Impact of RARα and miR-138 on retinoblastoma etoposide resistance

BACKGROUND: Retinoblastoma (RB) is the most common childhood eye cancer. Chemotherapeutic drugs such as etoposide used in RB treatment often cause massive side effects and acquired drug resistances. Dysregulated genes and miRNAs have a large impact on cancer progression and development of chemotherapy resistances. OBJECTIVE: This study was designed to investigate the involvement of retinoic acid receptor alpha (RARα) in RB progression and chemoresistance as well as the impact of miR-138, a potential RARα regulating miRNA. METHODS: RARα and miR-138 expression in etoposide resistant RB cell lines and chemotherapy treated patient tumors compared to non-treated tumors was revealed by Real-Time PCR. Overexpression approaches were performed to analyze the effects of RARα on RB cell viability, apoptosis, proliferation and tumorigenesis. Besides, we addressed the effect of miR-138 overexpression on RB cell chemotherapy resistance. RESULTS: A binding between miR-138 and RARα was shown by dual luciferase reporter gene assay. The study presented revealed that RARα is downregulated in etoposide resistant RB cells, while miR-138 is endogenously upregulated. Opposing RARα and miR-138 expression levels were detectable in chemotherapy pre-treated compared to non-treated RB tumor specimen. Overexpression of RARα increases apoptosis levels and reduces tumor cell growth of aggressive etoposide resistant RB cells in vitro and in vivo. Overexpression of miR-138 in chemo-sensitive RB cell lines partly enhances cell viability after etoposide treatment. CONCLUSIONS: Our findings show that RARα acts as a tumor suppressor in retinoblastoma and is downregulated upon etoposide resistance in RB cells. Thus, RARα may contribute to the development and progression of RB chemo-resistance.

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MiR-203 Determines Poor Outcome and Suppresses Tumor Growth by Targeting TBK1 in Osteosarcoma

Cellular Physiology and Biochemistry ◽

10.1159/000438556 ◽

2015 ◽

Vol 37 (5) ◽

pp. 1956-1966 ◽

Cited By ~ 11

Author(s):

Shiping Liu ◽

Peng Feng

Keyword(s):

Cell Growth ◽

Cell Lines ◽

Cancer Progression ◽

Human Cancer ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

Poor Overall Survival ◽

Gene Assay

Background/Aims: Increasing evidence has shown that miR-203 plays important role in human cancer progression. However, little is known about the function of miR-203 in osteosarcoma (OS). Methods: The expression of miR-203 in OS tissues and cell lines were examined by qRT-PCR. The biological role of miR-20 in OS cell proliferation was examined in vitro and in vivo. The targets of miR-203 were identified by a luciferase reporter gene assay. Results: miR-203 was down regulated in OS tissues and cell lines; decreased miR-203 was associated with a poor overall survival in OS patients. Restoration of miR-203 expression reduced cell growth in vitro and suppressed tumorigenicity in vivo. In contrast, inhibition of miR-203 stimulated OS cell growth both in vitro and in vivo. In addition, TANK binding kinase 1 (TBK1) was identified as a direct target of miR-203; overexpression of TBK1 partly reversed the suppressive effects of miR-203. Furthermore, TBK1 was found up-regulated and inversely correlated with miR-203 in OS tissues. Conclusion: Taken together, these findings suggest that miR-203 acts as a tumor suppressor via regulation of TBK1 expression in OS progression, and miR-203 may be a promising therapeutic target for OS.

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NR2F2-AS1 Suppressed Trastuzumab Effects on Esophageal Cancer By Inhibiting miR-4429 and miR-425-5p Expression Through Targeting IGF1R

10.21203/rs.3.rs-401449/v1 ◽

2021 ◽

Author(s):

Zhibin Zhang ◽

Xu Zhao ◽

Rui Zhu ◽

Hong Ren

Keyword(s):

Esophageal Cancer ◽

Cell Viability ◽

Rna Sequencing ◽

Inhibitory Effect ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

Luciferase Reporter Gene Assay ◽

Gene Assay

Abstract Aim:In this manuscript, we aimed to investigate the involvement of non-coding RNAs in mediating trastuzumab effects in EAC. Background: Scarce evidences supported that targeted drugs, like Trastuzumab, can be applied to esophageal adenocarcinoma patients (EAC). Objective: Evaluating the role and mechanism of NR2F2-AS1 in regulating Trastuzumab effects in EAC patients. Method: RNA sequencing to screen IGF1R related lncRNAs. qRT-PCR and western blot were used to evaluate the expression level of genes. CCK-8 was used to test the cell proliferation ability. Dual luciferase reporter gene assay and RNA pull-down were used for crosstalk evaluation. Results: NR2F2-AS1 was identified to be associated with HER2 expression by RNA sequencing and its expression related to worse prognosis and advanced T and N stage.NR2F2-AS1 expression induced by Trastuzumab through mediating H3K27ac. Furtherly, miR-4429 and miR-425-5p, which were predicted and proved to interact with NR2F2-AS1 and IGF1R, expressed lowly in esophageal cancer both in vivo and in vitro and suppressed cell viability. Most importantly, miR-4429 and miR-425-5p overexpression could increase trastuzumab’s inhibitory effect on cell viability. Conclusion: Trastuzumab has the potential to suppress EAC progression mainly in the presence of miR-4429 and miR-425-5p overexpression targeting HER2. However, Trastuzumab induces exosomal NR2F2-AS1 expression, which binds to miR-4429 and miR-425-5p to suppress their expression, resulting in the failure of trastuzumab treatment. Therefore, targeting exosomes might be a novel way to develop auxiliary drugs for trastuzumab in EAC.

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Mechanism of MicroRNA-375 Promoter Methylation in Promoting Ovarian Cancer Cell Malignancy

Technology in Cancer Research & Treatment ◽

10.1177/1533033820980115 ◽

2021 ◽

Vol 20 ◽

pp. 153303382098011

Author(s):

Junjun Shu ◽

Ling Xiao ◽

Sanhua Yan ◽

Boqun Fan ◽

Xia Zou ◽

...

Keyword(s):

Ovarian Cancer ◽

Protein Expression ◽

Cell Lines ◽

Cell Invasion ◽

Promoter Methylation ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

Gene Assay ◽

Cell Malignancy

Objective: Ovarian cancer (OC) ranks one of the most prevalent fatal tumors of female genital organs. Aberrant promoter methylation triggers changes of microRNA (miR)-375 in OC. Our study aimed to evaluate the mechanism of methylated miR-375 promoter region in OC cell malignancy and to seek the possible treatment for OC. Methods: miR-375 promoter methylation level in OC tissues and cells was detected. miR-375 expression in OC tissues and cell lines was compared with that in demethylated cells. Role of miR-375 in OC progression was measured. Dual-luciferase reporter gene assay was utilized to verify the targeting relationship between miR-375 and Yes-associated protein 1 (YAP1). Then, Wnt/β-catenin pathway-related protein expression was tested. Moreover, xenograft transplantation was applied to confirm the in vitro experiments. Results: Highly methylated miR-375 was seen in OC tissues and cell lines, while its expression was decreased as the promoter methylation increased. Demethylation in OC cells brought miR-375 back to normal level, with obviously declined cell invasion, migration and viability and improved apoptosis. Additionally, miR-375 targeted YAP1 to regulate the Wnt/β-catenin pathway protein expression. Overexpressed YAP1 reversed the protein expression, promoted cell invasion, migration and viability while reduced cell apoptosis. Overexpressed miR-375 in vivo inhibited OC progression. Conclusion: Our study demonstrated that demethylated miR-375 inhibited OC growth by targeting YAP1 and downregulating the Wnt/β-catenin pathway. This investigation may offer novel insight for OC treatment.

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Sp1-Induced Upregulation of LBX2-AS1 Aggravates The Progression of Glioblastoma By Targeting The miR-491-5p/LIF Axis

10.21203/rs.3.rs-242079/v1 ◽

2021 ◽

Author(s):

Wentao Li ◽

Ismatullah Soufiany ◽

Xiao Lyu ◽

Lin Zhao ◽

Chenfei Lu ◽

...

Keyword(s):

Cell Proliferation ◽

Cell Lines ◽

Cancer Progression ◽

Epithelial To Mesenchymal Transition ◽

Luciferase Reporter ◽

Mesenchymal Transition ◽

Stat3 Signaling ◽

Regulatory Effects

Abstract Background: Mounting evidences have shown the importance of lncRNAs in tumorigenesis and cancer progression. LBX2-AS1 is an oncogenic lncRNA that has been found abnormally expressed in gastric cancer and lung cancer samples. Nevertheless, the biological function of LBX2-AS1 in glioblastoma (GBM) and potential molecular mechanism are largely unclear. Methods: Relative levels of LBX2-AS1 in GBM samples and cell lines were detected by qRT-PCR and FISH. In vivo and in vitro regulatory effects of LBX2-AS1 on cell proliferation, epithelial-to-mesenchymal transition (EMT) and angiogenesis in GBM were examined through xenograft models and functional experiments, respectively. The interaction between Sp1 and LBX2-AS1 was assessed by ChIP. Through bioinformatic analyses, dual-luciferase reporter assay, RIP and Western blot, the regulation of LBX2-AS1 and miR-491-5p on the target gene leukemia Inhibitory factor (LIF) was identified. Results: LBX2-AS1 was upregulated in GBM samples and cell lines, and its transcription was promoted by binding to the transcription factor Sp1. As a lncRNA mainly distributed in the cytoplasm, LBX2-AS1 upregulated LIF, and activated the LIF/STAT3 signaling by exerting the miRNA sponge effect on miR-491-5p, thus promoting cell proliferation, EMT and angiogenesis in GBM. Besides, LBX2-AS1 was unfavorable to the progression of glioma and the survival. Conclusion: Upregulated by Sp1, LBX2-AS1 promotes the progression of GBM by targeting the miR-491-5p/LIF axis. It is suggested that LBX2-AS1 may be a novel diagnostic biomarker and therapeutic target of GBM.

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Contribution of microRNA-30d to the prevention of the thyroid cancer progression: mechanism and implications

10.21203/rs.3.rs-25018/v1 ◽

2020 ◽

Author(s):

Yuan Shao ◽

Shaoqiang Zhang ◽

Xiaoxia Wang ◽

Xin Sun ◽

Jie Wu ◽

...

Keyword(s):

Thyroid Cancer ◽

Cancer Progression ◽

Endocrine Tumor ◽

Luciferase Reporter ◽

Tunel Staining ◽

Loss Of Function ◽

Luciferase Reporter Gene ◽

Gene Assay ◽

Low Expression

Abstract Background Thyroid cancer is a major endocrine tumor and represents an emerging health problem worldwide. MicroRNAs (miRNAs) have been addressed to be associated with the pathogenesis and progression of thyroid cancer. However, it remains largely unknown what functions miR-30d may exert on thyroid cancer. This study herein aimed to identify the functional significance and mechanism of miR-30d in the progression of thyroid cancer. Methods The expression of miR-30d and ubiquitin-specific protease 22 (USP22) in cancerous tissues of patients with thyroid cancer was measured using RT-qPCR and Western blot analysis. In response to the gain- or loss-of-function of miR-30d and USP22, cell apoptosis was evaluated by flow cytometry and TUNEL staining in combination with the measurement of apoptosis-related proteins. The interactions among miR-30d, USP22, SIRT1, FOXO3a and PUMA were explored using a series of assays, including dual-luciferase reporter gene assay, Co-IP and ChIP assay. The effects of miR-30d and USP22 on thyroid tumorigenesis were finally validated in vivo. Results MiR-30b presented aberrant low expression in thyroid cancer tissues and this low expression correlated with poor prognosis of thyroid cancer patients. miR-30d promoted apoptosis of thyroid cancer cells through targeting USP22, an up-regulated gene in thyroid cancer. USP22 could enhance the stability of SIRT1 by inducing deubiquitination which consequently contributed to FOXO3a deacetylation-induced PUMA repression. It was verified that this regulatory mechanism was responsible for the pro-apoptotic effect of miR-30d by the in vivo tumorigenicity assay. Conclusion To conclude, the progression of thyroid cancer can be suppressed by miR-30d-mediated inhibition of USP22, provides a promising therapeutic target for thyroid cancer treatment.

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PRPF6 Induces the Alternative Splicing of lncRNA SNHG16 To Promote Progression and Paclitaxel Resistance of Ovarian Cancer via Regulating GATA3 Transcription

10.21203/rs.3.rs-1045802/v1 ◽

2021 ◽

Author(s):

Han Wang ◽

Yingying Zhou ◽

Siyang Zhang ◽

Ya Qi ◽

Min Wang

Keyword(s):

Ovarian Cancer ◽

Alternative Splicing ◽

Epithelial To Mesenchymal Transition ◽

Binding Protein ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

Mesenchymal Transition ◽

Gene Assay

Abstract Background Small nucleolar RNA host gene 16 (SNHG16) and pre-mRNA processing factor 6(PRPF6) play vital roles in regulatory mechanisms of multiple cancers, but the mechanisms in ovarian cancer (OC) remains poorly understood. Methods The expression of SNHG16 transcripts-SNHG16-L/S in OC tissues were analyzed by real-time PCR (RT-PCR). The expression of PRPF6 in OC tissues were detected by Immunohistochemistry (IHC). Tumorigenesis, epithelial-to-mesenchymal transition (EMT) and PTX-resistance were detected by western blot, transwell, CCK-8 assays, colony formation assays and flow cytometry analyses. Molecular interactions were examined by dual-luciferase reporter gene assay, RNA immunoprecipitation (RIP) and chromatin immunoprecipitation (ChIP). Results The results indicated the expression of SNHG16-L/S was opposite in chemo-resistance and chemo-sensitivity tissues of OC. And SNHG16-L/S had different effects on the progression and PTX-resistance of OC cells. SNHG16-L inhibited GATA binding protein 3 (GATA3) transcription through CCAAT/enhancer-binding protein b (CEBPB) to further promote tumorigenesis, EMT and PTX-resistance of OC. Moreover, PRPF6 was upregulated in chemo-resistance tissues of OC. PRPF6 promoted tumorigenesis and PTX-resistance in vitro and in vivo. Mechanistically, PRPF6 induced the alternative splicing of SNHG16 to downregulate SNHG16-L, which further mediated progression and PTX-resistance through upregulating GATA3 in OC. Conclusions Totally, the results demonstrated that PRPF6 promoted progression and PTX-resistance in OC through SNHG16-L/CEBPB/GATA3 axis. Thus, PRPF6 may become a valuable target for OC therapy.

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miR-495-3p Depresses Cell Proliferation and Migration By Down-Regulating HMGB1 in Colorectal Cancer

10.21203/rs.3.rs-663678/v1 ◽

2021 ◽

Author(s):

Zhang Jieling ◽

Li Kai ◽

Zheng Huifen ◽

Zhu Yiping

Keyword(s):

Colorectal Cancer ◽

Cell Proliferation ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

Gene Assay ◽

And Migration ◽

Cancer Tissues

Abstract Background: MicroRNAs play an important role in the genesis and progression of tumors, including colorectal cancer (CRC), which has a high morbidity and mortality rate. In this research, the role of miR-495-3p and HMGB1 in CRC was investigated.Methods: We performed qRT-PCR to detect the expression of miR-495-3p in colorectal cancer tissues and cell lines. Functional experiments such as CCK-8 assay, EDU assay, Transwell assay and apoptosis assay were conducted to explore the effects of miR-495-3p on the proliferation, migration and apoptosis of CRC cells in vitro. Then, the use of database prediction, dual-luciferase reporter gene assay and functional experiments verified the role of miR-495-3p target gene HMGB1 in CRC. Finally, rescue experiments was performed to investigate whether overexpression of HMGB1 could reverse the inhibitory effect of miR-495-3p on CRC cell proliferation in vivo and in vitro.Results: miR-495-3p was down-regulated in colorectal cancer tissues and cell lines, and could inhibit the proliferation and migration of colorectal cancer cells, and promote cell apoptosis. The database prediction and dual-luciferase reporter gene assay showed that HMGB1 was the downstream target gene of miR-495-3p. We finally demonstrated that miR-495-3p inhibited CRC cell proliferation by targeting HMGB1 in vitro and in vivo.Conclusion: Our research shows that miR-495-3p inhibits the progression of colorectal cancer by down-regulating the expression of HMGB1, which indicates that miR-495-3p may become a potential therapeutic target for colorectal cancer.

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Linc00473 potentiates cholangiocarcinoma progression by modulation of DDX5 expression via miR-506 regulation

10.21203/rs.3.rs-25881/v2 ◽

2020 ◽

Author(s):

Lining Huang ◽

Xingming Jiang ◽

Zhenglong Li ◽

Jinglin Li ◽

Xuan Lin ◽

...

Keyword(s):

Cell Lines ◽

Cancer Progression ◽

Luciferase Reporter ◽

Loss Of Function ◽

Qrt Pcr ◽

Competitive Endogenous Rnas ◽

Rna Immunoprecipitation ◽

Underlying Mechanisms

Abstract Background: Cholangiocarcinoma (CCA) is a mortal cancer with high mortality, whereas the function and mechanism of occurrence and progression of CCA are still mysterious. Long non-coding RNAs (lncRNAs) could function as important regulators in carcinogenesis and cancer progression. Growing evidences have indicated that the novel lncRNA linc00473 plays an important role in cancer progression and metastasis. However, its function and molecular mechanism in CCA remain unknown. Methods: The linc00473 expression in CCA tissues and cell lines was analyzed using qRT-PCR. Gain- and loss-of-function experiments were conducted to investigate the biological functions of linc00473 both in vitro and in vivo. Insights into the underlying mechanisms of competitive endogenous RNAs (ceRNAs) were determined by bioinformatics analysis, dual-luciferase reporter assays, qRT-PCR arrays, RNA immunoprecipitation (RIP) and rescue experiments. Results: Linc00473 was highly expressed in CCA tissues and cell lines. Linc00473 knockdown inhibited CCA growth and metastasis. Furthermore, linc00473 acted as miR-506 sponge and regulated its target gene DDX5 expression. Rescue assays verified that linc00473 modulated the tumorigenesis of CCA by regulating miR-506. Conclusions: The data indicated that linc00473 played an oncogenic role in CCA growth and metastasis, and could serve as a novel molecular target for treating CCA.

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IFN-γ Enhances the Efficacy of Exosomes Derived from Mesenchymal Stem Cells in Myocardial Infarction Rats via miR-21 Stimulated by STAT1

10.21203/rs.3.rs-1044001/v1 ◽

2021 ◽

Author(s):

Jian Zhang ◽

Yao Lu ◽

Yangming Mao ◽

Yue Yu ◽

Tianyu Wu ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Umbilical Vein ◽

Luciferase Reporter ◽

H9c2 Cells ◽

Luciferase Reporter Gene ◽

Gene Assay ◽

Ifn Γ

Abstract Background: Mesenchymal stem cells (MSCs) activated with IFN-γ elicit more powerful physical effects. Exosomes (Exos) secreted from MSCs have protective against myocardial injury. The aim of this study was to investigate whether Exsos derived from IFN-γ-pretreated MSCs exhibit more potent cardioprotective function and the underlying mechanisms. Methods: Exos were isolated from MSCs (Ctrl-Exo) and IFN-γ-primed MSCs (IFN-γ-Exo) and were then delivered to H9c2 cells or human umbilical vein endothelial cells (HUVECs) in vitro under oxygen and glucose deprivation (OGD) condition or in vivo in an infarcted rat heart. RNA sequencing was to identify the different expressed functional transcription factor (TF). Quantitative reverse transcription-PCR (qPCR) was to confirm the upregulated TF and miRNA in IFN-γ-primed MSCs. Dual-luciferase reporter gene assay were to analyze the transcriptional regulation of miRNAs by STAT1. The target of miR-21-5p (miR-21) was disclosed by luciferase reporter assays and qPCR. The function of BTG2 was verified in vitro under OGD condition.Result: IFN-γ-Exo accelerated migration, tube-like structure formation, and prevented H9c2 from OGD-induced apoptosis. Similarly, IFN-γ-Exo leaded to further reduction in fibrosis size, reduced cardiomyocyte apoptosis and improved cardiac function compared to Ctrl-Exo. miR-21 was significantly upregulated in both IFN-γ-primed MSCs and IFN-γ-Exo. STAT1 transcriptionally induced miR-21 expression. Up-regulated miR-21 can inhibit the expression of BTG2. BTG2 promoted H9c2 cells apoptosis and reversed the protective effect of miR-21 under OGD environment.Conclusion: IFN-γ-Exo have enhanced therapeutic efficacy against acute MI possibly through promoting angiogenesis and anti-apoptotic effect through increasing the level of miR-21, which directly targeted on BTG2.

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Circular RNA circANKS1B as the Sponge of miR-152-3p Promotes Prostate Cancer Progression by up-Regulating TGF-α Expression

10.21203/rs.3.rs-66869/v1 ◽

2020 ◽

Author(s):

Liangjun Tao ◽

Xinyuan Pan ◽

Jiawei Wang ◽

Li Zhang ◽

Lingsong Tao ◽

...

Keyword(s):

Prostate Cancer ◽

Cancer Progression ◽

Circular Rna ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Prostate Cancer Progression ◽

Luciferase Reporter Gene ◽

Gene Assay ◽

And Function ◽

The Impact

Abstract Background: Growing studies indicate that circRNAs play critical roles in human diseases, and show great potential as biomarkers and therapeutic targets. This study aims to investigate the expression and function of circANKS1B in prostate cancer (PC).Methods: The expression of circANKS1B and miRNA-152-3p were determined by real-time qRT-PCR. The cell migration and invasion were measured by transwell assay. The interaction between circANKS1B and miR-152-3p was confirmed by dual-luciferase reporter gene assay. Rescue experiments were conducted to demonstrate whether circANKS1B regulated the migration and invasion of PC cells by the circANKS1B-miR-152-3p-TGF-α pathway.Results: The expression of circANKS1B was dramatically up-regulated both in PC cells and tissues. Moreover, high circANKS1B expression was associated with a poor prognosis of PC patients. Dual-luciferase reporter assay indicated that circABKS1B directly bound to miRNA-152-3p. Furthermore, circANKS1B negatively regulated miR-152-3p expression. Knockdown of circANKS1B remarkably suppressed PC cells invasion and TGF-α expression, while the effects of circANKS1B silencing were reversed by miR-152-3p deficiency. In addition, the impact of miR-152-3p silencing on PC cell invasion was also abrogated by TGF-α deficiency. In all, circANKS1B as the sponge of miR-152-3p promotes prostate cancer progression by up-regulating TGF-α expression.Conclusion: Our findings reveal that circANKS1B could be a potential prognostic biomarker and therapeutic target of PC.

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