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Comparing MicroRNA Profilings of Purified HER-2-Negative and HER-2-Positive Cells Validates miR-362-5p/Sema3A as Characteristic Molecular Change in Triple-Negative Breast Cancers.

ABSTRACT: Background:HER-2 is a key molecule serving as the therapeutic target, prognostic biomarker, and classification marker in breast cancer. Accurate microRNA profilings had not been conducted in purified tumor cells of HER-2-negative and HER-2-positive tissue specimens obtained from breast cancer patients. Methods:(i) Differential expression microRNA discovery using laser capture microdissection- (LCM-) assisted specimen preparation and microRNA array chips on HER-2 overexpressing and triple-negative breast carcinoma (TNBC) subtype tissues, (ii) differential expression microRNA validation by quantitative real-time PCR, and (iii) independent validation on tissue microarray. Results:Five microRNAs (miR-20a-5p, miR-221-3p, miR-362-5p, miR-502-3p, and miR-222-3p) were screened and validated as upregulated microRNAs in TNBC cells comparing to HER-2 overexpressing cells using a microRNA array (5 cases in each group) and quantitative real-time PCR (20 cases in each group). The expression difference of miR-362-5p had the most significant statistical significance (p = 0.0016) among the five microRNAs. The expression of miR-362-5p and its target gene Sema3A was further analyzed using in situ hybridization (ISH) and immunohistochemistry on standard tissue sections (n = 150). 70.8% of HER-2-negative cells showed moderate expression of miR-362-5p whereas 20.4% HER-2-negative cells correlated with strong expression of miR-362-5p (p < 0.0001). The proportion of patients with moderate/strong miR-362-5p expression in luminal, HER-2 overexpressing, and TNBC subtypes were 53.2%, 22.2%, and 74.3%, respectively (p = 0.0002). High miR-362-5p expressers had shorter overall survival in the univariate analysis (p = 0.046). There was a significant negative correlation between miR-362-5p and Sema3A expression (p < 0.0001). The patients with negative/weak Sema3A protein expression had poorer prognosis than those with moderate (HR: 3.723, p = 0.021) or strong (HR: 3.966, p = 0.013) Sema3A protein expression in the multivariate analysis. Conclusions:miR-362-5p/Sema3A might provide a promising therapeutic pathway and represents a candidate therapeutic target of the TNBC subtype.

SUBMITTER: Zhang X

PROVIDER: S-EPMC6935799 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Comparing MicroRNA Profilings of Purified HER-2-Negative and HER-2-Positive Cells Validates miR-362-5p/Sema3A as Characteristic Molecular Change in Triple-Negative Breast Cancers.

Zhang Xiaoqing X He Qi Q Sun Leiqin L Zhang Yanfei Y Qin Shengying S Fan Junwei J Wang Jianfeng J

Disease markers 20191216

<h4>Background</h4>HER-2 is a key molecule serving as the therapeutic target, prognostic biomarker, and classification marker in breast cancer. Accurate microRNA profilings had not been conducted in purified tumor cells of HER-2-negative and HER-2-positive tissue specimens obtained from breast cancer patients.<h4>Methods</h4>(i) Differential expression microRNA discovery using laser capture microdissection- (LCM-) assisted specimen preparation and microRNA array chips on HER-2 overexpressing and ...[more]

PMID: 31929841

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Project description:BackgroundDNA damage and DNA damage repair (DDR) are important therapeutic targets for triple-negative breast cancer (TNBC), a subtype with limited chemotherapy efficiency and poor outcome. However, the role of microRNAs in the therapy is emerging. In this study, we explored whether miR-26a-5p could act as BRCAness and enhance chemotherapy sensitivity in TNBC.MethodsQuantitative reverse transcription polymerase chain reaction (RT-qPCR) was used to detect the expression of miR-26a-5p in breast cancer tissues and cell lines. CCK-8 was used to measure drug sensitivity in concentration gradient and time gradient. Comet assay was used to detect DNA damage. Flow cytometry was performed to examine apoptosis. Moreover, we used western blot and immunofluorescence to detect biomarkers. Luciferase reporter assay was performed to verify the combination of miR-26a-5p and 3'UTR of target gene. Hormone deprivation and stimulation assay were used to validate the effect of hormone receptors on the expression of miR-26a-5p. Chromatin immunoprecipitation (ChIP) assays were used to verify the binding sites of ER-a or PR with the promoter of miR-26a-5p. Animal experiments were performed to the effect of miR-26a-5p on Cisplatin treatment.ResultsThe expression of miR-26a-5p was significantly downregulated in TNBC. Overexpressing miR-26a-5p enhanced the Cisplatin-induced DNA damage and following apoptosis. Interestingly, miR-26a-5p promoted the expression of Fas without Cisplatin stimulating. It suggested that miR-26a-5p provided a hypersensitivity state of death receptor apoptosis and promoted the Cisplatin sensitivity of TNBC cells in vitro and in vivo. Besides, miR-26a-5p negatively regulated the expression of BARD1 and NABP1 and resulted in homologous recombination repair defect (HRD). Notably, overexpressing miR-26a-5p not only facilitated the Olaparib sensitivity of TNBC cells but also the combination of Cisplatin and Olaparib. Furthermore, hormone receptors functioned as transcription factors in the expression of miR-26a-5p, which explained the reasons that miR-26a-5p expressed lowest in TNBC.ConclusionsTaken together, we reveal the important role of miR-26a-5p in Cisplatin sensitivity and highlight its new mechanism in DNA damage and synthetic lethal.

Dataset Information

Comparing MicroRNA Profilings of Purified HER-2-Negative and HER-2-Positive Cells Validates miR-362-5p/Sema3A as Characteristic Molecular Change in Triple-Negative Breast Cancers.

Publications

Comparing MicroRNA Profilings of Purified HER-2-Negative and HER-2-Positive Cells Validates miR-362-5p/Sema3A as Characteristic Molecular Change in Triple-Negative Breast Cancers.

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