title: Buformin inhibits the stemness of erbB-2-overexpressing breast cancer cells and premalignant mammary tissues of MMTV-erbB-2 transgenic mice.
abstract: Metformin, an FDA-approved drug for the treatment of Type II diabetes, has emerged as a promising anti-cancer agent. Other biguanide analogs, including buformin and phenformin, are suggested to have similar properties. Although buformin was shown to reduce mammary tumor burden in carcinogen models, the anti-cancer effects of buformin on different breast cancer subtypes and the underlying mechanisms remain unclear. Therefore, we aimed to investigate the effects of buformin on erbB-2-overexpressing breast cancer with in vitro and in vivo models.
  
  
  
  MTT, cell cycle, clonogenic/CFC, ALDEFLUOR, tumorsphere, and Western blot analyses were used to determine the effects of buformin on cell growth, stem cell populations, stem cell-like properties, and signaling pathways in SKBR3 and BT474 erbB-2-overexpressing breast cancer cell lines. A syngeneic tumor cell transplantation model inoculating MMTV-erbB-2 mice with 78617 mouse mammary tumor cells was used to study the effects of buformin (1.2 g buformin/kg chow) on tumor growth in vivo. MMTV-erbB-2 mice were also fed buformin for 10 weeks, followed by analysis of premalignant mammary tissues for changes in morphological development, mammary epithelial cell (MEC) populations, and signaling pathways.
  
  
  
  Buformin significantly inhibited SKBR3 and BT474 cell growth, and in vivo activity was demonstrated by considerable growth inhibition of syngeneic tumors derived from MMTV-erbB-2 mice. In particular, buformin suppressed stem cell populations and self-renewal in vitro, which was associated with inhibited receptor tyrosine kinase (RTK) and mTOR signaling. Consistent with in vitro data, buformin suppressed mammary morphogenesis and reduced cell proliferation in MMTV-erbB-2 mice. Importantly, buformin decreased MEC populations enriched with mammary reconstitution units (MRUs) and tumor-initiating cells (TICs) from MMTV-erbB-2 mice, as supported by impaired clonogenic and mammosphere formation in primary MECs. We further demonstrated that buformin-mediated in vivo inhibition of MEC stemness is associated with suppressed activation of mTOR, RTK, ER, and β-catenin signaling pathways.
  
  
  
  Overall, our results provide evidence for buformin as an effective anti-cancer drug that selectively targets TICs, and present a novel prevention and/or treatment strategy for patients who are genetically predisposed to erbB-2-overexpressing breast cancer.
authors.affiliation: Julius L. Chambers Biomedical/Biotechnology Research Institute, Department of Biological and Biomedical Sciences, North Carolina Central University, 500 Laureate Way, NRI 4301, Kannapolis, North Carolina, 28081, USA.
authors.forename: Amanda B
authors.initials: AB
authors.lastname: Parris
authors.affiliation: Julius L. Chambers Biomedical/Biotechnology Research Institute, Department of Biological and Biomedical Sciences, North Carolina Central University, 500 Laureate Way, NRI 4301, Kannapolis, North Carolina, 28081, USA.
authors.forename: Qingxia
authors.initials: Q
authors.lastname: Zhao
authors.affiliation: Julius L. Chambers Biomedical/Biotechnology Research Institute, Department of Biological and Biomedical Sciences, North Carolina Central University, 500 Laureate Way, NRI 4301, Kannapolis, North Carolina, 28081, USA.
authors.forename: Erin W
authors.initials: EW
authors.lastname: Howard
authors.affiliation: Julius L. Chambers Biomedical/Biotechnology Research Institute, Department of Biological and Biomedical Sciences, North Carolina Central University, 500 Laureate Way, NRI 4301, Kannapolis, North Carolina, 28081, USA.
authors.forename: Ming
authors.initials: M
authors.lastname: Zhao
authors.affiliation: Julius L. Chambers Biomedical/Biotechnology Research Institute, Department of Biological and Biomedical Sciences, North Carolina Central University, 500 Laureate Way, NRI 4301, Kannapolis, North Carolina, 28081, USA.|College of Medicine, Henan University of Sciences and Technology, Luoyang, China.
authors.forename: Zhikun
authors.initials: Z
authors.lastname: Ma
authors.affiliation: Julius L. Chambers Biomedical/Biotechnology Research Institute, Department of Biological and Biomedical Sciences, North Carolina Central University, 500 Laureate Way, NRI 4301, Kannapolis, North Carolina, 28081, USA. xyang@nccu.edu.|College of Medicine, Henan University of Sciences and Technology, Luoyang, China. xyang@nccu.edu.
authors.forename: Xiaohe
authors.initials: X
authors.lastname: Yang
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage* / Q000494:pharmacology; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000378:metabolism; D002026:Buformin / Q000008:administration & dosage* / Q000494:pharmacology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D058492:Drug Repositioning; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D014411:Neoplastic Stem Cells / Q000187:drug effects*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects; D023041:Xenograft Model Antitumor Assays
chemical_list: D000970:Antineoplastic Agents; D002026:Buformin; D011505:Protein-Tyrosine Kinases; D018719:Erb-b2 Receptor Tyrosine Kinases; D058570:TOR Serine-Threonine Kinases; C546842:MTOR protein, human; C508053:ERBB2 protein, human
keywords: Breast cancer; Buformin; Cancer stem cells; ErbB-2; Mammary epithelial cells