases of Langerhans cell histiocytosis were positive for platelet-derived growth factor receptor α, whereas absent/weak expression was seen in 10 cases and all controls. All cases were negative for platelet-derived growth factor receptor β and c-KIT. The fluorescence in situ hybridization studies were also negative in all 8 cases with adequate quality DNA. Our findings suggest that a subset of cases of Langerhans cell histiocytosis may be treated with tyrosine kinase inhibitors due to the expression of platelet-derived growth factor receptor α. Clinical trials that evaluate the use of tyrosine kinase inhibitors in Langerhans cell histiocytosis seem warranted and should evaluate these markers.

Assessing the utility of confirmatory studies following identification of large-scale genomic imbalances by microarray

Purpose:The identification of clinically relevant genomic dosage anomalies assists in accurate diagnosis, prognosis, and medical management of affected individuals. Technological advancements within the field, such as the advent of microarray, have markedly increased the resolution of detection; however, clinical laboratories have maintained conventional techniques for confirmation of genomic imbalances identified by microarray to ensure diagnostic accuracy. In recent years the utility of this confirmatory testing of large-scale aberrations has been questioned but has not been scientifically addressed.Methods:We retrospectively reviewed 519 laboratory cases with genomic imbalances meeting reportable criteria by microarray and subsequently confirmed with a second technology, primarily fluorescence in situ hybridization.Results:All genomic imbalances meeting reportable criteria detected by microarray were confirmed with a second technology. Microarray analysis generated no false-positive results.Conclusion:Confirmatory testing of large-scale genomic imbalances (deletion of ≥150 kb, duplication of ≥500 kb) solely for the purpose of microarray verification may be unwarranted. In some cases, however, adjunct testing is necessary to overcome limitations inherent to microarray. A recommended clinical strategy for adjunct testing following identified genomic imbalances using microarray is detailed.Genet Med 17 11, 875–879.

Breast Cancer and Non-Hodgkin Lymphoma in a Young Male with Cowden Syndrome.

Male breast cancer (MBC) is unusual, especially in young adults. Most cases of MBC as a secondary malignancy relate to the previous treatment with ionizing radiation. MBC can be associated with mutations in hereditary cancer predisposition syndrome genes (i.e., BRCA2); however, no such association has been reported in patients with Cowden syndrome (involving the phosphatase and tensin homolog [PTEN] gene). We describe a patient with Cowden syndrome who was initially diagnosed with B‐cell lymphoblastic lymphoma at the age of 7 years, then MBC at the age of 31 years, and never received radiation therapy.