title: High-throughput screens identify microRNAs essential for HER2 positive breast cancer cell growth.
abstract: MicroRNAs (miRNAs) are non-coding RNAs regulating gene expression post-transcriptionally. We have characterized the role of miRNAs in regulating the human epidermal growth factor receptor 2 (HER2)-pathway in breast cancer. We performed miRNA gain-of-function assays by screening two HER2 amplified cell lines (KPL-4 and JIMT-1) with a miRNA mimic library consisting of 810 human miRNAs. The levels of HER2, phospho-AKT, phospho-ERK1/2, cell proliferation (Ki67) and apoptosis (cPARP) were analyzed with reverse-phase protein arrays. Rank product analyses identified 38 miRNAs (q < 0.05) as inhibitors of HER2 signaling and cell growth, the most effective being miR-491-5p, miR-634, miR-637 and miR-342-5p. We also characterized miRNAs directly targeting HER2 and identified seven novel miRNAs (miR-552, miR-541, miR-193a-5p, miR-453, miR-134, miR-498, and miR-331-3p) as direct regulators of the HER2 3'UTR. We demonstrated the clinical relevance of the miRNAs and identified miR-342-5p and miR-744* as significantly down-regulated in HER2-positive breast tumors as compared to HER2-negative tumors from two cohorts of breast cancer patients (101 and 1302 cases). miR-342-5p specifically inhibited HER2-positive cell growth, as it had no effect on the growth of HER2-negative control cells in vitro. Furthermore, higher expression of miR-342-5p was associated with better survival in both breast cancer patient cohorts. In conclusion, we have identified miRNAs which are efficient negative regulators of the HER2 pathway that may play a role in vivo during breast cancer progression. These results give mechanistic insights in HER2 regulation which may open potential new strategies towards prevention and therapeutic inhibition of HER2-positive breast cancer.
authors.affiliation: Department of Genetics, Institute for Cancer Research, Oslo University Hospital, The Norwegian Radiumhospital, N-0310 Oslo, Norway; The K.G. Jebsen Center for Breast Cancer Research, Institute for Clinical Medicine, Faculty of Medicine, University of Oslo, N-0424 Oslo, Norway; Medical Biotechnology, VTT Technical Research Centre of Finland, FI-20520 Turku, Finland. Electronic address: suvi-katri.leivonen@rr-research.no.
authors.forename: Suvi-Katri
authors.initials: SK
authors.lastname: Leivonen
authors.affiliation: Department of Genetics, Institute for Cancer Research, Oslo University Hospital, The Norwegian Radiumhospital, N-0310 Oslo, Norway; The K.G. Jebsen Center for Breast Cancer Research, Institute for Clinical Medicine, Faculty of Medicine, University of Oslo, N-0424 Oslo, Norway; Department of Research, Vestre Viken Hospital Trust, N-3004 Drammen, Norway.
authors.forename: Kristine Kleivi
authors.initials: KK
authors.lastname: Sahlberg
authors.affiliation: Medical Biotechnology, VTT Technical Research Centre of Finland, FI-20520 Turku, Finland.
authors.forename: Rami
authors.initials: R
authors.lastname: Mäkelä
authors.affiliation: Department of Genetics, Institute for Cancer Research, Oslo University Hospital, The Norwegian Radiumhospital, N-0310 Oslo, Norway; The K.G. Jebsen Center for Breast Cancer Research, Institute for Clinical Medicine, Faculty of Medicine, University of Oslo, N-0424 Oslo, Norway.
authors.forename: Eldri Undlien
authors.initials: EU
authors.lastname: Due
authors.affiliation: Institute for Molecular Medicine Finland, University of Helsinki, FI-00014 Helsinki, Finland.
authors.forename: Olli
authors.initials: O
authors.lastname: Kallioniemi
authors.affiliation: Department of Genetics, Institute for Cancer Research, Oslo University Hospital, The Norwegian Radiumhospital, N-0310 Oslo, Norway; The K.G. Jebsen Center for Breast Cancer Research, Institute for Clinical Medicine, Faculty of Medicine, University of Oslo, N-0424 Oslo, Norway.
authors.forename: Anne-Lise
authors.initials: AL
authors.lastname: Børresen-Dale
authors.affiliation: Medical Biotechnology, VTT Technical Research Centre of Finland, FI-20520 Turku, Finland.
authors.forename: Merja
authors.initials: M
authors.lastname: Perälä
mesh_terms: D001940:Breast / Q000378:metabolism / Q000473:pathology; D001943:Breast Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D040081:Protein Array Analysis; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D035683:MicroRNAs; D018719:Erb-b2 Receptor Tyrosine Kinases; C508053:ERBB2 protein, human
keywords: Breast cancer; EGFR; ER; Functional screen; HER2; MAPK; MicroRNA; RPPA; Reverse-phase protein array; UTR; cPARP; cleaved PARP; epidermal growth factor receptor; estrogen receptor; human epidermal growth factor receptor 2; miRNA; microRNA; mitogen-activated protein kinase; reverse-phase protein array; untranslated region