Evaluation of circulating microRNAs as non-invasive biomarkers in the diagnosis of ovarian cancer: a case–control study

Abstract Purpose Ovarian cancer is the seventh most frequent form of malignant diseases in women worldwide and over 150,000 women die from it every year. More than 70 percent of all ovarian cancer patients are diagnosed at a late-stage disease with poor prognosis necessitating the development of sufficient screening biomarkers. MicroRNAs displayed promising potential as early diagnostics in various malignant diseases including ovarian cancer. The presented study aimed at identifying single microRNAs and microRNA combinations detecting ovarian cancer in vitro and in vivo. Methods Intracellular, extracellular and urinary microRNA expression levels of twelve microRNAs (let-7a, let-7d, miR-10a, miR-15a, miR-15b, miR-19b, miR-20a, miR-21, miR-100, miR-125b, miR-155, miR-222) were quantified performing quantitative real-time-PCR. Therefore, the three ovarian cancer cell lines SK-OV-3, OAW-42, EFO-27 as well as urine samples of ovarian cancer patients and healthy controls were analyzed. Results MiR-15a, miR-20a and miR-222 showed expression level alterations extracellularly, whereas miR-125b did intracellularly across the analyzed cell lines. MicroRNA expression alterations in single cell lines suggest subtype specificity in both compartments. Hypoxia and acidosis showed scarce effects on single miRNA expression levels only. Furthermore, we were able to demonstrate the feasibility to clearly detect the 12 miRNAs in urine samples. In urine, miR-15a was upregulated whereas let-7a was down-regulated in ovarian cancer patients. Conclusion Intracellular, extracellular and urinary microRNA expression alterations emphasize their great potential as biomarkers in liquid biopsies. Especially, miR-15a and let-7a qualify for possible circulating biomarkers in liquid biopsies of ovarian cancer patients.

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Evaluation of Circulating microRNAs as Non-Invasive Biomarkers in the Diagnosis of Ovarian Cancer – A Case-Control Study

10.21203/rs.3.rs-28389/v1 ◽

2020 ◽

Author(s):

Kai Berner ◽

Marc Hirschfeld ◽

Daniela Weiß ◽

Gerta Rücker ◽

Jasmin Asberger ◽

...

Keyword(s):

Ovarian Cancer ◽

Cancer Patients ◽

Cell Lines ◽

Microrna Expression ◽

Urine Samples ◽

Malignant Diseases ◽

Liquid Biopsies ◽

Expression Levels

Abstract Background Ovarian cancer is the seventh most frequent form of malignant diseases in women worldwide and over 150.000 women die from it every year. More than 70 percent of all ovarian cancer patients are diagnosed at a late stage disease with poor prognosis necessitating the development of sufficient screening biomarkers. MicroRNAs displayed promising potential as early diagnostics in various malignant diseases including ovarian cancer. The presented study aimed at identifying single microRNAs and microRNA combinations detecting ovarian cancer in vitro and in vivo.Methods Intracellular, extracellular and urinary microRNA expression levels of twelve microRNAs (let-7a, let-7d, miR-10a, miR-15a, miR-15b, miR-19b, miR-20a, miR-21, miR-100, miR-125b, miR-155, miR-222) were quantified performing quantitative real-time-PCR. Therefore, the three ovarian cancer cell lines SK-OV-3, OAW-42, EFO-27 as well as urine samples of ovarian cancer patients and healthy controls were analyzed.Results MiR-15a, miR-20a and miR-222 showed expression level alterations extracellularly, whereas miR-125b did intracellularly across the analyzed cell lines. MicroRNA expression alterations in single cell lines suggest subtype specificity in both compartments. Hypoxia and acidosis showed scarce effects on single miRNA expression levels only. Furthermore, we were able to demonstrate the feasibility to clearly detect the 12 miRNAs in urine samples. In urine, miR-15a was upregulated whereas let-7a was down-regulated in ovarian cancer patients.Conclusion Intracellular, extracellular and urinary micoRNA expression alterations emphasize their great potential as biomarkers in liquid biopsies. Especially miR-15a and let-7a qualify for possible circulating biomarkers in liquid biopsies of ovarian cancer patients.

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Proline-rich protein 11 overexpression is associated with a more aggressive phenotype and poor overall survival in ovarian cancer patients

10.21203/rs.3.rs-35747/v2 ◽

2020 ◽

Author(s):

yu zhan ◽

xueyuan wu ◽

gang zheng ◽

jingjing jin ◽

chaofu li ◽

...

Keyword(s):

Ovarian Cancer ◽

Overall Survival ◽

Curative Surgery ◽

Poor Overall Survival ◽

Expression Levels ◽

Prognostic Assessment ◽

Therapy Strategies ◽

Proline Rich Protein

Abstract Background: The proline-rich protein 11 (PRR11) is a newly identified oncogene associated with a poor prognosis in several human cancers. Nonetheless, research on its role in ovarian cancer (OC) remains largely understudied. Therefore, this study aims to evaluate the expression levels of PRR11 protein and its role in human ovarian cancer. Methods: Immunohistochemistry analysis was used to evaluate the expression levels of PRR11 protein in human samples obtained from 49 patients diagnosed with OC and subjected to curative surgery in the First Affiliated Hospital of Wenzhou Medical University between 2007 and 2015. Results: In total, 57.1% of the primary OC tumor tissue evaluated demonstrated overexpression of PRR11. Meanwhile, survival analysis showed that the overall survival (OS) of patients presenting overexpression of PRR11 was significantly lower than the OS of the patients with negative PRR11. In subsequent experiments, it was found that silencing the expression of PRR11 expression inhibited the proliferation of tumor cells and the migration of cells in vitro. Further, cells subjected to PRR11 knockdown exhibited a decrease in tumor growth in vivo. The downregulation of PRR11 was coupled with a decrease in N-cadherin and downregulation in the expression of early growth response protein 1 (EGR1).Conclusions: The findings suggest that PRR11 might be considered as a potential target for prognostic assessment and gene therapy strategies for patients diagnosed with OC.

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Long non-coding RNA PTPRG-AS1 promotes cell tumorigenicity in epithelial ovarian cancer by decoying microRNA-545-3p and consequently enhancing HDAC4 expression

10.21203/rs.3.rs-51729/v3 ◽

2020 ◽

Author(s):

Juanjuan Shi ◽

Xijian Xu ◽

Dan Zhang ◽

Jiuyan Zhang ◽

Hui Yang ◽

...

Keyword(s):

Ovarian Cancer ◽

Cell Proliferation ◽

Tumor Growth ◽

Epithelial Ovarian Cancer ◽

Cell Lines ◽

Luciferase Reporter ◽

Non Coding Rna ◽

Long Non Coding Rna

Abstract Background: Long non-coding RNA PTPRG antisense RNA 1 (PTPRG-AS1) deregulation has been reported in various human malignancies and identified as an important modulator of cancer development. Few reports have focused on the detailed role of PTPRG-AS1 in epithelial ovarian cancer (EOC) and its underlying mechanism. This study aimed to determine the physiological function of PTPRG-AS1 in EOC. A series of experiments were also performed to identify the mechanisms through which PTPRG-AS1 exerts its function in EOC.Methods: Reverse transcription-quantitative polymerase chain reaction was used to determine PTPRG-AS1 expression in EOC tissues and cell lines. PTPRG-AS1 was silenced in EOC cells and studied with respect to cell proliferation, apoptosis, migration, and invasion in vitro and tumor growth in vivo. The putative miRNAs that target PTPRG-AS1 were predicted using bioinformatics analysis and further confirmed in luciferase reporter and RNA immunoprecipitation assays.Results: Our data verified the upregulation of PTPRG-AS1 in EOC tissues and cell lines. High PTPRG-AS1 expression was associated with shorter overall survival in patients with EOC. Functionally, EOC cell proliferation, migration, invasion in vitro, and tumor growth in vivo were suppressed by PTPRG-AS1 silencing. In contrast, cell apoptosis was promoted by loss of PTPRG-AS1. Regarding the mechanism, PTPRG-AS1 could serve as a competing endogenous RNA in EOC cells by decoying microRNA-545-3p (miR-545-3p), thereby elevating histone deacetylase 4 (HDAC4) expression. Furthermore, rescue experiments revealed that PTPRG-AS1 knockdown-mediated effects on EOC cells were, in part, counteracted by the inhibition of miR-545-3p or restoration of HDAC4.Conclusions: PTPRG-AS1 functioned as an oncogenic lncRNA that aggravated the malignancy of EOC through the miR-545-3p/HDAC4 ceRNA network. Thus, targeting the PTPRG-AS1/miR-545-3p/HDAC4 pathway may be a novel strategy for EOC anticancer therapy.

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Proline-rich protein 11 overexpression is associated with a more aggressive phenotype and poor overall survival in ovarian cancer patients

10.21203/rs.3.rs-35747/v3 ◽

2020 ◽

Author(s):

yu zhan ◽

xueyuan wu ◽

gang zheng ◽

jingjing jin ◽

chaofu li ◽

...

Keyword(s):

Ovarian Cancer ◽

Overall Survival ◽

Curative Surgery ◽

Poor Overall Survival ◽

Expression Levels ◽

Prognostic Assessment ◽

Therapy Strategies ◽

Proline Rich Protein

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LY2157299 Monohydrate, a TGF-βR1 Inhibitor, Suppresses Tumor Growth and Ascites Development in Ovarian Cancer

Cancers ◽

10.3390/cancers10080260 ◽

2018 ◽

Vol 10 (8) ◽

pp. 260 ◽

Cited By ~ 9

Author(s):

Qing Zhang ◽

Xiaonan Hou ◽

Bradley Evans ◽

Jamison VanBlaricom ◽

Saravut Weroha ◽

...

Keyword(s):

Ovarian Cancer ◽

Tumor Growth ◽

Cell Lines ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Migration And Invasion ◽

Ascites Formation ◽

Tgf Β1

Transforming growth factor beta (TGF-β) signaling has pleiotropic functions regulating cancer initiation, development, and metastasis, and also plays important roles in the interaction between stromal and cancer cells, making the pathway a potential therapeutic target. LY2157299 monohydrate (LY), an inhibitor of TGF-β receptor I (TGFBRI), was examined for its ability to inhibit ovarian cancer (OC) growth both in high-grade serous ovarian cancer (HGSOC) cell lines and xenograft models. Immunohistochemistry, qRT-PCR, and Western blot were performed to study the effect of LY treatment on expression of cancer- and fibroblast-derived genes. Results showed that exposure to TGF-β1 induced phosphorylation of SMAD2 and SMAD3 in all tested OC cell lines, but this induction was suppressed by pretreatment with LY. LY alone inhibited the proliferation, migration, and invasion of HGSOC cells in vitro. TGF-β1-induced fibroblast activation was blocked by LY. LY also delayed tumor growth and suppressed ascites formation in vivo. In addition, independent of tumor inhibition, LY reduces ascites formation in vivo. Using OVCAR8 xenograft specimens we confirmed the inhibitory effect of LY on TGF-β signaling and tumor stromal expression of collagen type XI chain 1 (COL11A1) and versican (VCAN). These observations suggest a role for anti-TGF-β signaling-directed therapy in ovarian cancer.

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Targeting RNF8 Effectively Reverse Cisplatin and Doxorubicin Resistance in Endometrial Cancer

10.21203/rs.3.rs-57263/v1 ◽

2020 ◽

Author(s):

Ben Yang ◽

Wang Ke ◽

Yingchun Wan ◽

Tao Li

Keyword(s):

Endometrial Cancer ◽

Cell Lines ◽

Quantitative Polymerase Chain Reaction ◽

Mrna Levels ◽

Mice Model ◽

Expression Levels ◽

Gynecological Malignancy ◽

Ec Cells

Abstract Background Endometrial cancer (EC) is one of the most frequent gynecological malignancy worldwide. However, resistance to chemotherapy remains one of the major difficulties in the treatment of EC. Thus, there is an urgent requirement to understand mechanisms of chemoresistance and identify novel regimens for patients with EC. Methods Cisplatin and doxorubicin resistant cell lines were acquired by continuous exposing parental EC cells to cisplatin or doxorubicin for 3 months. Cell viability was determined by using MTT assay. Protein Expression levels of protein were examined by western blotting assay. mRNA levels were measured by quantitative polymerase chain reaction (qPCR) assay. Ring finger protein 8 (RNF8) knockout cell lines were generated by clustered regularly interspaced short palindromic repeats (CRISPR)–Cas9 gene editing assay. Nonhomologous end joining (NHEJ) efficiency were quantified by plasmid based NHEJ assay. DNA double strand breaks (DSB) were generated using laser micro-irradiation. Protein recruitment to DSB was analyzed by immunofluorescent assay. Tumor growth was examined by AN3CA xenograft mice model. Results We found that protein and mRNA expression levels of RNF8 were significantly increased in both cisplatin and doxorubicin resistant EC cells. Cell survival assay showed that RNF deficiency significantly enhanced the sensitivity of resistant EC cells to cisplatin and doxorubicin (P < 0.01). In addition, chemoresistant EC cells exhibited increased NHEJ efficiency. Knockout of RNF8 in chemoresistant EC cells significantly reduced NHEJ efficiency and prolonged Ku80 retention on DSB. Moreover, cisplatin resistant AN3CA xenograft showed that RNF8 deficiency overcame cisplatin resistance. Conclusions Our in vitro and in vivo assays provide evidence for RNF8, which is a NHEJ factor, serving as a promising, novel target in EC chemotherapy.

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Functional Studies on Viable Circulating Tumor Cells

Clinical Chemistry ◽

10.1373/clinchem.2015.242537 ◽

2016 ◽

Vol 62 (2) ◽

pp. 328-334 ◽

Cited By ~ 53

Author(s):

Klaus Pantel ◽

Catherine Alix-Panabières

Keyword(s):

Cancer Patients ◽

Tumor Cells ◽

Circulating Tumor Cells ◽

Cancer Diagnostics ◽

Published Data ◽

Liquid Biopsies ◽

In Vivo Models ◽

Functional Studies

AbstractBACKGROUNDResearch on circulating tumor cells (CTCs) as new biomarkers has received great attention over the past decade. In particular, the capture and analysis of CTCs as “liquid biopsies” provides the possibility to avoid invasive tissue biopsies, with obvious implications in cancer diagnostics.CONTENTThe focus of this review is to describe and discuss how functional studies on viable CTCs can enlarge the spectrum of applications of liquid biopsies, with emphasis on breast, prostate, colon, and lung cancer as the major tumor entities in industrialized countries. The low number of CTCs in the peripheral blood of most cancer patients makes challenging the in vitro culture of CTCs. Epithelial tumor cells are difficult to culture, even when starting with millions of tumor cells. Recently, several groups have achieved important advances in the in vitro and in vivo expansion of CTCs from cancer patients at very advanced stages with higher amounts of CTCs. Here, we present current technologies to enrich and detect viable human CTCs, including positive and negative enrichment strategies that are based on antigen expression and physical properties of CTCs. We also discuss published data about functional studies on CTCs that use in vitro and in vivo models.SUMMARYFunctional analyses on CTCs offer the possibility to identify the biological properties of metastatic cells, including the identification of metastasis-initiating cells. Moreover, CTC-derived cell lines and xenografts might reveal new therapeutic targets and can be used for drug screening.

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Quantitative Expression and Virus Transmission Analysis of DC-SIGN on Monocyte-Derived Dendritic Cells

Journal of Virology ◽

10.1128/jvi.76.18.9135-9142.2002 ◽

2002 ◽

Vol 76 (18) ◽

pp. 9135-9142 ◽

Cited By ~ 86

Author(s):

Frédéric Baribaud ◽

Stefan Pöhlmann ◽

George Leslie ◽

Frank Mortari ◽

Robert W. Doms

Keyword(s):

Dendritic Cells ◽

Cell Lines ◽

Ebola Virus ◽

Virus Transmission ◽

Virus Binding ◽

Expression Levels ◽

Quantitative Expression ◽

Immunodeficiency Virus

ABSTRACT The C-type lectins DC-SIGN and DC-SIGNR efficiently bind human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) strains and can transmit bound virus to adjacent CD4-positive cells. DC-SIGN also binds efficiently to the Ebola virus glycoprotein, enhancing Ebola virus infection. DC-SIGN is thought to be responsible for the ability of dendritic cells (DCs) to capture HIV and transmit it to T cells, thus promoting HIV dissemination in vitro and perhaps in vivo as well. To investigate DC-SIGN function and expression levels on DCs, we characterized a panel of monoclonal antibodies (MAbs) directed against the carbohydrate recognition domain of DC-SIGN. Using quantitative fluorescence-activated cell sorter technology, we found that DC-SIGN is highly expressed on immature monocyte-derived DCs, with at least 100,000 copies and often in excess of 250,000 copies per DC. There was modest variation (three- to fourfold) in DC-SIGN expression levels between individuals and between DCs isolated from the same individual at different times. Several MAbs efficiently blocked virus binding to cell lines expressing human or rhesus DC-SIGN, preventing HIV and SIV transmission. Interactions with Ebola virus pseudotypes were also blocked efficiently. Despite their ability to block virus-DC-SIGN interactions on cell lines, these antibodies only inhibited transmission of virus from DCs by approximately 50% or less. These results indicate that factors other than DC-SIGN may play important roles in the ability of DCs to capture and transmit HIV.

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LncRNA SNHG20 promotes migration and invasion of ovarian cancer via modulating the microRNA-148a/ROCK1 axis

Journal of Ovarian Research ◽

10.1186/s13048-021-00889-8 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Qi Yang ◽

Yu-Jie Dong

Keyword(s):

Ovarian Cancer ◽

Cell Lines ◽

Noncoding Rna ◽

Tumor Xenograft ◽

Migration And Invasion ◽

Epithelial Cell Line ◽

Invasion And Migration ◽

And Migration

Abstract Background Ovarian cancer (OC) is characterized by early metastasis and poor prognosis, which threatens the health of women worldwide. Small nucleolar RNA host gene 20 (SNHG20), a long noncoding RNA (lncRNA), has been verified to be significantly up-regulated in several tumors, including OC. MicroRNA-148a (miR-148a)/rho-kinase1 (ROCK1) axis plays an important role in the modulation of tumor development. However, whether SNHG20 can regulate OC progression through miR-148a/ROCK1 axis remains unclear. Normal human ovarian epithelial cell line and four OC cell lines were adopted for in vitro experiments. Real-time PCR was performed to assess the levels of SNHG20 and miR-148a. OC cell proliferation, apoptosis, invasion and migration were detected using clone formation, flow cytometry, transwell, and wound healing assays, respectively. Tumor xenograft assay was applied to evaluate the effect of SNHG20 on tumor growth in vivo. Results Significant higher expression of SNHG20 was observed in OC cell lines. SNHG20 markedly promoted the invasion, migration, proliferation and inhibited the apoptosis of OC cells. SNHG20 enhanced ROCK1 expression by sponging miR-148a, and the direct binding between SNHG20/ROCK1 and miR-148a was identified. Conclusion SNHG20 promoted invasion and migration of OC via targeting miR-148a/ROCK1 axis. The present research may provide a novel insight for the therapeutic strategies of OC.

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Long Non-coding RNA PTPRG-AS1 Promotes Cell Tumorigenicity in Epithelial Ovarian Cancer by Decoying microRNA-545-3p and Consequently Enhancing HDAC4 Expression

10.21203/rs.3.rs-51729/v1 ◽

2020 ◽

Author(s):

Juanjuan Shi ◽

Xijian Xu ◽

Dan Zhang ◽

Jiuyan Zhang ◽

Hui Yang ◽

...

Keyword(s):

Ovarian Cancer ◽

Cell Proliferation ◽

Tumor Growth ◽

Epithelial Ovarian Cancer ◽

Cell Lines ◽

Luciferase Reporter ◽

Non Coding Rna ◽

Long Non Coding Rna

Abstract Background: Long non-coding RNA PTPRG antisense RNA 1 (PTPRG-AS1) deregulation has been reported in various human malignancies and identified as an important modulator of cancer development. Few reports have focused on the detailed role of PTPRG-AS1 in epithelial ovarian cancer (EOC) and its underlying mechanism. This study aimed to determine the physiological function of PTPRG-AS1 in EOC. A series of experiments were also done to identify the mechanisms through which PTPRG-AS1 exerts its function in EOC.Methods: Reverse transcription-quantitative polymerase chain reaction was used to determine PTPRG-AS1 expression in EOC tissues and cell lines. PTPRG-AS1 was silenced in EOC cells and studied with respect to cell proliferation, apoptosis, migration, invasion in vitro, and tumor growth in vivo. The putative miRNAs that target PTPRG-AS1 were predicted using bioinformatics analysis and further confirmed in luciferase reporter and RNA immunoprecipitation assays.Results: Our data verified the upregulation of PTPRG-AS1 in EOC tissues and cell lines. High PTPRG-AS1 expression was associated with shorter overall survival in patients with EOC. Functionally, EOC cell proliferation, migration, invasion in vitro and tumor growth in vivo were suppressed by PTPRG-AS1 silencing. In contrast, cell apoptosis was promoted by loss of PTPRG-AS1. For the mechanism part, PTPRG-AS1 could serve as a competing endogenous RNA in EOC cells by decoying microRNA-545-3p (miR-545-3p), thereby elevating histone deacetylase 4 (HDAC4) expression. Furthermore, rescue experiments revealed that PTPRG-AS1 knockdown-mediated effects on EOC cells were, in part, counteracted by the inhibition of miR-545-3p or restoration of HDAC4.Conclusions: PTPRG-AS1 functioned as an oncogenic lncRNA that aggravated the malignancy of EOC through the miR-545-3p/HDAC4 ceRNA network. Thus, targeting the PTPRG-AS1/miR-545-3p/HDAC4 pathway may be a novel strategy for EOC anticancer therapy.

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