[Familial paraganglioma syndrome: phenotype and relevance of a new SDHB mutation]. / Síndrome de paraganglioma familiar: expresividad clínica y relevancia de una nueva mutación en SDHB.

BACKGROUND AND OBJECTIVE: Advances in molecular biology have discovered new genes involved in the development of familial paraganglioma syndrome (PGL) including those encoding mitochondrial succinate dehydrogenase complex (SDH). We describe the diagnosis, clinical expression and genetic counselling in a family diagnosed of PGL due to a new SDHB mutation. PATIENTS AND METHOD: Genetic study by PCR-direct sequencing SDHB gene and biochemical determination in blood/urine fractionated catecholamine 24h, metanephrines and conventional (computed tomography/magnetic resonance imaging) and functional imaging ((123)I-MIBG) in all members of a family diagnosed of PGL. RESULT: DNA sequencing showed a non-described SDHB heterozygous mutation (c.287-3C>G intron3/exon4) in 5 of the subjects (71%). The estimated penetrance of the mutation's carriers was 40%, with a mean age of 35 years at diagnosis. All patients with active illness required surgical treatment after imaging and laboratory confirmation. CONCLUSIONS: We describe the pathogenicity, diagnostic algorithm, genetic counselling and clinical expression of a new SDHB mutation (c.287-3C>G) in a family diagnosed of PGL.

A SDHB malignant paraganglioma with dramatic response to temozolomide-capecitabine.

Ten percent of paragangliomas are malignant and one-third occurs in a genetic background. We report a case of succinate dehydrogenase subunit B (SDHB)-related malignant paraganglioma with dramatic response to temozolomide and capecitabine regimen (decrease in tumor size of 70% with RECIST criteria). Tumor cells harbored a new mutation in SDHB gene and showed aberrant hypermethylation of O6-methylguanine-DNA-methyltransferase promoter. Our report suggests the importance of molecular predictive factors of response for the selection of chemotherapeutic as well as targeted agents. This observation points to a possible genotype response to treatment relationships, which could help to design tailor-made treatments in the future.

Core 2 N-acetylglucosaminyltransferase-1 expression induces aggressive potential of testicular germ cell tumor.

We studied orchiectomy specimens from 130 patients immuhistochemically with testicular germ cell tumor (TGCT) using anti-core 2 N-acetylglucosaminyltransferase-1 (C2GnT-1) antibody. The incidence of C2GnT-1 positivity in stage I disease (29.5%, 21/71) was significantly lower than that in higher stages (84.7%, 50/59) (P < 0.001, chi(2) test). This significant difference was also found when the cases were divided into seminoma and NSGCT according to histopathological classification. Kaplan-Meier plots and the log rank test showed that in the patients with stage I seminoma, C2GnT-1-positive cases had a higher risk for recurrence (P < 0.001). This was also the case with the patients with stage I NSGCT (P < 0.001). To determine whether C2GnT-1 promotes aggressive behavior of cancer cells, a C2GnT-1-negative human TGCT cell line, JKT-1, was stably transfected with a mammalian expression vector containing C2GnT-1 cDNA. In vitro assays revealed that JKT-1-C2 cells are more invasive than mock transfectants, although there are no differences in proliferation activity. When orthotopically inoculated into athymic nude mice, JKT-1-C2 cells produced larger testicular tumors extending to the retroperitoneum with mesenteric metastasis, while mock transfectants produced small tumors without metastasis (P < 0.01, Mann-Whitney's U-test). When injected via the tail vein, JKT-1-C2 cells produced a number of metastatic lung foci. In contrast, mock transfectants produced a small number of nodules (p < 0.01, Mann-Whitney's U-test). These results strongly suggest that C2GnT-1 enhances the metastatic potential of TGCT and may be a reliable biomarker for aggressive potential of TGCT.

Expression and clinical significance of heparanase in neuroblastoma.

BACKGROUND: Previous studies indicate that heparanase (HPA), an endoglycosidase involved in tumor angiogenesis and metastasis, is up-regulated in a variety of malignancies. However, the expression of HPA in neuroblastoma (NB), one of the most common extra cranial solid tumors in children, remains unknown. This study was undertaken to explore the expression and clinical significance of HPA in NB. METHODS: Immunohistochemical staining was applied to detect the expression of HPA in 42 cases of NB. The relationships among HPA expression, international neuroblastoma staging system (INSS) stages, histopathological classification, and postoperative survival of the NB patients were analyzed. RESULTS: The expression rate of HPA in NB was 61.9% (26/42), mainly in the cytoplasm of neuroblastoma cells. The expression rates of stage 1-2, stage 3-4 and stage 4S were 35.7%, 80.0% and 62.5%, respectively. The differences between stage 1-2 and stage 3-4 were significant (P<0.01). The expression of HPA was significantly higher in the NB cases that had one of the histopathological factors: age more than 1 year (P<0.01), poorer differentiation (P<0.01), and higher mitosis karyorrhexis index (P<0.01). The survival time of HPA-negative patients was significantly longer than that of HPA-positive patients (P<0.05). CONCLUSION: Although these results indicate that heparanase might be correlated with development and progression of NB, a larger series of patients with a longer follow-up are probably needed to strengthen its role in assessment of NB prognosis.

C-kit expression in dedifferentiated and well-differentiated liposarcomas; immunohistochemistry and genetic analysis.

BACKGROUND: c-kit expression by immunohistochemistry has been utilized to identify cancer patients who can be treated with imatinib-mesylate. In gastrointestinal stromal tumors (GISTs), an activating mutation in c-kit predicts treatment response; its presence in other soft tissue tumors is unexplored. MATERIALS AND METHODS: We evaluated seven cases of dedifferentiated liposarcomas (DDLS) and compared those with seven well-differentiated liposarcomas (WDLS). Immunohistochemical staining for c-kit was performed using a polyclonal antibody. Using PCR, exons 9, 10-11, 12-13 and 17 of c-kit were amplified and direct DNA sequencing performed. RESULTS: Two out of 7 (30%) DDLS showed focal weak immunoreactivity with c-kit; no (0%) WDLS stained with c-kit. Seven out of 7 (100%) DDLS showed an allelic variation in exon 10, with a single base pair substitution (A >C) at codon 541; 3/7 (43%) WDLS showed the same change. CONCLUSION: c-kit immunoreactivity did not correlate with the change in DNA sequence; DDLS showed a consistent allelic variation in c-kit that may have significant prognostic, diagnostic and therapeutic implications.

[Correlation between expression of MMP-2, MMP-9, TIMP-2, TIMP-1 and metastasis of neuroblastoma].

OBJECTIVE: To study MMP-2, MMP-9, TIMP-2 and TIMP-1 expression, and their association to invasion and metastasis of neuroblastoma (NB). METHODS: The staining status was compared of MMP-2, MMP-9, TIMP-2 and TIMP-1 in cryostat sections of tumor tissue in 35 NB patients by immunohistochemistry. RESULTS: Strong expression of MMP-2 was detected only in 2 patients with early stage NB (group A without metastasis), but in 9 and 10 respectively with advanced stage NB (group B with local metastasis and group C with distant metastasis) (compared to group A, P < 0.01). Strong MMP-9 staining was found in 4, 8 and 11 patients for group A, B and C patients (group A vs group C, P < 0.05). The expression of TIMP-2 was the strongest in 4 group A patients, but it decreased with progression of the disease. There was no statistical difference in TIMP-1 expression among the three groups of patients. CONCLUSION: MMP-2, MMP-9 expression may be related to metastasis and progression of neuroblastoma, while TIMP-2 may have an inhibitory effect.

ASK1 (MAP3K5) as a potential therapeutic target in malignant fibrous histiocytomas with 12q14-q15 and 6q23 amplifications.

Malignant fibrous histiocytomas (MFHs) are aggressive tumors without any definable line of differentiation. We recently demonstrated that about 20% of them are characterized by high-level amplifications of the 12q14-q15 chromosome region, associated with either 1p32 or 6q23 band amplification. This genetic finding, very similar to that in well-differentiated liposarcomas, strongly suggests that these tumors actually correspond to undifferentiated liposarcomas. It also suggests that the lack of differentiation could be the consequence of amplification of target genes localized in the 1p32 or 6q23 bands. We report here the characterization by array CGH of the 6q23 minimal region of amplification. Our findings demonstrate that amplification and overexpression of ASK1 (MAP3K5), a gene localized in the 6q23 band and encoding a mitogen-activated protein kinase kinase kinase of the JNK-MAPK signaling pathway, could inhibit the adipocytic differentiation process of the tumor cells. Treatment of a cell line with specific inhibitors of ASK1 protein resulted in the bypass of the differentiation block and induction of a strong adipocytic differentiation. These observations indicate that ASK1 is a target for new therapeutic management of these aggressive tumors.

Novel succinate dehydrogenase subunit B (SDHB) mutations in familial phaeochromocytomas and paragangliomas, but an absence of somatic SDHB mutations in sporadic phaeochromocytomas.

Phaeochromocytomas arising in adrenal or extra-adrenal sites and paragangliomas of the head and neck, in particular of the carotid bodies, occur sporadically and also in a familial setting. In addition to mutations in RET and VHL in familial disease, germline mutations in SDHD and SDHB genes that encode subunits of mitochondrial complex II have also been associated with the development of familial phaeochromocytomas. To further investigate the role of SDHD and SDHB in the development of these tumours we determined the occurrence of germline SDHD and SDHB mutations in four patients with a family history of phaeochromocytoma with associated head and neck paraganglioma, one patient with a family history of phaeochromocytoma only and two patients with apparently sporadic extra-adrenal phaeochromocytoma, one of whom had early onset disease. Secondly, we investigated whether somatic SDHB mutations correlated with loss of heterozygosity at 1p36 in a subgroup of 11 sporadic and three MEN 2-associated RET-mutation-positive phaeochromocytomas. Novel SDHB mutations were identified in the probands from four families and two apparently sporadic cases (six of seven probands studied), including two missense mutations, a single nonsense and frameshift mutation, as well as two splice site mutations, one of which was shown to have partial penetrance resulting in 'leaky' splicing. Further, five intronic polymorphisms in SDHB were found. No SDHD mutations were identified. In addition, no somatic SDHB mutations were found in the remaining allele of the 11 sporadic adrenal phaeochromocytomas with allelic loss at 1p36 or the three MEN 2-associated RET-mutation-positive phaeochromocytomas. Therefore, we conclude that SDHB has a major role in the pathogenesis of familial phaeochromocytomas, but the possible role of SDHB in sporadic tumours showing allelic loss at 1p36 has yet to be ascertained.

Metastatic retroperitoneal paraganglioma in a 16-year-old girl. Case report, molecular pathological and cytogenetic findings.

Retroperitoneal paraganglioma is a rare tumor, especially occurring in childhood and adolescence, with a marked tendency to become biologically malignant. It has not been possible to predict the clinical outcome of paraganglioma patients by conventional histology, hence malignancy can only be demonstrated by the occurrence of metastatic lesions. Currently, only limited information on the genetics of this tumor is available. We report on a 16-year-old girl with a large retroperitoneal paraganglioma and an osseous metastasis to the first lumbar vertebra. In addition to morphological and immunohistochemical examinations, a molecular cytogenetic analysis was performed. Comparative genomic hybridization (CGH) revealed imbalanced chromosomal aberrations with a loss of chromosome 1p and a gain of 1q, indicating isochromosome 1q. A loss of chromosome 3 as well as low-level gains of chromosomes 4, 5, 6q, 11q and 13q were detected. A PCR-based microsatellite analysis of 1p confirmed the loss of heterozygosity, including NB1 and NB2 putative tumor-suppressor gene regions. Telomerase activity, which is found in the majority of malignant tumors, could not be detected. The case presented here is the first more comprehensive molecular genetic analysis of a sporadic malignant paraganglioma.

Chronic increased serum lipase without evidence of pancreatitis: tumor-derived lipase?

A 51-year-old man developed a large retroperitoneal tumor with liver and lymph node metastases; there was no radiological evidence of pancreatic involvement. Despite the progression of disease, results of laboratory tests, notably serum amylase, were normal except for minor increases in aspartate aminotransferase and gamma-glutamyltransferase and a marked increase in lipase. The increased lipase was not attributable to formation of macroenzyme. To determine the source of the lipase, we fractionated serum and a tumor biopsy homogenate, using electrophoresis. The lipase pattern obtained from the patient's serum differed from that seen in serum from a patient with acute pancreatitis. Additionally, the lipase pattern obtained from a homogenate of biopsy sample from the retroperitoneal tumor did not match the pattern observed for normal pancreas. Apparently, the source of this increased serum lipase activity was the nonpancreatic tumor.

Neuron-specific enolase as a marker for neuroblastoma and small-cell carcinoma of the lung.

The use of neuron-specific enolase (NSE, E.C. 4.2.1.11) as a clinical marker for neuroblastoma and small-cell carcinoma of lung (SCCL) is presented. Both tumors have a high content of NSE as demonstrated enzymatically or by immunocytochemistry. Other retroperitoneal tumors in children and other lung tumors had insignificant NSE concentrations. NSE can thus be used in the differential diagnosis of neuroblastoma and SCCL. 73% of patients with SCCL had elevated serum NSE levels. The corresponding figure for patients with other types of lung cancer was 3%. There was a good correlation between serum NSE levels and the clinical course of patients with SCCL.

Neuron-specific enolase as a differential marker in a case of retroperitoneal tumour.

In some cases of retroperitoneal tumour, preoperative investigations fail to conclusively differentiate between neuroblastoma and Wilms' tumour. Diagnostic difficulties can be encountered also intraoperatively, and even the histopathological presentation can be equivocal. According to earlier results, neuron-specific enolase (NSE) in tumour extract can be used as a marker for neuronal tissue and thus offers a new diagnostic criterion differing neuroblastoma from e.g. Wilms' tumour. This paper presents a case of Wilms' tumour where a low content of NSE in the tumour extract correctly indicated a non-neuronal tumour, whereas preoperative presentation and primary histopathology suggested a neuroblastoma.

Papillary serous carcinoma of the retroperitoneum.

We describe a retroperitoneal neoplasm in an 11-year-old girl which had a light microscopic appearance identical to that of papillary serous carcinoma of the ovary. There was no evidence of ovarian involvement. Immunohistochemical staining for amylase was positive within the cytoplasm of tumor cells. Since amylase is a marker for serous ovarian tumors, this finding supports the belief that "ovarian-type" neoplasms that occur at ectopic locations are essentially identical to their ovarian counterparts. We believe they originate from metaplasia of mesothelium. Our findings support the concept that these tumors should exhibit a biologic behavior and therapeutic response which are similar to those of an ovarian tumor of the same grade and comparable stage. The demonstration of intracytoplasmic amylase also may prove useful in differentiating peritoneal serous tumors from non-metaplastic mesothelial proliferations. We are unaware of a prior report of an extra-ovarian serous carcinoma in a child.

Retroperitoneal germinoma marked by elevated serum lactic dehydrogenase.

A case of primary retroperitoneal germinoma is presented in which markedly elevated serum lactic dehydrogenase levels were present before diagnosis, but declined to normal with abdominal irradiation. It is the first report of lactic dehydrogenase serving as a tumor marker in germinoma.