Rhabdomyosarcoma of the parotid region occurring in childhood and adolescence. A report from the Intergroup Rhabdomyosarcoma Study Group.

BACKGROUND: Rhabdomyosarcoma (RMS) of the parotid region is rare and to the authors' knowledge little information is available regarding the site of tumor origin, clinical presentation, and outcome in these patients. Therefore, the authors reviewed the files of all patients with RMS of the parotid region who were registered on the Intergroup Rhabdomyosarcoma Studies (IRS) I-IV. METHODS: Patient charts and the Intergroup Rhabdomyosarcoma Study Group (IRSG) database were reviewed. RESULTS: Sixty-two patients presenting with a mass in the parotid region were identified. None of the tumors was localized exclusively to the parotid gland, so the primary site was referred to as the "parotid region." The tumor invaded a parameningeal site in 30 patients. These cases have been designated as parameningeal-parotid tumors to distinguish them from 32 cases that did not invade a parameningeal site and were designated as nonparameningeal-parotid tumors. The majority of patients had Group III tumors in both the nonparameningeal-parotid and parameningeal-parotid subgroups. However, although there were 16 patients with Group I or II tumors in the nonparameningeal-parotid subgroup, no patients with Group I or II tumors were found in the parameningeal-parotid subgroup (P = 0.001). Fifty-six of 62 patients (90%) received radiotherapy. The parameningeal primary site designation resulted in intensification of both chemotherapy and radiotherapy for patients with parameningeal-parotid RMS. The 5-year failure-free survival rate was 81% and the 5-year survival rate was 84%. There were no deaths reported among patients with Group I or II tumors. The 5-year failure-free survival did not appear to differ when comparing patients with parameningeal-parotid tumors with patients with nonparameningeal-parotid tumors (P = 0.21). CONCLUSIONS: Treatment as defined by the IRS protocols has been reported to be highly effective for patients with RMS of the parotid region. Outcome for the more aggressively treated patients with parameningeal-parotid RMS appears similar to that for patients with nonparameningeal-parotid RMS.

GLI1 localization in the germinal epithelial cells alternates between cytoplasm and nucleus: upregulation in transgenic mice blocks spermatogenesis in pachytene.

The zinc finger transcription factor GLI1 is the mediator of signaling by members of the Hedgehog (Hh) family. Male mice in which Desert hedgehog (Dhh), an Hh homologue expressed in Sertoli cells of the testis, was knocked out are sterile, suggesting that the Dhh/GLI1 pathway plays a role in spermatogenesis. Using an antiserum raised against human GLI1, we found that during the first round of spermatogenesis, GLI1 expression is initially cytoplasmic, then shifts to the nuclei of Sertoli and germ cells, and finally shifts back to the cytoplasm. In the adult mouse testis, GLI1 expression localized to the nuclei of germ cells, beginning with pachytene cells and persisting through round spermatids. Localization of GLI1 in elongating spermatids shifted from the nucleus to the cytoplasm and became associated with microtubules. We also examined a line of transgenic mice that overexpressed human GLI1. Male mice in this line were sterile. Spermatogenesis was blocked at the pachytene stage, and a subset of the morphologically indistinguishable pachytene cells underwent apoptosis. Patched-2, which is a Dhh receptor, and Fused, another component of the signal transduction pathway, are expressed in Leydig cells and in primary and secondary spermatocytes. Expression of GLI1 in the same cell types as Patched-2 and Fused and the disruption of spermatogenesis by GLI1 overexpression suggest that GLI1 is the mediator of the Dhh signal in the testis, and that it may be a regulator of spermatogenesis.

Detection of EBV DNA in the cord blood donor for a patient developing Epstein-Barr virus-associated lymphoproliferative disorder following mismatched unrelated umbilical cord blood transplantation.

Epstein-Barr virus-associated post-transplant lymphoproliferative disorder (PTLD) has been well described as a complication following allogeneic stem cell transplantation but has only recently been reported following umbilical cord blood (UCB) transplant. We report the case of a child transplanted with unrelated mismatched UCB for juvenile chronic myelogenous leukemia (JCML) who developed EBV-associated PTLD, which was confirmed pathologically, 139 days following stem cell infusion. There was no clinical response to reduction of immune suppression, high-dose acyclovir, or alpha interferon. The patient died 160 days after transplantation. EBV was detected by polymerase chain reaction in the cord blood unit used for transplantation. This case demonstrates that EBV-associated PTLD can occur following mismatched unrelated UCB transplant and may be related to transmission of EBV infection by donor lymphocytes.

Developmental pathways: Sonic hedgehog-Patched-GLI.

Developmental pathways are networks of genes that act coordinately to establish the body plan. Disruptions of genes in one pathway can have effects in related pathways and may result in serious dysmorphogenesis or cancer. Environmental exposures can be associated with poor pregnancy outcomes, including dysmorphic offspring or children with a variety of diseases. An important goal of environmental science should be reduction of these poor outcomes. This will require an understanding of the genes affected by specific exposures and the consequence of alterations in these genes or their products, which in turn will require an understanding of the pathways critical in development. The ligand Sonic hedgehog, the receptors Patched and Smoothened, and the GLI family of transcription factors represent one such pathway. This pathway illustrates several operating principles important in the consideration of developmental consequences of environmental exposures to toxins.

High-dose chemotherapy followed by peripheral blood stem cell rescue for metastatic rhabdomyosarcoma: the experience at Chicago Children's Memorial Hospital.

BACKGROUND: Because outcome for metastatic rhabdomyosarcoma remains poor with standard therapy, and because some patients with extensive unresectable metastatic rhabdomyosarcoma are unable to tolerate standard therapy with the associated large radiation fields, peripheral blood stem cell rescue (PBSCR) following high-dose chemotherapy was offered as consolidative therapy for patients with Stage 4/Group IV rhabdomyosarcoma. PATIENTS AND METHODS: Eight patients with Stage 4/Group IV rhabdomyosarcoma were diagnosed from May, 1992, through November, 1994. Consolidative PBSCR following thiotepa 300 mg/M2 on days -7, -6, and -5; cyclophosphamide 1,500 mg/M2 on days -5, -4, -3, and -2; and carboplatin 600 mg/M2 on days -3 and -2 was offered to those patients who achieved a complete remission with multimodality therapy. Patients with extensive metastatic disease who did not receive full doses of radiation to all sites of disease remained eligible for high-dose chemotherapy and PBSCR. RESULTS: Five of eight patients achieved a complete response. Four patients underwent PBSCR. One of the four patients is alive without evidence of disease 53 months post-PBSCR. All other patients died of progressive disease. CONCLUSIONS: These results, along with the existing literature, show no advantage of high-dose chemotherapy followed by PBSCR as consolidative therapy for patients with Stage 4/Group IV rhabdomyosarcoma over standard dose chemotherapy, radiation, and surgery. For patients with extensive, unresectable disease at diagnosis who cannot receive radiation to all areas of disease based on concerns of marrow reserve, high-dose chemotherapy followed by PBSCR does not appear to provide adequate local control and cannot be offered as curative therapy.

Prolonged survival of a patient with sickle cell trait and metastatic renal medullary carcinoma.

PURPOSE: The treatment and outcome of a patient with sickle cell trait and metastatic renal medullary carcinoma is described. PATIENT AND METHODS: A 12-year-old boy with sickle cell trait had metastatic renal medullary carcinoma. After surgical resection of the primary tumor, he received chemotherapy with methotrexate, vinblastine, doxorubicin, and cisplatin. The carcinoma progressed after a 6-month period of stable disease. At that time, he received chemotherapy including ifosfamide, etoposide, carboplatin, and topotecan. RESULTS: The patient died of progressive disease 15 months from diagnosis. The patient's tumor in this report showed no progression while he was receiving methotrexate, vinblastine, doxorubicin, and cisplatin, but eventually became refractory to these and other cytotoxic agents. CONCLUSION: Renal medullary carcinoma is a highly chemotherapy-resistant tumor. Average survival after diagnosis is 15 weeks; the longest survival reported in the literature is 12 months from diagnosis. The patient in this report survived longer than the previously described patients before dying from progressive disease.

Characterization of the promoter region and genomic organization of GLI, a member of the Sonic hedgehog-Patched signaling pathway.

GLI is the prototype for the Gli-Kruppel gene family characterized by a consensus C2-H2 zinc finger domain and is believed to function as a transcription activator in the vertebrate Sonic hedgehog-Patched signal transduction pathway. Understanding GLI gene regulation may be of importance to understanding causes of human birth defects and cancer. To begin to understand the regulation of this developmentally important gene we have cloned the human GLI gene and functionally characterized its 5' flanking region. The GLI gene is composed of 12 exons and 11 introns and in the zinc finger coding region shares a highly conserved splicing pattern with several other Gli family members in both vertebrates and C. elegans. A major transcription initiation site was identified upstream of the GLI translation start site along with three minor transcription initiation sites. The region surrounding the transcription initiation sites lacks TATA and CCAAT consensus sequences, has a high GC content, includes a CpG island, and contains several GC boxes. A 487bp segment surrounding the transcription initiation sites increased expression of a luciferase reporter gene 15-fold in Tera-1 cells and was defined as the core promoter region of human GLI. In transgenic mice this region directed beta-galactosidase expression to the central nervous system on embryonic days 10.5-12.5 and to sites of endochondral ossification on embryonic days 12.5 and 13.5 in a pattern comparable to the endogenous expression pattern of mouse gli within these tissues. The previously identified gastrointestinal expression of gli was not driven by this region and may require elements outside of the core promoter. Sequence analysis of the 5' flanking region of the mouse gli gene and the full-length mouse gli cDNA demonstrated high homology with human GLI, suggesting conservation of GLI regulation and function.

GLI activates transcription through a herpes simplex viral protein 16-like activation domain.

Three proteins have been identified in mammals, GLI, GLI2, and GLI3, which share a highly conserved zinc finger domain with Drosophila Cubitus interruptus and are believed to function as transcription factors in the vertebrate Sonic hedgehog-Patched signaling pathway. To understand the role GLI plays in the Sonic hedgehog-Patched pathway and mechanisms of GLI-induced transcriptional regulation, we have characterized its transcriptional regulatory properties and contributions of specific domains to transcriptional regulation. We have demonstrated that GLI activates expression of reporter constructs in HeLa cells in a concentration-dependent manner through the GLI consensus binding motif and that a GAL4 binding domain-GLI fusion protein activates reporter expression through the GAL4 DNA binding site. GLI-induced transcriptional activation requires the carboxyl-terminal amino acids 1020-1091, which includes an 18-amino acid region highly similar to the alpha-helical herpes simplex viral protein 16 activation domain, including the consensus recognition element for the human TFIID TATA box-binding protein-associated factor TAFII31 and conservation of all three amino acid residues believed to contact directly chemically complementary residues in TAFII31. The presence of this region in the GLI activation domain provides a mechanism for GLI-induced transcriptional regulation.

Hemangiopericytoma of the liver: immunohistochemical observations, expression of angiogenic factors, and review of the literature.

PURPOSE: We describe an unusual case of a hemangiopericytoma in the liver of a child, review the literature, and characterize the tumor by immunohistochemistry and electron microscopy. We study the expression of basic fibroblast growth factor (bFGF) and of vascular endothelial growth factor (VEGF) in the tumor. MATERIALS AND METHODS: Clinical history and pathology were reviewed; sections of the tumor were studied by histology, electron microscopy, and immunohistochemistry using antibodies directed towards factor-XIIIa, HAM-56, bFGF and VEGF, among others. RESULTS: The expression of VEGF resembled that of "proliferating" hemangiomas; however, despite being markedly elevated in the urine, bFGF could not be unequivocally detected in the tumor. A subpopulation of factor XIIIa positive cells was identified, similar to the "interstitial" cells of the cellular hemangiomas of infancy. The nature and function of these cells remains speculative. CONCLUSIONS: Hemangiopericytomas are rare in the liver. When arising in this location in a child, they may clinically resemble a hemangioma, may express angiogenic factors in a similar fashion, and should be considered in the differential diagnosis.

Conservation of the C.elegans tra-2 3'UTR translational control.

The Caenorhabditis elegans sex-determination gene, tra-2, is translationally regulated by two 28 nt elements (DREs) located in the 3'UTR that bind a factor called DRF. This regulation requires the laf-1 gene activity. We demonstrate that the nematode Caenorhabditis briggsae tra-2 gene and the human oncogene GLI are translationally regulated by elements that are functionally equivalent to DREs. Here, we rename the DREs to TGEs (tra-2 and GLI elements). Similarly to the C.elegans tra-2 TGEs, the C.briggsae tra-2 and GLI TGEs repress translation of a reporter transgene in a laf-1 dependent manner. Furthermore, they regulate poly(A) tail length and bind DRF. We also find that the C.elegans TGEs control translation and poly(A) tail length in C.briggsae and rodent cells. Moreover, these same organisms contain a factor that specifically associates with the C.elegans TGEs. These findings are consistent with the TGE control being present in C.briggsae and rodent cells. Three lines of evidence indicate that C.briggsae tra-2 and GLI are translationally controlled in vivo by TGEs. First, like C.elegans tra-2 TGEs, the C.briggsae tra-2 and GLI TGEs control translation and poly(A) tail lengths in C.briggsae and rodent cells, respectively. Second, the same factor in C.briggsae and mammalian cells that binds to the C.elegans tra-2 TGEs binds the C.briggsae tra-2 and GLI TGEs. Third, deletion of the GLI TGE increases GLI's ability to transform cells. These findings suggest that TGE control is conserved and regulates the expression of other mRNAs.

Expression of human GLI in mice results in failure to thrive, early death, and patchy Hirschsprung-like gastrointestinal dilatation.

BACKGROUND: GLI is an oncodevelopmental gene in the vertebrate hedgehog/patched signaling pathway that is spatiotemporally regulated during development and is amplified in a subset of human cancers. GLI is the prototype for the Gli-Kruppel family of transcription factors, which includes the Drosophila segment polarity gene ci, the C. elegans sex-determining gene tra-1, and human and mouse GLI3, all of which contain a conserved domain of five C2-H2 zinc fingers. GLI3 mutations have been implicated in the mouse mutant extra toes, as well as in human Greig cephalopolydactaly syndrome and the autosomal dominant form of Pallister-Hall syndrome. As such, GLI and the vertebrate hedgehog/patched signaling pathway appear to play important roles in both normal development and neoplasia. MATERIALS AND METHODS: Since it is not known whether aberrant GLI expression is similarly linked to developmental disorders, we developed gain-of-function transgenic mice which express human GLI ectopically. RESULTS: Affected transgenic mice exhibit a phenotype of failure to thrive, early death, and Hirschsprung-like patches of gastrointestinal dilatation. The colons of affected mice have greatly attenuated smooth muscle layers and abnormal overlying epithelium. The density of myenteric plexuses is reduced in the colonic walls. The severity of the phenotype is related to the level of transgene expression. CONCLUSIONS: The transgenic mouse model supports a role for GLI in gastrointestinal development. As part of the vertebrate hedgehog/patched signaling pathway, GLI is essential to mesoderm and CNS ectoderm development and transgenic GLI expression affects neuronal, muscular, and epithelial cell differentiation in the gut. Expression of human GLI in mice results in impairment of enteric neuronal development and a Hirschsprung-like phenotype.

Management of an acute hemorrhagic ovarian cyst in a female patient with hemophilia A.

PURPOSE: This case report represents a description of a hemorrhagic ovarian cyst in a patient with hemophilia A. PATIENT: An 18-year-old female patient with severe factor VIII deficiency presented with the acute onset of a hemorrhagic ovarian cyst, meeting the standard indications for surgical intervention. Instead, because of the underlying coagulopathy, factor VIII therapy was instituted. RESULT: Within 1 day of starting the factor VIII therapy, the hemorrhage had clinically resolved and surgical intervention was avoided. CONCLUSIONS: Hemorrhagic ovarian cysts in the setting of hemophilia may respond to factor VIII replacement therapy without the need for surgical intervention. Because the incidence of functional ovarian cysts is high in the general population, female patients with hemophilia should be counseled regarding its possibility of occurrence. Moreover, if a female patient with hemophilia displays a propensity toward the development of ovarian cysts, the administration of prophylactic oral contraceptive pills should strongly be considered.

A soluble form of the human Fc receptor Fc gamma RIIA: cloning, transcript analysis and detection.

The Fc gamma receptors (Fc gamma R) are glycoproteins that bind the Fc region of immunoglobulin G. Human hematopoietic cells express three biochemically distinct classes of Fc gamma receptors: Fc gamma RI (CD64), Fc gamma RII (CD32) and Fc gamma RIII (CD16). Complementary DNA (cDNA) clones for each of the human Fc gamma receptors have been isolated from myeloid and lymphoid cells. We describe the isolation and characterization of four Fc gamma RII clones from a cDNA library obtained from a megakaryocyte-like cell line, human erythroleukemia (HEL). Three clones encode the Fc gamma RIIA transmembrane (TM) form, while one novel clone lacks the TM region but retains the cytoplasmic domain. By conducting reverse transcription coupled to polymerase chain reaction (PCR), we found transcripts coding for this unique form of receptor in RNA from platelets, HEL cells and a second megakaryocyte-like cell line, CHRF-288-11. These results were confirmed by RNase protection analysis of RNA from HEL cells. The structure of the novel cDNA suggested that it codes for a soluble form of Fc gamma RIIA. A soluble Fc gamma RII protein was detected in the conditioned medium from HEL cells but not from the Fc gamma RII-negative T cell line, Jurkat, by immunoprecipitation with the anti-Fc gamma RII monoclonal antibody (mAb), IV.3. The immunoprecipitated protein was of the expected size for a soluble Fc gamma RII lacking the TM region but retaining the cytoplasmic domain. Soluble Fc gamma RIIA may be important in modulating the interaction between immune complexes and membrane-associated Fc gamma RII.

gli, a zinc finger transcription factor and oncogene, is expressed during normal mouse development.

The oncogene GLI is amplified and expressed in some cases of human malignant glioma and undifferentiated childhood sarcoma and is the prototype for a gene family characterized by a highly conserved set of five tandem zinc fingers and a consensus cysteine-histidine link. This zinc finger motif has been shown to bind DNA with sequence specificity and may mediate transcriptional regulation. Since GLI is expressed in embryonal carcinoma cell lines but not in most normal adult tissues and shows significant sequence similarity within its zinc finger domain to cubitus interruptus dominant (ciD), a Drosophila segmentation gene known to be important in the morphogenesis of the posterior portion of each larval segment, we established the temporal and tissue expression patterns of the mouse homologue of human GLI in day 10 through 18 mouse embryos with Northern blotting, reverse transcriptase coupled PCR, and in situ hybridization. gli transcripts were demonstrated on days 10 through 18 of mouse embryonic development as well as in normal adult uterus, brain, testis, and limb. Tissue expression of gli during gestation was demonstrated in Meckel's precartilage mesenchyme, the basis occipitus, rib mesenchymal condensations, primordial vertebral bodies, digital mesenchymal condensations in forefoot and hindfoot plates, the ependymal layer of the spinal cord, and the mesoderm of the gastrointestinal tract. Expression persisted throughout gestation in developing bone and cartilage of the extremities, the ribs, and the vertebral bodies, as well as the gastrointestinal tract mesoderm. These findings support a role for gli family genes in normal craniofacial and digital development in mammals first suggested by the demonstration of translocation breakpoints within the GLI3 gene in families with the Greig cephalopolysyndactylyl syndrome and subsequently by reduced gli3 expression in the mouse mutant extra toes. It is surprising that a single gene would be expressed in such a wide range of mesenchymal structures.

Aortic stiffness in the DOCA-hypertensive pig.

Pulsatile changes in aortic diameter and pressure were monitored in order to obtain an index of aortic stiffness in the conscious pig during the development of mineralocorticoid hypertension. The aortic stiffness index increased but to a much lesser extent than it did in response to a similar pressure elevation produced by phenylephrine infusion. This observation suggests that chronic exposure to an elevated pulsatile pressure reduces the stiffness index at the elevated pressure.