关键词: 乳腺癌, 人表皮生长因子受体2, circ-0007766, miR-1972, 转移

Abstract:

Background and purpose: Human epidermal growth factor receptor 2 (HER2) serves as one of the paramount drivers of breast cancer metastasis, with roughly 20%-30% of breast cancer patients exhibiting high expression of HER2. The expression level of HER2 is regulatable at multiple molecular levels and determines the metastatic potential of breast cancer cells; however, the manner in which HER2 expression is modulated at the mRNA level remains ambiguous. Circ-0007766 is a circRNA originated from the coding gene ERBB2 for HER2, and whether circ-0007766 can regulate HER2 expression via the ceRNA mechanism has not been reported. This study aimed to analyze whether circ-0007766 acts as a miR-1972 sponge to promote breast cancer cell migration and invasion via upregulation of HER2 expression. Methods: In this study, a high-throughput circRNA chip was employed to screen for circRNAs that exhibited highly specific expression in HER2-positive breast cancer cells. RNA fluorescence in situ hybridization (FISH) was utilized to detect the subcellular localization of circ-0007766. The BaseScope experiment was conducted to analyze the expression level of circ-0007766 in breast cancer tissues and its clinical diagnostic significance. Breast cancer cell models with overexpression and knockdown of circ-0007766 were constructed by transfecting cloning plasmids and siRNA in vitro. The effect of circ-0007766 on the migration and invasion of breast cancer cells was assessed using transwell migration and invasion experiments, and the migration and invasion abilities of MDA-MB-231 and SK-BR-3 cells were measured. Additionally, it was evaluated whether circ-0007766 could promote the migration and invasion of breast cancer cells through miR-1972. A dual luciferase reporter gene assay was used to verify whether circ-0007766 could regulate HER2 expression by binding to miR-1972. The direct interaction between circ-0007766 and miR-1972 was further verified through the RAP experiment. RIP detection was performed in MDA-MB-231 cells, and the relative 3'UTR of HER2 mRNA was measured by real-time fluorescence quantitative polymerase chain reaction (RTFQ-PCR). Western blot was used to detect the protein expressions. Results: Circ-0007766 was conspicuously highly expressed in HER2-positive breast cancer cells and distributed in both the cytoplasm and nucleus of cells, with the preponderance being in the cytoplasm. The expression level of circ-0007766 was strikingly higher in breast cancer tissues than in para-cancerous tissues. The expression of circ-0007766 was significantly elevated in HER2-positive breast cancer samples compared with HER2-negative samples. The overexpression (knockdown) of circ-0007766 in HER2-negative breast cancer cells (in HER2-positive breast cancer cells) was capable of promoting (inhibiting) the migration and invasion of breast cancer cells. Circ-0007766 directly bound to miR-1972, which inhibited breast cancer cell migration and invasion, thereby forming an endogenous competitive RNA (ceRNA) regulatory network and impeding the downregulation of HER2 mRNA and protein expression mediated by miR-1972. Circ-0007766 could potentiate the inhibitory effect of miR-1972 on HER2-mediated breast cancer cell migration and invasion that was negatively regulated by miR-1972. CircRNAs sequestered miRNAs to function as ceRNAs, thereby regulating gene expression at both the transcriptional and translational levels. Finally, we discovered that the expressions of circ-0007766 and HER2 were positively correlated in breast cancer cell and tissue samples, while the expression levels of miR-1972 and HER2 were negatively correlated. Circ-0007766 could specifically target miR-1972 to hinder its regulatory effect on HER2 expression. Conclusion: This study discovers that circ-0007766 facilitates the migration and invasion of breast cancer cells via the miR-1972/HER2 signal axis, offering a novel biomarker and potential therapeutic target for patients with metastatic HER2-positive breast cancer.

Key words: Breast cancer, Human epidermal growth factor receptor 2, circ-0007766, miR-1972, Metastasis