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1.
Neuroendocrinology ; 111(3): 273-287, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-32241015

RESUMEN

Molecular mechanisms underlying the development and progression of pancreatic neuroendocrine tumors (PanNETs) are still insufficiently understood. Efficacy of currently approved PanNET therapies is limited. While novel treatment options are being developed, patient stratification permitting more personalized treatment selection in PanNET is yet not feasible since no predictive markers are established. The lack of representative in vitro and in vivo models as well as the rarity and heterogeneity of PanNET are prevailing reasons for this. In this study, we describe an in vitro 3-dimensional (3-D) human primary PanNET culture system as a novel preclinical model for more personalized therapy selection. We present a screening platform allowing multicenter sample collection and drug screening in 3-D cultures of human primary PanNET cells. We demonstrate that primary cells isolated from PanNET patients and cultured in vitro form islet-like tumoroids. Islet-like tumoroids retain a neuroendocrine phenotype and are viable for at least 2 weeks in culture with a high success rate (86%). Viability can be monitored continuously allowing for a per-well normalization. In a proof-of-concept study, islet-like tumoroids were screened with three clinically approved therapies for PanNET: sunitinib, everolimus and temozolomide. Islet-like tumoroids display varying in vitro response profiles to distinct therapeutic regimes. Treatment response of islet-like tumoroids differs also between patient samples. We believe that the presented human PanNET screening platform is suitable for personalized drug testing in a larger patient cohort, and a broader application will help in identifying novel markers predicting treatment response and in refining PanNET therapy.


Asunto(s)
Antineoplásicos , Ensayos de Selección de Medicamentos Antitumorales , Islotes Pancreáticos , Tumores Neuroendocrinos , Neoplasias Pancreáticas , Cultivo Primario de Células , Antineoplásicos/farmacología , Línea Celular Tumoral , Criopreservación , Everolimus/farmacología , Humanos , Prueba de Estudio Conceptual , Sunitinib/farmacología , Temozolomida/farmacología
2.
NPJ Precis Oncol ; 8(1): 59, 2024 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-38429350

RESUMEN

There are no therapeutic predictive biomarkers or representative preclinical models for high-grade gastroenteropancreatic neuroendocrine neoplasms (GEP-NEN), a highly aggressive, fatal, and heterogeneous malignancy. We established patient-derived (PD) tumoroids from biobanked tissue samples of advanced high-grade GEP-NEN patients and applied this model for targeted rapid ex vivo pharmacotyping, next-generation sequencing, and perturbational profiling. We used tissue-matched PD tumoroids to profile individual patients, compared ex vivo drug response to patients' clinical response to chemotherapy, and investigated treatment-induced adaptive stress responses.PD tumoroids recapitulated biological key features of high-grade GEP-NEN and mimicked clinical response to cisplatin and temozolomide ex vivo. When we investigated treatment-induced adaptive stress responses in PD tumoroids in silico, we discovered and functionally validated Lysine demethylase 5 A and interferon-beta, which act synergistically in combination with cisplatin. Since ex vivo drug response in PD tumoroids matched clinical patient responses to standard-of-care chemotherapeutics for GEP-NEN, our rapid and functional precision oncology approach could expand personalized therapeutic options for patients with advanced high-grade GEP-NEN.

3.
Virchows Arch ; 478(6): 1215-1219, 2021 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-33005981

RESUMEN

Recently, the term mixed neuroendocrine non-neuroendocrine neoplasms (MiNEN) has been proposed as an umbrella definition covering different possible combinations of mixed neuroendocrine-exocrine neoplasms. Among these, the adenoma plus neuroendocrine tumor (NET) combination is among the rarest and not formally recognized by the 2019 WHO Classification. In this setting, the debate between either collision tumors or true mixed neoplasms is still unsolved. In this report, a pancreatic intraductal papillary mucinous neoplasm (IPMN) plus a NET is described, and the molecular investigations showed the presence in both populations of the same KRAS, GNAS, and CDKN2A mutations and the amplification of the CCND1 gene. These data prove clonality and support a common origin of both components, therefore confirming the true mixed nature. For this reason, mixed neuroendocrine-exocrine neoplasms, in which the exocrine component is represented by a glandular precursor lesion (adenoma/IPMN) only, should be included into the MiNEN family.


Asunto(s)
Carcinoma Ductal Pancreático/patología , Tumores Neuroendocrinos/patología , Neoplasias Intraductales Pancreáticas/patología , Neoplasias Pancreáticas/patología , Adenocarcinoma Mucinoso/diagnóstico , Adenocarcinoma Mucinoso/patología , Biomarcadores de Tumor/genética , Carcinoma Ductal Pancreático/diagnóstico , Carcinoma Ductal Pancreático/genética , Carcinoma Papilar/patología , Análisis Mutacional de ADN , Humanos , Masculino , Persona de Mediana Edad , Tumores Neuroendocrinos/complicaciones , Tumores Neuroendocrinos/diagnóstico , Neoplasias Intraductales Pancreáticas/diagnóstico , Neoplasias Intraductales Pancreáticas/genética , Neoplasias Pancreáticas/complicaciones
4.
Cancers (Basel) ; 13(19)2021 Oct 07.
Artículo en Inglés | MEDLINE | ID: mdl-34638497

RESUMEN

Pancreatic neuroendocrine neoplasms are epigenetically driven tumors, but therapies against underlying epigenetic drivers are currently not available in the clinical practice. We aimed to investigate EZH2 (Enhancer of Zest homolog) expression in PanNEN and the impact of EZH2 inhibition in three different PanNEN preclinical models. EZH2 expression in PanNEN patient samples (n = 172) was assessed by immunohistochemistry and correlated with clinico-pathological data. Viability of PanNEN cell lines treated with EZH2 inhibitor (GSK126) was determined in vitro. Lentiviral transduction of shRNA targeting EZH2 was performed in QGP1 cells, and cell proliferation was measured. Rip1TAG2 mice underwent GSK126 treatment for three weeks starting from week 10 of age. Primary cells isolated from PanNEN patients (n = 6) were cultivated in 3D as islet-like tumoroids and monitored for 10 consecutive days upon GSK126 treatment. Viability was measured continuously for the whole duration of the treatment. We found that high EZH2 expression correlated with higher tumor grade (p < 0.001), presence of distant metastases (p < 0.001), and shorter disease-free survival (p < 0.001) in PanNEN patients. Inhibition of EZH2 in vitro in PanNEN cell lines and in patient-derived islet-like tumoroids reduced cell viability and impaired cell proliferation, while inhibition of EZH2 in vivo in Rip1TAG2 mice reduced tumor burden. Our results show that EZH2 is highly expressed in high-grade PanNENs, and during disease progression it may contribute to aberrations in the epigenetic cellular landscape. Targeting EZH2 may represent a valuable epigenetic treatment option for patients with PanNEN.

5.
Commun Biol ; 3(1): 740, 2020 12 07.
Artículo en Inglés | MEDLINE | ID: mdl-33288854

RESUMEN

Recent data suggest that Pancreatic Neuroendocrine Tumours (PanNETs) originate from α- or ß-cells of the islets of Langerhans. The majority of PanNETs are non-functional and do not express cell-type specific hormones. In the current study we examine whether tumour DNA methylation (DNAme) profiling combined with genomic data is able to identify cell of origin and to reveal pathways involved in PanNET progression. We analyse genome-wide DNAme data of 125 PanNETs and sorted α- and ß-cells. To confirm cell identity, we investigate ARX and PDX1 expression. Based on epigenetic similarities, PanNETs cluster in α-like, ß-like and intermediate tumours. The epigenetic similarity to α-cells progressively decreases in the intermediate tumours, which present unclear differentiation. Specific transcription factor methylation and expression vary in the respective α/ß-tumour groups. Depending on DNAme similarity to α/ß-cells, PanNETs have different mutational spectra, stage of the disease and prognosis, indicating potential means of PanNET progression.


Asunto(s)
Epigénesis Genética , Regulación Neoplásica de la Expresión Génica/fisiología , Tumores Neuroendocrinos/metabolismo , Neoplasias Pancreáticas/metabolismo , Variaciones en el Número de Copia de ADN , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Tumores Neuroendocrinos/genética , Neoplasias Pancreáticas/genética
6.
Thyroid ; 29(7): 979-992, 2019 07.
Artículo en Inglés | MEDLINE | ID: mdl-30938231

RESUMEN

Background: Anaplastic thyroid carcinoma (ATC) is one of the most aggressive human cancers, with a median survival of only three to six months. Standard treatment options and even targeted therapies have so far failed to improve long-term overall survival. Thus, novel treatment modalities for ATC, such as immunotherapy, are urgently needed. CD47 is a "don't eat me" signal, which prevents cancer cells from phagocytosis by binding to signal regulatory protein alpha on macrophages. So far, the role of macrophages and the CD47-signal regulatory protein alpha signaling axis in ATC is not well understood. Methods: This study analyzed 19 primary human ATCs for macrophage markers, CD47 expression, and immune checkpoints by immunohistochemistry. ATC cell lines and a fresh ATC sample were assessed by flow cytometry for CD47 expression and macrophage infiltration, respectively. CD47 was blocked in phagocytosis assays of co-cultured macrophages and ATC cell lines. Anti-CD47 antibody treatment was administered to ATC cell line xenotransplanted immunocompromised mice, as well as to tamoxifen-induced ATC double-transgenic mice. Results: Human ATC samples were heavily infiltrated by CD68- and CD163-expressing tumor-associated macrophages (TAMs), and expressed CD47 and calreticulin, the dominant pro-phagocytic molecule. In addition, ATC tissues expressed the immune checkpoint molecules programmed cell death 1 and programmed death ligand 1. Blocking CD47 promoted the phagocytosis of ATC cell lines by macrophages in vitro. Anti-CD47 antibody treatment of ATC xenotransplanted mice increased the frequency of TAMs, enhanced the expression of macrophage activation markers, augmented tumor cell phagocytosis, and suppressed tumor growth. In double-transgenic ATC mice, CD47 was expressed on tumor cells, and blocking CD47 increased TAM frequencies. Conclusions: Targeting CD47 or CD47 in combination with programmed cell death 1 may potentially improve the outcomes of ATC patients and may represent a valuable addition to the current standard of care.


Asunto(s)
Antígenos de Diferenciación/inmunología , Antígeno CD47/inmunología , Macrófagos/inmunología , Fagocitosis/inmunología , Receptores Inmunológicos/inmunología , Carcinoma Anaplásico de Tiroides/inmunología , Neoplasias de la Tiroides/inmunología , Escape del Tumor/inmunología , Anciano , Anciano de 80 o más Años , Animales , Antígenos de Diferenciación/metabolismo , Antígeno B7-H1/inmunología , Antígeno B7-H1/metabolismo , Antígeno CD47/antagonistas & inhibidores , Antígeno CD47/metabolismo , Línea Celular Tumoral , Femenino , Humanos , Inmunohistoquímica , Inmunoterapia , Técnicas In Vitro , Macrófagos/metabolismo , Masculino , Ratones , Ratones Transgénicos , Persona de Mediana Edad , Terapia Molecular Dirigida , Trasplante de Neoplasias , Receptor de Muerte Celular Programada 1/inmunología , Receptor de Muerte Celular Programada 1/metabolismo , Receptores Inmunológicos/metabolismo , Carcinoma Anaplásico de Tiroides/metabolismo , Neoplasias de la Tiroides/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto
7.
Cancer Cell ; 32(3): 342-359.e10, 2017 09 11.
Artículo en Inglés | MEDLINE | ID: mdl-28898696

RESUMEN

Concomitant hepatocyte apoptosis and regeneration is a hallmark of chronic liver diseases (CLDs) predisposing to hepatocellular carcinoma (HCC). Here, we mechanistically link caspase-8-dependent apoptosis to HCC development via proliferation- and replication-associated DNA damage. Proliferation-associated replication stress, DNA damage, and genetic instability are detectable in CLDs before any neoplastic changes occur. Accumulated levels of hepatocyte apoptosis determine and predict subsequent hepatocarcinogenesis. Proliferation-associated DNA damage is sensed by a complex comprising caspase-8, FADD, c-FLIP, and a kinase-dependent function of RIPK1. This platform requires a non-apoptotic function of caspase-8, but no caspase-3 or caspase-8 cleavage. It may represent a DNA damage-sensing mechanism in hepatocytes that can act via JNK and subsequent phosphorylation of the histone variant H2AX.


Asunto(s)
Carcinogénesis/metabolismo , Carcinogénesis/patología , Caspasa 8/metabolismo , Daño del ADN , Neoplasias Hepáticas/enzimología , Neoplasias Hepáticas/patología , Animales , Apoptosis , Carcinoma Hepatocelular/patología , Proliferación Celular , Senescencia Celular , Enfermedad Crónica , Cruzamientos Genéticos , Reparación del ADN , Proteína de Dominio de Muerte Asociada a Fas/metabolismo , Femenino , Inestabilidad Genómica , Hepatectomía , Hepatocitos/patología , Histonas/metabolismo , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Hígado/metabolismo , Hígado/patología , Regeneración Hepática , Masculino , Ratones , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Fosforilación , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Receptores Tipo I de Factores de Necrosis Tumoral/metabolismo , Factores de Riesgo
8.
Nat Commun ; 7: 12662, 2016 09 07.
Artículo en Inglés | MEDLINE | ID: mdl-27601008

RESUMEN

The most frequently encountered symbiont on tree roots is the ascomycete Cenococcum geophilum, the only mycorrhizal species within the largest fungal class Dothideomycetes, a class known for devastating plant pathogens. Here we show that the symbiotic genomic idiosyncrasies of ectomycorrhizal basidiomycetes are also present in C. geophilum with symbiosis-induced, taxon-specific genes of unknown function and reduced numbers of plant cell wall-degrading enzymes. C. geophilum still holds a significant set of genes in categories known to be involved in pathogenesis and shows an increased genome size due to transposable elements proliferation. Transcript profiling revealed a striking upregulation of membrane transporters, including aquaporin water channels and sugar transporters, and mycorrhiza-induced small secreted proteins (MiSSPs) in ectomycorrhiza compared with free-living mycelium. The frequency with which this symbiont is found on tree roots and its possible role in water and nutrient transport in symbiosis calls for further studies on mechanisms of host and environmental adaptation.


Asunto(s)
Ascomicetos/genética , Ecosistema , Genoma Fúngico , Micorrizas/genética , Acuaporinas/metabolismo , Basidiomycota/genética , ADN de Hongos/genética , Proteínas Fúngicas , Regulación Fúngica de la Expresión Génica , Micorrizas/fisiología , Filogenia , Pinus sylvestris/microbiología , Raíces de Plantas/microbiología , Transcriptoma , Agua
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