Radical Resection in Entero-Pancreatic Neuroendocrine Tumors: Recurrence-Free Survival Rate and Definition of a Risk Score for Recurrence.

BACKGROUND: Surgery with radical intent is the only potentially curative option for entero-pancreatic neuroendocrine tumors (EP-NETs) but many patients develop recurrence even after many years. The subset of patients at high risk of disease recurrence has not been clearly defined to date. OBJECTIVE: The aim of this retrospective study was to define, in a series of completely resected EP-NETs, the recurrence-free survival (RFS) rate and a risk score for disease recurrence. PATIENTS AND METHODS: This was a multicenter retrospective analysis of sporadic pancreatic NETs (PanNETs) or small intestine NETs (SiNETs) [G1/G2] that underwent R0/R1 surgery (years 2000-2016) with at least a 24-month follow-up. Survival analysis was performed using the Kaplan-Meier method and risk factor analysis was performed using the Cox regression model. RESULTS: Overall, 441 patients (224 PanNETs and 217 SiNETs) were included, with a median Ki67 of 2% in tumor tissue and 8.2% stage IV disease. Median RFS was 101 months (5-year rate 67.9%). The derived prognostic score defined by multivariable analysis included prognostic parameters, such as TNM stage, lymph node ratio, margin status, and grading. The score distinguished three risk categories with a significantly different RFS (p < 0.01). CONCLUSIONS: Approximately 30% of patients with EP-NETs recurred within 5 years after radical surgery. Risk factors for recurrence were disease stage, lymph node ratio, margin status, and grading. The definition of risk categories may help in selecting patients who might benefit from adjuvant treatments and more intensive follow-up programs.

Systemic therapeutic strategies for GEP-NETS: what can we expect in the future?

Over the last few years, there have been important advances in the understanding of the molecular biology of neuroendocrine tumors (NETs) that have already translated into relevant advances in the clinic. Several studies have extensively assessed the mutational profile of NETs, and have shown the key roles that angiogenesis and the PI3K-AKT-mTOR pathway play in the pathogenesis of these tumors. Recent data has also revealed the potential relevance of transcription factors such as death domain-associated protein, x-linked mental retardation, and α-thalassemia syndrome protein or ataxia telangiectasia-mutated in NETs of pancreatic origin. This fast progress is leading to a rapidly increasing number of new agents being explored in the field of NETs. However, and despite some unquestionable success, objective remission rates remain low, and evidence of a substantial survival impact is lacking. Thus, there is an important need to improve our ability to identify patients most likely to benefit from specific therapies, and to incorporate biomarkers in the management of NETs. In addition, further efforts to understand mechanisms of escape and acquired resistance to the different available agents is of utmost importance, and will likely require performing paired tumor biopsies (prior and after treatment) or sequential sampling of surrogate tissues. Combinations of approved agents with new agents, either in a rational or biomarker-driven manner, are certainly warranted in this field. Likewise, sequential strategies to modulate and compensate for escape phenomenons are also of great interest. It should also be noted, however, that targeted agents are not innocuous and frequently yield toxicities that need to be adequately addressed by experienced specialists, particularly when drug combinations are considered. This review summarizes the salient data on biomarker and new agent development for the treatment of NETs.

A rare cause of peripheral arthralgia in inflammatory bowel disease: multifocal osteonecrosis.

Osteonecrosis (ON) is characterized by an infarction of osseous tissue in the subchondral regions of the bone. We report the case of a young male patient with ulcerative colitis (UC) developing severe and multifocal, large joint ON resulting in severe disability. Since typical symptoms of ON, like joint pain, might be misinterpreted as common extraintestinal manifestations, ON might easily be overlooked in patients with inflammatory bowel disease (IBD). Plain radiographs detect only advanced cases, MRI is the diagnostic method of choice with a specificity and sensitivity of > 90 %. We discuss the incidence of ON specifically in IBD and provide an update on risk factors like treatment with corticosteroids (CS), although ON has been reported in IBD patients without previous CS treatment. Apart from that, underlying inflammation, thromboembolic events and genetic risk factors might be involved in ON development supporting the hypothesis of a complex cascade. Causative therapies for ON are not available, and surgical interventions like trepanning, core decompression and prosthetic replacement are often necessary. Our intention is to direct attention to this severe complication in the differential work-up of joint pain in IBD patients.

[Neuroendocrine neoplasms of the distal jejunum and ileum]. / Neuroendokrine Neoplasien des distalen Jejunums und Ileums.

Neuroendocrine neoplasms (NEN) of the distal jejunum and ileum derive from serotonin-producing enterochromaffin (EC) cells. Due to their low proliferation rate and their infiltrative growth, they are often discovered at an advanced disease stage when metastasis has already occurred. The biology of these tumours is different from other NEN of the digestive tract. In order to standardise and improve diagnosis and therapy, the guidelines for the diagnosis and clinical management of jejuno-ileal NEN as well as for the management of patients with liver and other distant metastases from NEN were revised by the European Neuroendocrine Tumour Society (ENETS) in 2012. This review focuses on aspects relevant for surgical pathology.

Glucagon increases circulating fibroblast growth factor 21 independently of endogenous insulin levels: a novel mechanism of glucagon-stimulated lipolysis?

AIMS/HYPOTHESIS: Glucagon reduces body weight by modifying food intake, glucose/lipid metabolism and energy expenditure. All these physiological processes are also controlled by fibroblast growth factor 21 (FGF-21), a circulating hepatokine that improves the metabolic profile in obesity and type 2 diabetes. Animal experiments have suggested a possible interaction between glucagon and FGF-21 however, the metabolic consequences of this crosstalk are not understood. METHODS: The effects of exogenous glucagon on plasma FGF-21 levels and lipolysis were evaluated in healthy volunteers and humans with type 1 diabetes, as well as in rodents with streptozotocin (STZ)-induced insulinopenic diabetes. In vitro, the role of glucagon on FGF-21 secretion and lipolysis was studied using isolated primary rat hepatocytes and adipocytes. Fgf-21 expression in differentiated rat pre-adipocytes was suppressed by small interfering RNA and released FGF-21 was immunoneutralised by polyclonal antibodies. RESULTS: Glucagon induced lipolysis in healthy human volunteers, patients with type 1 diabetes, mice and rats with STZ-induced insulinopenic diabetes, and in adipocytes isolated from diabetic and non-diabetic animals. In addition, glucagon increased circulating FGF-21 in healthy humans and rodents, as well as in patients with type 1 diabetes, and insulinopenic rodents. Glucagon stimulated FGF-21 secretion from isolated primary hepatocytes and adipocytes derived from animals with insulinopenic diabetes. Furthermore, FGF-21 stimulated lipolysis in primary adipocytes isolated from non-diabetic and diabetic rats. Reduction of Fgf-21 expression (by approximately 66%) or immunoneutralisation of released FGF-21 markedly attenuated glucagon-stimulated lipolysis in adipocytes. CONCLUSIONS/INTERPRETATION: These results indicate that glucagon increases circulating FGF-21 independently of endogenous insulin levels. FGF-21 participates in glucagon-induced stimulation of lipolysis.

Impact of octreotide long-acting release on tumour growth control as a first-line treatment in neuroendocrine tumours of pancreatic origin.

BACKGROUND: Somatostatin analogues (SSA) are widely used in the treatment of patients with functioning and non-functioning neuroendocrine tumours (NET). The aim of our investigation was to evaluate the antiproliferative effect of SSA in patients with pancreatic NET. METHODS: We retrospectively analysed records of 43 patients with pancreatic NET treated at our clinic with octreotide long-lasting release as a first-line therapy. The aim of our study was to investigate the overall best response according to the RECIST criteria, overall best response defined as disease control rate (SD+PR), response and disease control rate at 12 months, and time to tumour progression (TTP). RESULTS: The mean age (± SD) of the patients (16 female/27 male) at initial diagnosis was 54.7 ± 11.86 years. At the start of therapy, 39 of 43 patients were classified as stage IV according to ENETS-TNM. Tumours were graded, based on MiB-1/Ki67 staining, as G1 (n = 8), G2 (n = 30) or unknown (n = 5). The octreoscan was positive in 37 patients, negative in 2 and unknown in 4 cases. Nineteen patients had functioning tumours, 24 patients had non-functioning tumours. Median overall survival was 98 months, and median TTP was 13 months. Analysis of grading showed a statistically significant influence on TTP when comparing the median TTP for Ki67 >10% with Ki67 <5% (p = 0.009) and Ki67 5-10% (p = 0.036). CONCLUSION: SSA may be considered as a first-line treatment for antiproliferative purposes in metastatic NET of the pancreas. Patients with a proliferation index <10% displayed a more durable response compared to those with a higher proliferation index.

Assessment and clinical implications of RANK/RANKL/OPG pathway as markers of bone tumor progression in patients with NET harboring bone metastases.

INTRODUCTION: The impact on the survival of bone metastases (BM) in patients with neuroendocrine tumor (NET) is a matter of debate. BM have a key role in causing symptoms and in decreasing patients' quality of life. Although the mechanisms of the development of BM are not completely clear, it is now well understood that the Receptor Activator of Nuclear factor Kappa-B-/Ligand (RANK/RANKL)/osteoprotegerin (OPG) pathway plays a relevant role. AIM: To characterize the RANK/RANKL/OPG pathway in patients affected with NET. PATIENTS AND METHODS: Two cohorts of 15 patients each were enrolled in the study; one cohort was affected with NET without BM and the second cohort was affected with NET with BM. The serum RANK/RANKL/OPG pathway was assessed in both the groups. RESULTS: Serum OPG levels and RANKL/OPG ratio were lower and higher, respectively, in NET patients harboring BM than in those without BM. During the ROC analysis, a cut-off value of 1071 pg/ml for OPG and 0.62 for RANKL/OPG ratio were able to significantly distinguish between the two groups. CONCLUSIONS: This study indicates that RANK/RANKL/OPG pathway is imbalanced in patients with NET harboring BM. Specific alterations of this pathway could predict an early development of BM.

The proline TP53 variant stimulates likely lymphangiogenesis in an orthotopic mouse model of pancreatic cancer.

BACKGROUND: Pancreatic cancer is a deadly disease characterised by high incidence of TP53 mutations. Restoration of TP53 function is perceived as a highly attractive therapeutic strategy, whose effects are not well characterised. METHODS: The current work adapted an inducible strategy of stage-specific reexpression of wild-type (wt) TP53 in an in vivo orthotopic mouse model of pancreatic cancer. RESULTS: The reconstitution of wt TP53 function in TP53-mutant DanG and MiaPaCa-2 cells caused G1 cell cycle arrest but no evidence of apoptosis induction. Consistent with subcutaneous xenograft models, we found that wt TP53 reduced primary tumour growth. Wt TP53 reexpression during early tumour growth led to significant increase in vascularisation. This correlated with an unexpectedly high rate of micro-metastases in lymph nodes of animals with wt TP53 induction, despite the 90% decrease in median primary tumour weight. Reexpression of wt TP53 later in tumour development did not significantly affect the number of CD31-reactive vessels, but increased lymphatic vessel density. CONCLUSION: The increased number of lymphatic vessels and micro-metastases suggests that wt TP53 induction complexly affected the biology of different tumour constituents of pancreatic cancer. Our observation suggests that combination of the inducible system with an orthotopic model can yield important insights into in vivo pancreatic cancer biology.

[Gastroenteropancreatic neuro-endocrine neoplasms]. / Neuroendokrine Neoplasiendes gastroenteropankreatischen Systems.

Neuroendocrine neoplasms (NEN) occur in the entire gastrointestinal tract. The updated classification of the WHO (2010) and current TNM-classification provide a basis for the therapeutic decision and assess the prognosis. Endoscopy and different imaging techniques are important for the localization of the primary tumor as well as local and distant metastases. The most important diagnostic imaging technique is somatostatin receptor scintigraphy. Therapeutic strategy should be discussed within the scope of a multidisciplinary tumor board. Surgery of NEN is the sole curative option. The treatment options for liver metastases include surgical resection as well as radiofrequency ablation and hepatic artery embolization. In advanced stage, systemic therapy should be used. Recent studies demonstrated a significantly prolonged progression-free survival using octreotide in well differentiated NEN. Besides the well established steptozotocine based chemotherapy regimens for pancreatic NEN, novel agents such as the mTOR-inhibitor everolimus and multityrosine kinase inhibitor sunitinib have recently also shown a prolonged progression-free survival. Moreover, temozolomide-based chemotherapy appears to be effective in pancreatic NEN. Finally, somatostatin receptor targeted radionuclide therapy can be effective in progressing gastroenteropancreatic NEN.

Orexin A stimulates glucose uptake, lipid accumulation and adiponectin secretion from 3T3-L1 adipocytes and isolated primary rat adipocytes.

AIMS/HYPOTHESIS: Orexin A (OXA) modulates body weight, food intake and energy expenditure. In vitro, OXA increases PPARγ (also known as PPARG) expression and inhibits lipolysis, suggesting direct regulation of lipid metabolism. Here, we characterise the metabolic effects and mechanisms of OXA action in adipocytes. METHODS: Isolated rat adipocytes and differentiated murine 3T3-L1 adipocytes were exposed to OXA in the presence or absence of phosphoinositide 3-kinase (PI3K) inhibitors. Pparγ expression was silenced using small interfering RNA. Glucose uptake, GLUT4 translocation, phosphatidylinositol (3,4,5)-trisphosphate production, lipogenesis, lipolysis, and adiponectin secretion were measured. Adiponectin plasma levels were determined in rats treated with OXA for 4 weeks. RESULTS: OXA PI3K-dependently stimulated active glucose uptake by translocating the glucose transporter GLUT4 from cytoplasm into the plasma membrane. OXA increased cellular triacylglycerol content via PI3K. Cellular triacylglycerol accumulation resulted from increased lipogenesis as well as from a decrease of lipolysis. Adiponectin levels in chow- and high-fat diet-fed rats treated chronically with OXA were increased. OXA stimulated adiponectin expression and secretion in adipocytes. Both pharmacological blockade of peroxisome proliferator-activated receptor γ (PPARγ) activity or silencing Pparγ expression prevented OXA from stimulating triacylglycerol accumulation and adiponectin production. CONCLUSIONS/INTERPRETATION: Our study demonstrates that OXA stimulates glucose uptake in adipocytes and that the evolved energy is stored as lipids. OXA increases lipogenesis, inhibits lipolysis and stimulates the secretion of adiponectin. These effects are conferred via PI3K and PPARγ2. Overall, OXA's effects on lipids and adiponectin secretion resemble that of insulin sensitisers, suggesting a potential relevance of this peptide in metabolic disorders.

Impact of the coxsackievirus and adenovirus receptor on the adenoma-carcinoma sequence of colon cancer.

BACKGROUND: Coxsackie and adenovirus receptor (CAR) has been suggested to function as a tumour suppressor. Its impact on the adenoma-carcinoma sequence of the colon, however, is unclear. METHODS: Coxsackie and adenovirus receptor was analysed in non-cancerous and neoplastic colon samples using immunohistochemistry and quantitative RT-PCR. The function of CAR in colon cancer cell lines was determined following application of CAR siRNA or ectopic expression of a human full-length CAR cDNA. RESULTS: Compared with healthy mucosa, increased CAR-mRNA expression was found in adenomas, whereas primary cancers and metastases displayed a marked decline. At the plasma membrane, CAR was present in normal mucosa samples (93%), adenomas, and metastases (100% ea.), whereas in colon cancers, it was found less frequently (49%, P<0.0001). Cytoplasmic CAR immunopositivity increased from normal mucosa (22%), to adenomas (73%, P=0.0006), primary cancers (83%, P<0.0001), and metastases (67%, P=0.0019). In cancer cell lines, CAR inhibition resulted in increased proliferation, whereas enforced ectopic CAR expression led to opposite results. Blocking the extracellular portion of CAR increased cell invasion in vitro. In mice, xenotransplants of colon cancer cells with enforced CAR expression formed significantly smaller tumours, whereas CAR inhibition increased the formation of liver metastases. CONCLUSION: We conclude that CAR facilitates complex effects during colon carcinogenesis, potentially mediated by its stage-dependent subcellular distribution; high CAR expression potentially prevents apoptosis in adenomas, loss of CAR at the plasma membrane promotes growth, and dissemination of primary cancers, and high membranous CAR presence may support the establishment of distant metastases.

CD154, a marker of antigen-specific stimulation of CD4 T cells, is associated with response to treatment in patients with chronic HCV infection.

CD4 T-cell function is crucial for the eradication of HCV, and insufficient function is observed in chronic carriers. The monitoring of T-cell responses is complicated by the scarcity of antigen-specific T cells and the relative inefficiency of virus-specific T cells to produce effector cytokines. CD154 is a marker of activation expressed on T cells induced through their T-cell receptor. We analysed CD4 T-cell responses in 72 patients with chronic or resolved HCV infection (23 treatment naïve, 49 treatment experienced, including 16 who had achieved a sustained response). In an additional prospective protocol, 20 of the chronically infected patients were analysed before and after 8-12 weeks of combination therapy with peg-interferon-α and ribavirin. T-cell responses were measured by detecting the expression of CD154 and Th1 cytokines after stimulation with recombinant HCV proteins and were correlated with pretreatment status and outcome of therapy. Broader T-cell responses were observed in treatment naïve than in experienced patients, while the outcome of a preceding therapy regimen did not influence T-cell responses. In the prospective cohort, an on-treatment increase in CD154+ cytokine- T-cell activity was associated with response to treatment, while a decrease was observed in nonresponders. Stronger antigen-independent activity of CD154+ cytokine+ T cells was observed in responders than in nonresponders. Our data indicate that CD154 as a marker of activation of CD4 T cells is a suitable tool for the analysis of T-cell responses in patients with HCV infection.

Management and outcomes of haemorrhage after pancreatogastrostomy versus pancreatojejunostomy.

BACKGROUND: Postpancreatectomy haemorrhage (PPH) is a major cause of morbidity and mortality after pancreaticoduodenectomy (PD). It remains unclear whether performance of a pancreatogastrostomy (PG) instead of a pancreatojejunostomy (PJ) improves outcomes owing to better endoscopic accessibility. METHODS: A large retrospective analysis was undertaken to compare outcomes of PPH, depending on whether a PG or PJ was performed. The primary outcome was the rate of successful endoscopy. A secondary outcome was the therapeutic success after adding surgery. RESULTS: Of 944 patients who had a PD, 8·4 per cent developed PPH. Endoscopy was the primary intervention in 21 (81 per cent) of 26 patients with a PG and 34 (64 per cent) of 53 with a PJ; it identified the bleeding site in 35 and 25 per cent respectively (P = 0·347). Successful endoscopic treatment was more common in the PG group (31 versus 9 per cent; P = 0·026). Surgery was performed for PPH in 15 patients (58 per cent) with a PG and 35 (66 per cent) with a PJ (P = 0·470). The majority of haemorrhages that required surgery were non-anastomotic intra-abdominal haemorrhages (12 of 15 versus 21 of 35; P = 0·171). Endoscopic or conservative treatment for PPH was successful in 42 per cent of patients with a PG and 32 per cent with a PJ (P = 0·520). The success rate increased to 85 and 91 per cent respectively when surgery was included in the algorithm (P = 0·467). CONCLUSION: The type of pancreatic anastomosis and its inherent effect on endoscopic accessibility had very little impact on the outcome of PPH. This was because haemorrhage frequently occurred from intra-abdominal or non-anastomotic intraluminal lesions.

Novel therapeutic agents for the treatment of gastroenteropancreatic neuroendocrine tumors.

Neuroendocrine tumors (NET) are frequently diagnosed late and not amenable to curative surgery due to metastatic disease to the liver and lymph nodes. The disease is complex and heterogeneous given the various functionalities, distinct tumor growth patterns, and tumor spread upon diagnosis. Established therapies include somatostatin analogues, alpha-interferon, systemic chemotherapy, and loco-regional therapies of the liver. The availability of novel agents and expression of targets, such as growth factor receptors, different subtypes of somatostatin receptors, and the mammalian target of rapamycin (mTOR) have led to the exploration of different classes of drugs and offer new treatment opportunities in neuroendocrine tumors. This review provides an overview on novel drugs, focus on the impact of recently approved drugs on the management of NET disease, and outline future perspectives.

A matter of the heart: myocardial metastases in neuroendocrine tumors.

The aim of the present study was to evaluate frequency, clinical spectrum, and treatment of myocardial metastases in patients with histologically proven neuroendocrine tumors by analysis of our database and literature review. The literature on cardiac metastases in patients with neuroendocrine tumors published from 1973 to the present was reviewed for age, sex, primary tumor localization, metastases, symptoms, complications, treatment, diagnostic methods, and histology. Patient records from our institution were analyzed retrospectively for cardiac metastases detected by any diagnostic means and detailed patient histories are given. 4 patients with myocardial metastases could be identified in our database (n=550) while literature review identified 41 published cases. Mean age at initial diagnosis was 57.5 years (females=13, males=28), primary tumor localizations were foregut (n=7), midgut (n=28), hindgut (n=1), or unknown (n=3). Carcinoid syndrome was reported for 28 patients. Cardiac involvement was right-ventricular only (n=10), left-ventricular only (n=11), or biventricular (n=10). Diagnosis was obtained by echocardiography (n=21), CT/MRI (n=12) and other methods (n=9), or by autopsy (n=9). We describe visualization of cardiac metastases by (68)Ga-DOTATOC-PET/CT for the first time. Clinical presentation ranged from asymptomatic patients to cardiac arrest. Follow-up times ranged from <1 month up to 12 years. Clinicians treating patients with neuroendocrine tumors should be aware of the heart as a possible site of metastatic disease. Echocardiography and MRI are the methods of choice for follow-up, while PET/CT might contribute to earlier and more frequent detection. Management of cardiac metastases requires close cooperation between specialists of internal medicine, nuclear medicine, and cardiac surgery.

The oral multitarget tumour growth inhibitor, ZK 304709, inhibits growth of pancreatic neuroendocrine tumours in an orthotopic mouse model.

BACKGROUND AND AIMS: Current systemic therapies for neuroendocrine tumours (NETs) do not provide sufficient control of tumour growth. However, efficient evaluation of novel drugs is hindered by the lack of a suitable preclinical animal model. Here an orthotopic mouse model of pancreatic NET is established and used to study the action of ZK 304709, a first in class, oral multitarget tumour growth inhibitor. ZK 304709 is an inhibitor of cyclin-dependent kinases (Cdks) 1, 2, 4, 7 and 9, vascular endothelial growth factor receptor-type kinases (VEGF-RTKs) 1-3 and platelet-derived growth factor receptor-type kinase beta (PDGF-RTKss). METHODS: BON and QGP-1 human NET cells were used to study proliferation, survival and cell cycle distribution in vitro. For induction of orthotopic NETs, BON cells were injected into the pancreas of NMRI(nu/nu) mice. Primary tumour growth and metastatic spread were recorded after 9 weeks, and apoptosis, microvessel density and lymphatic vessel density were determined. RESULTS: ZK 304709 dose-dependently suppressed proliferation and colony formation of NET cells. Direct effects on NET cells were consistent with Cdk inhibition and involved G(2) cell cycle arrest and apoptosis induction, which was associated with reduced expression of MCL1 (myeloid cell leukaemia sequence 1), survivin and hypoxia-inducible factor 1alpha (HIF1alpha). Apoptosis similarly occurred in vivo in ZK 304709-treated orthotopic BON tumours, resulting in a 80% reduction of primary tumour growth. In contrast, treatment with lanreotide or 5-fluorouracil and streptozotocin failed to inhibit tumour gowth. ZK 304709 also reduced tumour microvessel density, implicating antiangiogenic mechanisms. CONCLUSION: BON orthotopic tumours provide an informative model for preclinical drug evaluation in NETs. In this model, ZK 304709 achieved efficacious tumour growth control via induction of apoptosis and inhibition of tumour-induced angiogenesis.

Loss of Coxsackie and adenovirus receptor downregulates alpha-catenin expression.

BACKGROUND: The Coxsackie and adenovirus receptor (CAR) has been shown to inhibit cancer cell proliferation, migration, and invasion. The underlying mechanisms, however, are poorly understood. METHODS: The differential gene expression in the human colon cancer cell line DLD1 on RNAi-mediated functional CAR knockdown was analysed using oligo-array technology. Expression of alpha-catenin was determined by quantitative RT-PCR and western blotting. Proliferation, migration, and invasion after CAR knockdown were assessed by in vitro assays, and cell morphology in a three-dimensional context was evaluated using matrigel. RESULTS: Oligo-array technology identified alpha-catenin as the strongest downregulated gene after CAR knockdown. Western blotting and quantitative RT-PCR confirmed a reduced alpha-catenin expression after CAR knockdown in DLD1 cells and in the rat intestinal cell line IEC-6. Functionally, both cell lines showed a marked increase in proliferation, migration, and invasion on CAR knockdown. In matrigel, both cell lines formed amorphous cell clusters in contrast to well-organised three-dimensional structures of CAR-expressing vector controls. Ectopic 're'-expression of alpha-catenin in DLD1 and IEC-6 CAR knockdown cells reversed these functional and morphological effects. CONCLUSION: These data suggest that an interaction of CAR and alpha-catenin mediates the impact of CAR on cell proliferation, migration, invasion, and morphology.

Loss of the coxsackie and adenovirus receptor contributes to gastric cancer progression.

Loss of the coxsackie and adenovirus receptor (CAR) has previously been observed in gastric cancer. The role of CAR in gastric cancer pathobiology, however, is unclear. We therefore analysed CAR in 196 R(0)-resected gastric adenocarcinomas and non-cancerous gastric mucosa samples using immunohistochemistry and immunofluorescence. Coxsackie and adenovirus receptor was found at the surface and foveolar epithelium of all non-neoplastic gastric mucosa samples (n=175), whereas only 56% of gastric cancer specimens showed CAR positivity (P<0.0001). Loss of CAR correlated significantly with decreased differentiation, increased infiltrative depths, presence of distant metastases, and was also associated with reduced carcinoma-specific survival. To clarify whether CAR impacts the tumorbiologic properties of gastric cancer, we subsequently determined the role of CAR in proliferation, migration, and invasion of gastric cancer cell lines by application of specific CAR siRNA or ectopic expression of a human full-length CAR cDNA. These experiments showed that RNAi-mediated CAR knock down resulted in increased proliferation, migration, and invasion of gastric cancer cell lines, whereas enforced ectopic CAR expression led to opposite effects. We conclude that the association of reduced presence of CAR in more severe disease states, together with our findings in gastric cancer cell lines, suggests that CAR functionally contributes to gastric cancer pathogenesis, showing features of a tumour suppressor.