title: A chemical screen identifies small molecules that regulate hepcidin expression.
abstract: Hepcidin, a peptide hormone produced in the liver, decreases intestinal iron absorption and macrophage iron release via effects on ferroportin. Bone morphogenic protein and Stat3 signaling regulate Hepcidin's transcription. Hepcidin is a potential drug target for patients with iron overload syndromes because its levels are inappropriately low in these individuals. To generate a tool for identifying small molecules that modulate Hepcidin expression, we stably transfected human hepatocytes (HepG2) cells with a reporter construct containing 2.7kb of the human Hepcidin promoter upstream of a firefly reporter gene. We used high throughput methods to screen 10,169 chemicals in duplicate for their effect on Hepcidin expression and cell viability. Regulators were identified as chemicals that caused a change >3 standard deviations above or >1 standard deviation below the mean of the other chemicals (z-score >3 or <1), while not adversely affecting cell viability, quantified by fluorescence assay. Following validation assays, we identified 16 chemicals in a broad range of functional classes that promote Hepcidin expression. All of the chemicals identified increased expression of bone morphogenic protein-dependent and/or Stat3-dependent genes, however none of them strongly increased phosphorylation of Smad1,5,8 or Stat3.
authors.affiliation: Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, 330 Brookline Avenue, Boston, MA 02215, United States; Department of Medicine, Harvard Medical School, 330 Brookline Avenue, Boston, MA 02215, United States.
authors.forename: Vera
authors.initials: V
authors.lastname: Gaun
authors.affiliation: Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, 330 Brookline Avenue, Boston, MA 02215, United States; Department of Medicine, Harvard Medical School, 330 Brookline Avenue, Boston, MA 02215, United States.
authors.forename: Bonnie
authors.initials: B
authors.lastname: Patchen
authors.affiliation: Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, 330 Brookline Avenue, Boston, MA 02215, United States; Department of Medicine, Harvard Medical School, 330 Brookline Avenue, Boston, MA 02215, United States.
authors.forename: Josephine
authors.initials: J
authors.lastname: Volovetz
authors.affiliation: Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, 330 Brookline Avenue, Boston, MA 02215, United States; Department of Medicine, Harvard Medical School, 330 Brookline Avenue, Boston, MA 02215, United States.
authors.forename: Aileen W
authors.initials: AW
authors.lastname: Zhen
authors.affiliation: Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, 330 Brookline Avenue, Boston, MA 02215, United States; Department of Medicine, Harvard Medical School, 330 Brookline Avenue, Boston, MA 02215, United States.
authors.forename: Aleksandr
authors.initials: A
authors.lastname: Andreev
authors.affiliation: Department of Chemistry and Chemical Biology, Northeastern University, 417 Egan Research Center, 120 Forsyth Street, Boston, MA 02115, United States.
authors.forename: Michael P
authors.initials: MP
authors.lastname: Pollastri
authors.affiliation: Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, 330 Brookline Avenue, Boston, MA 02215, United States; Department of Medicine, Harvard Medical School, 330 Brookline Avenue, Boston, MA 02215, United States. Electronic address: pfraenke@bidmc.harvard.edu.
authors.forename: Paula G
authors.initials: PG
authors.lastname: Fraenkel
mesh_terms: D019485:Bone Morphogenetic Proteins / Q000235:genetics / Q000378:metabolism; D002470:Cell Survival; D062126:Databases, Chemical; D055808:Drug Discovery; D005786:Gene Expression Regulation / Q000187:drug effects*; D017930:Genes, Reporter; D056945:Hep G2 Cells; D064451:Hepcidins / Q000819:agonists / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D057166:High-Throughput Screening Assays; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000737:chemistry / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D051785:Smad Proteins / Q000235:genetics / Q000378:metabolism; D054852:Small Molecule Libraries / Q000737:chemistry / Q000494:pharmacology*; D014162:Transfection
chemical_list: D019485:Bone Morphogenetic Proteins; C578702:HAMP protein, human; D064451:Hepcidins; D011993:Recombinant Fusion Proteins; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; D051785:Smad Proteins; D054852:Small Molecule Libraries; D008156:Luciferases
keywords: Bone morphogenic protein; Hemochromatosis; Interleukin-6; Stat3; Thalassemia