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Strategy for Biobanking of Ovarian Cancer Organoids: Addressing the Interpatient Heterogeneity across Histological Subtypes and Disease Stages

While the establishment of an ovarian cancer biobank from patient-derived organoids along with their clinical background information promises advances in ...

University Hospital, Ludwig-Maximilian-University (LMU) Munich

Malignant ascites in ovarian carcinoma patients is associated with poor prognosis and reduced quality of life. The trifunctional antibody catumaxomab (anti-EpCAM x anti-CD3) enhances the antitumor activity by redirecting T cells and Fcgamma receptor I/III--positive accessory cells to the tumor. This multicenter phase I/II dose-escalating study investigated tolerability and efficacy of i.p. catumaxomab application in ovarian cancer patients with malignant ascites containing epithelial cell adhesion molecule (EpCAM)--positive tumor cells.

Effective relief of malignant ascites in patients with advanced ovarian cancer by a trifunctional anti-EpCAM x anti-CD3 antibody: a phase I/II study.

Patients with ovarian cancer usually present to a family physician with nonspecific symptoms, most often abdominal pain. The outcome depends above all on the stage of the disease when it is diagnosed and on the quality of treatment.

Ovarian cancer: diagnosis and treatment.

Nuclear factor erythroid 2-related factor 2 (NRF2) regulates cytoprotective antioxidant processes. In this study, the prognostic potential of NRF2 and its interactions with the estrogen receptor α (ERα) in ovarian cancer cells was investigated. NRF2 and ERα protein expression in ovarian cancer tissue was analyzed as well as mRNA expression of NRF2 () and ERα () in four ovarian cancer and one benign cell line.  silencing was carried out to evaluate a potential interplay between NRF2 and ERα. Cytoplasmic NRF2 expression as inactive form had significantly higher expression in patients with low-grade histology ( = 0.03). In the serous cancer subtype, high cytoplasmic NRF2 expression (overall survival (OS), median 50.6 vs. 29.3 months;  = 0.04) and high ERα expression (OS, median 74.5 vs. 27.1 months;  = 0.002) was associated with longer overall survival as well as combined expression of both inactive cytoplasmic NRF2 and ERα in the whole cohort (median 74.5 vs. 37.7 months;  = 0.04). Cytoplasmic NRF2 expression showed a positive correlation with ERα expression ( = 0.004).  was found to be highly expressed in the ovarian cancer cell lines OVCAR3, UWB1.289, and TOV112D. Compared with the benign cell line HOSEpiC,  expression was reduced in all ovary cancer cell lines (all  < 0.001). Silencing of  induced a higher mRNA expression of  in the  downregulated cancer cell lines OVCAR3 ( = 0.003) and ES2 ( < 0.001), confirming genetic interactions of NRF2 and ERα. In this study, both inactive cytoplasmic NRF2 and high ERα expression were demonstrated to be associated with improved survival in ovarian cancer patients. Further understanding of interactions within the estradiol⁻ERα⁻NRF2 pathway could better predict the impact of endocrine therapy in ovarian cancer.

Interaction of ERα and NRF2 Impacts Survival in Ovarian Cancer Patients.

Expression of the aryl hydrocarbon receptor (AhR) has been described in various tumor entities from different organs. However, its role in ovarian cancer has not been thoroughly investigated. We aimed to elucidate the prognostic impact of AhR, its correlation with the follicle-stimulating hormone receptor (FSHR), and their functional role in ovarian cancer. By immunohistochemistry, AhR staining was analyzed in a subset of 156 samples of ovarian cancer patients. AhR staining was assessed in the nucleus and the cytoplasm using the semi-quantitative immunoreactive score (IRS), and the scores were grouped into high- and low-level expression. AhR expression was detected in all histological subtypes, with clear cell ovarian cancer displaying the highest staining intensity. Low cytoplasmic expression of AhR was associated with longer overall survival (median 183.46 vs. 85.07 months;  = 0.021). We found a positive correlation between AhR and FSHR ( = 0.005). Ovarian cancer patients with high cytoplasmic AhR and concurrent FSHR expression had the worst outcome (median 69.72 vs. 43.32 months;  = 0.043). Consequently, low cytoplasmic AhR expression seems to be associated with improved survival in ovarian cancer patients. Our data suggest that AhR and FSHR levels correlate with each other, and their concurrent expression was observed in ovarian cancer patients with the worst outcome. Further investigation of the interaction of both receptors and their functional role might better predict the impact of endocrine therapy in ovarian cancer.

Correlation of the Aryl Hydrocarbon Receptor with FSHR in Ovarian Cancer Patients.

Multi drug resistance protein 1 (MDR1) expression on tumor cells has been widely investigated in context of drug resistance. However, the role of MDR1 on the immune cell infiltrate of solid tumors remains unknown. The aim of this study was to analyze the prognostic significance of a MDR1+ immune cell infiltrate in epithelial ovarian cancer (EOC) and to identify the MDR1+ leucocyte subpopulation. MDR1 expression was analyzed by immunohistochemistry in 156 EOC samples. In addition to MDR1+ cancer cells, we detected a MDR1+ leucocyte infiltrate (high infiltrate >4 leucocytes per field of view). Correlations and survival analyses were calculated. To identify immune cell subpopulations immunofluorescence double staining was performed. The MDR1+ leucocyte infiltrate was associated with human epidermal growth factor receptor 2 (HER2) (cc = 0.258,  = 0.005) and tumor-associated mucin 1 (TA-MUC1) (cc = 0.202,  = 0.022) expression on cancer cells. A high MDR1+ leucocyte infiltrate was associated with impaired survival, especially in patients whose carcinoma showed either serous histology (median OS 28.80 vs. 50.64 months,  = 0.027, n = 91) or TA-MUC1 expression (median OS 30.60 vs. 63.36 months,  = 0.015, n = 110). Similar findings for PFS suggest an influence of MDR1+ immune cells on the development of chemoresistance. A Cox regression analysis confirmed the independency of a high MDR1+ leucocyte infiltrate as prognostic factor. M2 macrophages were identified as main part of the MDR1+ leucocyte infiltrate expressing MDR1 as well as the M2 marker CD163 and the pan-macrophage marker CD68. Infiltration of MDR1+ leucocytes, mostly M2 macrophages, is associated with poor prognosis of EOC patients. Further understanding of the interaction of M2 macrophages, MDR1 and TA-MUC1 appears to be a key aspect to overcome chemoresistance in ovarian cancer.

M2 Macrophages Infiltrating Epithelial Ovarian Cancer Express MDR1: A Feature That May Account for the Poor Prognosis.

The vitamin D receptor (VDR), primarily known as a crucial mediator of calcium homeostasis and metabolism, has been shown to play a significant role in various cancer entities. Previous studies have focused on vitamin D and its receptor in gynecological cancers, noting that the receptor is upregulated in epithelial ovarian cancer (EOC). The aim of this study is to analyze the prognostic impact of VDR and its functional significance in ovarian cancer. Through immunohistochemistry, VDR staining was examined in 156 ovarian cancer samples. Evaluation of VDR staining was conducted in the nucleus and the cytoplasm using the semi-quantitative immunoreactive score, and the scores were classified into high- and low-level expressions. Expression levels were correlated with clinical and pathological parameters as well as with overall survival to assess for prognostic impact. Differences in cytoplasmic VDR expression were identified between the histological subtypes (p = 0.001). Serous, clear cell, and endometrioid subtypes showed the highest staining, while the mucinous subtype showed the lowest. Cytoplasmic VDR correlated with higher FIGO stage (p = 0.013; Cc = 0.203), positive lymph node status (p = 0.023; Cc = 0.236), high-grade serous histology (p = 0.000; Cc = 0.298) and grading from the distinct histological subtypes (p = 0.006; Cc = - 0.225). Nuclear VDR did not correlate with clinicopathological data. High cytoplasmic expression of VDR was associated with impaired overall survival (HR 2.218, 32.5 months vs. median not reached; p < 0.001) and was confirmed as a statistically independent prognostic factor in the Cox regression multivariate analysis. Additional knowledge of VDR as a biomarker and its interactions within the mitogen-activated protein kinase (MAPK) signaling pathway could potentially improve the prognosis of therapeutic approaches for specific subgroups in EOC.

Cytoplasmic VDR expression as an independent risk factor for ovarian cancer.

Establishing reliable prognostic factors as well as specific targets for new therapeutic approaches is an urgent requirement in advanced ovarian cancer. For several tumor entities, the ubiquitously spread scaffold protein β-arrestin 2, a multifunctional scaffold protein regulating signal transduction and internalization of activated G protein-coupled receptors (GPCRs), has been considered with rising interest for carcinogenesis. Therefore, we aimed to elucidate the prognostic impact of β-arrestin 2 and its functional role in ovarian cancer. β-arrestin 2 expression was analyzed in a subset of 156 samples of ovarian cancer patients by immunohistochemistry. Cytoplasmic expression levels were correlated with clinical as well as pathological characteristics and with prognosis. The biologic impact of β-arrestin 2 on cell proliferation and survival was evaluated, . Following transient transfection by increasing concentrations of plasmid encoding β-arrestin 2, different cell lines were evaluated in cell viability and death. β-arrestin 2 was detected in all histological ovarian cancer subtypes with highest intensity in clear cell histology. High β-arrestin 2 expression levels correlated with high-grade serous histology and the expression of the gonadotropin receptors FSHR and LHCGR, as well as the membrane estrogen receptor GPER and hCGβ. Higher cytoplasmic β-arrestin 2 expression was associated with a significantly impaired prognosis (median 29.88 vs. 50.64 months;  = 0.025). Clinical data were confirmed in transfected HEK293 cells, human immortalized granulosa cell line (hGL5) and the ovarian cancer cell line A2780 , where the induction of β-arrestin 2 cDNA expression enhanced cell viability, while the depletion of the molecule by siRNA resulted in cell death. Reflecting the role of β-arrestin 2 in modulating GPCR-induced proliferative and anti-apoptotic signals, we propose β-arrestin 2 as an important prognostic factor and also as a promising target for new therapeutic approaches in advanced ovarian cancer.

β-arrestin 2 Is a Prognostic Factor for Survival of Ovarian Cancer Patients Upregulating Cell Proliferation.

Enolase-1, primarily known for its role in glucose metabolism, is overexpressed in various cancer entities. In contrast its alternative spliced nuclear isoform MBP-1 acts as a tumor suppressor. The aim of this study is to analyze the prognostic impact of Enolase-1/ MBP-1 and its functional significance in epithelial ovarian cancer (EOC).

Nuclear Enolase-1/ MBP-1 expression and its association with the Wnt signaling in epithelial ovarian cancer.

Aberrantly activated Wnt/β-catenin signaling pathway, as well as platelet-activating factor (PAF), contribute to cancer progression and metastasis of many cancer entities. Nonetheless, the role of the degradation enzyme named platelet-activating factor acetylhydrolase (PLA2G7/PAF-AH) in ovarian cancer etiology is still unclear. This study investigated the functional impact of platelet-activating factor acetylhydrolase on BRCA1 mutant ovarian cancer biology and its crosstalk with the Wnt signaling pathway. PAF-AH, pGSK3β, and β-catenin expressions were analyzed in 156 ovarian cancer specimens by immunohistochemistry. PAF-AH expression was investigated in ovarian cancer tissue, serum of BRCA1-mutated patients, and in vitro in four ovarian cancer cell lines. Functional assays were performed after PLA2G7 silencing. The association of PAF-AH and β-catenin was examined by immunocytochemistry. In an established ovarian carcinoma collective, we identified PAF-AH as an independent positive prognostic factor for overall survival (median 59.9 vs. 27.4 months;  = 0.016). PAF-AH correlated strongly with the Wnt signaling proteins pGSK3β (Y216; nuclear: cc = 0.494,  < 0.001; cytoplasmic: cc = 0.488,  < 0.001) and β-catenin (nuclear: cc = 0.267,  = 0.001; cytoplasmic: cc = 0.291,  < 0.001). In particular, high levels of PAF-AH were found in tumor tissue and in the serum of BRCA1 mutation carriers. By in vitro expression analysis, a relevant gene and protein expression of PLA2G7/PAF-AH was detected exclusively in the BRCA1-negative ovarian cancer cell line UWB1.289 ( < 0.05). Functional assays showed enhanced viability, proliferation, and motility of UWB1.289 cells when PLA2G7/PAF-AH was downregulated, which underlines its protective character. Interestingly, by siRNA knockdown of PLA2G7/PAF-AH, the immunocytochemistry staining pattern of β-catenin changed from a predominantly membranous expression to a nuclear one, suggesting a negative regulatory role of PAF-AH on the Wnt/β-catenin pathway. Our data provide evidence that PAF-AH is a positive prognostic factor with functional impact, which seems particularly relevant in BRCA1 mutant ovarian cancer. For the first time, we show that its protective character may be mediated by a negative regulation of the Wnt/β-catenin pathway. Further studies need to specify this effect. Potential use of PAF-AH as a biomarker for predicting the disease risk of BRCA1 mutation carriers and for the prognosis of patients with BRCA1-negative ovarian cancer should be explored.

Platelet-Activating Factor Acetylhydrolase Expression in BRCA1 Mutant Ovarian Cancer as a Protective Factor and Potential Negative Regulator of the Wnt Signaling Pathway.

The platelet-activating factor receptor (PAFR) and its ligand (PAF) are important inflammatory mediators that are overexpressed in ovarian cancer. The receptor is an important player in ovarian cancer development. In this study, we aimed to evaluate the prognostic value of PAFR in epithelial ovarian cancer (EOC) and the potential use of its antagonist, rupatadine, as an experimental treatment. Tissue microarrays of ovarian cancer patients, most markedly those with a non-mucinous subtype, immunohistochemically overexpressed PAFR. Elevated cytoplasmic PAFR expression was found to significantly and independently impair patients' overall and recurrence-free survival (OS: median 83.48 vs. 155.03 months;  = 0.022; RFS: median 164.46 vs. 78.03 months;  = 0.015). In vitro, the serous ovarian cancer subtypes especially displayed an elevated PAFR gene and protein expression. siRNA knockdown of PAFR decreased cell proliferation significantly, thus confirming the receptor's protumorigenic effect on ovarian cancer cells. The clinically approved PAFR antagonist rupatadine effectively inhibited in vitro cell proliferation and migration of ovarian cancer cells. PAFR is a prognostic marker in ovarian cancer patients and its inhibition through rupatadine may have important therapeutic implications in the therapy of ovarian cancer patients.

The Platelet-Activating Factor Receptor's Association with the Outcome of Ovarian Cancer Patients and Its Experimental Inhibition by Rupatadine.

There are different studies worldwide, which have shown a higher risk of mental disorders due to the COVID-19 pandemic. One aim of this study was to identify influencing factors of the psychological burden related to the COVID-19 pandemic and the impact on the development of postpartum depression. Further, the role of individual stress and coping strategies was analyzed in this context.

Risk of postpartum depressive symptoms is influenced by psychological burden related to the COVID-19 pandemic and dependent of individual stress coping.

Past studies have confirmed that aberrant activation of the Wnt/β-catenin signaling is associated with tumorigenesis and metastasis in breast cancer, while the role of platelet-activating factor acetylhydrolase (PLA2G7/PAF-AH) in this signaling pathway remains unclear. In this study, we analyze the functional impact of PAF-AH on BRCA1 mutant breast cancer and explore its relationship to the Wnt signaling pathway. By performing immunohistochemistry, PAF-AH expression and β-catenin expression were examined in both BRCA1 WT and BRCA1 mutant breast cancer specimens. The BRCA1 mutant breast cancer cell line HCC1937 was used for in vitro experiments to assess the impact of PAF-AH on cellular functions. The intracellular distribution of β-catenin depending on PLA2G7/PAF-AH expression was investigated by immunocytochemistry. Significantly higher nuclear expression levels of PAF-AH were found in BRCA1 mutant tissue specimens than in BRCA1 WT samples. Cell viability, proliferation, and the motility rate of HCC1937 were significantly enhanced after PLA2G7 silencing, which indicated a protective role of PAF-AH in breast cancer. Nuclear PAF-AH expressed correlatedly with membranous β-catenin. PLA2G7 silencing provoked the β-catenin translocation from the membrane to the nucleus and activated Wnt signaling downstream genes. Our data showed a protective effect of high PAF-AH expression in BRCA1 mutant breast cancer. PAF-AH may achieve its protective effect by negatively regulating the Wnt pathway. In conclusion, our research sheds new light on the regulatory pathways in BRCA1 mutant breast cancer.

PLA2G7/PAF-AH as Potential Negative Regulator of the Wnt Signaling Pathway Mediates Protective Effects in BRCA1 Mutant Breast Cancer.

The significance of the non-classical G-protein-coupled estrogen receptor (GPER) as positive or negative prognostic factor for ovarian cancer patients remains still controversial. Recent results indicate that an imbalance of both co-factors and co-repressors of nuclear receptors regulates ovarian carcinogenesis by altering the transcriptional activity through chromatin remodeling. The present study aims to investigate whether the expression of the nuclear co-repressor NCOR2 plays a role in GPER signaling which thereby could positively impact overall survival rates of ovarian cancer patients.

Article	Year
Effective relief of malignant ascites in patients with advanced ovarian cancer by a trifunctional anti-EpCAM x anti-CD3 antibody: a phase I/II study. Clinical cancer research : an official journal of the American Association for Cancer Research\| PubMed ID: 17606723	2007
Ovarian cancer: diagnosis and treatment. Deutsches Arzteblatt international\| PubMed ID: 22025930	2011
Interaction of ERα and NRF2 Impacts Survival in Ovarian Cancer Patients. International journal of molecular sciences\| PubMed ID: 30597961	2018
Correlation of the Aryl Hydrocarbon Receptor with FSHR in Ovarian Cancer Patients. International journal of molecular sciences\| PubMed ID: 31212758	2019
M2 Macrophages Infiltrating Epithelial Ovarian Cancer Express MDR1: A Feature That May Account for the Poor Prognosis. Cells\| PubMed ID: 32429133	2020
Cytoplasmic VDR expression as an independent risk factor for ovarian cancer. Histochemistry and cell biology\| PubMed ID: 32572587	2020
β-arrestin 2 Is a Prognostic Factor for Survival of Ovarian Cancer Patients Upregulating Cell Proliferation. Frontiers in endocrinology\| PubMed ID: 33042017	2020
Nuclear Enolase-1/ MBP-1 expression and its association with the Wnt signaling in epithelial ovarian cancer. Translational oncology\| PubMed ID: 33069100	2021
Platelet-Activating Factor Acetylhydrolase Expression in BRCA1 Mutant Ovarian Cancer as a Protective Factor and Potential Negative Regulator of the Wnt Signaling Pathway. Biomedicines\| PubMed ID: 34206491	2021
The Platelet-Activating Factor Receptor's Association with the Outcome of Ovarian Cancer Patients and Its Experimental Inhibition by Rupatadine. Cells\| PubMed ID: 34571986	2021
Risk of postpartum depressive symptoms is influenced by psychological burden related to the COVID-19 pandemic and dependent of individual stress coping. Archives of gynecology and obstetrics\| PubMed ID: 36482113	2023
PLA2G7/PAF-AH as Potential Negative Regulator of the Wnt Signaling Pathway Mediates Protective Effects in BRCA1 Mutant Breast Cancer. International journal of molecular sciences\| PubMed ID: 36614323	2023
Nuclear receptor co-repressor NCOR2 and its relation to GPER with prognostic impact in ovarian cancer. Journal of cancer research and clinical oncology\| PubMed ID: 37131060	2023

Alexander Burges

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