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Engineering tumor-derived small extra cellular vesicles to encapsulate miR-34a, effectively inhibits 4T1 cell proliferation, migration, and gene expression

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Abstract

Tumor cells produce small extra cellular vesicles-(tsEV) massively, which act as cancer messengers that may also have anti-cancer effects. Based on this knowledge, we hypothesized that we can benefit from 4T1-derived sEVs to amplify the anti-cancer effects of miR-34a-replacement therapy in 4T1 cells. Supernatant of 4T1 cultured cells gathered after 24 h of exposure to serum-free media. tsEVs purified by commercial kit and characterized by transmission and scanning electron microscopy, dynamic light scattering, and bicinchoninic acid assay. Modified CaCl2 method applied for miR-34a loading in tsEV (tsEV-miR) and loading confirmation evaluated by the relative expression of miR-34a. MTT, annexin V/PI, cell cycle, scratch test, and real-time PCR were performed for proliferation, apoptosis, invasion, and relative expression of miR-34a target genes after treatment with tsEV/tsEV-miR, respectively. The results indicated that tsEV-miR provides a time–dose–dependent anti-proliferative effect versus tsEV/control group. tsEV-miR could induce apoptosis and arrest the cell cycle at G0/G1 phase, and moreover, it effectively halted the invasion capability of 4T1 cells. Treatment with tsEV-miR down-regulated miR-34a target genes, including B-cell lymphoma-2, vascular endothelial growth factor and its receptor, matrix metalloproteinase-2 and -9, and interleukin-6. Engineered tsEVs can affect different aspects of 4T1 cancer cells including proliferation, apoptosis, cell cycle, migration, and cancer-related gene expression profile. In this regard, tsEV could be considered a proper vehicle for miR-34a replacement therapy and could exacerbate its anti-cancer effects in triple-negative breast cancer. Indeed, TNBC can be targeted by multiple angles by its weapon.

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Data availability

The datasets related to findings of this research are available upon reasonable request from the corresponding author.

Abbreviations

miR:

MicroRNA

tsEV:

Tumor-derived small extra cellular vesicle

tsEV-miR:

MiR-34a loaded tsEV

DLS:

Dynamic light scattering

TEM:

Transmission electron microscopy

BCA:

Bicinchoninic acid assay

BCL2:

B-cell lymphoma-2

MMP-2,MMP-9:

Matrix metalloproteinase-2 and -9

IL-6:

Interleukin-6

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Acknowledgements

This article has been extracted from the thesis written by Ms. Mahsa Hajivalili in School of Medicine, Shahid Beheshti University of Medical Sciences (Registration No: 391). Ethics committee approval ID was IR.SBMU.MSP.REC.1398.581

Funding

This research was financially supported by the Shahid Beheshti University of Medical Sciences under Grant [No. 20846].

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Authors and Affiliations

  1. Department of Immunology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran

    Mahsa Hajivalili, Nariman Mosaffa, Bahare Niknam & Davar Amani

  2. Gastroenterology and Liver Diseases Research Center, Research Institute for Gastroenterology and Liver Diseases, Shahid Beheshti University of Medical Sciences, Tehran, Iran

    Kaveh Baghaei & Bahare Niknam

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  1. Mahsa Hajivalili
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  2. Kaveh Baghaei
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Contributions

MH involved in investigation, visualization, formal analysis, data curation, and writing -original draft. KB participated in visualization and writing -review and editing. NM took part in resources, writing-review & editing. BN performed visualization. DA carried out conceptualization, project administration, funding acquisition, writing-review & editing, supervision, and validation. All authors contributed to the article and approved the submitted version.

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Correspondence to Davar Amani.

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Hajivalili, M., Baghaei, K., Mosaffa, N. et al. Engineering tumor-derived small extra cellular vesicles to encapsulate miR-34a, effectively inhibits 4T1 cell proliferation, migration, and gene expression. Med Oncol 39, 93 (2022). https://doi.org/10.1007/s12032-022-01685-0

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