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EGF-SNX3-EGFR axis drives tumor progression and metastasis in triple-negative breast cancers

Oncogene volume 41, pages 220–232 (2022)Cite this article

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Abstract

Epidermal growth factor receptor (EGFR) has critical roles in epithelial cell physiology. Over-expression and over-activation of EGFR have been implicated in diverse cancers, including triple-negative breast cancers (TNBCs), prompting anti‐EGFR therapies. Therefore, developing potent therapies and addressing the inevitable drug resistance mechanisms necessitates deciphering of EGFR related networks. Here, we describe Sorting Nexin 3 (SNX3), a member of the recycling retromer complex, as a critical player in the epidermal growth factor (EGF) stimulated EGFR network in TNBCs. We show that SNX3 is an immediate and sustained target of EGF stimulation initially at the protein level and later at the transcriptional level, causing increased SNX3 abundance. Using a proximity labeling approach, we observed increased interaction of SNX3 and EGFR upon EGF stimulation. We also detected colocalization of SNX3 with early endosomes and endocytosed EGF. Moreover, we show that EGFR protein levels are sensitive to SNX3 loss. Transient RNAi models of SNX3 downregulation have a temporary reduction in EGFR levels. In contrast, long-term silencing forces cells to recover and overexpress EGFR mRNA and protein, resulting in increased proliferation, colony formation, migration, invasion in TNBC cells, and increased tumor growth and metastasis in syngeneic models. Consistent with these results, low SNX3 and high EGFR mRNA levels correlate with poor relapse-free survival in breast cancer patients. Overall, our results suggest that SNX3 is a critical player in the EGFR network in TNBCs with implications for other cancers dependent on EGFR activity.

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Fig. 1: EGF induction of SNX3.
Fig. 2: Effect of blocking of EGFR and downstream pathways on SNX3 protein levels.
Fig. 3: Proximity-based labeling by SNX3-TurboID.
Fig. 4: Time-dependent interactions of SNX3.
Fig. 5: Effects of transient SNX3 silencing.
Fig. 6: Effects of long-term silencing of SNX3.
Fig. 7: Effect of Snx3 loss on tumor progression and metastasis in vivo.
Fig. 8: Schematic diagram showing the EGF-SNX3-EGFR axis.

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Acknowledgements

We thank current and past members of the laboratory for critical reading of the manuscript and discussions. We thank METABRIC Consortium for sharing the breast cancer data. TUBITAK 116Z257 (AEEB), and 119Z672 (EC). EMBO short term fellowship (MO). National Institutes of Health Grant 2P20GM109091-06 (OS), 1R01CA251374-01A1 (OS). The Turkish Ministry of Development project #KanSil 2016K121540 (CanSyL).

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Author notes

  1. These authors contributed equally: Esra Cicek, Ayca Circir, Merve Oyken.

  2. Deceased 19 May 2020: Huib Ovaa.

Authors and Affiliations

  1. Department of Biological Sciences, Middle East Technical University (METU), Dumlupinar Blv No:1, Universiteler Mah., Cankaya, 06800, Ankara, Turkey

    Esra Cicek, Ayca Circir, Merve Oyken, Didem Naz Dioken & Ayse Elif Erson-Bensan

  2. Oncode Institute & Department of Cell and Chemical Biology, Leiden University Medical Center, Einthovenweg 20, 2333 ZC, Leiden, The Netherlands

    Merve Oyken, Aysegul Sapmaz & Huib Ovaa

  3. Department of Molecular Biology and Genetics, Bilkent University, 06800, Ankara, Turkey

    Ozge Akbulut Caliskan

  4. Cancer System Biology Laboratory, CanSyL, Graduate School of Informatics, Middle East Technical University, 06800, Ankara, Turkey

    Sezen Guntekin Ergun, Rengul Cetin-Atalay & Ayse Elif Erson-Bensan

  5. Drug Discovery and Biomedical Sciences, College of Pharmacy, University of South Carolina, Columbia, SC, 29208, USA

    Ozgur Sahin

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Contributions

EC performed EGF induction experiments, AC developed and characterized RNAi models of SNX3, MO performed siRNA transfections, biotinylation, and microscopy experiments, OAC performed in vivo experiments, DND performed survival analysis, SGE and RCA performed and guided NanoString hybridizations, respectively, AS and HO oversaw protein interaction and localization experiments, OS managed mouse experiments, AEEB designed and supervised experiments, and wrote the paper. All authors, except late HO read and commented on the paper.

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Correspondence to Ayse Elif Erson-Bensan.

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Cicek, E., Circir, A., Oyken, M. et al. EGF-SNX3-EGFR axis drives tumor progression and metastasis in triple-negative breast cancers. Oncogene 41, 220–232 (2022). https://doi.org/10.1038/s41388-021-02086-9

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