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Background <p>Triple-negative breast cancer (TNBC) is highly aggressive tumor with limited therapeutic options. Studying the molecular mechanisms underlying TNBC is necessary to address the unmet need in novel therapeutic targets. TNBC is demonstrated to have robust fatty acid (FA) metabolism activity, and recent studies proposed the linkage of FA metabolism with ferroptosis sensitivity. Hence, this study aimed to explore the targets that may regulate FA metabolism to sensitize TNBC cells to ferroptosis.</p> Methods <p>RNA-sequencing data in The Cancer Genome Atlas (TCGA) and four microarray datasets in Gene Expression Omnibus (GEO) database were analyzed to identify key target <i>RACGAP1</i>, followed by a series of functional experiments to explore the exact role of <i>RACGAP1</i> in two TNBC cell lines (human MDA-MB-231 and mouse 4T1) and Xenograft tumor model. Dual-luciferase and chromatin immunoprecipitation (ChIP) assay was utilized to verify the binding of <i>RACGAP1</i> and <i>MAZ</i>. RNA sequencing on 4T1 cells transfecting with sh-NC and sh-<i>RACGAP1</i> was performed to validate the actions of <i>RACGAP1</i>.</p> Results <p><i>RACGAP1</i> was highly expressed in breast cancer, and associated with poor prognosis and ferroptosis activity. <i>RACGAP1</i> silencing could inhibit tumor cells survival and promote ferroptosis, and such anti-tumor activity could be blocked by ferroptosis inhibitors. RNA-sequencing analysis suggested that <i>RACGAP1</i> silencing could inhibit FA metabolism activity, which was further confirmed by metabolic analysis and the reduced level of ATP, triglyceride and FA oxidation. <i>CPT1A</i> overexpression reversed such changes, indicating that the regulation of <i>RACGAP1</i> on FA metabolism was <i>CPT1A</i>-dependent. Activation of FA metabolism activity or <i>CPT1A</i> overexpression blocked the ferroptosis sensitivity induced by <i>RACGAP1</i> silencing. Transcription factor <i>MAZ</i> was identified to directly up-regulate the expression of <i>RACGAP1</i>.</p> Conclusion <p>Inhibition of <i>RACGAP1</i> sensitized TNBC cells to ferroptosis by inhibiting <i>CPT1A</i>-mediated FA metabolism. Targeting <i>RACGAP1</i> might be feasible strategy for TNBC management.</p> Graphical Abstract <p></p>

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Inhibition of RACGAP1 sensitizes triple-negative breast cancer cells to ferroptosis by regulating CPT1A-dependent fatty acid metabolism

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年12月24日

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  • Zhike Zhou,
  • Ye Hua,
  • Jun Ma,
  • Wenqiang Cong,
  • Rui Zhan,
  • Kexin Kang,
  • Lu Wang,
  • Hongyi Wei

摘要

Background

Triple-negative breast cancer (TNBC) is highly aggressive tumor with limited therapeutic options. Studying the molecular mechanisms underlying TNBC is necessary to address the unmet need in novel therapeutic targets. TNBC is demonstrated to have robust fatty acid (FA) metabolism activity, and recent studies proposed the linkage of FA metabolism with ferroptosis sensitivity. Hence, this study aimed to explore the targets that may regulate FA metabolism to sensitize TNBC cells to ferroptosis.

Methods

RNA-sequencing data in The Cancer Genome Atlas (TCGA) and four microarray datasets in Gene Expression Omnibus (GEO) database were analyzed to identify key target RACGAP1, followed by a series of functional experiments to explore the exact role of RACGAP1 in two TNBC cell lines (human MDA-MB-231 and mouse 4T1) and Xenograft tumor model. Dual-luciferase and chromatin immunoprecipitation (ChIP) assay was utilized to verify the binding of RACGAP1 and MAZ. RNA sequencing on 4T1 cells transfecting with sh-NC and sh-RACGAP1 was performed to validate the actions of RACGAP1.

Results

RACGAP1 was highly expressed in breast cancer, and associated with poor prognosis and ferroptosis activity. RACGAP1 silencing could inhibit tumor cells survival and promote ferroptosis, and such anti-tumor activity could be blocked by ferroptosis inhibitors. RNA-sequencing analysis suggested that RACGAP1 silencing could inhibit FA metabolism activity, which was further confirmed by metabolic analysis and the reduced level of ATP, triglyceride and FA oxidation. CPT1A overexpression reversed such changes, indicating that the regulation of RACGAP1 on FA metabolism was CPT1A-dependent. Activation of FA metabolism activity or CPT1A overexpression blocked the ferroptosis sensitivity induced by RACGAP1 silencing. Transcription factor MAZ was identified to directly up-regulate the expression of RACGAP1.

Conclusion

Inhibition of RACGAP1 sensitized TNBC cells to ferroptosis by inhibiting CPT1A-mediated FA metabolism. Targeting RACGAP1 might be feasible strategy for TNBC management.

Graphical Abstract