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Objective: The aim of the study was to compareimpact on survival after resection of primary tumors (PTs) after peptide receptor radionuclide therapy (PRRT). Background: PRRT is a highly effective therapeutic option to treat locally advanced or metastatic neuroendocrine neoplasms (NENs). Methods: We retrospectively analyzed the data of 889 patients with advanced NEN (G1-G3, stage IV) treated with at least 1 cycle of PRRT. In 486 of 889 patients (55%, group 1), PT had been removed before PRRT. Group 2 constituted 403 patients (45%) with no prior PT resection. Progression-free survival (PFS) and overall survival (OS) was determined by 68Ga SSTR-PET/ CT in all patients applying RECIST and EORTC. Results: Most patients had their PT in pancreas (n ¼ 335; 38%) and small intestine (n ¼ 284; 32%). Both groups received a mean of 4 cycles of PRRT (P ¼ 0.835) with a mean cumulative administered radioactivity of 21.6 11.7 versus 22.2 11.2 GBq (P ¼ 0.407). Median OS in group 1 was 134.0 months [confidence interval (CI): 118–147], whereas OS in group 2 was 67.0 months (CI: 60–80; hazard ratio 2.79); P < 0.001. Likewise, the median progressionfree survival after first PRRT was longer in group 1 with 18.0 (CI: 15–20) months as compared to group 2 with 14.0 (CI: 15–18; hazard ratio 1.21) months; P ¼ 0.012. Conclusions: A previous resection of the PT before PRRT provides a significant survival benefit in patients with NENs stage IV. [ABSTRACT FROM AUTHOR]

Background/Aims: The tumor suppressor p53 is rarely mutated in gastroenteropancreatic neuroendocrine neoplasms (GEP-NEN) but they frequently show a strong expression of negative regulators of p53, rendering these tumors excellent targets for a p53 recovery therapy. Therefore, we analyzed the mechanisms of a p53 recovery therapy on intestinal neuroendocrine tumors in vitro and in vivo.Methods: By Western blot and immunohistochemistry, we found that in GEP-NEN biopsy material overexpression of MDM2 was present in intestinal NEN. Therefore, we analyzed the effect of a small-molecule inhibitor, nutlin-3a, in p53 wild-type and mutant GEP-NEN cell lines by proliferation assay, flow cytometry, immunofluorescence, Western blot, and by multiplex gene expression analysis. Finally, we analyzed the antitumor effect of nutlin-3a in a xenograft mouse model in vivo. During the study, the tumor volume was determined. Results: The midgut wild-type cell line KRJ-I responded to the treatment with cell cycle arrest and apoptosis. By gene expression analysis, we could demonstrate that nutlins reactivated an antiproliferative p53 response. KRJ-I-derived xenograft tumors showed a significantly decreased tumor growth upon treatment with nutlin-3a in vivo. Furthermore, our data suggest that MDM2 also influences the expression of the oncogene FOXM1 in a p53-independent manner. Subsequently, a combined treatment of nutlin-3a and cisplatin (as chemoresistance model) resulted in synergistically enhanced antiproliferative effects. Conclusion: In summary, MDM2 overexpression is a frequent event in p53 wild-type intestinal neuroendocrine neoplasms and therefore recovery of a p53 response might be a novel personalized treatment approach in these tumors. [ABSTRACT FROM AUTHOR]

Background/Aims: Neuroendocrine tumors of the small intestine (SI-NETs) exhibit an increasing incidence and high mortality rate. Until now, no fundamental molecular event has been linked to the tumorigenesis and progression of these tumors. Only the loss of chromosome 18 (Chr18) has been shown in up to two thirds of SI-NETs, whereby the significance of this alteration is still not understood. We therefore performed the first comprehensive study to identify Chr18-related events at the genetic, epigenetic and gene/protein expression levels. Methods: We did expression analysis of all seven putative Chr18-related tumor suppressors by quantitative real-time PCR (qRT-PCR), Western blot and immunohistochemistry. Next-generation exome sequencing and SNP array analysis were performed with five SI-NETs with (partial) loss of Chr18. Finally, we analyzed all microRNAs (miRNAs) located on Chr18 by qRT-PCR, comparing Chr18+/- and Chr18+/+ SI-NETs. Results: Only DCC (deleted in colorectal cancer) revealed loss of/greatly reduced expression in 6/21 cases (29%). No relevant loss of SMAD2, SMAD4, elongin A3 and CABLES was detected. PMAIP1 and maspin were absent at the protein level. Next-generation sequencing did not reveal relevant recurrent somatic mutations on Chr18 either in an exploratory cohort of five SI-NETs, or in a validation cohort (n = 30). SNP array analysis showed no additional losses. The quantitative analysis of all 27 Chr18-related miRNAs revealed no difference in expression between Chr18+/- and Chr18+/+ SI-NETs. Conclusion: DCC seems to be the only Chr18-related tumor suppressor affected by the monoallelic loss of Chr18 resulting in a loss of DCC protein expression in one third of SI-NETs. No additional genetic or epigenetic alterations were present on Chr18. [ABSTRACT FROM AUTHOR]

Novel molecular analytes are needed in small bowel neuroendocrine tumours (SBNETs) to better determine disease aggressiveness and predict treatment response. In this study, we aimed to profile the global miRNome of SBNETs, and identify microRNAs (miRNAs) involved in tumour progression for use as potential biomarkers. Two independent miRNA profiling experiments were performed (n = 90), including primary SBNETs (n = 28), adjacent normal small bowel (NSB; n = 14), matched lymph node (LN) metastases (n = 24), normal LNs (n = 7), normal liver (n = 2) and liver metastases (n = 15). We then evaluated potentially targeted genes by performing integrated computational analyses. We discovered 39 miRNAs significantly deregulated in SBNETs compared with adjacent NSB. The most upregulated (miR-204-5p, miR-7-5p and miR-375) were confirmed by qRT-PCR. Two miRNAs (miR-1 and miR-143-3p) were significantly downregulated in LN and liver metastases compared with primary tumours. Furthermore, we identified upregulated gene targets for miR-1 and miR-143-3p in an existing SBNET dataset, which could contribute to disease progression, and show that these miRNAs directly regulate FOSB and NUAK2 oncogenes. Our study represents the largest global miRNA profiling of SBNETs using matched primary tumour and metastatic samples. We revealed novel miRNAs deregulated during SBNET disease progression, and important miRNA-mRNA interactions. These miRNAs have the potential to act as biomarkers for patient stratification and may also be able to guide treatment decisions. Further experiments to define molecular mechanisms and validate these miRNAs in larger tissue cohorts and in biofluids are now warranted. [ABSTRACT FROM AUTHOR]

Aims Due to the growing number of somatostatin receptor ( SSTR) targeting analogues and radiopeptides used for the diagnosis and therapy of neuroendocrine neoplasms ( NEN), the assessment of SSTR subtype status has increasingly gained predictive value. In pathology, the SSTR protein levels are detected routinely by immunohistochemistry ( IHC); however, a lack of a standardized evaluation system persists. Thus, in the present investigation, three well-established semi-quantitative scoring systems [immunoreactive score ( IRS), human epidermal growth factor receptor 2 ( HER2)/ neu score, H score] used commonly for SSTR- IHC evaluation in NEN were compared. Methods and results A total of 240 formalin-fixed, paraffin-embedded tumour samples from 90 patients with bronchopulmonary NEN were examined by IHC and quantitative reverse transcription-polymerase chain reaction ( qRT- PCR) for SSTR1, 2A, 3, 4 and 5 expression. Using both methods, SSTR1, 2A and 5 were the most frequently expressed subtypes. For all SSTR subtypes, all three scores correlated well with each other and with qRT-PCR data. However, the IRS was the most meaningful score with the best correlation to mRNA levels. Conclusions Because a unified IHC scoring system for SSTR analysis is needed urgently to optimize the theranostics of NEN, among the scores tested, the IRS seems to be the most suitable according to our results. It provides sufficient accuracy combined with high practicability. [ABSTRACT FROM AUTHOR]

Purpose: Somatostatin receptors (SSTR) are known for an overexpression in gastroenteropancreatic neuroendocrine tumours (GEP-NET). The aim of the present study was to find out if the receptor density predicted by the semi-quantitative parameters generated from the static positron emission tomography (PET/CT) correlated with the in vitro immunohistochemistry using a novel rabbit monoclonal anti-SSTR2A antibody (clone UMB-1) for specific SSTR2A immunohistochemistry and polyclonal antibodies for SSTR1 and 3-5. Methods: Overall 14 surgical specimens generated from 34 histologically documented GEP-NET patients were correlated with the preoperative Ga-DOTA-NOC PET/CT. Quantitative assessment of the receptor density was done using the immunoreactive score (IRS) of Remmele and Stegner; the additional 4-point IRS classification for immunohistochemistry and standardized uptake values (SUV and SUV) were used for PET/CT. Results: The IRS for SSTR2A and SSTR5 correlated highly significant with the SUV on the PET/CT ( p < 0.001; p < 0.05) and the IRS for SSTR2A with the SUV ( p < 0.013). The level of SSTR2A score correlated significantly with chromogranin A staining and indirectly to the tumour grading. Conclusion: The highly significant correlation between SSTR2A and SSTR5 and the SUV on the Ga-DOTA-NOC PET/CT scans is concordant with the affinity profile of Ga-DOTA-NOC to the SSTR subtypes and demonstrates the excellent qualification of somatostatin analogues in the diagnostics of NET. This study correlating somatostatin receptor imaging using Ga-DOTA-NOC PET/CT with immunohistochemically analysed SSTR also underlines the approval of therapy using somatostatin analogues, follow-up imaging as well as radionuclide therapy. [ABSTRACT FROM AUTHOR]

Introduction: Peptide receptor radionuclide therapy (PRRT) of patients with somatostatin receptor expressing neuroendocrine neoplasms has shown promising results in clinical trials and a recently published phase III study., Methods: In our center, 2294 patients were screened between 2004 and 2014 by 68 Ga somatostatin receptor (SSTR) PET/CT. Intention to treat analysis included 1048 patients, who received at least one cycle of 90 Yttrium or 177 Lutetium-based PRRT. Progression free survival was determined by 68 Ga SSTR-PET/CT and EORTC response criteria. Adverse events were determined by CTCAE criteria., Results: Overall survival (95% confidence interval) of all patients was 51 months (47.0-54.9) and differed significantly according to radionuclide, grading, previous therapies, primary site and functionality. Progression free survival (based on PET/CT) of all patients was 19 months (16.9-21), which was significantly influenced by radionuclide, grading, and origin of neuroendocrine neoplasm. Progression free survival after initial progression and first and second resumption of PRRT after therapy-free intervals of more than 6 months were 11 months (9.4-12.5) and 8 months (6.4-9.5), respectively. Myelodysplastic syndrome or leukemia developed in 22 patients (2.1%) and 5 patients required hemodialysis after treatment, other adverse events were rare., Conclusion: PRRT is effective and overall survival is favorable in patients with neuroendocrine neoplasms depending on the radionuclide used for therapy, grading and origin of the neuroendocrine neoplasm which is not exactly mirrored in progression free survival as determined by highly sensitive 68 Ga somatostatin receptor PET/CT using EORTC criteria for determining response to therapy., Competing Interests: CONFLICTS OF INTEREST R.P. Baum received honoraria from Ipsen Pharma, ROTOP Pharmaka, OctreoPharm Sciences GmbH and Advanced Accelerator Applications outside the submitted work; H.R. Kulkarni has nothing to disclose; K. Niepsch has nothing to disclose; D. Kaemmerer received travel support by the companies IPSEN Pharma GmbH and Pfizer; D. Mueller is inventor of the herein mentioned NaCl based 68Ga labeling method. In some countries, patent applications have been registered; V. Prasad has nothing to disclose; M. Hommann has nothing to disclose; F. Robiller has nothing to disclose; A. Singh has nothing to disclose; H. Franz reports personal fees from Zentralklinik Bad Berka, during the conduct of the study; A. Jochems has nothing to disclose; P. Lambin is member is the Scientific advisory board of the companies DNAmito and Oncoradiomics and is inventor of several radiomics patents; D. Hörsch reports personal fees from Lexicon Pharma Inc, grants and personal fees from Ipsen Pharma Inc, during the conduct of the study; grants and personal fees from Novartis Pharma Inc, personal fees from Pfizer Pharma Inc, grants and personal fees from Ipsen Pharma Inc, personal fees from Lexicon Pharma Inc, outside the submitted work.

Background: Carcinoid tumor is a slow-growing type of neuroendocrine tumor, originating from enterochromaffin cells and secreting mainly serotonin. The diagnosis is based on clinical symptoms, hormone blood levels, radiological and nuclear imaging, and histological confirmation. However, most patients have metastases at the time of diagnosis because the clinical signs often remain unnoticed or are attributed to other abdominal conditions. In up to 50% of patients the endocardium is affected due to a hormonally active tumor profile. The study aim was to report the outcome of surgical treatment in patients with carcinoid heart disease, including the data of radiological and nuclear imaging, histological diagnosis, and follow up information., Methods: Between 2008 and 2014, a total of 39 consecutive patients (28 males, 11 females; mean age 66 years; range: 28-84 years) with carcinoid heart syndrome were operated on at the authors' institution. Valvular heart disease was diagnosed with two-dimensional echocardiography. The study population included 26 patients (67%) with severe metastatic disease, who underwent radiotherapy preoperatively, and 13 patients (33%) who were metastasis-free and did not receive preoperative systemic therapy. Follow up was available for all hospital survivors, all of whom underwent serial echocardiographic follow up postoperatively. Adverse cardiac events were defined as cardiac-related death, a need for valvular reintervention, the occurrence of valve prosthesis-related complications, or echocardiographic evidence of new, high-degree valvular dysfunction during follow up., Results: The majority of patients (n = 34; 87%) underwent isolated tricuspid valve replacement, while simultaneous pulmonary valve replacement was performed in five patients (13%). Postoperative complications included reoperation for bleeding in five patients (13%) and new heart block requiring pacemaker implantation in 10 (25%). The in-hospital mortality was 5% (n = 2). The overall survival was 43% at six years postoperatively. At the latest follow up, 12 of the 17 survivors were in NYHA class I, and five in NYHA class II. The adverse cardiac event rate was 71%. Echocardiographically, 46% of patients (6/13) showed at least stationary or mild improvement in the right ventricular ejection fraction at follow up, with no evidence of paravalvular leak, infective endocarditis, or progressive other native valvular carcinoid affection. Postoperatively, the right atrial dimensions were preserved as normal in 23 patients (59%), mildly dilated in six (15%), moderately dilated in three (8%), and severely dilated in seven (18%). Valve-in-valve transcatheter aortic valve implantation was performed in two patients (12%) due to structural degeneration of the valve bioprosthesis and native valve disease progression., Conclusions: Despite advanced systemic disease, the surgical treatment of patients with carcinoid heart syndrome is associated with an acceptable perioperative risk and satisfactory mid-term survival. Those patients who survived valve surgery benefited from a significant improvement in their functional capacity. Percutaneous procedures may represent a useful tool to reduce the risk of late valvular reinterventions.

Introduction: For many tumors, the overexpression of the chemokine receptor CXCR4 is associated with increased malignancy and poor patient outcomes. However, comprehensive data for neuroendocrine neoplasms of the lung are still lacking., Methods: CXCR4 expression was evaluated in a panel of bronchopulmonary neuroendocrine neoplasms (BP-NEN) comprising typical carcinoids (n = 26), atypical carcinoids (n = 30), and small cell lung cancers (SCLC, n = 34). Samples were analyzed by immunohistochemistry using the novel monoclonal rabbit anti-human CXCR4 antibody UMB-2 and by qRT-PCR. The expression was correlated with clinical data and overall patient survival., Results: CXCR4 was predominantly localized at the plasma membrane of the tumor cells. CXCR4 was expressed with a high intensity in almost all of the 30 SCLC samples. In contrast, it was detected infrequently and with low intensity in the typical carcinoid and atypical carcinoid samples. There was a significant correlation between the immunohistochemistry and qRT-PCR data. Additionally, there was a significant negative relationship between CXCR4 expression and overall survival., Conclusions: With increasing malignancy, BP-NEN clearly differ in the extent of CXCR4 expression. As in other tumor entities, CXCR4 overexpression significantly correlates with negative patient outcome. Due to its particular high expression rate in SCLC, CXCR4 may serve as a promising new target for diagnostic and pharmacological intervention as well as for peptide receptor-based radionuclide therapy.

Background: Manual evaluation of somatostatin receptor (SSTR) immunohistochemistry (IHC) is a time-consuming and cost-intensive procedure. Aim of the study was to compare manual evaluation of SSTR subtype IHC to an automated software-based analysis, and to in-vivo imaging by SSTR-based PET/CT., Methods: We examined 25 gastroenteropancreatic neuroendocrine tumor (GEP-NET) patients and correlated their in-vivo SSTR-PET/CT data (determined by the standardized uptake values SUVmax,-mean) with the corresponding ex-vivo IHC data of SSTR subtype (1, 2A, 4, 5) expression. Exactly the same lesions were imaged by PET/CT, resected and analyzed by IHC in each patient. After manual evaluation, the IHC slides were digitized and automatically evaluated for SSTR expression by Definiens XD software. A virtual IHC score "BB1" was created for comparing the manual and automated analysis of SSTR expression., Results: BB1 showed a significant correlation with the corresponding conventionally determined Her2/neu score of the SSTR-subtypes 2A (rs: 0.57), 4 (rs: 0.44) and 5 (rs: 0.43). BB1 of SSTR2A also significantly correlated with the SUVmax (rs: 0.41) and the SUVmean (rs: 0.50). Likewise, a significant correlation was seen between the conventionally evaluated SSTR2A status and the SUVmax (rs: 0.42) and SUVmean (rs: 0.62)., Conclusion: Our data demonstrate that the evaluation of the SSTR status by automated analysis (BB1 score), using digitized histopathology slides ("virtual microscopy"), corresponds well with the SSTR2A, 4 and 5 expression as determined by conventional manual histopathology. The BB1 score also exhibited a significant association to the SSTR-PET/CT data in accordance with the high affinity profile of the SSTR analogues used for imaging.

Aims: To evaluate the frequency of somatostatin-receptor 5 (SSTR 5) in pancreatic neuroendocrine tumors by using monoclonal and polyclonal antibodies., Material and Method: we analyzed 66 proven pancreatic neuroendocrine tumors immunohistochemically with monoclonal (clone UMB-4) and polyclonal SSTR 5-antibodies. Immunoreactive score (IRS) and DAKO-score Her2/neu were evaluated., Results: Immunohistochemistry analysis demonstrated for the IRS a significant higher staining of all specimen using the monoclonal antibodies ( IRS SSTR5 poly vs IRS SSTR 5 mono; 20.0% vs 30.3% p < 0.001) by a correlation of 0.21; p = 0.04. For the HER2 score there was also a significant higher staining in the monoclonal group (Her2 SSTR 5 poly vs Her2 SSTR 5 mono; 21.5% vs 28.8% p < 0.001) by a correlation of 0.20; p = 0.08., Conclusion: Both antibodies are useful in staining of SSTR, although UMB-4 demonstrated a 10% higher SSTR 5 staining. Due to the previous underestimated expression rate of SSTR 5, current standards in diagnostics and therapy should be reconsidered. The increasing usage of long-acting pansomatostatin receptor analogues will rise the adverse effects connected to SSTR5 binding.

Unlabelled: Neuroendocrine tumors (NET) are known for an overexpression of somatostatin receptors (SSTR). In light of very few and partially contradictory publications, the present study aims to achieve a definite immunohistochemical (IHC) quantification and assessment of the distribution of all five SSTR-subtypes on NET and to evaluate an implementable scoring system, comparing the immunoreactive score of Remmele and Stegner (IRS) to the Her2-score. In 21 patients 40 different tumor tissues were IHC analysed using polyclonal antibodies for SSTR1 and 3-5 and the monoclonal antibody UMB-1 for SSTR2A. SSTR expression was quantitatively evaluated according to HER2-score and IRS, correlated among each other and to the maximum standardized uptake value (SUVmax) in tumor lesions as measured by PET/CT using 68Ga-DOTA-NOC., Results: According to the IRS, the expression of SSTR2A and 3 predominated equally with 84%, followed by SSTR4 (44%) and SSTR1 and 5 (32%). With the Her2-scoring system the most frequent subtype was found to be SSTR2A (68%), followed by SSTR3 (64%), SSTR1 (44%), SSTR5 (40%), and SSTR4 (36%). The IRS-classification and the Her2-score were found to be statistically comparable, and their correlation is highly significant for each SSTR assessment (p<0.01)., Conclusion: The results of the analyses revealed heterogeneous expression patterns. SSTR2A and 3 were highly expressed, demonstrating the importance of SSTR for diagnostics and therapy. Relatively high frequency of SSTR3 and 4 on NET give reasons to try pansomatostatin analogues for therapy rather than concentrating only on the SSTR2A. Statistically, none of the immunohistochemical scores was superior. However, due the heterogeneity of the cytoplasmic staining justice we propose the IRS as a uniform scoring scheme for IHC NET diagnostic.

Pancreatic endocrine tumors are rare but are among the most common neuroendocrine neoplasms of the abdomen. At diagnosis many of them are already advanced and difficult to treat. We report on an initially inoperable malignant pancreatic endocrine tumor in a 33-year-old woman, who received neoadjuvant peptide receptor radionuclide therapy (PRRT) as first-line treatment. This resulted in a significant downstaging of the tumor and allowed its subsequent complete surgical removal. Follow-up for eighteen months revealed a complete remission. This is the first report on neoadjuvant PRRT in a neuroendocrine neoplasm with subsequent successful complete resection.