Outcome of hepatoblastoma treated with the JPLT-1 (Japanese Study Group for Pediatric Liver Tumor) Protocol-1: A report from the Japanese Study Group for Pediatric Liver Tumor.

PURPOSE: Hepatoblastoma is the most common malignant liver tumor in childhood. Multicenter studies elucidate the optimal pre- or postoperative chemotherapeutic regimens. This report reviews the results of the Japanese Study Group for Pediatric Liver Tumor Protocol-1 (JPLT-1) and compares its outcomes with published reports of other studies. METHODS: From March 1991 to December 1999, 154 patients with malignant liver tumor including 145 cases of hepatoblastomas were enrolled in the JPLT study. Data from 134 cases were analyzed in this study. JPLT-1 protocol 91A was used for patients with stage I or II hepatoblastoma. The chemotherapy regimen consisted of repeated courses of cisplatin (CDDP), 40 mg/m(2), and tetrahydropyranyl (THP)-Adriamycin, 30 mg/m(2). JPLT-1 protocol 91B was administered to patients with stage IIIA, IIIB, or IV hepatoblastoma. The chemotherapy regimen consisted of repeated courses of CDDP, 80 mg/m(2), and THP-Adriamycin, 30 mg/m(2)/day for 2 days. Courses were repeated every 4 weeks as tolerated. RESULTS: Seven patients died of chemotherapy-related side effects. Six of them died of sepsis caused by leukopenia and 1 case of liver failure. Overall survival rate (3-year/6-year) was 100%/100% for stage I (n = 9), 100%/95.7% for stage II (n = 32), 76.6%/73.8% for stage IIIA (n = 48), 50.3%/50.3% for stage IIIB (n = 25), 64.8%/38.9% for stage IV (n = 20), and 77.8%/73.4% overall. For stage IIIA and B disease, intravenous chemotherapy was better than intraarterial chemotherapy (66.4% v 38.1% for event-free survival and 69.3% v. 57.1% for overall survival). Patients less than 1 year of age had a better prognosis than older patients, but age was not a significant prognostic factor by multivariate analysis. CONCLUSIONS: The overall and event-free survival rates of the JPLT-1 study of hepatoblastoma were comparable with the results of other multicenter studies in Europe and the United States. The event-free survival rate at 3 years for stage IIIB and IV disease was under 50%. New treatment strategies are needed for patients with advanced hepatoblastoma.

Ewing's sarcoma family of tumor arising in the adrenal gland--possible diagnostic pitfall in pediatric pathology: histologic, immunohistochemical, ultrastructural, and molecular study.

We present an adrenal Ewing's sarcoma family of tumor (ESFT) arising in an 11-year-old Japanese boy. Although intensive chemoradiotherapy and radical surgery were performed, the patient died of obstinate disease 1 year and 3 months after the initial presentation. The primary site (adrenal gland) with radiologic findings (with foci of calcification), high titer of serum neuron specific enolase, and sheets of monotonous primitive rounded cells on histology mostly favored neuroblastoma. However, a diagnosis of ESFT was confirmed by immunohistochemical profile, including MIC2-positivity and molecular study disclosing EWS-FLI1 chimera gene verified by direct sequencing. Recognition of adrenal ESFT and use of newly developed diagnostic techniques are required for differential diagnosis of undifferentiated small round cell tumor of the adrenal gland.

Clinicopathological study of a hilar nodule in the livers of long-term survivors with biliary atresia.

With the application of liver transplantation for patients with biliary atresia (BA), we have had the opportunity to review the clinicopathologic features of the native livers from 10 transplanted BA patients. A single large nodule at porta hepatis (hilar nodule) was noted in three of 10 patients, and an ill-defined nodule-like lesion at porta hepatis was present in two other patients. The three BA patients with hilar nodules were long-term survivors, compared to the patients with nodule-like and those without nodules. The hilar nodules measured between 5.0 cm and 8.0 cm and histologically, they were partly surrounded by fibrous septa with relatively well-preserved liver architectures and fewer inflammatory cells at the portal triads when compared to the surrounding cirrhotic lesions. No nuclear or cellular atypia was observed. Proliferating cell nuclear antigen labeling index was higher in the surrounding cirrhotic lesions than the hilar nodules. The nodule-like lesions at porta hepatis also showed similar light microscopic and immunohistochemical features as the hilar nodules. These hilar nodules did not seem to contain any malignant potential. The benign histology with relatively well-preserved liver architecture and the preferential site of occurrence at porta hepatis where bile seemed to flow more smoothly, suggested possible residues of less-affected hepatic tissues.

HMB-45/melan-A and smooth muscle actin-positive clear-cell epithelioid tumor arising in the ligamentum teres hepatis: additional example of clear cell 'sugar' tumors.

HMB-45-positive clear-cell epithelioid tumor arising in the ligamentum teres hepatis of a 13-year-old Japanese girl is described. The well-defined tumor was completely removed and measured 9 x 7 x 6 cm. Cut sections showed a tan-white, homogeneous appearances with no hemorrhage or necrosis. The tumor was composed of nests or sheets of polygonal or oval-shaped cells rich in clear or finely granular cytoplasm. Capillary network was well developed, and sinusoid vessels were often seen with occasional perivascular hyalinization. There was moderate nuclear atypia but mitotic figures were absent. Periodic acid-Schiff stain showed a large amount of glycogen digested by diastase. Immunohistochemical stains for smooth muscle actin, Melan-A, and HMB-45 were positive in most of the tumor cells. Stains for vimentin, muscle actin, and HAM56 were focally positive, whereas stains for desmin, cytokeratin, epithelial membrane antigen, S-100, CD34, CD68, CD99, neurofilament proteins, and estrogen/progesterone receptors were negative. Ultrastructurally, the cytoplasm contained a considerable number of mitochondria, monoparticipate or membrane-bound glycogen, and longitudinally oriented thin filaments with focal condensations and subplasmalemmal densities. The histopathology of the present case, originally interpreted as epithelioid leiomyoma, was consistent with clear cell "sugar" tumors. The present case may indicate ubiquitous distribution of clear cell "sugar tumors" of which histogenesis remains unknown but is presumed to be of perivascular epithelioid cell origin.

Clinicopathologic study of mass-screened neuroblastoma with special emphasis on untreated observed cases: a possible histologic clue to tumor regression.

Spontaneous regression and maturation of neuroblastoma (NB) are well documented and occur frequently in infants, including those detected by mass screening. To seek histologic clues for regression/maturation in mass-screened NB, clinicopathologic features of 12 tumors that were resected after 2 to 18 months of untreated observation were reviewed. Unobserved screened and age-matched unscreened patients were also studied. To evaluate the possible important role of apoptosis, apoptotic cells were detected by in situ deoxyribonucleic acid (DNA) nick end labeling and immunohistochemical stain for activated caspase-3. Nests with a varying degree of reduced cellularity ("less cellular" and "hypocellular" nests) were common in patients younger than 18 months of age, and were rare in older patients. Two characteristic cells, which have not been focused previously, were frequent, especially in the hypocellular nests. One showed amorphic eosinophilic cytoplasm with pyknotic nuclei and the other contained plump cytoplasm with well-maintained nuclei. These cells were also observed in 89% of the unobserved screened NBs and 79% of the age-matched unscreened patients with good outcome, whereas they could not be confirmed in any of the age-matched unscreened NBs with poor outcome. The amorphic and plump cells were negative for activated caspase-3 and in situ DNA nick end labeling. From these results, the authors hypothesize that these cells most likely represent a degenerative process, in either a state before the activation of caspase-3 or a caspase-independent form of cell death. The presence of less cellular and hypocellular nests with amorphic/plump cells may serve as one of the important clues in predicting tumor prognosis.

Congenital primitive epithelial tumor of the liver showing focal rhabdoid features, placental involvement, and clinical features mimicking multifocal hemangioma or stage 4S neuroblastoma.

We describe an unusual case of congenital primitive epithelial tumor of the liver with focal rhabdoid features. The present case is unique and informative in the following aspects: (1) a first case of congenital epithelial tumor of the liver with no hepatocytic differentiation but focal rhabdoid features, (2) clinical similarities to multicentric hemangioma or stage 4S neuroblastoma, (3) diagnosis obtained from histological examination of the placenta immediately after birth.

Ossifying fibromyxoid tumor of soft parts in a child: a case report.

Ossifying fibromyxoid tumor (OFMT) is a relatively rare tumor, most of which occur in adults with preferential localization in subcutis or muscles of the extremeties. Although Schwannian or cartilage origin has been suggested, histogenesis of this tumor still is unclear. The authors present a 6-year-old girl with retroperitoneal OFMT showing paraspinal extension, who is alive and tumor free 9 years after excision. The current case is the youngest reported patient showing unusual deep trunk site with surgically identified association with the spinal nerve. Both the clinical and histopathologic features strongly suggested Schwannian origin of this tumor.

Chromophobe renal cell carcinoma: an immunohistochemical study of 21 Japanese cases.

Chromophobe renal cell carcinoma (RCC) is a newly established category of RCC composed histologically of characteristic "chromophobe" tumor cells. Although ultrastructural and immunohistochemical studies showed that these tumor cells present several features similar to those found in the intercalated cells of the collecting duct, immunohistochemical studies using antibody panels on a large number of cases are limited. We performed an immunohistochemical study of 21 Japanese cases of chromophobe RCC, along with cases of clear RCC and renal oncocytoma, to find hallmarks useful for precise differential diagnosis of these tumors. Chromophobe RCC was positive for epithelial membrane antigen but negative for vimentin. Cytokeratins did not show constant immunoreactivity in the three types of renal tumors. Furthermore, all of the chromophobe RCCs and renal oncocytomas were positive for E-cadherin but not for N-cadherin, whereas all of the clear RCCs were negative for E-cadherin, and 58% were positive for N-cadherin. The Ki-67 labeling indices were significantly lower in cases classified as (pT1) or Grade 2 chromophobe RCC than in cases of clear RCC. Immunoreaction for E-cadherin was demonstrated to be useful for distinguishing chromophobe RCC from clear RCC, and a low Ki-67 labeling index might indicate a favorable prognosis, as reported in several previous studies.

Segmental mediolytic arteritis [correction of arteries]: a case report with review of the literature.

Segmental mediolytic arteritis is a very rare vascular disease which causes sudden intraabdominal hemorrhage. The disease is characterized by degeneration of the arterial media, followed by aneurysmal dilatation and rupture of the involved artery. Up to now, only 13 cases have been reported, and this unique disease is not fully recognized among general pathologists and physicians. Here, we present a case of segmental mediolytic arteritis involving the propria hepatic artery, which resulted in intraabdominal hemorrhage, and consequently hypovolemic circulatory disturbance. Histologically, the rupture focus showed degeneration and desquamation of the intima and media with fibrin-like material covering the exposed adventitia. Inflammatory infiltrates were only noted in the rupture focus as a secondary reactive change. Other than the rupture focus, there were two foci showing similar findings. This disease has rarely been reported and is seldom recognized as a cause of arterial rupture. In cases of sudden intraabdominal hemorrhage, segmental mediolytic arteritis should be considered as a possible cause in addition to atherosclerotic and mycotic aneurysm, traumatic injury and vasculitis syndromes.

Pancreatoblastoma: optically clear nuclei in squamoid corpuscles are rich in biotin.

Pancreatoblastoma is a rare pancreatic neoplasm that generally occurs in children and demonstrates unique histopathologic features, including squamoid corpuscles. We encountered five cases of pancreatoblastoma and unexpectedly found the presence of optically clear nuclei (OCN) only in tumor cells of squamoid corpuscles. Recent studies showed that OCN seen in some types of neoplastic and non-neoplastic tissues were rich in biotin and revealed false-positive immunostaining by the avidin-biotin peroxidase complex method. We conducted an investigation to see if the same were true for pancreatoblastoma. As a result, the OCN were positively stained when the tissue section was incubated with peroxidase-labeled avidin or streptavidin and developed in a chromogen solution. The reaction was completely blocked by pretreatment with free avidin. An identical nuclear staining pattern was observed by the indirect immunoperoxidase method with the antibiotin antibody. Ultrastructurally, the OCN were occupied by filamentous substructures of chromatin measuring approximately 10 to 12 nm in diameter. These results suggested to us that the OCN in pancreatoblastomas were part of the unique nuclear change in which abundant biotin accumulated in association with the characteristically arranged chromatin substructure. When immunostaining with biotin as the linkage agent is performed in pancreatoblastomas, the false positivity in the OCN should be kept in mind to avoid any misinterpretation of the results.

Specific expression of PP5/TFPI2 mRNA by syncytiotrophoblasts in human placenta as revealed by in situ hybridization.

Placental protein 5 (PP5) is a placenta-derived glycoprotein with serine proteinase-inhibiting activity. To date its physiological functions have not been well elucidated. Recently, cDNA sequence analysis revealed that PP5 belongs to the Kunitz-type proteinase inhibitor family and it is identical to tissue factor pathway inhibitor-2 (TFPI-2), homologous to TFPI. Northern blot analysis demonstrated that placental tissue is extremely rich in the transcripts. This study localized PP5/TFPI-2 mRNA in placental tissues at three different gestational periods using in situ hybridization. PP5/TFPI-2 mRNA was specifically detected in syncytiotrophoblast at any gestational period examined, suggesting that syncytiotrophoblast is the principal production site of PP5/TFPI-2 in developing placental tissues. This mRNA expression pattern of PP5/TFPI-2 is quite different from that of TFPI, which is mainly found in vascular endothelial cells. The results indicated possible roles of PP5/TFPI-2 in the trophoblast differentiation and in the maintenance of intervillous blood flow. Also, Northern analysis demonstrated no or little expression of PP5/TFPI-2 in four choriocarcinoma cell lines, in contrast to its abundant expression in syncytiotrophoblast.

Stimulation of cellular growth and adhesion to fibronectin and vitronectin in culture and tumorigenicity in nude mice by overexpression of trypsinogen in human gastric cancer cells.

It has previously been reported that the trypsinogen gene is expressed in various human cancers. To investigate the possible role of trypsin in tumor malignancy, trypsinogen-1 cDNA was introduced into the human gastric carcinoma cell line MKN-1. The overexpression of trypsinogen-1 in MKN-1 cells stimulated cellular growth and adhesion to fibronectin and vitronectin when the trypsinogen activator enterokinase was added into the culture. Enterokinase treatment of the conditioned medium of the MKN-1 transfectants partially converted the proforms of gelatinases B and A to their apparent active forms. When the MKN-1 transfectants expressing trypsinogen-1 were intraperitoneally transplanted into nude mice, the mice frequently produced tumors in the colon, spleen and liver. However, the mice implanted with control MKN-1 cells produced no tumors. These results strongly suggest that tumor-derived trypsin contributes to the disseminated growth of some types of cancer cells including gastric cancer.

Identification of integrin-dependent and -independent cell adhesion domains in COOH-terminal globular region of laminin-5 alpha 3 chain.

Laminin-5 is an isoform of laminin that consists of alpha 3, beta 3, and gamma 2 chains and has potent cell adhesion- and cell migration-promoting activities. In this study, five subdomains in the COOH-terminal globular (G) domain of human laminin alpha 3 chain were individually expressed in Escherichia coli, and their biological activities were investigated. Recombinant G2, G4, and G5 domains promoted adhesion to plastic plates of HT1080 fibrosarcoma cells, A431 epidermoid carcinoma cells, and ECV304 vascular endothelial cells. For the cell adhesion activity, the G2 domain required a divalent cation and heat-sensitive conformation more strongly than G4 and G5. The cell adhesion to G2 but not G4 and G5 was effectively inhibited by an anti-integrin alpha 3 antibody. A cell adhesion sequence of 22 amino acids, alpha 3G2A, that was homologous to the integrin alpha 3 beta 1-binding sequence GD-6 of laminin alpha 1 chain was identified within the G2 structure. The cell adhesion to alpha 3G2A peptide was also inhibited by the anti-integrin alpha 3 antibody. The cell adhesion to G2, alpha 3G2A, G4, and G5 was strongly inhibited by heparin, but that to native laminin-5 was inhibited less effectively. Moreover, G5 potently stimulated chemotactic migration of rat liver epithelial cells in Boyden chambers, but G2 and G4 did not. These results indicate that the G domain of laminin alpha 3 contains multiple cell binding sites with different mechanisms and different functions. The G2 domain seems to recognize integrin alpha 3 beta 1, whereas G4 and G5 may interact with heparin-like molecules on cell surface.

Expression of matrilysin in vascular endothelial cells adjacent to matrilysin-producing tumors.

Matrilysin is believed to play important roles in tumor progression and metastasis. In the present study, we analyzed matrilysin-producing cells in various human cancer tissues by immunohistochemistry and in situ hybridization. Tumor cells in colorectal carcinomas, pancreatic carcinomas, transitional-cell carcinomas of the kidney and small-cell lung carcinomas were frequently positive for matrilysin. In addition, we found that endothelial cells of arterioles and venules adjacent to matrilysin-positive tumors expressed matrilysin mRNA and protein. The endothelial cells adjacent to matrilysin-negative tumors and those in normal tissues were negative for matrilysin. Furthermore, analyses by casein zymography, Western blotting and RT-PCR showed that matrilysin was weakly expressed by cultured human umbilical vein endothelial cells. Our results suggest that the expression of matrilysin in vascular endothelial cells and in tumor cells may be regulated by common soluble factors, and that endothelial cell-derived matrilysin may contribute to tumor angiogenesis and tumor metastasis.

Adult ganglioneuroblastoma of the anterior mediastinum.

A case of ganglioneuroblastoma occurring in the anterior mediastinum of a 79 year-old man is reported. The tumor was mainly composed of neuroblasts with occasional ganglion cells. Foci of melanin-laden cells were also identified. Immunohistochemistry revealed that the tumor cells showed both schwannian and melanocytic differentiation with immunoreactivity to anti-S100 protein and anti-HMB45 antibodies. In addition, the tumor contained several microcysts lined by squamous epithelial and one lymphoid tissue abundant in T lymphocytes, which appeared to be derived from thymic tissue. This case is unique in that neuroblastoma group tumors including ganglioneuroblastoma is uncommon in the elderly and in the thymic region, and rarely shows melanocytic differentiation. To the best of our knowledge, this case is a tumor of neuroblastoma group occurring in the eldest patient.

Differential expression of laminin-5/ladsin subunits in human tissues and cancer cell lines and their induction by tumor promoter and growth factors.

We previously reported a new laminin variant containing laminin gamma 2 (or B2t) chain, ladsin, which exerted prominent cell-scattering, cell-adhesion, and cell-migration activities. In the present study, this laminin was further characterized, and gene expression of its three subunits in various human tissues and cancer cell lines was examined by Northern blotting. cDNA cloning of the largest subunit of ladsin and partial amino acid sequencing of its beta (or B1) subunit revealed that ladsin was identical to laminin-5 (kalinin/epiligrin/ nicein). Among various human tissues, placenta, lung, and fetal kidney expressed high levels of mRNAs for the three subunits of laminin-5 (laminin alpha 3EPA, beta 3, and gamma 2 chains). Most gastric and squamous carcinoma cell lines constitutively expressed all of the three subunit mRNAs, while other types of carcinoma cell lines expressed one or two of them. The tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) and epidermal growth factor (EGF) strongly enhanced the gene expression of the three subunits, increasing 2 to 8-fold the secretion of laminin-5 from carcinoma cells into culture medium. However, TPA treatment did not increase the secretion of laminin beta 1 chain, a subunit of laminins-1, -3, and -6. The unique properties and inducibility by TPA and EGF of laminin-5 suggest that it is associated with growth and migration of cancer cells.

Cloning of the cDNA encoding mouse PP5/TFPI-2 and mapping of the gene to chromosome 6.

Placental protein 5 (PP5)/tissue factor pathway inhibitor-2 (TFPI-2) is a new homologue of TFPI, which contains three tandemly repeated Kunitz-type proteinase inhibitory (KPI) domains and potently inhibits the extrinsic blood coagulation cascade. In this study, mouse PP5/TFPI-2 cDNA was cloned using a human PP5/TFPI2 cDNA fragment as a probe. The characteristic three KPI domains with short spacer sequences and a basic amino acid stretch in the carboxyl-terminal region present in human PP5/TFPI-2 were well conserved in mouse PP5/TFPI-2. In general, the P1 reactive site residues of active KPI domains are basic amino acids. However, the putative P1 residues of the first, second, and third KPI domains were glutamine, aspartic acid, and serine, respectively. Mouse PP5/TFPI-2 mRNA was highly expressed in developing placenta as in humans. Adult liver and kidney also contained a significant amount of its transcripts. The mouse PP5/TFPI-2 gene was found to be located in the R-positive A2 band by the direct R-banding FISH and identified at 2.7 cM proximal to D6Mit 1 by interspecific backcross analysis.

Von Hippel-Lindau tumour suppressor gene. Localization of expression by in situ hybridization.

Inactivation of the von Hippel-Lindau (VHL) tumour suppressor gene is responsible not only for VHL disease, but also for sporadic renal cell carcinoma and cerebellar haemangioblastoma. The distribution of VHL gene expression in the mouse embryo was recently studied by in situ hybridization, along with human VHL in 14-week-old fetal kidney: there was widely distributed expression in the former and expression in the tubules and blastema in the latter. Adult human tissue and other fetal organs were not examined. The present paper describes an in situ hybridization study to assess the function of the VHL gene in adult human tissues and in tissues of human fetus at 28 weeks of gestation. The expression of the VHL gene was limited to the adult and fetal brain and kidney, and the adult prostate. Nerve cells in adult and fetal brain were positive, including the cerebellar Purkinje cells. In adult and fetal kidney, the proximal tubular epithelium, the putative origin of the common type of renal cell carcinoma, showed intense signal, whereas the distal nephron, glomeruli, and nephrogenic blastema showed no significant signal. The prostate showed significant signal in the basal epithelium. The adrenal, pancreas, and epidydimis showed no significant signal, in spite of the frequent occurrence at these sites of neoplastic or hamartomatous lesions in VHL disease.