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Targeting specificity protein 1 with miR-128-3p overcomes TGF-β1 mediated epithelial-mesenchymal transition in breast cancer: An in vitro study

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Abstract

Background

Specificity protein 1 (SP1) was found to play a critical role in the regulation of TGF-β1 driven epithelial-mesenchymal transition (EMT). Recent clinical findings demonstrated a significant drop in the expression of miR-128-3p with the cancer progression in breast cancer patients. However, the impact of miR-128-3p on the SP1 expression in breast cancer remains unknown. Herein, we evaluated the role of miR-128-3p mimics in suppressing EMT of breast cancer cell lines by regulating the TGF-β1/SP1 axis.

Methods

miR-128-3p interaction with SP1 was detected by in silico tools and dual-luciferase reporter assay. qPCR, western blot, and immunocytochemistry experiments were conducted for determining the expression levels of miR-128-3p and EMT markers with and without the treatment of miR-128-3p mimics. Further, to understand the effect of miR-128-3p mimics on cancer progression, experiments such as wound healing assay, transwell assay, adhesion assay, and cell cycle analysis were performed.

Results

A significant inverse relation between SP1 and miR-128-3p levels was found in MCF-7 and MDA-MB-231 cell lines. miR-128-3p overexpression impeded the SP1 mediated EMT markers in TGF-β1 stimulated cells by inhibiting the SP1 nuclear function. Further, treatment with miR-128-3p mimics significantly reduced the migration, invasion and spreading capability of TGF-β1 stimulated cells. Flow cytometry results showed the impeding role of miR-128-3p on the cell cycle progression.

Conclusions

Upregulated miR-128-3p inhibited SP1, thereby limiting the TGF-β1 induced EMT in MCF-7 and MDA-MB-231 cell lines for the first time. This study may pave the path to explore novel miRNA therapeutics for eradicating advanced breast cancer cases.

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Data availability

All data generated or analysed during this study are included in this published article.

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Acknowledgements

Amit Khairnar gratefully acknowledges the support of the Ramalingaswami fellowship (No. BT/RLF/Re-entry/24/2017) from the Department of Biotechnology, India.

Funding

This supplement was supported by the seed fund of the National Institute of Pharmaceutical Education and Research (NIPER)-Ahmedabad, Department of Pharmaceutics, Ministry of Chemicals and Fertilizers, Government of India.

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Authors and Affiliations

  1. Department of Pharmacology and Toxicology, National Institute of Pharmaceutical Education and Research-Ahmedabad (NIPER-A), Palaj, Gandhinagar, Gujarat, 382355, India

    Lakshmi Vineela Nalla & Amit Khairnar

  2. Department of Biotechnology, National Institute of Pharmaceutical Education and Research-Ahmedabad (NIPER-A), Gandhinagar, Gujarat, 382355, India

    Piyush Gondaliya & Kiran Kalia

Authors
  1. Lakshmi Vineela Nalla
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  2. Piyush Gondaliya
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Correspondence to Amit Khairnar.

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Nalla, L.V., Gondaliya, P., Kalia, K. et al. Targeting specificity protein 1 with miR-128-3p overcomes TGF-β1 mediated epithelial-mesenchymal transition in breast cancer: An in vitro study. Mol Biol Rep 49, 6987–6996 (2022). https://doi.org/10.1007/s11033-022-07466-2

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