title: A cell-based high-throughput screen for novel chemical inducers of fetal hemoglobin for treatment of hemoglobinopathies.
abstract: Decades of research have established that the most effective treatment for sickle cell disease (SCD) is increased fetal hemoglobin (HbF). Identification of a drug specific for inducing γ-globin expression in pediatric and adult patients, with minimal off-target effects, continues to be an elusive goal. One hurdle has been an assay amenable to a high-throughput screen (HTS) of chemicals that displays a robust γ-globin off-on switch to identify potential lead compounds. Assay systems developed in our labs to understand the mechanisms underlying the γ- to β-globin gene expression switch during development has allowed us to generate a cell-based assay that was adapted for a HTS of 121,035 compounds. Using chemical inducer of dimerization (CID)-dependent bone marrow cells (BMCs) derived from human γ-globin promoter-firefly luciferase β-globin promoter-Renilla luciferase β-globin yeast artificial chromosome (γ-luc β-luc β-YAC) transgenic mice, we were able to identify 232 lead chemical compounds that induced γ-globin 2-fold or higher, with minimal or no β-globin induction, minimal cytotoxicity and that did not directly influence the luciferase enzyme. Secondary assays in CID-dependent wild-type β-YAC BMCs and human primary erythroid progenitor cells confirmed the induction profiles of seven of the 232 hits that were cherry-picked for further analysis.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America; Department of Anatomy and Cell Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Kenneth R
authors.initials: KR
authors.lastname: Peterson
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Flávia C
authors.initials: FC
authors.lastname: Costa
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Halyna
authors.initials: H
authors.lastname: Fedosyuk
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Renee Y
authors.initials: RY
authors.lastname: Neades
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Allen M
authors.initials: AM
authors.lastname: Chazelle
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Lesya
authors.initials: L
authors.lastname: Zelenchuk
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Andrea H
authors.initials: AH
authors.lastname: Fonteles
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Parmita
authors.initials: P
authors.lastname: Dalal
authors.affiliation: High Throughput Screening Laboratory, University of Kansas, Lawrence, Kansas, United States of America.
authors.forename: Anuradha
authors.initials: A
authors.lastname: Roy
authors.affiliation: High Throughput Screening Laboratory, University of Kansas, Lawrence, Kansas, United States of America.
authors.forename: Rathnam
authors.initials: R
authors.lastname: Chaguturu
authors.affiliation: Department of Pediatrics, Georgia Regents University, Augusta, Georgia, United States of America.
authors.forename: Biaoru
authors.initials: B
authors.lastname: Li
authors.affiliation: Department of Pediatrics, Georgia Regents University, Augusta, Georgia, United States of America; Department of Molecular and Cell Biology, Georgia Regents University, Augusta, Georgia, United States of America.
authors.forename: Betty S
authors.initials: BS
authors.lastname: Pace
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D001854:Bone Marrow Cells / Q000187:drug effects* / Q000378:metabolism*; D018244:Chromosomes, Artificial, Yeast; D055808:Drug Discovery; D004353:Drug Evaluation, Preclinical; D015672:Erythroid Precursor Cells / Q000187:drug effects / Q000378:metabolism; D005319:Fetal Hemoglobin / Q000096:biosynthesis / Q000235:genetics*; D005786:Gene Expression Regulation / Q000187:drug effects*; D018390:Gene Targeting; D017930:Genes, Reporter; D056426:Genetic Loci; D005822:Genetic Vectors / Q000235:genetics; D006453:Hemoglobinopathies / Q000188:drug therapy / Q000235:genetics; D057166:High-Throughput Screening Assays; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D055544:beta-Globins / Q000096:biosynthesis / Q000235:genetics; D055545:gamma-Globins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D018952:Antigens, CD34; D055544:beta-Globins; D055545:gamma-Globins; D005319:Fetal Hemoglobin