HIF-1 promotes murine breast cancer brain metastasis by increasing production of integrin β3–containing extracellular vesicles
Yongkang Yang, … , Kathleen L. Gabrielson, Gregg L. Semenza
Yongkang Yang, … , Kathleen L. Gabrielson, Gregg L. Semenza
Published July 15, 2025
Citation Information: J Clin Invest. 2025;135(14):e190470. https://doi.org/10.1172/JCI190470.
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Research Article Oncology Vascular biology

HIF-1 promotes murine breast cancer brain metastasis by increasing production of integrin β3–containing extracellular vesicles

Abstract

Brain metastasis is a major cause of breast cancer (BC) mortality, but the cellular and molecular mechanisms have not been fully elucidated. BC cells must breach the blood-brain barrier in order to colonize the brain. Here, we determined that integrin β3 (ITGB3) expression mediated by hypoxia-inducible factor 1 (HIF-1) plays a critical role in metastasis of BC cells to the brain. Hypoxia stimulated BC cell migration and invasion ex vivo and brain colonization in vivo. Knockdown of either HIF-1α or ITGB3 expression impaired brain colonization by human or mouse BC cells injected into the cardiac left ventricle. Exposure of BC cells to hypoxia increased expression of ITGB3 and its incorporation into small extracellular vesicles (EVs). EVs harvested from the conditioned medium of hypoxic BC cells showed increased retention in the brain after intracardiac injection that was HIF-1α and ITGB3 dependent. EVs from hypoxic BC cells showed binding to brain endothelial cells (ECs), leading to increased EC–BC cell interaction, increased vascular endothelial growth factor receptor 2 signaling, increased EC permeability, and increased transendothelial migration of BC cells. Taken together, our studies implicate HIF-1–stimulated production of ITGB3+ EVs as a key mechanism by which hypoxia promotes BC brain metastasis.

Authors

Yongkang Yang, Chelsey Chen, Yajing Lyu, Olesia Gololobova, Xin Guo, Tina Yi-Ting Huang, Vijay Ramu, Varen Talwar, Elizabeth E. Wicks, Shaima Salman, Daiana Drehmer, Dominic Dordai, Qiaozhu Zuo, Kenneth W. Witwer, Kathleen L. Gabrielson, Gregg L. Semenza

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Figure 1

Identification of HIF target genes in brain-metastatic BC cells by RNA-Seq and ChIP-Seq.

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Identification of HIF target genes in brain-metastatic BC cells by RNA-S...
(A and B) RNA-Seq analysis of MDA231-BrM2 cells exposed to 20% or 1% O2 for 24 hours was performed. Volcano plot (A) and Gene Ontology (GO) analysis (B) of HIF target genes are shown. (C) Gene set enrichment analysis (GSEA) revealed that expression of the “breast cancer metastasis” gene set was significantly correlated with the expression of HIF target genes in MDA231-BrM2 cells. (D) HIF-1α binding profiles at significantly called peak summits ± 1 kb in MDA231-BrM2 cells exposed to 20% or 1% O2, as determined by ChIP-Seq. (E) Venn analysis shows the overlap among 2,234 HIF-dependent hypoxia-induced genes as determined by RNA-Seq; 1,474 genes with HIF-1α binding sites by ChIP-Seq; and 1,957 genes overexpressed in MDA231-BrM2 cells, as compared with MDA231 cells (from ref. 16). (F) Heatmaps showing RNA expression of 44 shared genes in NTC versus HIF-double-knockdown cells at 20% or 1% O2 (left), and in MDA231 versus MDA231-BrM2 cells at 20% O2 (right).