H. Scherübl

Endoscopic Ultrasonography of Neuroendocrine Tumours

Neuroendocrine tumours (NETs) of the upper gastrointestinal tract are mainly located in the pancreas, stomach or duodenum. The aims of preoperative work-up are the localization of primary tumour(s), determination of local tumour invasion, of lymph node metastases and of the hormones secreted by the tumour. Endoscopic ultrasonography (EUS) offers ideal conditions to localize and stage NETs of the foregut. We report our results in localizing and staging NETs of the foregut in 40 patients examined between 1990 and 1997 by EUS, somatostatin receptor scintigraphy (SRS), computed tomography (CT), magnetic resonance imaging (MRI) and transabdominal ultrasound (US). EUS shows the highest sensitivity in localizing insulinomas compared with SRS, US, CT and MRI. US and EUS should be the first-line diagnostics if insulinoma has been proven by a fasting test. Further diagnostic procedures are unnecessary in most cases. Further diagnostics such as CT or MRI to search for distant metastases are necessary in large tumours or local invasive tumours. EUS shows the highest accuracy to detect or exclude pancreatic gastrinomas, but fails to detect extrapancreatic gastrinomas in about 50%. The combination of EUS and SRS gives additional information. First-line diagnostics in gastrinoma patients should be SRS and CT or MRI. If no metastases are detected, EUS should be the next preoperative imaging procedure. In nonfunctional NETs, EUS provides the best information on local tumor invasion and regional lymph node involvement.

Peripheral-type benzodiazepine receptors in the regulation of proliferation of MCF-7 human breast carcinoma cell line

Peripheral-type benzodiazepine receptors (PBR) have been implicated in cell proliferation. The aim of the present study was to test the effect of the PBR ligands PK 11195 and Ro 5-4864 and the central-type benzodiazepine receptor ligand clonazepam on breast carcinoma cell proliferation, using [3H] thymidine incorporation. We then carried out a study to identify where the PBR-specific ligands Ro 5-4864 and PK 11195 act in the cell cycle, using flow cytometric analysis. We found PBR expression in the malignant breast cancer tumors, representing various levels of estrogen and/or progesterone receptors, as well as in the MCF-7 breast carcinoma cell line. PK 11195 and Ro 5-4864 inhibited cell proliferation at concentrations of 10(-5) to 10(-4) M, while clonazepam (the central-type benzodiazepine receptor-specific ligand) had no effect. In this same concentration range, PK 11195 and Ro 5-4864, in contrast to clonazepam, induced an accumulation of MCF-7 cells in both the G0-G1 and G2-M phases of the cell cycle. The present study demonstrates that PBR ligands play a role in regulating cell proliferation in the human breast carcinoma cell line MCF-7.

Expression of neuroendocrine markers: a signature of human undifferentiated carcinoma of the colon and rectum

Abstract. The frequency and prognostic significance of neuroendocrine marker expression in undifferentiated colorectal cancers has not yet been studied in great detail. Therefore, the survival of 20 patients with small cell undifferentiated colorectal cancers, treated at our institution between 1982 and 1997 (0.8% of all operated colorectal carcinomas), was correlated with the extent of neuroendocrine differentiation. Chromogranin A, synaptophysin, syntaxin1, VAMP2, SNAP25 and α/β-SNAP were used as neuroendocrine markers. Based on the degree of immunoreactivity for these marker proteins, tumors were separated into group 0 (<2% cells stained positive for neuroendocrine markers) and group 1 (>2% cells stained positive). Patients were followed up for at least 5xa0years or until death. Of 20 (45%) undifferentiated colorectal tumors, 9 expressed neuroendocrine markers (group 1). Only one patient of this group survived for 2xa0years (11%), whereas the 2-year-survival rate was 45.4% in group 0. Of the 11 patients in group 0, 9 were diagnosed with UICC stages I–III, whereas 8 of 9 tumors with expression of neuroendocrine markers were diagnosed with UICC stage IV (P=0.002). Our results show that neuroendocrine differentiation is often seen in small cell undifferentiated colorectal cancer. It correlates with a more aggressive course of the disease.

Somatostatin-receptor imaging of neuroendocrine gastroenteropancreatic tumors

BACKGROUNDnGastroenteropancreatic neuroendocrine tumors are often difficult to localize. This study was conducted to examine the value of somatostatin-receptor scintigraphy for visualization of gastroenteropancreatic neuroendocrine tumors.nnnMETHODSnApplying the recently developed indium-labeled somatostatin analogue 111In-pentetreotide to 40 patients with gastroenteropancreatic neuroendocrine tumors, the diagnostic power of pentreotide-receptor scintigraphy was evaluated in comparison with conventional imaging techniques.nnnRESULTSnExpression of somatostatin receptors was observed in the majority of patients (11 of 17 in the foregut, 14 of 16 in the midgut, and 7 of 7 in metastatic neuroendocrine tumors with unknown primary). Comparative imaging by computerized tomography, magnetic resonance imaging, and transabdominal ultrasonography yielded false-negative results for somatostatin-receptor scintigraphy in 8 of 40 patients; however, in 16 patients, tumor tissue that had escaped conventional imaging techniques was detected by 111In-pentetreotide scintigraphy.nnnCONCLUSIONSn111In-pentetreotide scintigraphy is a practical, safe, and sensitive procedure for in vivo imaging of gastroenteropancreatic neuroendocrine tumors.

Specific ligands of the peripheral benzodiazepine receptor induce apoptosis and cell cycle arrest in human esophageal cancer cells

Esophageal cancer is the most markedly increasing tumor entity in Western countries. Due to very poor 5‐year‐survival, new therapeutic approaches are mandatory. Peripheral benzodiazepine receptors (PBR) have been implicated in growth control of various tumor models, but they have not been studied yet in esophageal cancer. We used esophageal cancer cell lines and primary cell cultures of human esophageal cancers and evaluated (i) expression and localization of PBR; (ii) PBR‐ligand‐induced inhibition of cell growth; (iii) induction of apoptosis; and (iv) alterations in cell cycle. Expression of PBR was detected both in cell lines and in primary cell cultures of human esophageal cancers. PBR was localized in the mitochondria. The PBR‐specific ligands FGIN‐1‐27 and PK 11195, but not the centrally acting benzodiazepine clonazepam or the indolacetamide FGIN‐1‐52, neither of which displaying any affinity to the PBR, inhibited cell proliferation. FGIN‐1‐27 and PK 11195, but not clonazepam, potently induced apoptosis. FGIN‐1‐27 was shown to sequentially decrease the mitochondrial membrane potential, then to activate caspase‐3 and finally to cause DNA fragmentation. In addition, PBR‐specific ligands induced cell cycle arrest in the G1/G0 phase. Our data qualify PBR‐specific ligands as innovative proapoptotic and antiproliferative substances. They might prove suitable for the treatment of esophageal cancer.

Expression of dopamine receptors and transporter in neuroendocrine gastrointestinal tumor cells.

C-11- or F-18-DOPA positron emission tomography (DOPA PET) is a new sensitive imaging technique for small neuroendocrine gastrointestinal tumors which evaluates the decarboxylase activity. To further characterize the dopaminergic system in neuroendocrine gastrointestinal tumor cells, we investigated the expression of both dopamine receptors and the transmembrane dopamine transporter (DAT) in the human neuroendocrine pancreatic cell line BON and in the neuroendocrine gut cell line STC-1. Both BON and STC-1 cells expressed mRNA of the dopamine receptors D2-D5 and DAT. mRNA of the dopamine receptor D1 was detected in BON cells only. Both in BON and STC-1 cells, expression of D2 and D5 receptors and DAT was also demonstrated immunocytochemically. For functional receptor characterization intracellular cAMP levels ([cAMP]i) were determined. Whereas in STC-1 cells dopamine and the D1-like (D1/D5) receptor agonist SKF 38393 increased [cAMP]i, [cAMP]i was decreased by dopamine or the D2-like (D2-D4) receptor agonist quinpirole in BON cells. Functional DAT activity was, however, not detected in either cell line. The presence of both dopamine receptors and of the DAT suggests an autocrine and/or paracrine function of dopamine in neuroendocrine gastrointestinal tumor cells. Yet neither the transmembrane dopamine transporter nor dopamine receptors are likely to contribute to positive DOPA PET imaging of neuroendocrine gastrointestinal tumors. However, these molecules may be of diagnostic importance when applying other dopaminergic system tracers.

A somatostatin receptor 1 selective ligand inhibits Ca2+ currents in rat insulinoma 1046-38 cells

Rat insulinoma 1046‐38 cells represent a model system to study β‐cell function. The mRNAs for sst1 and sst2, two of the five somatostatin receptors, were detected by reverse transcription polymerase chain reaction amplification in these cells. Displacement binding analysis suggested that sst1 represents the major somatostatin receptor subtype. The sst1 selective compound CH‐275 did not inhibit adenylyl cyclases while compounds that activated sst2 did. In contrast, CH‐275 caused a marked inhibition of voltage‐operated Ca2+ channels while the sst2 specific analog octreotide elicited a less pronounced effect suggesting that in rat insulinoma 1046‐38 cells sst1 preferably mediates the inhibition of Ca2+ channels.

Expression of functional GABAA receptors in cholecystokinin‐secreting gut neuroendocrine murine STC‐1 cells

1 Gastrointestinal neuroendocrine (NE) cells synthesize, store and secrete γ‐aminobutyric acid (GABA). Recently, an autocrine‐paracrine function of GABA has been proposed for secretion from NE cells. 2 To search for functional GABAA receptors in NE gut cells, we performed whole‐cell and perforated‐patch‐clamp studies in the intestinal cholecystokinin (CCK)‐secreting NE cell line STC‐1. 3 Application of GABA evoked currents in STC‐1 cells. These effects were mimicked by muscimol, an agonist of GABAA receptors, and blocked by picrotoxin or bicuculline, antagonists of GABAA receptors. The GABA‐ or muscimol‐activated currents reversed near 0 mV, which under the recording conditions used was consistent with the activation of the GABAA receptor‐Cl− channel complex. 4 In contrast to the effect on most neurons, GABA as well as muscimol led to a (reversible) depolarization of the membrane potential of STC‐1 cells. Membrane depolarization in turn activated voltage‐gated Ca2+ channels and increased intracellular Ca2+ concentrations in STC‐1 cells. 5 In accordance with the observed membrane depolarization and activation of voltage‐gated Ca2+ channels, both GABA and muscimol stimulated Ca2+‐dependent CCK release. In contrast, bicuculline inhibited the GABA‐induced secretion of CCK. 6 Using the reverse transcription‐polymerase chain reaction (RT‐PCR), mRNA of the GABAA receptor subunits α2, α3, α5, β1, β3 and δ could be detected in STC‐1 cells. 7 In summary, we have shown that the CCK‐secreting gut NE cell line STC‐1 expresses functional GABAA receptors and that GABA stimulates CCK release. Thus, GABA is involved in the fine tuning of CCK secretion from the gut NE cell line STC‐1.

Expression of SIALYL-Lex antigen defined by MAb AM-3 is an independent prognostic marker in colorectal carcinoma patients

Expression of mucin‐bound sialyl‐Lex antigen during the progression of colorectal carcinoma and its potential prognostic value were analysed in sections of tumours from 182 patients with a documented follow‐up by immunohistochemistry using the monoclonal antibody (MAb) AM‐3. Two groups of colonic carcinomas with weak (n = 79) and strong (n = 103) sialyl‐Lex expression were discerned. The percentage of strongly expressing tumours increased with the progression of the disease (UICC stage I = 10%, stage II = 46%, stage III = 63%, stage IV = 68%, p < 0.0001). Seventy‐four percent of patients with carcinomas exhibiting a strong sialyl‐Lex expression but only 34% of patients with weak sialyl‐Lex expression died of the disease (p = 0.0026). In multivariate analysis, strong sialyl‐Lex expression increased the relative risk of cancer‐related death 3.8‐fold (95% CI = 1.8–7.9, p = 0.00034). The separate analyses of patients in UICC stage II (n = 56), III (n =5 9) and IV (n = 57) revealed that strong sialyl‐Lex expression was associated with a reduction of the 5‐year overall survival rate in UICC stage II (84% vs. 54%, p = 0.0013) and in stage III patients (86% vs. 35%, p = 0.0008) after curative resection but was not relevant in patients with distant metastases. In conclusion, the strong expression of sialyl‐Lex antigen defined by the MAb AM‐3 in colorectal carcinomas is an independent unfavourable prognostic factor after curative resection in stage II and III patients. The predictive power of the sialyl‐Lex expression may be helpful to define subgroups of patients at high risk for whom preventive adjuvant therapy can be selectively applied before the occurrence of detectable metastases. Int. J. Cancer 88:281–286, 2000.

Drug Therapy in Metastatic Neuroendocrine Tumors of the Gastroenteropancreatic System

Successful treatment of neuroendocrine tumor disease of the gastroenteropancreatic system requires a multimodal approach. Radical tumor surgery is required before other therapies are initiated. So far, only surgery has proven to be curative. If surgical intervention is not possible or a tumor-free state cannot be achieved, biotherapy with the somatostatin analogues octreotide or lanreotide should then be preferably carried out in patients with functional tumors. Interferon-alpha can alternatively be given. In patients with gastrinoma, therapy with proton pump inhibitors (e.g., omeprazol) is the initial treatment of choice. In patients with nonfunctional tumors, indication for treatment is only given in cases of documented tumor progress. In case of progressive tumor disease or functionality under the above-mentioned therapies, treatment with somatostatin analogues can be intensified by dose escalation or alternatively by a combination therapy with interferon-alpha and a somatostatin analogue. On the basis of the less favorable response of neuroendocrine foregut tumors to biotherapy, chemotherapy should be initiated after failure of biotherapy in documented tumor progression. A combination of streptozotocin and 5-fluorouracil, possibly combined with D,L-folinic acid, is the treatment of choice, considering the response and side effect rates. In case of predominantly anaplastic neuroendocrine tumors in advanced stages, good tumor response rates with a chemotherapeutic scheme consisting of cisplatin and etoposide can be achieved. Since the chemotherapy scheme is less effective in patients with midgut or hindgut tumors, chemoembolization of liver metastases should follow biotherapy. The response to chemoembolization may be increased by simultaneous systemic chemotherapy. Attention should always be paid to an adequate analgesic drug administration.