Neutrophil-rich anaplastic large cell lymphoma of the skull presenting after head trauma.

The presentation of anaplastic large cell lymphoma in bone is uncommon. We report a case of anaplastic large cell lymphoma of the skull that was diagnosed after head trauma. Biopsy revealed significant destruction of the outer table of the frontal bone. Histopathologically, the initial evaluation suggested osteomyelitis because of a mixed inflammatory infiltrate with large numbers of neutrophils. However, several clusters and individual mononuclear cells were atypical. The tumor cells had large, pleomorphic nuclei; these cells stained positively with antibodies to Ki-1 (CD 30), ALK-1, and EMA. Fluorescence in situ hybridization (FISH) showed rearrangement of the ALK gene, which usually results from the t(2;5) translocation, present in most anaplastic large cell lymphomas. There was no evidence of systemic disease. The patient has tolerated chemotherapy and is free of disease 12 months later.

Neutrophil-rich anaplastic large cell lymphoma of T-cell lineage. A report of two cases arising in HIV-positive patients.

Neutrophil-rich anaplastic large cell lymphoma (ALCL) is an uncommon morphologic variant of ALCL. We report 2 cases of neutrophil-rich T-cell ALCL that presented as scalp masses in HIV-positive men. Histologically, the neoplastic cells extensively infiltrated the dermis and subcutaneous tissue. The neoplastic cells strongly expressed CD30 and were of T-cell lineage, positive for CD3 and CD45RO, and negative for CD20. The neoplastic cells were negative for anaplastic lymphoma kinase-1. Numerous admixed neutrophils also were present, representing up to 70% of all cells in some microscopic fields. Neither patient had peripheral blood leukocytosis. One patient had relative neutrophilia, 79% (0.79; reference range, 50%-70% [0.50-0.70]). The absolute CD4 counts were 160 cells/microL (160 x 10(6)/L) and 150 cells/microL (150 x 10(6)/L), respectively (reference range, 431-1,623/microL [431-1,623 x 10(6)/L]). Both patients were treated with multiagent chemotherapy but died of Pneumocystis carinii pneumonia within 6 months of diagnosis. In our review of the literature, we identified 5 similar T-cell cases, including 1 in an HIV-positive patient. Neutrophil-rich T-cell ALCL is a rare morphologic variant of ALCL that should be considered in the histologic evaluation of neutrophil-rich biopsy specimens.

Oncocytic mucoepidermoid carcinoma of the trachea.

We report a rare case of an oncocytic mucoepidermoid carcinoma of the trachea, which presented in a 78-year-old woman with hemoptysis. Oncocytic cells comprised the majority of this low-grade lesion and demonstrated granular cytoplasmic phosphotungstic acid-hematoxylin staining as well as strong immunohistochemical reactivity to antimitochondrial antibody. Most tracheobronchial tumors with oncocytic change are carcinoid tumors. To our knowledge, this is the first oncocytic mucoepidermoid carcinoma of the trachea reported. This diagnosis was facilitated by histochemical and immunohistochemical studies.

Documentation of EWS gene rearrangements by fluorescence in-situ hybridization (FISH) in frozen sections of Ewing's sarcoma-peripheral primitive neuroectodermal tumor.

Prompt and accurate diagnosis of small round cell tumors warrants ancillary studies. Recently, two-color fluorescence in-situ hybridization (FISH) using probes for specific gene rearrangements has gained wide acceptance. EWS gene rearrangements, present in essentially 100% of Ewing's Sarcoma/peripheral primitive neuroectodermal tumor, were evaluated by FISH on frozen sections (FS) of tumor biopsies from 10 patients, plus a negative control, and in seven other malignant neoplasms of childhood. 4mu FS were hybridized overnight, using a single EWS gene-specific probe spanning the EWS breakpoint. We identified EWS rearrangements in 8 of 10 cases (80%) of Ewing's Sarcoma/pPNET. There are no known false positives in diploid or near-diploid tumors, or in any of the non-EWS tumors tested; the uncommon false negative can be confirmed by RT-PCR. Hyperdiploid cases with multiple copies of chromosome 22 may be better evaluated by two-color FISH. This is the first use on FS biopsy material of a single probe for EWS, capable of detecting all known EWS rearrangements, in ES and other tumors. Utilization of this ancillary technique on FS for ES/pPNET and other tumors with distinctive chromosomal translocation is highly specific, reliable, expeditious (24-36 hours) and cost-effective.

Posttransplant lymphoproliferative disorder in pediatric bone marrow transplant recipients: disseminated disease of donor origin demonstrated by fluorescence in situ hybridization.

BACKGROUND: Posttransplant lymphoproliferative disorders in bone marrow transplantation are typically rapidly progressive and fatal B-cell lymphoid proliferations associated with Epstein-Barr virus, and are mostly of donor origin. We report three pediatric bone marrow transplant cases in which posttransplant lymphoproliferative disorder was diagnosed at postmortem examination. Epstein-Barr virus in these cases was identified by a combined in situ hybridization-immunoperoxidase technique and donor origin was identified by fluorescence in situ hybridization. METHODS: Tissues obtained from postmortem examination were evaluated by light microscopy, immunohistochemistry, combined in situ hybridization-immunoperoxidase technique with Epstein-Barr virus-encoded RNA probe, and fluorescence in situ hybridization with X and Y centromeric probes. RESULTS: Three pediatric patients underwent sex-mismatched, T-cell-depleted bone marrow transplants complicated by graft versus host disease, rapidly progressive multiple organ failure, and postmortem diagnosis of posttransplant lymphoproliferative disorder. Histologic examination and immunohistochemistry studies demonstrated immunoblastic lymphoma (one case) or polymorphic B-cell lymphoma (two cases). In all cases, Epstein-Barr virus-encoded RNA was detected by a combined in situ hybridization-immunoperoxidase technique. Fluorescence in situ hybridization for X and Y chromosomes in paraffin sections demonstrated donor origin in two cases (one case was indeterminate). CONCLUSION: Fluorescence in situ hybridization was used to prove donor derivation of Epstein-Barr virus-associated posttransplant lymphoproliferative disorders in pediatric bone marrow transplant recipients. Many features of posttransplant lymphoproliferative disorders in pediatric bone marrow transplant recipients are very similar to adult cases, although a higher proportion of children appear to be diagnosed postmortem and have a fatal outcome.

Absence of detectable EWS/FLI1 expression after therapy-induced neural differentiation in Ewing sarcoma.

Ewing sarcoma and other peripheral primitive neuroectodermal tumors (pPNETs) display limited neural differentiation and are thought to have a neural crest origin Greater than 95% of these tumors share common t(11;22)(q24;q12) ort(21;22)(q22;q12) chromosomal translocations leading to ES/FLI1 or EWS/ERG gene fusions, respectively. The resulting chimeric oncoproteins seem to function as aberrant transcription factors. However, whether these molecules contribute to the limited neural differentiation observed in pPNETs or actually inhibit differentiation remains unclear. We report a Ewing sarcoma case from the forearm of a 10-year-old girl which expressed EWS/FLI1 fusion transcripts. The tumor was treated with surgery, chemotherapy, and local radiation, but residual tumor was detected within a year as a well-differentiated peripheral neural tumor lacking detectable EWS/FLI1 expression. Further studies suggested that the primary and residual tumors were clonally related. This association between apparent therapy-induced differentiation in Ewing sarcoma and absence of detectable fusion transcripts in the residual tumor provides presumptive evidence that EWS/FLI1 expression may inhibit differentiation in tumour cells.

Donor origin of posttransplant lymphoproliferative disorder localized to a liver allograft: demonstration by fluorescence in situ hybridization.

SETTING: Posttransplant lymphoproliferative disorders in solid organ transplantation are mostly of recipient origin. We report an unusual case of posttransplant lymphoproliferative disorder following liver transplantation with localized limited involvement of the solid organ allograft. DESIGN: Tissues were obtained at the time of surgery and evaluated by immunohistochemistry, in situ hybridization, and fluorescence in situ hybridization with chromosome X and Y centromeric probes. PATIENT: A 53-year-old Hispanic man with hepatic failure due to hepatitis C virus who underwent orthotopic liver transplant from a female donor and developed posttransplant lymphoma in the transplanted liver. INTERVENTION: Withdrawal of immunosuppression, resection of liver allograft, and second transplant. RESULTS: This posttransplant lymphoproliferative disorder was clearly shown to be derived from Epstein-Barr virus-infected donor lymphoid cells. This was demonstrated by fluorescence in situ hybridization for X and Y chromosomes in paraffin sections in a sex-mismatched transplant. Despite aggressive histology (monoclonal B-cell immunoblastic lymphoma) and lack of response to withdrawal of immunosuppression, the posttransplant lymphoproliferative disorder was successfully managed by repeat liver transplantation without recurrence. CONCLUSION: Fluorescence in situ hybridization was used to prove donor derivation in a posttransplant lymphoma of the liver. Allograft-localized donor posttransplant lymphoproliferative disorder may represent a unique category with more favorable prognosis requiring different clinical management from other cases.

Molecular alterations of the RB1, TP53, and MDM2 genes in primary and xenografted human osteosarcomas.

We report the status of the RB1, TP53, and MDM2 genes in human osteosarcomas and cell lines established from surgical specimens and transplanted into athymic naked mice. By using reverse transcriptase-polymerase chain reaction (RT-PCR) as a prescreening technique and posterior sequencing, we observe new mutations in the RB1 gene, notably a duplication in tandem of exons 3 through 6. TP53 mutations appear in codons most frequently mutated in osteosarcomas. We have not seen MDM2 gene amplification in any reported case. These molecular alterations appear in different osteosarcomas not simultaneously present in the same tumor sample. A link has been described between these three genes in the pathways that control the cell cycle and the tumoral progression, but their functions are probably independent in the development of osteosarcomas. TP53 mutations appear in adult patients, whereas RB1 alterations occur mostly in younger patients.

Molecular genetics of small round cell tumors.

The small round cell tumors of children and young adults constitute part of a group of undifferentiated tumors, the precise diagnosis of which is often a challenge for the pathologist because of their uniform morphological appearance. Diagnostic cytogenetic analysis and identification of specific chromosomal abnormalities have been especially useful for the classification of some of these tumors. The cloning of the genes and molecular characterization of the associated genetic anomalies have led to the discovery of the mechanisms involved in their neoplastic transformation and identified a variety of tumor-specific molecular genetic markers. Information has also been provided regarding the histogenetic origin and mechanisms of differentiation in these tumors. This review focuses on the tumor-specific genetic markers, particularly those of clinical relevance, and the recently identified genes and deregulation mechanisms associated with them. The availability of these markers provides auxiliary methods with increasingly improved resolution for primary diagnosis and classification of histologically similar tumors and tools for monitoring patients and identifying potential antineoplastic therapy targets.

Olfactory neuroblastoma is a peripheral primitive neuroectodermal tumor related to Ewing sarcoma.

Olfactory neuroblastoma (ONB) is a malignant tumor of the nasal mucosa whose histogenesis is unclear. A relationship to neuroblastoma (NB), a pediatric tumor of the sympathetic nervous system, is based on morphologic similarities and the expression of similar neural antigens. However, the clinical presentation of ONB differs from that of NB, and MYCN amplification characteristic of NB is not observed. We have therefore examined the relationship of this malignancy to other classes of neural tumors. In previous studies, two ONB cell lines demonstrated cytogenetic features and patterns of protooncogene expression suggestive of a relationship to the Ewing sarcoma family of childhood peripheral primitive neuroectodermal tumors (pPNETs). The pPNETs show t(11;22)(q24;q12) or t(21;22)(q22;q12) chromosomal translocations fusing the EWS gene from 22q12 with either the FL11 gene on 11q24 or the ERG gene on 21q22. We therefore analyzed ONBs for the presence of pPNET-associated gene fusions. Both cell lines showed rearrangement of the EWS gene, and fluorescence in situ hybridization (FISH) of each case demonstrated fusion of EWS and FL11 genomic sequences. Moreover, both lines expressed EWS/FL11 fusion transcripts with in-frame junctions between exon 7 of EWS and exon 6 of FL11 as described for pPNETs. We identified similar gene fusions in four of six primary ONB cases. None of the cases expressed tyrosine hydroxylase, a catecholamine biosynthetic enzyme widely expressed in NB. Our studies indicate that ONB is not a NB but is a member of the pPNET family.

Promoters containing ATF-binding sites are de-regulated in cells that express the EWS/ATF1 oncogene.

Chromosomal translocations that fuse the N-terminal region of the Ewings sarcoma oncogene (EWS) to the C-terminal region (including the DNA-binding domain) of the cellular transcription factor ATF1 are associated with a tumour type termed malignant melanoma of soft parts (MMSP). It is envisioned that transformation by the EWS/ATF1 fusion protein results from aberrant transcriptional regulation of genes that are normally regulated by ATF1. To examine this hypothesis we have expressed exogenous EWS-ATF1 in JEG3 cells and tested its ability to activate several promoters that contain binding sites for ATF1. We show that EWS-ATF1 is a strong constitutive activator of some promoters tested but represses others. Significantly, the ability of particular promoters to be activated by EWS/ATF1 in JEG3 cells correlates with promoter activity in two MMSP-derived cell lines (SU-CCS-1 and DTC1). Our results therefore provide evidence that endogenous EWS/ATF1 can de-regulate transcription and that this capacity may contribute to transformation in MMSP.

A second Ewing's sarcoma translocation, t(21;22), fuses the EWS gene to another ETS-family transcription factor, ERG.

The t(11;22)(q24;q12), present in 85% of Ewing's sarcoma and related tumours, fuses the EWS gene from chromosome 22q12 and the ETS family member, FLI-1. This results in the expression of a chimaeric protein containing the amino-terminal portion of EWS fused to the ETS DNA-binding domain of FLI-1. We have identified a second Ewing's sarcoma translocation, t(21;22)(q22;q12), that fuses EWS to a different ETS family member, the ERG gene located on band 21q22. Identical EWS nucleotide sequences found in the EWS/FLI-1 fusion transcripts are fused to portions of ERG encoding an ETS DNA-binding domain resulting in expression of a hybrid EWS/ERG protein. These findings suggest that fusion of EWS to different members of the ETS family of transcription factor genes may result in the expression of similar disease phenotypes.