Mechanism of ferroptosis mediated by the bromodomain protein subfamily 4/nuclear factor-kappa B signaling pathway in chemotherapy resistance of triple-negative breast cancer

Zhang Shuowen; Li Dan; He Jing; Du Zhixing; Pei Yongbin

doi:10.13406/j.cnki.cyxb.003536

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Mechanism of ferroptosis mediated by the bromodomain protein subfamily 4/nuclear factor-kappa B signaling pathway in chemotherapy resistance of triple-negative breast cancer

Basic research | 更新时间：2025-10-21

- Mechanism of ferroptosis mediated by the bromodomain protein subfamily 4/nuclear factor-kappa B signaling pathway in chemotherapy resistance of triple-negative breast cancer
- Journal of Chongqing Medical University Vol. 49, Issue 7, Pages: 844-852(2024)
- 作者机构：
  
  1.河北医科大学第一医院健康管理中心，石家庄 050000
  2.河北医科大学第一医院普外科，石家庄 050000
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.003536
  CLC： R737.9
- Received：31 August 2023，
  
  Revised：2024-03-04，
  
  Accepted：12 March 2024，
  
  Published：28 July 2024
- 稿件说明：
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张硕稳,李丹,贺静,等.BRD4/NF-κB信号通路介导的铁死亡参与三阴性乳腺癌化疗耐药的机制[J].重庆医科大学学报,2024,49(7):844-852.

Zhang Shuowen,Li Dan,He Jing,et al.Mechanism of ferroptosis mediated by the bromodomain protein subfamily 4/nuclear factor-kappa B signaling pathway in chemotherapy resistance of triple-negative breast cancer[J].Journal of Chongqing Medical University,2024,49(7):844-852.
张硕稳,李丹,贺静,等.BRD4/NF-κB信号通路介导的铁死亡参与三阴性乳腺癌化疗耐药的机制[J].重庆医科大学学报,2024,49(7):844-852. DOI： 10.13406/j.cnki.cyxb.003536.

Zhang Shuowen,Li Dan,He Jing,et al.Mechanism of ferroptosis mediated by the bromodomain protein subfamily 4/nuclear factor-kappa B signaling pathway in chemotherapy resistance of triple-negative breast cancer[J].Journal of Chongqing Medical University,2024,49(7):844-852. DOI： 10.13406/j.cnki.cyxb.003536.

摘要

目的

探讨溴域蛋白亚家族4（bromodomain protein subfamily 4，BRD4）/核因子κB（nuclear factor kappa-B，NF-κB）信号通路介导的铁死亡参与乳腺癌恶性生物学行为的机制。

方法

多柔比星（Doxorubicin，DOX）抗性MDA-MB-231细胞（MDA-MB-231/DOX）分为对照（Con）组、DOX组和si-BRD4+DOX组。si-BRD4+DOX组在用si-BRD4转染MDA-MB-231/DOX细胞48 h后，用1 μmol/L DOX处理细胞24 h。通过集落形成试验和5-乙炔基-2′-脱氧尿苷（5-Ethynyl-2'-deoxyuridine，EdU）分析评估细胞增殖能力。通过FerroOrange、liperfloo检测细胞亚铁离子浓度和脂质过氧化水平。将15只雌性BALB/c-nu小鼠随机分为3组：对照组、DOX组、DOX+si-BRD4组，每组5只，用于建立MDA-MB-231/DOX细胞皮下接种模型。通过qRT-PCR、蛋白质印迹、免疫组化分析BRD4/NF-κB信号通路表达。

结果

与si-NC组相比，si-BRD4#1和si-BRD4#2组MDA-MB-231、BT549细胞的细胞克隆数、EDU阳性染色、谷胱甘肽（glutathione，GSH）水平均降低（

0.001），和MDA-MB-231、BT549细胞亚铁离子水平、活性氧（reactive oxygen species，ROS）水平、氧化型谷胱甘肽（Oxidized glutathione，GSSG）/GSH比值升高（

0.01）。与亲代细胞（MDA-MB-231）相比，在MDA-MB-231/DOX中BRD4的mRNA和蛋白质表达水平升高。与Con组相比，DOX组细胞中IKβ-α、NF-κB、BRD4表达和ROS水平增加（

0.05），和细胞克隆数和EDU阳性染色均降低（

0.05）；与DOX组相比，si-BRD4+DOX组细胞中核因子κB抑制蛋白α（Anti-IKB alpha，IKβ-α）、NF-κB表达、细胞克隆数、EDU阳性染色降低（

0.05），ROS水平升高（

0.05）。与对照组相比，DOX组肿瘤重量和体积均减少（

0.05），并且肿瘤组织中TUNEL染色细胞增加（

0.05）。此外，BRD4+DOX组肿瘤重量和体积较DOX组进一步降低（

0.001），TUNEL染色细胞进一步增加（

0.001）。BRD4+DOX组肿瘤组织中Ki-67、BRD4、NF-κB较DOX组下调（

0.01），4-羟基壬烯醛（4-Hydroxynonenal，4-HNE）上调（

0.01）。

结论

BRD4是一个重要的耐药因子，它可以通过促进NF-κB信号通路的激活来抑制乳腺癌化疗诱导的铁死亡。

Abstract

Objective

To investigate the mechanism of ferroptosis mediated by the bromodomain protein subfamily 4（BRD4）/nuclear factor-kappa B（NF-κB） signaling pathway in the malignant biological behavior of breast cancer.

Methods

Doxorubicin-resistant MDA-MB-231 cells（MDA-MB-231/DOX） were divided into control group（Con group），DOX group，and si-BRD4+DOX group. In the si-BRD4+DOX group，MDA-MB-231/DOX cells were transfected with si-BRD4 for 48 hours and were then treated with 1 μmol/L DOX for 24 hours. Colony formation assay and 5-ethynyl-2'-deoxyuridine（EdU） analysis were used to evaluate the proliferation ability of cells，and FerroOrange and liperfloo were used to measure the concentration of ferrous ion and the level of lipid peroxidation. A total of 15 female BALB/c-nu mice were randomly divided into control group，DOX group，and DOX+Si-BRD group，with 5 mice in each group，and a subcutaneous inoculation model of MDA-MB-231/DOX cells was established. The methods of qRT-PCR，Western blotting，and immunohistochemistry were used to measure the expression of the BRD4/NF-κB signaling pathway.

Results

Compared with the si-NC group，the si-BRD4#1 group and the si-BRD4#2 group had significant reductions in the number of cell clones，positive EDU staining，and glutathione level in MDA-MB-231 and BT549 cells（

0.001），as well as significant increases in the level of ferrous ion，the level of reactive oxygen species（ROS），and oxidized glutathione/glutathione ratio（

0.01）. Compared with the parental cells（MDA-MB-231），there were increases in the mRNA and protein expression levels of BRD4 in MDA-MB-231/DOX cells. Compared with the Con group，the DOX group had significant increases in the expression levels of IKβ-α，NF-κB，and BRD4 and the level of ROS（

0.05） and significant reductions in the number of cell clones and positive EDU staining（

0.05）； compared with the DOX group，the si-BRD4+DOX group had significant reductions in the expression levels of IKβ-α and NF-κB，the number of cell clones，and positive EDU staining（

0.05） and a significant increase in the level of ROS（

0.05）. Compared with the control group，the DOX group had significant reductions in tumor weight and volume（

0.05） and a significant increase in the number of TUNEL-stained cells in tumor tissue（

0.05）. In addition，compared with the DOX group，the BRD4+DOX group had further reductions in tumor weight and volume（

0.001） and a significant increase in the number of TUNEL-stained cells（

0.001）. Compared with the DOX group，the BRD4+DOX group had significant reductions in Ki-67，BRD-4，and NF-κB（

0.01） and a significant increase in 4-hydroxynonenal（

0.01） in tumor tissue.

Conclusion

BRD4 is an important drug resistance factor，which can inhibit ferroptosis induced by chemotherapy in breast cancer by promoting the activation of the NF-κB signaling pathway.

关键词

Keywords

references

Sun JJ ， Zhao HY ， Xu WX ， et al . Recent advances in photothermal therapy-based multifunctional nanoplatforms for breast cancer [J ] . Front Chem ， 2022 ， 10 ： 1024177 ．

Derakhshan F ， Reis-Filho JS . Pathogenesis of triple-negative breast cancer [J ] . Annu Rev Pathol ， 2022 ， 17 ： 181 - 204 ．

Li Y ， Zhang HJ ， Merkher Y ， et al . Recent advances in therapeutic strategies for triple-negative breast cancer [J ] . J Hematol Oncol ， 2022 ， 15 （ 1 ）： 121 ．

Asleh K ， Riaz N ， Nielsen TO . Heterogeneity of triple negative breast cancer：current advances in subtyping and treatment implications [J ] . J Exp Clin Cancer Res ， 2022 ， 41 （ 1 ）： 265 ．

Zhang Z ， Lu ML ， Chen CL ， et al . Holo-lactoferrin：the link between ferroptosis and radiotherapy in triple-negative breast cancer [J ] . Theranostics ， 2021 ， 11 （ 7 ）： 3167 - 3182 ．

Xie YZ ， Wang BY ， Zhao YN ， et al . Mammary adipocytes protect triple-negative breast cancer cells from ferroptosis [J ] . J Hematol Oncol ， 2022 ， 15 （ 1 ）： 72 ．

Li K ， Xu K ， He Y ， et al . Oxygen self-generating nanoreactor mediated ferroptosis activation and immunotherapy in triple-negative breast cancer [J ] . ACS Nano ， 2023 ， 17 （ 5 ）： 4667 - 4687 ．

Li K ， Lin CC ， Li MH ， et al . Multienzyme-like reactivity cooperatively impairs glutathione peroxidase 4 and ferroptosis suppressor protein 1 pathways in triple-negative breast cancer for sensitized ferroptosis therapy [J ] . ACS Nano ， 2022 ， 16 （ 2 ）： 2381 - 2398 ．

Liu J ， Zhu S ， Zeng L ， et al . DCN released from ferroptotic cells ignites AGER-dependent immune responses [J ] . Autophagy ， 2022 ， 18 （ 9 ）： 2036 - 2049 ．

Ushijima T ， Clark SJ ， Tan P . Mapping genomic and epigenomic evolution in cancer ecosystems [J ] . Science ， 2021 ， 373 （ 6562 ）： 1474 - 1479 ．

Wu YQ ， Zhang SW ， Gong XX ， et al . The epigenetic regulators and metabolic changes in ferroptosis-associated cancer progression [J ] . Mol Cancer ， 2020 ， 19 （ 1 ）： 39 ．

Zhang JF ， Tang P ， Zou L ， et al . Discovery of novel dual-target inhibitor of bromodomain-containing protein 4/casein kinase 2 inducing apoptosis and autophagy-associated cell death for triple-negative breast cancer therapy [J ] . J Med Chem ， 2021 ， 64 （ 24 ）： 18025 - 18053 ．

Chang XS ， Sun DJ ， Shi DF ， et al . Design，synthesis，and biological evaluation of quinazolin-4（3 H ）-one derivatives co-targeting poly（ADP-ribose） polymerase-1 and bromodomain containing protein 4 for breast cancer therapy [J ] . Acta Pharm Sin B ， 2021 ， 11 （ 1 ）： 156 - 180 ．

Schmitt A ， Grimm M ， Kreienkamp N ， et al . BRD4 inhibition sensitizes diffuse large B-cell lymphoma cells to ferroptosis [J ] . Blood ， 2023 ， 142 （ 13 ）： 1143 - 1155 ．

Yang MH ， Liu K ， Chen P ， et al . Bromodomain-containing protein 4（BRD4） as an epigenetic regulator of fatty acid metabolism genes and ferroptosis [J ] . Cell Death Dis ， 2022 ， 13 （ 10 ）： 912 ．

Huang JF ， Wen CJ ， Zhao GZ ， et al . Overexpression of ABCB4 contributes to acquired doxorubicin resistance in breast cancer cells in vitro [J ] . Cancer Chemother Pharmacol ， 2018 ， 82 （ 2 ）： 199 - 210 ．

Zhou TQ ， Zhou MM ， Tong CC ， et al . Cauliflower bioactive compound sulforaphane inhibits breast cancer development by suppressing NF-κB/MMP-9 signaling pathway expression [J ] . Cell Mol Biol ， 2022 ， 68 （ 4 ）： 134 - 143 ．

Yang GJ ， Song YQ ， Wang WH ， et al . An optimized BRD4 inhibitor effectively eliminates NF-κB-driven triple-negative breast cancer cells [J ] . Bioorg Chem ， 2021 ， 114 ： 105158 ．

Komohara Y ， Kurotaki D ， Tsukamoto H ， et al . Involvement of protumor macrophages in breast cancer progression and characterization of macrophage phenotypes [J ] . Cancer Sci ， 2023 ， 114 （ 6 ）： 2220 - 2229 ．

Greco S ， Fabbri N ， Spaggiari R ， et al . Update on classic and novel approaches in metastatic triple-negative breast cancer treatment：a comprehensive review [J ] . Biomedicines ， 2023 ， 11 （ 6 ）： 1772 ．

Sarnella A ， Ferrara Y ， Albanese S ， et al . Inhibition of bone marrow-mesenchymal stem cell-induced carbonic anhydrase IX potentiates chemotherapy efficacy in triple-negative breast cancer cells [J ] . Cells ， 2023 ， 12 （ 2 ）： 298 ．

Sen U ， Coleman C ， Sen T . Stearoyl coenzyme A desaturase-1：multitasker in cancer，metabolism，and ferroptosis [J ] . Trends Cancer ， 2023 ， 9 （ 6 ）： 480 - 489 ．

Li PY ， Lin QH ， Sun SY ， et al . Inhibition of cannabinoid receptor type 1 sensitizes triple-negative breast cancer cells to ferroptosis via regulating fatty acid metabolism [J ] . Cell Death Dis ， 2022 ， 13 （ 9 ）： 808 ．

Sha R ， Xu YQ ， Yuan CW ， et al . Predictive and prognostic impact of ferroptosis-related genes ACSL4 and GPX4 on breast cancer treated with neoadjuvant chemotherapy [J ] . EBioMedicine ， 2021 ， 71 ： 103560 ．

Hu JF ， Pan D ， Li G ， et al . Regulation of programmed cell death by Brd4 [J ] . Cell Death Dis ， 2022 ， 13 （ 12 ）： 1059 ．

Donati B ， Lorenzini E ， Ciarrocchi A . BRD4 and Cancer：going beyond transcriptional regulation [J ] . Mol Cancer ， 2018 ， 17 （ 1 ）： 164 ．

Sui SY ， Zhang J ， Xu SP ， et al . Ferritinophagy is required for the induction of ferroptosis by the bromodomain protein BRD4 inhibitor（+）-JQ1 in cancer cells [J ] . Cell Death Dis ， 2019 ， 10 （ 5 ）： 331 ．

Luo S ， Qin SJ ， Oudeng G ， et al . Iron-based hollow nanoplatforms for cancer imaging and theranostics [J ] . Nanomaterials ， 2022 ， 12 （ 17 ）： 3023 ．

Yang GJ ， Wang WH ， Lei PM ， et al . A 7-methoxybicoumarin derivative selectively inhibits BRD4 BD2 for anti-melanoma therapy [J ] . Int J Biol Macromol ， 2020 ， 164 ： 3204 - 3220 ．

Song MG ， Li JY ， Sun J ， et al . DNMT1-mediated DNA methylation in toll-like receptor 4（TLR4） inactivates NF-κB signal pathway-triggered pyroptotic cell death and cellular inflammation to ameliorate lipopolysaccharides（LPS）-induced osteomyelitis [J ] . Mol Cell Probes ， 2023 ， 71 ： 101922 ．

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