HPV16 E6 Promotes Breast Cancer Proliferation via Upregulation of COX-2 Expression

Background.Breast cancer remains the leading cause of cancer-related mortality worldwide. It has been indicated that human papillomaviruses 16 (HPV16) might participate in the pathogenesis and development of breast cancer. However, the detected rate of HPV16 varies with region. We will investigate HPV16 E6 expression in North China and explore the effects and mechanism of HPV16 E6 on breast cancer proliferation in this study.Methods.The expressions of HPV16 E6 and COX-2 in paraffin-embedded tissues of the invasive ductal breast cancer were detected by qPCR and IHC. The effects of HPV16 E6 on breast cancer proliferation were determined by function studies. The mechanism of HPV16 E6 in promoting breast cancer proliferation was explored by Western blot and Dual-Luciferase Reporter Assay.Results.HPV16 E6 was positive in 28% invasive ductal breast carcinoma in North China; HPV16 E6 promoted breast cancer proliferation. Inhibition of COX-2 by siCOX-2 or Celecoxib attenuated the proliferation of breast cancer cells with HPV16 E6 expression; and the upregulation of COX-2 could be suppressed by the inhibition of NF-κB activity.Conclusion.HPV16 E6 promotes breast cancer proliferation by activation of NF-κB signaling pathway and increase of COX-2 expression. COX-2 will be a potential target for HPV16 E6-associated breast cancer.

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MiR-4458 inhibits breast cancer cell growth, migration, and invasiveness by targeting CPSF4

Biochemistry and Cell Biology ◽

10.1139/bcb-2019-0008 ◽

2019 ◽

Vol 97 (6) ◽

pp. 722-730 ◽

Cited By ~ 6

Author(s):

Jianrong Wu ◽

Juan Miao ◽

Ye Ding ◽

Yayun Zhang ◽

Xiaohao Huang ◽

...

Keyword(s):

Breast Cancer ◽

Cell Proliferation ◽

Cell Growth ◽

Target Genes ◽

Human Lung ◽

Lung Tumor ◽

Luciferase Reporter ◽

Bioinformatics Analyses ◽

Cox 2

Numerous studies have reported that CPSF4 is over-expressed in a large percentage of human lung cancers, and CPSF4 has been identified as a potential oncogene of human lung tumor. Downregulation of CPSF4 inhibits the proliferation and promotes the apoptosis of lung adenocarcinoma cells. A previous study by our group also found overexpression of CPSF4 in breast cancer (BC), and was closely associated with a poor prognosis for the patient. This study investigates microRNAs (miRNAs) that target CPSF4 to modulate BC cell proliferation. We found that miR-4458 was noticeably reduced in BC tissues and cells. Using a miR-4458 mimic, we found that cell proliferation, migration, and invasiveness were suppressed by miR-4458 overexpression, and were enhanced by reducing the expression of miR-4458. Moreover, the results from bioinformatics analyses suggest a putative target site in the CPSF4 3′-UTR. Furthermore, using luciferase reporter assays and Western blotting, we verified that miR-4458 directly targets the 3′-UTR of CPSF4 and downregulates COX-2 and h-TERT, which are downstream target genes of CPSF4. Additionally, PI3K/AKT and ERK were shown to be inhibited by miR-4458 overexpression in BC cells. Moreover, miR-4458 suppresses BC cell growth in vivo. Consequently, these results suggest that the miR-4458–CPSF4–COX-2–hTERT axis might serve as a potential target for the treatment of BC patients.

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Cross-talk between SIM2s and NFκB regulates cyclooxygenase 2 expression in breast cancer

Breast Cancer Research ◽

10.1186/s13058-019-1224-y ◽

2019 ◽

Vol 21 (1) ◽

Cited By ~ 2

Author(s):

Garhett L. Wyatt ◽

Lyndsey S. Crump ◽

Chloe M. Young ◽

Veronica M. Wessells ◽

Cole M. McQueen ◽

...

Keyword(s):

Breast Cancer ◽

Cyclooxygenase 2 ◽

Luciferase Reporter ◽

Signaling Proteins ◽

Dependent Manner ◽

Mcf7 Cells ◽

Human Breast Carcinomas ◽

Cox 2

Abstract Background Breast cancer is a leading cause of cancer-related death for women in the USA. Thus, there is an increasing need to investigate novel prognostic markers and therapeutic methods. Inflammation raises challenges in treating and preventing the spread of breast cancer. Specifically, the nuclear factor kappa b (NFκB) pathway contributes to cancer progression by stimulating proliferation and preventing apoptosis. One target gene of this pathway is PTGS2, which encodes for cyclooxygenase 2 (COX-2) and is upregulated in 40% of human breast carcinomas. COX-2 is an enzyme involved in the production of prostaglandins, which mediate inflammation. Here, we investigate the effect of Singleminded-2s (SIM2s), a transcriptional tumor suppressor that is implicated in inhibition of tumor growth and metastasis, in regulating NFκB signaling and COX-2. Methods For in vitro experiments, reporter luciferase assays were utilized in MCF7 cells to investigate promoter activity of NFκB and SIM2. Real-time PCR, immunoblotting, immunohistochemistry, and chromatin immunoprecipitation assays were performed in SUM159 and MCF7 cells. For in vivo experiments, MCF10DCIS.COM cells stably expressing SIM2s-FLAG or shPTGS2 were injected into SCID mice and subsequent tumors harvested for immunostaining and analysis. Results Our results reveal that SIM2 attenuates the activation of NFκB as measured using NFκB-luciferase reporter assay. Furthermore, immunostaining of lysates from breast cancer cells overexpressing SIM2s showed reduction in various NFκB signaling proteins, as well as pAkt, whereas knockdown of SIM2 revealed increases in NFκB signaling proteins and pAkt. Additionally, we show that NFκB signaling can act in a reciprocal manner to decrease expression of SIM2s. Likewise, suppressing NFκB translocation in DCIS.COM cells increased SIM2s expression. We also found that NFκB/p65 represses SIM2 in a dose-dependent manner, and when NFκB is suppressed, the effect on the SIM2 is negated. Additionally, our ChIP analysis confirms that NFκB/p65 binds directly to SIM2 promoter site and that the NFκB sites in the SIM2 promoter are required for NFκB-mediated suppression of SIM2s. Finally, overexpression of SIM2s decreases PTGS2 in vitro, and COX-2 staining in vivo while decreasing PTGS2 and/or COX-2 activity results in re-expression of SIM2. Conclusion Our findings identify a novel role for SIM2s in NFκB signaling and COX-2 expression.

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NcRNAs-mediated High Expression of TIMM8A Correlates with Poor Prognosis and Act as an Oncogene in Breast Cancer

10.21203/rs.3.rs-1079496/v1 ◽

2021 ◽

Author(s):

Zhonglin Wang ◽

Shuqin Li ◽

Feng Xu ◽

Jingyue Fu ◽

Jie Sun ◽

...

Keyword(s):

Breast Cancer ◽

Poor Prognosis ◽

Molecular Mechanisms ◽

Human Cancer ◽

Luciferase Reporter ◽

Inner Mitochondrial Membrane ◽

Cancer Proliferation ◽

Bioinformatics Analyses

Abstract Background: Breast cancer is notorious for its increasing incidence for decades. Ascending evidence has demonstrated that translocase of inner mitochondrial membrane (TIMM) proteins play vital roles in progression of several types of human cancer. However, the biological behaviors and molecular mechanisms of TIMM8A in breast cancer remain not fully illustrated.Methods: Pan-cancer analysis was firstly performed for TIMM8A’s expression and prognosis by Oncomine database. Subsequently, TIMM8A-related noncoding RNAs (ncRNAs) were identified by a series of bioinformatics analyses and dual-luciferase reporter assay, including expression analysis, correlation analysis, and survival analysis. Moreover, the effect of TIMM8A on breast cancer proliferation and apoptosis was evaluated in vitro by CCK-8 assays, colony formation assays and Western blot assays and the in vivo effect was revealed through a patient-derived xenograft mouse model.Results: We found that TIMM8A showed higher expression level in breast cancer and the higher TIMM8A mRNA expression group had a poorer prognosis than the lower TIMM8A group. hsa-circ-0107314/hsa-circ-0021867/hsa-circ-0122013 might be the three most potential upstream circRNAs of hsa-miR-34c-5p/hsa-miR-449a-TIMM8A axis in breast cancer. TIMM8A promotes proliferation of breast cancer cells in vitro and tumor growth in vivo.Conclusion: Our results confirmed that ncRNAs-mediated upregulation of TIMM8A correlated with poor prognosis and act as an oncogene in breast cancer.

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Cross-talk between SIM2s and NFkB regulates cyclooxygenase 2 expression in breast cancer

10.1101/634113 ◽

2019 ◽

Author(s):

Garhett Wyatt ◽

Chloe Young ◽

Lyndsey Crump ◽

Veronica Wessells ◽

Tanya Gustafson ◽

...

Keyword(s):

Breast Cancer ◽

The United States ◽

Cyclooxygenase 2 ◽

Luciferase Reporter ◽

Signaling Proteins ◽

Dependent Manner ◽

Human Breast Carcinomas ◽

Reporter Assays ◽

Cox 2

AbstractBackgroundBreast cancer is a leading cause of cancer-related death for women in the United States. Thus, there a need to investigate novel prognostic markers and therapeutic strategies. Inflammation raises challenges to both treating and preventing the spread of breast cancer. Specifically, the nuclear factor kappa b (NFkB) pathway contributes to cancer progression by stimulating proliferation and preventing apoptosis. One target gene of this pathway is PTGS2, the gene that encodes for cyclooxygenase 2 (COX-2), which is upregulated in 40% of human breast carcinomas. COX-2 is an enzyme involved in inflammation. Here we investigate the effect of Singleminded 2s, a transcriptional tumor suppressor that is implicated in inhibition of tumor growth and metastasis, in regulating NFkB and COX-2.MethodsWe utilized in vitro reporter assays, immunoblot analyses, qPCR and immunohistochemical analysis to dissect the relationship between NFκB, SIM2s, and COX-2. Furthermore, we utilized COX-2 targeting strategies to determine tumor suppressive activities.ResultsOur results reveal that SIM2s attenuates the activation of a NFκB via luciferase reporter assays. Furthermore, immunostaining of lysates from breast cancer cells over expressing SIM2s showed reduction in various NFκB signaling proteins, whereas knockdown of SIM2 revealed increases in the same NFκB signaling proteins. Additionally, by increasing NFκB translocation to the nucleus in DCIS.COM cells, we show that NFκB signaling can act in a reciprocal manner to decrease expression of SIM2s. Likewise, suppressing NFκB translocation in DCIS.COM cells increases SIM2s expression. We also found that NFκB/p65 represses SIM2 in via dose-dependent manner and when NFκB is suppressed the effect on the SIM2 is negated. Additionally, our CHIP analysis confirms that NFκB/p65 binds directly to SIM2 promoter site and that the NFκB sites in the SIM2 promoter are required for NFkB-mediated suppression of SIM2s. Finally, over expression of SIM2s decreases PTGS2 in vitro and COX-2 staining in vivo while decreasing PTGS2 and/or Cox-2 activity results in re-expression of SIM2. Our findings identify a novel role for SIM2s in NFκB signaling and COX-2 expression.ConclusionsThese findings provide evidence for a mechanism where SIM2s may represses COX-2 expression to provide an overall better prognosis for breast cancer patients.

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miR-146a promoted breast cancer proliferation and invasion by regulating NM23-H1

The Journal of Biochemistry ◽

10.1093/jb/mvz079 ◽

2019 ◽

Vol 167 (1) ◽

pp. 41-48 ◽

Cited By ~ 8

Author(s):

Jun Chen ◽

Qiang Jiang ◽

Xue-Qin Jiang ◽

De-Quan Li ◽

Xiao-Cheng Jiang ◽

...

Keyword(s):

Breast Cancer ◽

Cancer Cell Line ◽

Luciferase Reporter ◽

Luciferase Reporter Gene ◽

Regulatory Relationship ◽

Cancer Proliferation ◽

Expression Levels ◽

Invasion And Migration ◽

Proliferation And Invasion

Abstract The study aimed to investigate the regulatory effect of miR-146a in proliferation, invasion and migration of breast cancer and its possible mechanism via NM23-H1. The expression levels of miR-146a in breast cancer with different pathological classification were significantly increased, while the expression levels of NM23-H1 were significantly decreased, which were closely correlated. Double luciferase reporter gene was used to verify the target regulatory relationship between miR-146 and NM23-H1 on a human breast cancer cell line. miR-146a was closely related to the proliferation and metastasis of breast cancer. miR-146a also promoted the growth of breast cancer in vivo via targeting NM23-H1. In conclusion, miR-146 can promote the proliferation and invasion of breast cancer by targeting NM23-H1.

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Evaluation of a Luciferase-Based Reporter Assay as a Screen for Inhibitors of Estrogen-ERα-Induced Proliferation of Breast Cancer Cells

CrossRef Listing of Deleted DOIs ◽

10.1177/1087057112442960 ◽

2012 ◽

Vol 17 (7) ◽

pp. 921-932 ◽

Cited By ~ 17

Author(s):

Neal Andruska ◽

Chengjian Mao ◽

Mathew Cherian ◽

Chen Zhang ◽

David J. Shapiro

Keyword(s):

Breast Cancer ◽

Cell Proliferation ◽

Cancer Cells ◽

Cell Lines ◽

Breast Cancer Cells ◽

Estrogen Receptor Α ◽

Luciferase Reporter ◽

Cancer Proliferation ◽

Estrogen Response ◽

A Cell

Estrogens, acting through estrogen receptor α (ERα), stimulate breast cancer proliferation, making ERα an attractive drug target. Since 384-well format screens for inhibitors of proliferation can be challenging for some cells, inhibition of luciferase-based reporters is often used as a surrogate end point. To identify novel small-molecule inhibitors of 17β-estradiol (E2)–ERα-stimulated cell proliferation, we established a cell-based screen for inhibitors of E2-ERα induction of an estrogen response element (ERE)3–luciferase reporter. Seventy-five “hits” were evaluated in tiered follow-up assays to identify where hits failed to progress and evaluate their effectiveness as inhibitors of E2-ERα-induced proliferation of breast cancer cells. Only 8 of 75 hits from the luciferase screen inhibited estrogen-induced proliferation of ERα-positive MCF-7 and T47D cells but not control ERα-negative MDA-MB-231 cells. Although 12% of compounds inhibited E2-ERα-stimulated proliferation in only one of the ERα-positive cell lines, 40% of compounds were toxic and inhibited growth of all the cell lines, and ~37% exhibited little or no ability to inhibit E2-ERα-stimulated cell proliferation. Representative compounds were evaluated in more detail, and a lead ERα inhibitor was identified.

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