title: Exosome derived from epigallocatechin gallate treated breast cancer cells suppresses tumor growth by inhibiting tumor-associated macrophage infiltration and M2 polarization.
abstract: BACKGROUND
  Tumor-associated macrophages (TAM) play an important role in tumor microenvironment. Particularly, M2 macrophages contribute to tumor progression, depending on the expression of NF-κB. Tumor-derived exosomes can modulate tumor microenvironment by transferring miRNAs to immune cells. Epigallocatechin gallate (EGCG) has well known anti-tumor effects; however, no data are available on the influence of EGCG on communication with cancer cells and TAM.
  
  
  METHODS
  Murine breast cancer cell lines, 4T1, was used for in vivo and ex vivo studies. Exosome was extracted from EGCG-treated 4T1 cells, and the change of miRNAs was screened using microarray. Tumor cells or TAM isolated from murine tumor graft were incubated with exosomes derived from EGCG-treated and/or miR-16 inhibitor-transfected 4T1 cells. Chemokines for monocytes (CSF-1 and CCL-2), cytokines both with high (IL-6 and TGF-β) and low (TNF-α) expression in M2 macrophages, and molecules in NF-κB pathway (IKKα and Iκ-B) were evaluated by RT-qPCR or western blot.
  
  
  RESULTS
  EGCG suppressed tumor growth in murine breast cancer model, which was associated with decreased TAM and M2 macrophage infiltration. Expression of chemokine for monocytes (CSF-1 and CCL-2) were low in tumor cells from EGCG-treated mice, and cytokines of TAM was skewed from M2- into M1-like phenotype by EGCG as evidenced by decreased IL-6 and TGF-β and increased TNF-α. Ex vivo incubation of isolated tumor cells with EGCG inhibited the CSF-1 and CCL-2 expression. Ex vivo incubation of TAM with exosomes from EGCG-treated 4T1 cells led to IKKα suppression and concomitant I-κB accumulation; increase of IL-6 and TGF-β; and, decrease of TNF-α. EGCG up-regulated miR-16 in 4T1 cells and in the exosomes. Treatment of tumor cells or TAM with exosomes derived from EGCG-treated and miR-16-knock-downed 4T1 cells restored the above effects on chemokines, cytokines, and NF-κB pathway elicited by EGCG-treated exosomes.
  
  
  CONCLUSIONS
  Our data demonstrate that EGCG up-regulates miR-16 in tumor cells, which can be transferred to TAM via exosomes and inhibits TAM infiltration and M2 polarization. We suggest a novel mechanism by which EGCG exerts anti-tumor activity via regulation of TAM in tumor microenvironment.
authors.affiliation: Tumor Immunity Medical Research Center, Cancer Research Institute, Seoul National University College of Medicine, 28 Yongon-dong, Jongno-gu, Seoul 110-799, Korea. cwkim@snu.ac.kr.
authors.forename: Ji-Young
authors.initials: JY
authors.lastname: Jang
authors.forename: Jong-Kuen
authors.initials: JK
authors.lastname: Lee
authors.forename: Yoon-Kyung
authors.initials: YK
authors.lastname: Jeon
authors.forename: Chul-Woo
authors.initials: CW
authors.lastname: Kim
mesh_terms: D000818:Animals; D016588:Anticarcinogenic Agents / Q000494:pharmacology; D001943:Breast Neoplasms / Q000235:genetics / Q000276:immunology* / Q000378:metabolism* / Q000473:pathology; D002392:Catechin / Q000031:analogs & derivatives / Q000494:pharmacology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D018932:Chemokine CCL2 / Q000235:genetics / Q000378:metabolism; D004195:Disease Models, Animal; D055354:Exosomes / Q000378:metabolism*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D051550:I-kappa B Kinase / Q000378:metabolism; D020672:I-kappa B Proteins / Q000378:metabolism; D016173:Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000378:metabolism; D008264:Macrophages / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism*; D051379:Mice; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism; D019899:Paracrine Communication / Q000187:drug effects
chemical_list: D016588:Anticarcinogenic Agents; D018932:Chemokine CCL2; D020672:I-kappa B Proteins; D035683:MicroRNAs; C522604:Mirn16 microRNA, mouse; D016173:Macrophage Colony-Stimulating Factor; D002392:Catechin; C045651:epigallocatechin gallate; D051550:I-kappa B Kinase
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