Top

Journal of Experimental & Clinical Cancer Research

Published in:

01-12-2021 | Breast Cancer | Research

Chemotherapy-elicited exosomal miR-378a-3p and miR-378d promote breast cancer stemness and chemoresistance via the activation of EZH2/STAT3 signaling

Authors: Qianxi Yang, Shaorong Zhao, Zhendong Shi, Lixia Cao, Jingjing Liu, Teng Pan, Dongdong Zhou, Jin Zhang

Published in: Journal of Experimental & Clinical Cancer Research | Issue 1/2021

Abstract

Background

Not all breast cancer (BC) patients who receive neoadjuvant chemotherapy achieve a pathologic complete response (pCR), but the reasons for this are unknown. Previous studies have shown that exosomes produced in the tumor microenvironment in response to chemotherapy promote a chemotherapy-resistant phenotype in tumors. However, the role of BC chemotherapy-elicited exosomes in regulating chemoresistance is poorly understood.

Methods

Using commercial kits, serum exosomes were extracted from patients before neoadjuvant chemotherapy, after one cycle of chemotherapy and after four cycles of chemotherapy consisting of doxorubicin (DOX) and paclitaxel (PTX). Their miRNAs were sequenced, and the correlation between the sequencing results and chemotherapy effects was further verified by RT-qPCR using patient serum exosomes. Cell Counting Kit-8 (CCK-8) was used to detect chemosensitivity. Stemness was assessed by CD44+/CD24- population analysis and mammosphere formation assays. Chromatin immunoprecipitation (ChIP) experiments were performed to verify the binding of signal transducer and activator of transcription 3 (STAT3) to the promoter of miRNAs.

Results

Here, we provide clinical evidence that chemotherapy-elicited exosomal miR-378a-3p and miR-378d are closely related to the chemotherapy response and that exosomes produced by BC cells after stimulation with DOX or PTX deliver miR-378a-3p and miR-378d to neighboring cells to activate WNT and NOTCH stemness pathways and induce drug resistance by targeting Dickkopf 3 (DKK3) and NUMB. In addition, STAT3, which is enhanced by zeste homolog 2 (EZH2), bound to the promoter regions of miR-378a-3p and miR-378d, thereby increasing their expression in exosomes. More importantly, chemotherapeutic agents combined with the EZH2 inhibitor tazemetostat reversed chemotherapy-elicited exosome-induced drug resistance in a nude mouse tumor xenograft model.

Conclusion

This study revealed a novel mechanism of acquired chemoresistance whereby chemotherapy activates the EZH2/STAT3 axis in BC cells, which then secrete chemotherapy-elicited exosomes enriched in miR-378a-3p and miR-378d. These exosomes are absorbed by chemotherapy-surviving BC cells, leading to activation of the WNT and NOTCH stem cell pathways via the targeting of DKK3 and NUMB and subsequently resulting in drug resistance. Therefore, blocking this adaptive mechanism during chemotherapy may reduce the development of chemotherapy resistance and maximize the therapeutic effect.

Available only for authorised users

Rastogi P, Anderson SJ, Bear HD, Geyer CE, Kahlenberg MS, Robidoux A, Margolese RG, Hoehn JL, Vogel VG, Dakhil SR, Tamkus D, King KM, Pajon ER, Wright MJ, Robert J, Paik S, Mamounas EP, Wolmark N, et al. Preoperative chemotherapy: updates of National Surgical Adjuvant Breast and bowel project protocols B-18 and B-27. J Clin Oncol. 2008;26(5):778–85. https://doi.org/10.1200/JCO.2007.15.0235.CrossRefPubMed

Foulkes WD, Smith IE, Reis-Filho JS. Triple-Negative Breast Cancer. N Engl J Med. 2010;363(20):1938–48. https://doi.org/10.1056/NEJMra1001389.CrossRefPubMed

Annette Becker BKT, Weiss JM, Kim HS, Peinado H, Lyden D. Extracellular vesicles in cancer: cell-to-cell mediators of metastasis. Cancer Cell. 2016;30(6):836–8.CrossRefPubMedPubMedCentral

Sharma A. Chemoresistance in cancer cells: exosomes as potential regulators of therapeutic tumor heterogeneity. Nanomedicine. 2017;12(17):2137–48. https://doi.org/10.2217/nnm-2017-0184.CrossRefPubMed

Ioanna Keklikoglou CC, Güç E, Squadrito ML, Spring LM, Tazzyman S, Lambein L, Poissonnier A, Ferraro GB, Baer C, Cassará A, Guichard A, Iruela-Arispe ML, Lewis CE, Coussens LM, Bardia A, Jain RK, Pollard JW, Palma d M. Chemotherapy elicits pro-metastatic extracellular vesicles in breast cancer models. Nat Cell Biol. 2019;21(2):190–202. https://doi.org/10.1038/s41556-018-0256-3.CrossRefPubMed

Meng Shen CD, Ruan X, Yan W, Cao M, Pizzo D, Wu X, Yang L, Liu L, Ren X, Wang SE. Chemotherapy-Induced Extracellular Vesicle miRNAs Promote Breast Cancer Stemness by Targeting ONECUT2. Cancer Res. 2019;79(14):3608–21. https://doi.org/10.1158/0008-5472.CAN-18-4055.CrossRefPubMedPubMedCentral

Chan T-S, Hsu CC, Pai VC, Liao W-Y, Huang S-S, Tan K-T, Yen CJ, Hsu S-C, Chen W-Y, Shan Y-S, Li C-R, Lee MT, Jiang K-Y, Chu JM, Lien G-S, Weaver VM, Tsai KK. Metronomic chemotherapy prevents therapy-induced stromal activation and induction of tumor-initiating cells. J Exp Med. 2016;213(13):2967–88. https://doi.org/10.1084/jem.20151665.CrossRefPubMedPubMedCentral

ElAyachi I, et al. The WNT10B Network is Associated with survival and metastases in ChemoresistantTriple-negative breast Cancer. Cancer Res. 2019;79(5):982–93. https://doi.org/10.1158/0008-5472.CAN-18-1069.CrossRef

Gardner EE, et al. Chemosensitive relapse in small cell lung cancer proceeds through an EZH2-SLFN11 axis. Cancer Cell. 2017;13:286–99.CrossRef

10.

Malta TM, Sokolov A, Gentles AJ, Burzykowski T, Poisson L, Weinstein JN, Kamińska B, Huelsken J, Omberg L, Gevaert O, Colaprico A, Czerwińska P, Mazurek S, Mishra L, Heyn H, Krasnitz A, Godwin AK, Lazar AJ, The Cancer Genome Atlas Research Network, Stuart JM, Hoadley KA, Laird PW, Noushmehr H, Wiznerowicz M. Machine Learning Identifies Stemness Features Associated with Oncogenic Dedifferentiation. Cell. 2019;173(2):338–54.CrossRef

11.

Wang T, Fahrmann JF, Lee H, Li Y-J, Tripathi SC, Yue C, Zhang C, Lifshitz V, Song J, Yuan Y, Somlo G, Jandial R, Ann D, Hanash S, Jove R, Yu H, et al. Correction--JAK/STAT3-regulated fatty acid β-oxidation is critical for breast Cancer stem Cell self-renewal and Chemoresistance. Cell Metab. 2018;27(1):136–50. https://doi.org/10.1016/j.cmet.2017.11.001.CrossRefPubMed

12.

Kim E, Kim M, Woo D-H, Shin Y, Shin J, Chang N, Oh YT, Kim H, Rheey J, Nakano I, Lee C, Joo KM, Rich JN, Nam D-H, Lee J, et al. Phosphorylation of EZH2 activates STAT3 signaling via STAT3 methylation and promotes tumorigenicity of glioblastoma stemlike cells. Cancer Cell. 2013;23(6):839–85. https://doi.org/10.1016/j.ccr.2013.04.008.CrossRefPubMedPubMedCentral

13.

Jingjing Liu YZ, Shi Z, Hu Y, Meng T, Zhang X, Zhang S, Zhang J. MicroRNA-497 Modulates Breast Cancer Cell Proliferation, Invasion, and Survival by Targeting SMAD7. DNA Cell Biol. 2016;35(9):521–9.CrossRefPubMed

14.

Yabing Zheng XL, Wang X, Wang B, Shao X, Huang Y, Shi L, Chen Z, Huang J, Huang P. MiR-181b promotes chemoresistance in breast cancer by regulating Bim expression. Oncol Rep. 2016;35(2):683–90. https://doi.org/10.3892/or.2015.4417.CrossRefPubMed

15.

Nina Pettersen Hessvik AL. Current knowledge on exosome biogenesis and release. Cell Mol LifeSci. 2018;75(2):193–208. https://doi.org/10.1007/s00018-017-2595-9.CrossRef

16.

Andel HV, et al. Aberrant Wnt signaling in multiple myeloma: molecular mechanisms and targeting options. Leukemia. 2019;33(5):1063–75. https://doi.org/10.1038/s41375-019-0404-1.CrossRefPubMedPubMedCentral

17.

Baron R, Gori F. Targeting WNT signaling in the treatment of osteoporosis. Curr Opin Pharmacol. 2018;40:134–41. https://doi.org/10.1016/j.coph.2018.04.011.CrossRefPubMed

18.

Angela N. Flores, N.M., Armelle Meunier and Laure Marignol, NUMB inhibition of NOTCH signalling as a therapeutic target in prostate cancer. Nat Rev Urol, 2014. 11: p. 499–507, 9, DOI: https://doi.org/10.1038/nrurol.2014.195.

19.

Deep Pokharel, M.P.P., Jamie F. Lu, Ritu Jaiswal, Steven P. Djordjevic, and Mary Bebawy. The Role of CD44 and ERM Proteins in Expression and Functionality of P-glycoprotein in Breast Cancer Cells. Molecules, 2016. 21(290): p. 1–14.

20.

Priya Samuel MF, Carter DRF. Mechanisms of Drug Resistance in Cancer: The Role of Extracellular Vesicles. Proteomics. 2017;17(1600375):1–10.

21.

Ling Mao JL, Zhong S-l, Chen W-x, Zhao J-h, Cai Y-q, Zhou J-w, Yu D-d, Tang J-h. Exosomes decrease sensitivity of breast cancer cells to adriamycin by delivering microRNAs. Tumor Biol. 2015;37(4):5247–56.

22.

Ingenito F, et al. The Role of Exo-miRNAs in Cancer: A Focus on Therapeutic and Diagnostic Applications. Int J Mol Sci. 2019;20:1–17.CrossRef

23.

Bethany N. Hannafon, Y.D.T., Cameron L. Calloway, Y. Daniel Zhao, David H. Lum, Alana L. Welm, Zhizhuang J. Zhao, Kenneth E. Blick, William C. Dooley, and W. Q. Ding, Plasma exosome microRNAs are indicative of breast cancer. Breast Cancer Res, 2016. 18(90): p. 1–14.

24.

Jerome Paggetti FH, Seiffert M, Janji B, Distler U, Ammerlaan W, Kim YJ, Adam J, Lichter P, Solary E, Berchem G, Moussay E. Exosomes released by chronic lymphocytic leukemia cells induce the transition of stromal cells into cancer-associated fibroblasts. Blood. 2015;126(9):1106–17.CrossRefPubMedPubMedCentral

25.

Mio Yoshikawa HI, Umemoto Y, Yanagisawa T, Matsumoto A, Jinno H. Exosome-encapsulated microRNA-223-3p as a minimally invasive biomarker for the early detection of invasive breast cancer. Oncol Lett. 2018;15:9584–92.PubMedPubMedCentral

26.

Ines Stevic VM, Weber K, Fasching PA, Karn T, Marmé F, Schem C, Stickeler E, Denkert C, van Mackelenbergh M, Salat C, Schneeweiss A, Pantel K, Loibl S, Untch M, Schwarzenbach H. Specific microRNA signatures in exosomes of triple-negative and HER2-positive breast cancer patients undergoing neoadjuvant therapy within the GeparSixto trial. BMC Med. 2018;16(179):1–16.

Title: Chemotherapy-elicited exosomal miR-378a-3p and miR-378d promote breast cancer stemness and chemoresistance via the activation of EZH2/STAT3 signaling
Authors: Qianxi Yang
Shaorong Zhao
Zhendong Shi
Lixia Cao
Jingjing Liu
Teng Pan
Dongdong Zhou
Jin Zhang
Publication date: 01-12-2021
Publisher: BioMed Central
Keywords: Breast Cancer
Breast Cancer
Cytostatic Therapy
Published in: Journal of Experimental & Clinical Cancer Research / Issue 1/2021
Electronic ISSN: 1756-9966
DOI: https://doi.org/10.1186/s13046-021-01901-1

Webinar | 19-02-2024 | 17:30 (CET)

Keynote webinar | Spotlight on antibody–drug conjugates in cancer

Watch now

Antibody–drug conjugates (ADCs) are novel agents that have shown promise across multiple tumor types. Explore the current landscape of ADCs in breast and lung cancer with our experts, and gain insights into the mechanism of action, key clinical trials data, existing challenges, and future directions.

Dr. Véronique Diéras

Prof. Fabrice Barlesi

Developed by: Springer Medicine

Keynote webinar | Spotlight on medication adherence

Springer Medicine

Abstract

Background

Methods

Results

Conclusion

Please log in to get access to this content

Other articles of this Issue 1/2021

Retraction Note: Dual loading miR-218 mimics and Temozolomide using AuCOOH@FA-CS drug delivery system: promising targeted antitumor drug delivery system with sequential release functions

M6A associated TSUC7 inhibition contributed to Erlotinib resistance in lung adenocarcinoma through a notch signaling activation dependent way

Correction to: Embedding similarities between embryos and circulating tumor cells: fundamentals of abortifacients used for cancer metastasis chemoprevention

USP1-dependent RPS16 protein stability drives growth and metastasis of human hepatocellular carcinoma cells

Elesclomol-induced increase of mitochondrial reactive oxygen species impairs glioblastoma stem-like cell survival and tumor growth

Gut microbiota influence tumor development and Alter interactions with the human immune system

Keynote webinar | Spotlight on antibody–drug conjugates in cancer