Molecular Oncology Research Center, Barretos Cancer Hospital, Barretos, São Paulo, 14784-400, Brazil.
Department of Mastology and Breast Reconstruction, Barretos Cancer Hospital, Barretos, São Paulo, 14784-400, Brazil.
BMC Cancer. 2021 Jan 18;21(1):76. doi: 10.1186/s12885-020-07731-2.
Breast cancer is the most frequently diagnosed malignancy among women. However, the role of microRNA (miRNA) expression in breast cancer progression is not fully understood. In this study we examined predictive interactions between differentially expressed miRNAs and mRNAs in breast cancer cell lines representative of the common molecular subtypes. Integrative bioinformatics analysis identified miR-193 and miR-210 as potential regulatory biomarkers of mRNA in breast cancer. Several recent studies have investigated these miRNAs in a broad range of tumors, but the mechanism of their involvement in cancer progression has not previously been investigated.
The miRNA-mRNA interactions in breast cancer cell lines were identified by parallel expression analysis and miRNA target prediction programs. The expression profiles of mRNA and miRNAs from luminal (MCF-7, MCF-7/AZ and T47D), HER2 (BT20 and SK-BR3) and triple negative subtypes (Hs578T e MDA-MB-231) could be clearly separated by unsupervised analysis using HB4A cell line as a control. Breast cancer miRNA data from TCGA patients were grouped according to molecular subtypes and then used to validate these findings. Expression of miR-193 and miR-210 was investigated by miRNA transient silencing assays using the MCF7, BT20 and MDA-MB-231 cell lines. Functional studies included, xCELLigence system, ApoTox-Glo triplex assay, flow cytometry and transwell inserts were performed to determine cell proliferation, cytotoxicity, apoptosis, migration and invasion, respectively.
The most evident effects were associated with cell proliferation after miR-210 silencing in triple negative subtype cell line MDA-MB-231. Using in silico prediction algorithms, TNFRSF10 was identified as one of the potential regulated downstream targets for both miRNAs. The TNFRSF10C and TNFRSF10D mRNA expression inversely correlated with the expression levels of miR-193 and miR210 in breast cell lines and breast cancer patients, respectively. Other potential regulated genes whose expression also inversely correlated with both miRNAs were CCND1, a known mediator on invasion and metastasis, and the tumor suppressor gene RUNX3.
In summary, our findings identify miR-193 and miR-210 as potential regulatory miRNA in different molecular subtypes of breast cancer and suggest that miR-210 may have a specific role in MDA-MB-231 proliferation. Our results highlight important new downstream regulated targets that may serve as promising therapeutic pathways for aggressive breast cancers.
乳腺癌是女性最常见的恶性肿瘤。然而，miRNA（microRNA）表达在乳腺癌进展中的作用尚不完全清楚。在这项研究中，我们研究了在代表常见分子亚型的乳腺癌细胞系中差异表达的 miRNA 和 mRNA 之间的预测性相互作用。综合生物信息学分析鉴定出 miR-193 和 miR-210 作为乳腺癌中 mRNA 的潜在调控生物标志物。最近有几项研究调查了这些 miRNA 在广泛的肿瘤中的作用，但它们在癌症进展中的参与机制尚未被研究。
通过平行表达分析和 miRNA 靶标预测程序鉴定乳腺癌细胞系中的 miRNA-mRNA 相互作用。使用 HB4A 细胞系作为对照，通过无监督分析可以清楚地将腔型（MCF-7、MCF-7/AZ 和 T47D）、HER2（BT20 和 SK-BR3）和三阴性亚型（Hs578T 和 MDA-MB-231）的 mRNA 和 miRNA 表达谱分开。根据分子亚型对来自 TCGA 患者的乳腺癌 miRNA 数据进行分组，然后用于验证这些发现。使用 MCF7、BT20 和 MDA-MB-231 细胞系通过 miRNA 瞬时沉默测定研究 miR-193 和 miR-210 的表达。功能研究包括使用 xCELLigence 系统、ApoTox-Glo 三联体测定、流式细胞术和 Transwell 插入物分别确定细胞增殖、细胞毒性、凋亡、迁移和侵袭。
miR-210 沉默后，三阴性亚型细胞系 MDA-MB-231 与细胞增殖的关联最明显。使用计算预测算法，TNFRSF10 被鉴定为这两种 miRNA 的潜在调控下游靶标之一。TNFRSF10C 和 TNFRSF10D mRNA 表达与乳腺癌细胞系和乳腺癌患者的 miR-193 和 miR210 表达水平呈负相关，分别。其他潜在调节基因的表达也与这两种 miRNA 呈负相关，这些基因包括已知参与侵袭和转移的 CCND1 和肿瘤抑制基因 RUNX3。
总之，我们的研究结果确定 miR-193 和 miR-210 是乳腺癌不同分子亚型的潜在调节 miRNA，并表明 miR-210 可能在 MDA-MB-231 增殖中具有特定作用。我们的结果强调了可能作为侵袭性乳腺癌有希望的治疗途径的重要新的下游调节靶标。
