Department of Breast, Longhua Hospital Affiliated to Shanghai University of TCM, Shanghai, China.
State Key Laboratory of Bioreactor Engineering and Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, Shanghai, China.
Aging (Albany NY). 2021 Jul 21;13(14):18191-18222. doi: 10.18632/aging.203156.
This investigation attempted to discern whether formononetin restrained progression of triple-negative breast cancer (TNBC) by blocking lncRNA AFAP1-AS1-miR-195/miR-545 axis. We prepared TNBC cell lines (i.e. MDA-MB-231 and BT-549) and normal human mammary epithelial cell line (i.e. MCF-10A) in advance, and the TNBC cell lines were, respectively, transfected by pcDNA3.1-lncRNA AFAP1-AS1, si-lncRNA AFAP1-AS1, pcDNA6.2/GW/EmGFP-miR-545 or pcDNA6.2/GW/EmGFP-miR-195. Resistance of TNBC cells in response to 5-Fu, adriamycin, paclitaxel and cisplatin was evaluated through MTT assay, while potentials of TNBC cells in proliferation, migration and invasion were assessed via CCK8 assay and Transwell assay. Consequently, silencing of lncRNA AFAP1-AS1 impaired chemo-resistance, proliferation, migration and invasion of TNBC cells (<0.05), and over-expression of miR-195 and miR-545, which were sponged and down-regulated by lncRNA AFAP1-AS1 (<0.05), significantly reversed the promoting effect of pcDNA3.1-lncRNA AFAP1-AS1 on proliferation, migration, invasion and chemo-resistance of TNBC cells (<0.05). Furthermore, CDK4 and Raf-1, essential biomarkers of TNBC progression, were, respectively, subjected to target and down-regulation of miR-545 and miR-195 (<0.05), and they were promoted by pcDNA3.1-lncRNA AFAP1-AS1 at protein and mRNA levels (<0.05). Additionally, formononetin significantly decreased expressions of lncRNA AFAP1-AS1, CDK4 and Raf-1, while raised miR-195 and miR-545 expressions in TNBC cells (<0.05), and exposure to it dramatically contained malignant behaviors of TNBC cells (<0.05). In conclusion, formononetin alleviated TNBC malignancy by suppressing lncRNA AFAP1-AS1-miR-195/miR-545 axis, suggesting that molecular targets combined with traditional Chinese medicine could yield significant clinical benefits in TNBC.
本研究旨在探讨芒柄花素是否通过阻断 lncRNA AFAP1-AS1- miR-195/miR-545 轴来抑制三阴性乳腺癌（TNBC）的进展。我们预先制备了 TNBC 细胞系（即 MDA-MB-231 和 BT-549）和正常的人乳腺上皮细胞系（即 MCF-10A），并分别用 pcDNA3.1-lncRNA AFAP1-AS1、si-lncRNA AFAP1-AS1、pcDNA6.2/GW/EmGFP-miR-545 或 pcDNA6.2/GW/EmGFP-miR-195 转染 TNBC 细胞系。通过 MTT 法评估 TNBC 细胞对 5-Fu、阿霉素、紫杉醇和顺铂的耐药性，通过 CCK8 法和 Transwell 法评估 TNBC 细胞的增殖、迁移和侵袭潜能。结果表明，沉默 lncRNA AFAP1-AS1 可削弱 TNBC 细胞的化疗耐药性、增殖、迁移和侵袭能力（<0.05），而过表达 lncRNA AFAP1-AS1 下调的 miR-195 和 miR-545（<0.05）显著逆转了 pcDNA3.1-lncRNA AFAP1-AS1 对 TNBC 细胞增殖、迁移、侵袭和化疗耐药性的促进作用（<0.05）。此外，CDK4 和 Raf-1 是 TNBC 进展的重要生物标志物，miR-545 和 miR-195 分别对其进行靶向和下调（<0.05），而 pcDNA3.1-lncRNA AFAP1-AS1 则在蛋白和 mRNA 水平上促进了它们的表达（<0.05）。此外，芒柄花素显著降低了 TNBC 细胞中 lncRNA AFAP1-AS1、CDK4 和 Raf-1 的表达，同时提高了 miR-195 和 miR-545 的表达（<0.05），并显著抑制了 TNBC 细胞的恶性行为（<0.05）。综上所述，芒柄花素通过抑制 lncRNA AFAP1-AS1-miR-195/miR-545 轴缓解 TNBC 的恶性程度，提示联合中药的分子靶点可能在 TNBC 中产生显著的临床获益。
