The Second People's Hospital of China Three Gorges University, Yichang 443002, China.
Hubei Key Laboratory of Tumor Microenvironment and Immunotherapy, Medical College, China Three Gorges University, Yichang 443002, China; Yichang Center for Disease Control and Prevention, Yichang 443002, China.
Life Sci. 2018 Nov 15;213:149-157. doi: 10.1016/j.lfs.2018.10.034. Epub 2018 Oct 20.
To study the underlying mechanisms of sulforaphane, a natural histone deacetylase (HDAC) inhibitor, in inhibiting triple negative breast cancer cells growth and the therapeutic effects of combination of sulforaphane and doxorubicin in TNBC treatment.
The antineoplastic activity of sulforaphane was evaluated in MDA-MB-231, BT549 and MDA-MB-468 cells with MTT assay. Cell apoptosis was detected with Annexin V/PI double staining by Flow cytometry. Cell autophagy was detected with fluorescence microscope. The effects of Sulforaphane and Doxorubicin combination treatments on cells growth were determined with Chou-Talalay median effect/combination index (CI) model. mRNA and protein expression of genes were assayed respectively with real-time PCR and Western bloting. Protein-protein interaction was detected with co-immunoprecipation. Gene knock-down was performed with small interfere RNA. In vivo assay of combinational treatment with sulforaphane and doxorubicin was investigated in athymic nude mice bearing MDA-MB-231 xenografts.
Results showed that sulforaphane inhibited cell growth and induced autophagy in MDA-MB-231, BT549 and MDA-MB-468 cells. Further study demonstrated that sulforaphane induced autophagy by down-regulating expression of HDAC6, which resulted in increased membrane translocation and acetylation modification of phosphatase and tensin homolog (PTEN). Sulforaphane and doxorubicin combination exhibited a synergistic inhibition on TNBC cells growth. In nude mice, the combination of sulforaphane and doxorubicin displayed a greater inhibitory effect on MDA-MB-231 xenografts growth as compared to either treatment alone.
Our study suggested that induction of autophagy by targeting HDAC6 in combination with chemotherapeutic reagent may provide a novel strategy for TNBC therapy.
研究天然组蛋白去乙酰化酶（HDAC）抑制剂萝卜硫素抑制三阴性乳腺癌细胞生长的潜在机制，以及萝卜硫素与多柔比星联合治疗三阴性乳腺癌的疗效。
采用 MTT 法检测萝卜硫素对 MDA-MB-231、BT549 和 MDA-MB-468 细胞的抗肿瘤活性。采用 Annexin V/PI 双染法通过流式细胞术检测细胞凋亡。荧光显微镜检测细胞自噬。采用 Chou-Talalay 中效/联合指数（CI）模型检测萝卜硫素和多柔比星联合处理对细胞生长的影响。分别采用实时 PCR 和 Western blot 检测基因的 mRNA 和蛋白表达。采用免疫共沉淀检测蛋白质-蛋白质相互作用。采用小干扰 RNA 进行基因敲低。在荷 MDA-MB-231 异种移植瘤的裸鼠体内进行萝卜硫素和多柔比星联合治疗的体内实验。
结果表明，萝卜硫素抑制 MDA-MB-231、BT549 和 MDA-MB-468 细胞的生长并诱导自噬。进一步研究表明，萝卜硫素通过下调 HDAC6 的表达诱导自噬，导致磷酸酶和张力蛋白同源物（PTEN）的膜转位和乙酰化修饰增加。萝卜硫素和多柔比星联合用药对三阴性乳腺癌细胞生长表现出协同抑制作用。在裸鼠中，与单独用药相比，萝卜硫素和多柔比星联合用药对 MDA-MB-231 异种移植瘤的生长具有更大的抑制作用。
本研究表明，通过靶向 HDAC6 诱导自噬与化疗药物联合可能为三阴性乳腺癌治疗提供新策略。
