BACKGROUND:Current therapies for neuroendocrine neoplasms (NENs) are limited, especially for fast-growing dedifferentiated NECs, which exhibit low somatostatin receptor (SSTR) expression and poor prognosis. Well-differentiated neuroendocrine tumors (NETs), on the other hand, retain SSTR expression, making them amenable to receptor-targeted therapies. In dedifferentiated NEC, C-X-C motif chemokine receptor 4 (CXCR4) has been shown to be abundantly expressed, making it a potential target for alternative treatment and diagnostic strategies. A major challenge in developing targeted therapies is the lack of primary patient-derived cell lines that maintain receptor profiles suitable for preclinical evaluation of established or innovative receptor-targeted approaches. METHODS:We established the MS-18 cell line from a metastatic rectal NEC. Neuroendocrine differentiation markers, SSTRs1-5, CXCR4, epithelial and mesenchymal markers, drug transporters (ABCB1, ABCG2), and Ki-67 were analyzed using qPCR and immunoblotting. Somatostatin receptor and CXCR4 function was evaluated by radiouptake assays. Electron microscopy, karyotyping, and CGH were performed, and the cell's in vivo engraftment rate was evaluated in a mouse NSG model. Viability studies on conventional therapeutic agents were performed. Extended molecular profiling of the primary tumor, liver metastasis, and MS-18 cell line was conducted. RESULTS:MS-18 cells showed strong expression of neuroendocrine markers (synaptophysin, neuron-specific enolase) and preserved epithelial differentiation (high E-cadherin, absence of mesenchymal markers). SSTR1, SSTR2, and SSTR5 were highly expressed, while SSTR3 and SSTR4 were absent. Uniquely, MS-18 cells exhibited strong CXCR4 expression. The proliferation index (Ki-67: 90%) matched that of the primary tumor. Elevated ABCG2 expression contributed to resistance to etoposide. Molecular profiling revealed no pathogenic mutations in key genes commonly altered in NENs, including MEN1, DAXX, ATRX, mTOR, PTEN, TP53, and RB1. The successful in vivo engraftment rate was high (4/5). CONCLUSION:The MS-18 cell line is the first patient-derived cell line with a transitional phenotype between differentiated NET and dedifferentiated NEC, showing strong expression of SSTR2 and CXCR4 and absence of driver mutations in key NET-/NEC-associated genes. It offers a unique platform for preclinical evaluation of targeted therapies.

Background Differentiated neuroendocrine tumors (NETs) express somatostatin receptors (SSTRs), targets for therapy with either unlabeled or radioactively labeled somatostatin analogs (SSA). Associated with worse prognosis, dedifferentiated NET loose SSTR expression, which may be linked to deregulation of Wnt/β-catenin signaling on an intracellular level. The aim of the present study was to investigate the effect of Wnt/β-catenin signaling pathway alterations on SSTR expression and its function in NET. Methods The NET cell lines BON-1 and QGP-1 were incubated with the Wnt-inhibitors 5-aza-2′-deoxycytidine (5-aza-CdR), Quercetin, or Niclosamide, or the Wnt activator lithium chloride (LiCl). Expression of SSTR1, SSTR2, and SSTR5 was determined by quantitative RT-PCR (qRT-PCR), immunocytomicroscopy and western blot. Changes in the Wnt pathway were analyzed by qRT-PCR of selected target genes and the TaqMan Array Human WNT Pathway. Receptor-associated function was determined by measuring the cellular uptake of [125I-Tyr3] octreotide. Results The mRNAs of SSTRs 1–5 were expressed in both cell lines. Wnt inhibitors caused downregulation of Wnt target genes, while 5-aza-CdR had the highest inhibitory effect. LiCl lead to an upregulation of Wnt genes, which was more marked in QGP-1 cells. SSTR expression increased in both cell lines upon Wnt inhibition. All three Wnt inhibitors lead to a marked increase in the specific uptake of [125I-Tyr3]octreotide, with 5-aza-CdR showing the greatest effect (increase by more than 50% in BON-1 cells), while a decreased uptake of [125I-Tyr3]octreotide was seen upon activation of Wnt signaling by LiCl. Conclusions We demonstrate here that Wnt signaling orchestrates SSTR expression and function in a preclinical NET model. Wnt inhibition increases [125I-Tyr3]octreotide uptake offering an opportunity to enhance the efficacy of SSTR-targeted theranostic approaches.

We aimed to elucidate the diagnostic potential of the C-X-C motif chemokine receptor 4 (CXCR4)-directed positron emission tomography (PET) tracer 68Ga-Pentixafor in patients with poorly differentiated neuroendocrine carcinomas (NEC), relative to the established reference standard 18F-FDG PET/computed tomography (CT). In our database, we retrospectively identified 11 treatment-naïve patients with histologically proven NEC, who underwent 18F-FDG and CXCR4-directed PET/CT for staging and therapy planning. The images were analyzed on a per-patient and per-lesion basis and compared to immunohistochemical staining (IHC) of CXCR4 from PET-guided biopsies. 68Ga-Pentixafor visualized tumor lesions in 10/11 subjects, while18F-FDG revealed sites of disease in all 11 patients. Although weak to moderate CXCR4 expression could be corroborated by IHC in 10/11 cases, 18F-FDG PET/CT detected significantly more tumor lesions (102 vs. 42; total lesions, n = 107; p < 0.001). Semi-quantitative analysis revealed markedly higher 18F-FDG uptake as compared to 68Ga-Pentixafor (maximum and mean standardized uptake values (SUV) and tumor-to-background ratios (TBR) of cancerous lesions, SUVmax: 12.8 ± 9.8 vs. 5.2 ± 3.7; SUVmean: 7.4 ± 5.4 vs. 3.1 ± 3.2, p < 0.001; and, TBR 7.2 ± 7.9 vs. 3.4 ± 3.0, p < 0.001). Non-invasive imaging of CXCR4 expression in NEC is inferior to the reference standard 18F-FDG PET/CT.

Loss of Somatostatin Receptor 2 (SSTR2) expression and rising CXC Chemokine Receptor Type 4 (CXCR4) expression are associated with dedifferentiation in neuroendocrine tumors (NET). In NET, CXCR4 expression is associated with enhanced metastatic and invasive potential and worse prognosis but might be a theragnostic target. Likewise, activation of Wnt/β-catenin signaling may promote a more aggressive phenotype in NET. We hypothesized an interaction of the Wnt/β-catenin pathway with CXCR4 expression and function in NET. The NET cell lines BON-1, QGP-1, and MS-18 were exposed to Wnt inhibitors (5-aza-CdR, quercetin, and niclosamide) or the Wnt activator LiCl. The expressions of Wnt pathway genes and of CXCR4 were studied by qRT-PCR, Western blot, and immunohistochemistry. The effects of Wnt modulators on uptake of the CXCR4 ligand [68Ga] Pentixafor were measured. The Wnt activator LiCl induced upregulation of CXCR4 and Wnt target gene expression. Treatment with the Wnt inhibitors had opposite effects. LiCl significantly increased [68Ga] Pentixafor uptake, while treatment with Wnt inhibitors decreased radiopeptide uptake. Wnt pathway modulation influences CXCR4 expression and function in NET cell lines. Wnt modulation might be a tool to enhance the efficacy of CXCR4-directed therapies in NET or to inhibit CXCR4-dependent proliferative signaling. The underlying mechanisms for the interaction of the Wnt pathway with CXCR4 expression and function have yet to be clarified.

Hintergrund Somatostatinrezeptoren sind der zentrale Angriffspunkt für Diagnostik und Therapie in der Behandlung neuroendokriner Tumoren. Die Somatostatin Rezeptoren SSTR2a und SSTR5 an der Zelloberfläche neuroendokriner Tumore (NET) weisen die höchste Bindungsaffinität für Somatostatinanaloga auf, jedoch treten insbesondere im Rahmen von Langzeittherapien mit zunehmender Dedifferenzierung der Tumorzellen Expressionsverluste auf. Ferner wurde gezeigt, dass eine Überexpression des kanonischen Wnt Signalweges in neuroendokrinen Tumorzelllinien Dedifferenzierung und epitheliale-mesenchymale Transitionen induziert. Ziel dieser Studie war es daher, den Einfluss von Wnt-Modulatoren auf die Expression von SSTR2a und SSTR5 zu untersuchen.

Hintergrund Die aktuell verfügbaren Therapieoptionen für neuroendokrine Karzinome sind eingeschränkt und die Prognose trist. Die Entwicklung neuer Behandlungsoptionen ist auch auf repräsentative in vitro Modelle angewiesen. In dedifferenzierten neuroendokrinen Tumoren mit niedriger Somatostatinrezeptorexpression wurde der CXC4 Rezeptor mit einer zunehmenden Dedifferenzierung und einer schlechteren Prognose assoziiert und wird daher als möglicher alternativer Ansatzpunkt für zielgerichtete Therapien diskutiert. Für die weitere Entwicklung eines solchen Therapieansatzes war bisher kein geeignetes präklinisches Model verfügbar. Von den wenigen beschriebenen Zelllinien werden BON und QGP-1 gängiger Weise verwendet. Keine der beiden Zelllinien weist eine nennenswerte Somatostatinrezeptor- oder CXC4 Rezeptorexpression auf. Wir berichten nun über die erfolgreiche Etablierung einer neuen Zelllinie aus einem neuroendokrinen Karzinom des Rektums.

ZusammenfassungNeuroendokrine Neoplasien (NEN) umfassen eine seltene Tumorentität mit heterogener Biologie, Prognose und therapeutischen Optionen. In Zusammenhang mit der kürzlichen Publikation der ersten deutschen Leitlinie zur Diagnostik und Therapie von NEN erfolgte die Analyse der Kohorte des Deutschen NET-Registers der Deutschen Gesellschaft für Endokrinologie (DGE). Hierzu wurden 2686 Fälle extrahiert und ihre Patientencharakteristika wie Alter, Geschlecht, Primärtumorlokalisation, Grading und Staging dargestellt sowie das Gesamtüberleben berechnet. Zusätzlich wurden die systemischen Behandlungsstrategien in den beiden größten Untergruppen, NEN des Dünndarms und Pankreas, im Stadium der Metastasierung analysiert.Die Verteilung der Primärtumoren, die histopathologische Charakterisierung, das Tumorstadium sowie das Gesamtüberleben waren vergleichbar mit den Ergebnissen internationaler Registerstudien. Somatostatinanaloga (SSA) und die Peptid-Rezeptor-Radionuklid-Therapie (PRRT) waren die häufigsten systemischen Therapieverfahren bei Dünndarm-NEN. Hingegen wurde eine Chemotherapie – in Übereinstimmung mit der neuen Leitlinie – vor allem bei pankreatischen NEN eingesetzt und kam in der Erstlinie in ähnlicher Frequenz wie die SSA-Therapie bzw. in der Zweitlinie ähnlich häufig wie eine PRRT zum Einsatz. Prognostisch relevante Parameter waren die WHO-Klassifikation 2010 und das TNM-Staging.Die aktuelle Analyse des deutschen NET-Registers charakterisiert damit eine multizentrische, interdisziplinäre, deutschlandweite Kohorte von NEN-Patienten und beschreibt die angewendeten systemischen Therapieverfahren, das Gesamtüberleben und die prognostische Bedeutung der WHO-Klassifikation 2010 sowie des TNM-Stadiums.

Background: Due to late diagnosis and resistance to chemotherapy, most patients with cholangiocarcinoma have an unfavorable prognosis. Despite the use of immunohistochemistry (IHC) in clinical routine, differentiation between intrahepatic cholangiocarcinoma (ICC) and secondary adenocarcinomas of the liver is frequently not clear, leading to false diagnosis and treatment decisions. Methods: Oligonucleotide microarrays (Affymetrix Hu133A©) were used for gene expression analysis of ICC (n = 11) and secondary adenocarcinomas (colorectal metastases; n = 6). By two-dimensional cluster analysis a specific gene expression profile of these tumors was established and confirmed by real-time polymerase chain reaction and IHC. Results: A total of 338 genes were significantly dysregulated (gene expression/fc ≥2; dysregulation in ≥60%) in both tumor groups. Using two-dimensional cluster analysis a fast, clear, and reproducible differentiation between ICC and colorectal metastases was possible in all cases. As potential biomarkers for differentiation, twelve genes (ICC: KRT7, DBN1, LCTB, LIF, STK17A, PIGF; metastases: TDGF1, HOXA9, TFF3, MYB, ABP1, BCL11A) were detected and will be used for further investigations. Conclusions: A specific gene expression profile for discrimination of primary and secondary adenocarcinoma of the liver could be established. In addition, marker genes for both cancers and their potential use as discrimination markers in clinical routine were also described partially for the first time.

Background/AimsThe failure to correctly differentiate between intrahepatic cholangiocarcinoma (CC) and hepatocellular carcinoma (HCC) is a significant clinical problem, particularly in terms of the different treatment goals for both cancers. In this study a specific gene expression profile to discriminate these two subgroups of liver cancer was established and potential diagnostic markers for clinical use were analyzed.Methods : To evaluate the gene expression profiles of HCC and intrahepatic CC, Oligonucleotide arrays (AffymetrixU133A) were used. Overexpressed genes were checked for their potential use as new markers for discrimination and their expression values were validated by reverse transcription polymerase chain reaction and immunohistochemistry analyses.Results : 695 genes/expressed sequence tags (ESTs) in HCC (245 up-/450 down-regulated) and 552 genes/ ESTs in CC (221 up-/331 down-regulated) were significantly dysregulated (p<0.05, fold change >2, ≥70%). Using a supervised learning method, and one-way analysis of variance a specific 270-gene expression profile that enabled rapid, reproducible differentiation between both tumors and non-malignant liver tissues was established. A panel of 12 genes (e.g., HSP90β, ERG1, GPC3, TKT, ACLY, and NME1 for HCC; SPT2, T4S3, CNX43, TTD1, HBD01 for CC) were detected and partly described for the first time as potential discrimination markers.Conclusion : sA specific gene expression profile for discrimination of primary liver cancer was identified and potential marker genes with feasible clinical impact were described.

Die Somatostatin Rezeptoren SSTR2a und SSTR5 an der Zelloberfläche neuroendokriner Tumore (NET) weisen die höchste Bindungsaffinität für Somatostatinanaloga auf, jedoch treten insbesondere i.R. von Langzeittherapien Expressionsverluste auf. Ziel dieser Studie war es daher, den Einfluss von Wnt-Modulatoren auf die Expression von SSTR2a und SSTR5 zu untersuchen.

Berichtet wird über eine 33-jährige Patientin, welche sich mit persistierender Übelkeit, Erbrechen und Bauchschmerzen zum wiederholten Male über unsere internistische Notaufnahme vorstellte. Klinisch waren zu keinem Zeitpunkt Abwehrspannung oder Peritonismus nachweisbar. Laborchemisch waren ein akutes Nierenversagen (AKIN II) mit Hypokaliämie, Hyponatriämie, ein erhöhtes CRP und ein diskret erhöhtes Serum-Lactat (2,6 mmol/l) nachzuweisen.

C-X-C motif chemokine receptor 4 (CXCR4) and somatostatin receptors (SSTR) are overexpressed in gastro-entero-pancreatic neuroendocrine tumors (GEP-NET). In this study, we aimed to elucidate the feasibility of non-invasive CXCR4 positron emission tomography/computed tomography (PET/CT) imaging in GEP-NET patients using [68Ga]Pentixafor in comparison to 68Ga-DOTA-D-Phe-Tyr3-octreotide ([68Ga]DOTATOC) and 18F-fluorodeoxyglucose ([18F]FDG). Twelve patients with histologically proven GEP-NET (3xG1, 4xG2, 5xG3) underwent [68Ga]DOTATOC, [18F]FDG, and [68Ga]Pentixafor PET/CT for staging and planning of the therapeutic management. Scans were analyzed on a patient as well as on a lesion basis and compared to immunohistochemical staining patterns of CXCR4 and somatostatin receptors SSTR2a and SSTR5. [68Ga]Pentixafor visualized tumor lesions in 6/12 subjects, whereas [18F]FDG revealed sites of disease in 10/12 and [68Ga]DOTATOC in 11/12 patients, respectively. Regarding sensitivity, SSTR-directed PET was the superior imaging modality in all G1 and G2 NET. CXCR4-directed PET was negative in all G1 NET. In contrast, 50% of G2 and 80% of G3 patients exhibited [68Ga]Pentixafor-positive tumor lesions. Whereas CXCR4 seems to play only a limited role in detecting well-differentiated NET, increasing receptor expression could be non-invasively observed with increasing tumor grade. Thus, [68Ga]Pentixafor PET/CT might serve as non-invasive read-out for evaluating the possibility of CXCR4-directed endoradiotherapy in advanced dedifferentiated SSTR-negative tumors.

Die beiden gegen CXCR4 gerichteten Radiopeptide [68Ga]Pentixafor und [177Lu]Pentixather wurden kürzlich im Rahmen eines Theranostik-Konzeptes entwickelt: [68Ga]Pentixafor zur bildgebenden Darstellung mittels PET-CT, und [177Lu]Pentixather für eine mögliche Chemokin-Rezeptor gerichtete Radiorezeptortherapie. Ziel der Studie war der Vergleich einer nicht invasiven CXCR4 PET/CT Bildgebung mit [68Ga]Pentixafor mit den etablierten, funktionalen Bildgebungen 68Ga-DOTA-D-Phe-Tyr3-octreotide ([68Ga]DOTATOC) und 18F-fluorodeoxyglucose ([18F]FDG) in gastroenteropankreatischen neuroendokrinen Tumoren (GEP-NET).

Etwa jede 7. Krebserkrankung in Deutschland betrifft das Kolon. Eine erhebliche Bedeutung im Hinblick auf die primäre Prävention des kolorektalen Karzinoms (CRC) kommt den Ernährungs- und Lebensgewohnheiten zu. Im Zusammenhang mit protektiven Ernährungsgewohnheiten taucht zunehmend der Begriff der sog. Chemoprävention auf. Bei Chemopräventiva handelt es sich um spezifische Agenzien natürlichen oder chemischen Ursprungs, welche die Karzinogenese unterdrücken, herauszögern oder rückgängig machen sollen. Der kurzkettigen Fettsäure Butyrat, welche als Fermentationsprodukt von über die Ernährung zugeführten Ballaststoffen im Kolon entsteht, wird Bedeutung als ein solches chemopräventives Agens zugesprochen.