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Article ; Online: Calcium Electroporation versus Electrochemotherapy with Bleomycin in an In Vivo CAM-Based Uveal Melanoma Xenograft Model.

Tsimpaki, Theodora / Anastasova, Ralitsa / Liu, Hongtao / Seitz, Berthold / Bechrakis, Nikolaos E / Berchner-Pfannschmidt, Utta / Kraemer, Miriam M / Fiorentzis, Miltiadis

International journal of molecular sciences

2024 Volume 25, Issue 2

Abstract: Despite recent advancements in the diagnosis and treatment of uveal melanoma (UM), its metastatic rate remains high and is accompanied by a highly dismal prognosis, constituting an unmet need for the development of novel adjuvant therapeutic strategies. ... ...

Abstract	Despite recent advancements in the diagnosis and treatment of uveal melanoma (UM), its metastatic rate remains high and is accompanied by a highly dismal prognosis, constituting an unmet need for the development of novel adjuvant therapeutic strategies. We established an in vivo chick chorioallantoic membrane (CAM)-based UM xenograft model from UPMD2 and UPMM3 cell lines to examine its feasibility for the improvement of selection of drug candidates. The efficacy of calcium electroporation (CaEP) with 5 or 10 mM calcium chloride (Ca) and electrochemotherapy (ECT) with 1 or 2.5 µg/mL bleomycin in comparison to monotherapy with the tested drug or electroporation (EP) alone was investigated on the generated UM tumors. CaEP and ECT showed a similar reduction of proliferation and melanocytic expansion with a dose-dependent effect for bleomycin, whereas CaEP induced a significant increase of the apoptosis and a reduction of vascularization with varying sensitivity for the two xenograft types. Our in vivo results suggest that CaEP and ECT may facilitate the adequate local tumor control and contribute to the preservation of the bulbus, potentially opening new horizons in the adjuvant treatment of advanced UM.
MeSH term(s)	Humans ; Animals ; Electrochemotherapy ; Calcium ; Bleomycin ; Chorioallantoic Membrane ; Heterografts ; Electroporation ; Calcium, Dietary ; Adjuvants, Immunologic ; Adjuvants, Pharmaceutic ; Chickens ; Disease Models, Animal ; Melanoma ; Uveal Neoplasms
Chemical Substances	Calcium (SY7Q814VUP) ; Bleomycin (11056-06-7) ; Calcium, Dietary ; Adjuvants, Immunologic ; Adjuvants, Pharmaceutic
Language	English
Publishing date	2024-01-11
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms25020938
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Article: Calcium Electroporation Reduces Viability and Proliferation Capacity of Four Uveal Melanoma Cell Lines in 2D and 3D Cultures.

Kraemer, Miriam M / Tsimpaki, Theodora / Berchner-Pfannschmidt, Utta / Bechrakis, Nikolaos E / Seitz, Berthold / Fiorentzis, Miltiadis

Cancers

2022 Volume 14, Issue 12

Abstract: Electrochemotherapy (ECT) is the combination of transient pore formation following electric pulse application with the administration of cytotoxic drugs, which enhances the cytotoxic effect of the applied agent due to membrane changes and ... ...

Abstract	Electrochemotherapy (ECT) is the combination of transient pore formation following electric pulse application with the administration of cytotoxic drugs, which enhances the cytotoxic effect of the applied agent due to membrane changes and permeabilization. Although EP represents an established therapeutic option for solid malignancies, recent advances shift to the investigation of non-cytotoxic agents, such as calcium, which can also induce cell death. The present study aims to evaluate the cytotoxic effect, the morphological changes in tumor spheroids, the effect on the cell viability, and the cell-specific growth rate following calcium electroporation (CaEP) in uveal melanoma (UM) 2D monolayer cell cultures as well as in 3D tumor spheroid models. The experiments were conducted in four cell lines, UM92.1, Mel270, and two primary UM cell lines, UPMD2 and UPMM3 (UPM). The 2D and 3D UM cell cultures were electroporated with eight rectangular pulses (100 µs pulse duration, 5 Hz repetition frequency) of a 1000 V/cm pulse strength alone or in combination with 0.11 mg/mL, 0.28 mg/mL, 0.55 mg/mL or 1.11 mg/mL calcium chloride or 1.0 µg/mL or 2.5 µg/mL bleomycin. The application of calcium chloride alone induced an ATP reduction only in the UM92.1 2D cell cultures. Calcium alone had no significant effect on ATP levels in all four UM spheroids. A significant decrease in the intracellular adenosine triphosphate (ATP) level was documented in all four 2D and 3D cell cultures for both CaEP as well as ECT with bleomycin. The results suggest a dose-dependent ATP depletion with a wide range of sensitivity among the tested UM cell lines, control groups, and the applied settings in both 2D monolayer cell cultures and 3D tumor spheroid models. The colony formation capacity of the cell lines after two weeks reduced significantly after CaEP only with 0.5 mg/mL and 1.1 mg/mL, whereas the same effect could be achieved with both applied bleomycin concentrations, 1.0 µg/mL and 2.5 µg/mL, for the ECT group. The specific growth rate on day 7 following CaEP was significantly reduced in UM92.1 cell lines with 0.5 and 1.1 mg/mL calcium chloride, while Mel270 showed a similar effect only after administration of 1.1 mg/mL. UM92.1 and Mel270 spheroids exhibited lower adhesion and density after CaEP on day three in comparison to UPM spheroids showing detachment after day 7 following treatment. CaEP and bleomycin electroporation significantly reduce cell viability at similar applied voltage settings. CaEP may be a feasible and inexpensive therapeutic option for the local tumor control with fewer side effects, in comparison to other chemotherapeutic agents, for the treatment of uveal melanoma. The limited effect on normal cells and the surrounding tissue has already been investigated, but further research is necessary to clarify the effect on the surrounding tissue and to facilitate its application in a clinical setting for the eye.
Language	English
Publishing date	2022-06-11
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2527080-1
ISSN	2072-6694
ISSN	2072-6694
DOI	10.3390/cancers14122889
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Article: Chick Chorioallantoic Membrane as a Patient-Derived Xenograft Model for Uveal Melanoma: Imaging Modalities for Growth and Vascular Evaluation.

Tsimpaki, Theodora / Bechrakis, Nikolaos E / Seitz, Berthold / Kraemer, Miriam M / Liu, Hongtao / Dalbah, Sami / Sokolenko, Ekaterina / Berchner-Pfannschmidt, Utta / Fiorentzis, Miltiadis

Cancers

2023 Volume 15, Issue 5

Abstract: Background: Patient-derived tumor xenografts (PDXs) have emerged as valuable preclinical in vivo models in oncology as they largely retain the polygenomic architecture of the human tumors from which they originate. Although animal models are accompanied ...

Abstract	Background: Patient-derived tumor xenografts (PDXs) have emerged as valuable preclinical in vivo models in oncology as they largely retain the polygenomic architecture of the human tumors from which they originate. Although animal models are accompanied by cost and time constraints and a low engraftment rate, PDXs have primarily been established in immunodeficient rodent models for the in vivo assessment of tumor characteristics and of novel therapeutic cancer targets. The chick chorioallantoic membrane (CAM) assay represents an attractive alternative in vivo model that has long been used in the research of tumor biology and angiogenesis, and can overcome some of these limitations. Methods: In this study, we reviewed different technical approaches for the establishment and monitoring of a CAM-based uveal melanoma PDX model. Forty-six fresh tumor grafts were acquired after enucleation from six uveal melanoma patients and were implanted onto the CAM on ED7 with Matrigel and a ring (group 1), with Matrigel (group 2), or natively without Matrigel or a ring (group 3). Real-time imaging techniques, such as various ultrasound modalities, optical coherence tomography, infrared imaging, and imaging analyses with Image J for tumor growth and extension, as well as color doppler, optical coherence angiography, and fluorescein angiography for angiogenesis, were performed on ED18 as alternative monitoring instruments. The tumor samples were excised on ED18 for histological assessment. Results: There were no significant differences between the three tested experimental groups regarding the length and width of the grafts during the development period. A statistically significant increase in volume ( Conclusion: The establishment of a CAM-PDX uveal melanoma model could elucidate the biological growth patterns and the efficacy of new therapeutic options in vivo. The methodological novelty of this study, investigating different implanting techniques and exploiting advances in real-time imaging with multiple modalities, allows precise, quantitative assessment in the field of tumor experimentation, underlying the feasibility of CAM as an in vivo PDX model.
Language	English
Publishing date	2023-02-24
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2527080-1
ISSN	2072-6694
ISSN	2072-6694
DOI	10.3390/cancers15051436
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Article ; Online: Blockade of ß-Adrenergic Receptors by Nebivolol Enables Tumor Control Potential for Uveal Melanoma in 3D Tumor Spheroids and 2D Cultures.

Farhoumand, Lina S / Liu, Hongtao / Tsimpaki, Theodora / Hendgen-Cotta, Ulrike B / Rassaf, Tienush / Bechrakis, Nikolaos E / Fiorentzis, Miltiadis / Berchner-Pfannschmidt, Utta

International journal of molecular sciences

2023 Volume 24, Issue 6

Abstract: Uveal melanoma (UM) is the most common primary cancer of the eye in adults. A new systemic therapy is needed to reduce the high metastasis and mortality rate. As β-blockers are known to have anti-tumor effects on various cancer entities, this study ... ...

Abstract	Uveal melanoma (UM) is the most common primary cancer of the eye in adults. A new systemic therapy is needed to reduce the high metastasis and mortality rate. As β-blockers are known to have anti-tumor effects on various cancer entities, this study focuses on investigating the effect of β1-selective blockers atenolol, celiprolol, bisoprolol, metoprolol, esmolol, betaxolol, and in particular, nebivolol on UM. The study was performed on 3D tumor spheroids as well as 2D cell cultures, testing tumor viability, morphological changes, long-term survival, and apoptosis. Flow cytometry revealed the presence of all three β-adrenoceptors with a dominance of β2-receptors on cell surfaces. Among the blockers tested, solely nebivolol concentration-dependently decreased viability and altered 3D tumor spheroid structure. Nebivolol blocked the repopulation of cells spreading from 3D tumor spheroids, indicating a tumor control potential at a concentration of ≥20 µM. Mechanistically, nebivolol induced ATP depletion and caspase-3/7 activity, indicating that mitochondria-dependent signaling is involved. D-nebivolol or nebivolol combined with the β2-antagonist ICI 118.551 displayed the highest anti-tumor effects, suggesting a contribution of both β1- and β2-receptors. Thus, the present study reveals the tumor control potential of nebivolol in UM, which may offer a perspective for co-adjuvant therapy to reduce recurrence or metastasis.
MeSH term(s)	Adult ; Humans ; Nebivolol/pharmacology ; Ethanolamines/pharmacology ; Benzopyrans/pharmacology ; Adrenergic beta-Antagonists/pharmacology ; Melanoma/drug therapy ; Receptors, Adrenergic, beta
Chemical Substances	Nebivolol (030Y90569U) ; Ethanolamines ; Benzopyrans ; Adrenergic beta-Antagonists ; Receptors, Adrenergic, beta
Language	English
Publishing date	2023-03-20
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms24065894
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Article ; Online: Lessons from mouse models of Graves' disease.

Eckstein, A / Philipp, S / Goertz, G / Banga, J P / Berchner-Pfannschmidt, U

Endocrine

2020 Volume 68, Issue 2, Page(s) 265–270

Abstract: Graves' disease (GD) is an autoimmune condition with the appearance of anti-TSH receptor (TSHR) autoantibodies in the serum. The consequence is the development of hyperthyroidism in most of the patients. In addition, in the most severe cases, patients ... ...

Abstract	Graves' disease (GD) is an autoimmune condition with the appearance of anti-TSH receptor (TSHR) autoantibodies in the serum. The consequence is the development of hyperthyroidism in most of the patients. In addition, in the most severe cases, patients can develop orbitopathy (GO), achropachy and dermopathy. The central role of the TSHR for the disease pathology has been well accepted. Therefore immunization against the TSHR is pivotal for the creation of in vivo models for the disease. However, TSHR is well preserved among the species and therefore the immune system is highly tolerant. Many differing attempts have been performed to break tolerance and to create a proper animal model in the last decades. The most successful have been achieved by introducing the human TSHR extracellular domain into the body, either by injection of plasmid or adenoviruses. Currently available models develop the whole spectrum of Graves' disease-autoimmune thyroid disease and orbitopathy and are suitable to study disease pathogenesis and to perform treatment studies. In recent publications new immunomodulatory therapies have been assessed and also diseaseprevention by inducing tolerance using small cyclic peptides from the antigenic region of the extracellular subunit of the TSHR.
MeSH term(s)	Animals ; Autoantibodies ; Disease Models, Animal ; Graves Disease ; Humans ; Immunization ; Mice ; Receptors, Thyrotropin
Chemical Substances	Autoantibodies ; Receptors, Thyrotropin
Language	English
Publishing date	2020-05-12
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Review
ZDB-ID	1194484-5
ISSN	1559-0100 ; 1355-008X ; 0969-711X
ISSN (online)	1559-0100
ISSN	1355-008X ; 0969-711X
DOI	10.1007/s12020-020-02311-7
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Article ; Online: Endokrine Orbitopathie.

Eckstein, A / Esser, J / Mattheis, S / Berchner-Pfannschmidt, U

Klinische Monatsblatter fur Augenheilkunde

2016 Volume 233, Issue 12, Page(s) 1385–1407

Title translation	Graves' Orbitopathy.
MeSH term(s)	Combined Modality Therapy/methods ; Evidence-Based Medicine ; Graves Ophthalmopathy/diagnosis ; Graves Ophthalmopathy/therapy ; Humans ; Immunosuppressive Agents/therapeutic use ; Radiotherapy, Conformal/methods ; Steroids/therapeutic use ; Treatment Outcome
Chemical Substances	Immunosuppressive Agents ; Steroids
Language	German
Publishing date	2016-12
Publishing country	Germany
Document type	Journal Article ; Review
ZDB-ID	80175-6
ISSN	1439-3999 ; 0344-6360 ; 0344-6387 ; 0023-2165
ISSN (online)	1439-3999
ISSN	0344-6360 ; 0344-6387 ; 0023-2165
DOI	10.1055/s-0042-118040
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Article: The Adrenergic Receptor Antagonist Carvedilol Elicits Anti-Tumor Responses in Uveal Melanoma 3D Tumor Spheroids and May Serve as Co-Adjuvant Therapy with Radiation.

Farhoumand, Lina S / Fiorentzis, Miltiadis / Kraemer, Miriam M / Sak, Ali / Stuschke, Martin / Rassaf, Tienush / Hendgen-Cotta, Ulrike / Bechrakis, Nikolaos E / Berchner-Pfannschmidt, Utta

Cancers

2022 Volume 14, Issue 13

Abstract: Uveal melanoma (UM) is the most common intraocular tumor in adults. Despite local tumor control, no effective therapy has been found to prevent metastasis, resulting in a high mortality rate. In the present study, we evaluated the anti-tumor potential of ...

Abstract	Uveal melanoma (UM) is the most common intraocular tumor in adults. Despite local tumor control, no effective therapy has been found to prevent metastasis, resulting in a high mortality rate. In the present study, we evaluated the anti-tumor potential of non-selective ß-blockers in 3D tumor spheroids grown from UM cell lines. Of the various ß-blockers tested, carvedilol and its enantiomers were most potent in decreasing the viability of Mel270 spheroids. Carvedilol at a concentration of 10-50 µM significantly elicited cytotoxicity and induced apoptosis in spheroid cells. In result, carvedilol inhibited tumor spheroid growth and compactness, and furthermore prevented the long-term survival and repopulation of spreading spheroid cells. The drug sensitivity of the different spheroids grown from Mel270, 92-1, UPMD2, or UPMM3 cell lines was dependent on 3D morphology rather than on high-risk cytogenetic profile or adrenergic receptor expression levels. In fact, the monosomy-3-containing UPMM3 cell line was most responsive to carvedilol treatment compared to the other cell lines. The concurrent treatment of UPMM3 spheroids with carvedilol and 5 or 10 Gy irradiation revealed additive cytotoxic effects that provided tumor control. Collectively, our data demonstrate the anti-tumor properties of carvedilol and its enantiomers, which may serve as candidates for the co-adjuvant therapy of UM.
Language	English
Publishing date	2022-06-23
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2527080-1
ISSN	2072-6694
ISSN	2072-6694
DOI	10.3390/cancers14133097
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Article ; Online: An Early Wave of Macrophage Infiltration Intertwined with Antigen-Specific Proinflammatory T Cells and Browning of Adipose Tissue Characterizes the Onset of Orbital Inflammation in a Mouse Model of Graves' Orbitopathy.

Philipp, Svenja / Horstmann, Mareike / Hose, Matthias / Daser, Anke / Görtz, Gina-Eva / Jesenek, Christoph / Flögel, Ulrich / Hansen, Wiebke / Bechrakis, Nikolaos / Banga, Jasvinder Paul S / Eckstein, Anja / Berchner-Pfannschmidt, Utta

Thyroid : official journal of the American Thyroid Association

2022 Volume 32, Issue 3, Page(s) 283–293

Abstract: Background: ...

Abstract	Background:
MeSH term(s)	Adipose Tissue ; Animals ; Antigens ; CD8-Positive T-Lymphocytes ; Disease Models, Animal ; Graves Disease ; Graves Ophthalmopathy/metabolism ; Humans ; Inflammation ; Macrophages/metabolism ; Mice ; Mice, Inbred BALB C ; Receptors, Thyrotropin ; Thyrotropin
Chemical Substances	Antigens ; Receptors, Thyrotropin ; Thyrotropin (9002-71-5)
Language	English
Publishing date	2022-01-11
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1086044-7
ISSN	1557-9077 ; 1050-7256
ISSN (online)	1557-9077
ISSN	1050-7256
DOI	10.1089/thy.2021.0464
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Zs.A 3307: Show issues			Location: Je nach Verfügbarkeit (siehe Angabe bei Bestand) bis Jg. 1994: Bestellungen von Artikeln über das Online-Bestellformular Jg. 1995 - 2021: Lesesall (2.OG) ab Jg. 2022: Lesesaal (EG)
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Article ; Online: Macrophage-Orbital Fibroblast Interaction and Hypoxia Promote Inflammation and Adipogenesis in Graves' Orbitopathy.

Görtz, Gina-Eva / Philipp, Svenja / Bruderek, Kirsten / Jesenek, Christoph / Horstmann, Mareike / Henning, Yoshiyuki / Oeverhaus, Michael / Daser, Anke / Bechrakis, Nikolaos E / Eckstein, Anja / Brandau, Sven / Berchner-Pfannschmidt, Utta

Endocrinology

2022 Volume 164, Issue 2

Abstract: The inflammatory eye disease Graves' orbitopathy (GO) is the main complication of autoimmune Graves' disease. In previous studies we have shown that hypoxia plays an important role for progression of GO. Hypoxia can maintain inflammation by attracting ... ...

Abstract	The inflammatory eye disease Graves' orbitopathy (GO) is the main complication of autoimmune Graves' disease. In previous studies we have shown that hypoxia plays an important role for progression of GO. Hypoxia can maintain inflammation by attracting inflammatory cells such as macrophages (MQ). Herein, we investigated the interaction of MQ and orbital fibroblasts (OF) in context of inflammation and hypoxia. We detected elevated levels of the hypoxia marker HIF-1α, the MQ marker CD68, and inflammatory cytokines TNFα, CCL2, CCL5, and CCL20 in GO biopsies. Hypoxia stimulated GO tissues to release TNFα, CCL2, and CCL20 as measured by multiplex enzyme-linked immunosorbent assay (ELISA). Further, TNFα and hypoxia stimulated the expression of HIF-1α, CCL2, CCL5, and CCL20 in OF derived from GO tissues. Immunofluorescence confirmed that TNFα-positive MQ were present in the GO tissues. Thus, interaction of M1-MQ with OF under hypoxia also induced HIF-1α, CCL2, and CCL20 in OF. Inflammatory inhibitors etanercept or dexamethasone prevented the induction of HIF-1α and release of CCL2 and CCL20. Moreover, co-culture of M1-MQ/OF under hypoxia enhanced adipogenic differentiation and adiponectin secretion. Dexamethasone and HIF-1α inhibitor PX-478 reduced this effect. Our findings indicate that GO fat tissues are characterized by an inflammatory and hypoxic milieu where TNFα-positive MQ are present. Hypoxia and interaction of M1-MQ with OF led to enhanced secretion of chemokines, elevated hypoxic signaling, and adipogenesis. In consequence, M1-MQ/OF interaction results in constant inflammation and tissue remodeling. A combination of anti-inflammatory treatment and HIF-1α reduction could be an effective treatment option.
MeSH term(s)	Humans ; Adipogenesis/physiology ; Cells, Cultured ; Dexamethasone/pharmacology ; Fibroblasts/metabolism ; Graves Ophthalmopathy/metabolism ; Graves Ophthalmopathy/pathology ; Hypoxia/metabolism ; Inflammation/metabolism ; Orbit/metabolism ; Orbit/pathology ; Tumor Necrosis Factor-alpha/pharmacology ; Tumor Necrosis Factor-alpha/metabolism ; Cell Communication/physiology ; Macrophages/metabolism
Chemical Substances	Dexamethasone (7S5I7G3JQL) ; Tumor Necrosis Factor-alpha
Language	English
Publishing date	2022-12-07
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	427856-2
ISSN	1945-7170 ; 0013-7227
ISSN (online)	1945-7170
ISSN	0013-7227
DOI	10.1210/endocr/bqac203
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Article ; Conference proceedings: Neue Einblicke in das Zytokinspektrum beim Morbus Basedow anhand eines autoimmunen Mausmodells mit Hyperthyreose

Daser, Anke / Horstmann, Mareike / Lang, Stephan / Mattheis, Stefan / Berchner-Pfannschmidt, Utta / Banga, J. Paul / Eckstein, Anja / Philipp, Svenja

Laryngo-Rhino-Otologie

(Abstract- und Posterband)

2022 Volume 101, Issue S 02

Event/congress	Abstract- und Posterband - 93. Jahresversammlung der Deutschen Gesellschaft für HNO-Heilkunde, Kopf- und Hals-Chirurgie e.V., Bonn Interface - Fokus Mensch im Zeitalter der technisierten Medizin, Deutsche Messe Hannover, 2022-05-25
Series title	Abstract- und Posterband
Language	German
Publishing date	2022-05-01
Publisher	Georg Thieme Verlag
Publishing place	Stuttgart ; New York
Document type	Article ; Conference proceedings
ZDB-ID	96005-6
ISSN	1438-8685 ; 0935-8943 ; 0340-1588
ISSN (online)	1438-8685
ISSN	0935-8943 ; 0340-1588
DOI	10.1055/s-0042-1747602
Database	Thieme publisher's database

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