title: miR-199a-5p confers tumor-suppressive role in triple-negative breast cancer.
abstract: Triple-negative breast cancer (TNBC) remains a poor prognostic factor for breast cancer since no effective targeted therapy is readily available. Our previous studies confirmed miR-199a-5p is a TNBC-specific circulating biomarker, however, its functional roles in breast cancer is largely unknown. Thus, we investigated the functional implication of miR-199a-5p in TNBC and its potential underlying mechanisms.
  
  
  
  MTT assay was performed to investigate the cell proliferation after transient transfection of miR-199a-5p in MDA-MB-231 cell line, followed by cell cycle analysis. Transwell invasion assay and wound healing assay were used to study the invasion and migration ability respectively. To further investigate the stemness-related characteristics of miR-199a-5p in breast cancer cells, single-cell clonogenic assay and aldehyde dehydrogenase (ALDH) assay were performed. 32 normal and 100 breast cancer patients' plasma were recruited to identify the potential circulating markers by qPCR.
  
  
  
  Cell proliferation assay revealed significant inhibition after miR-199a-5p ectopic expression (p < 0.0001), as a result of decreased S phase (p = 0.0284), increased G0/G1 phase (p = 0.0260) and apoptosis (p = 0.0374). Invasiveness (p = 0.0005) and wound healing ability were also decreased upon miR-199a-5p overexpression. It significantly altered EMT-related genes expression, namely CDH1, ZEB1 and TWIST. Single-cell clonogenic assay showed decreased colonies in miR-199a-5p (p = 0.0182). Significant downregulation (p = 0.0088) and inhibited activity (p = 0.0390) of ALDH was observed in miR-199a-5p. ALDH1A3, which is the dominant isoform of ALDH, is significantly upregulated in breast cancer plasma especially in TNBC (p = 0.0248). PIK3CD was identified as a potential downstream target of miR-199a-5p.
  
  
  
  Taken together, we unraveled, for the first time, the tumor-suppressive role of miR-199a-5p in TNBC, which attributed to EMT and cancer stemness properties, providing a novel therapeutic options towards this aggressive disease.
authors.affiliation: Breast Surgery Division, Department of Surgery, The University of Hong Kong, Hong Kong, SAR, China.
authors.forename: Jiawei
authors.initials: J
authors.lastname: Chen
authors.affiliation: Breast Surgery Division, Department of Surgery, The University of Hong Kong, Hong Kong, SAR, China.
authors.forename: Vivian Y
authors.initials: VY
authors.lastname: Shin
authors.affiliation: Breast Surgery Division, Department of Surgery, The University of Hong Kong, Hong Kong, SAR, China.
authors.forename: Man T
authors.initials: MT
authors.lastname: Siu
authors.affiliation: Breast Surgery Division, Department of Surgery, The University of Hong Kong, Hong Kong, SAR, China.
authors.forename: John C W
authors.initials: JC
authors.lastname: Ho
authors.affiliation: Breast Surgery Division, Department of Surgery, The University of Hong Kong, Hong Kong, SAR, China.
authors.forename: Isabella
authors.initials: I
authors.lastname: Cheuk
authors.affiliation: Breast Surgery Division, Department of Surgery, The University of Hong Kong, Hong Kong, SAR, China. akwong@asiabreastregistry.com.|Hong Kong Hereditary Breast Cancer Family Registry, Queen Mary Hospital, Room K1401, Pokfulam Road, Pok Fu Lam, Hong Kong. akwong@asiabreastregistry.com.|Department of Surgery, Hong Kong Sanatorium and Hospital, Hong Kong, SAR, China. akwong@asiabreastregistry.com.
authors.forename: Ava
authors.initials: A
authors.lastname: Kwong
mesh_terms: D000328:Adult; D000368:Aged; D000818:Animals; D017209:Apoptosis / Q000235:genetics; D014408:Biomarkers, Tumor; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics; D049109:Cell Proliferation; D058534:Class I Phosphatidylinositol 3-Kinases / Q000235:genetics; D004195:Disease Models, Animal; D005260:Female; D059585:G1 Phase Cell Cycle Checkpoints / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic; D016147:Genes, Tumor Suppressor; D064593:Heterografts; D006801:Humans; D051379:Mice; D035683:MicroRNAs / Q000235:genetics*; D008875:Middle Aged; D060787:Neoplasm Grading; D009362:Neoplasm Metastasis; D009367:Neoplasm Staging; D014411:Neoplastic Stem Cells / Q000378:metabolism; D034622:RNA Interference; D053781:Transforming Growth Factor beta2 / Q000235:genetics; D064726:Triple Negative Breast Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology
chemical_list: D014408:Biomarkers, Tumor; D035683:MicroRNAs; D053781:Transforming Growth Factor beta2; C531408:mirn199 microRNA, human; D058534:Class I Phosphatidylinositol 3-Kinases; C486018:PIK3CD protein, human
keywords: Triple negative breast cancer (TNBC); Tumor-suppressor; miR-199a-5p