title: Drug discovery for human African trypanosomiasis: identification of novel scaffolds by the newly developed HTS SYBR Green assay for Trypanosoma brucei.
abstract: Human African trypanosomiasis (HAT) is a vector-transmitted tropical disease caused by the protozoan parasite Trypanosoma brucei. High-throughput screening (HTS) of small-molecule libraries in whole-cell assays is one of the most frequently used approaches in drug discovery for infectious diseases. To aid in drug discovery efforts for HAT, the SYBR Green assay was developed for T. brucei in a 384-well format. This semi-automated assay is cost- and time-effective, robust, and reproducible. The SYBR Green assay was compared to the resazurin assay by screening a library of 4000 putative kinase inhibitors, revealing a superior performance in terms of assay time, sensitivity, simplicity, and reproducibility, and resulting in a higher hit confirmation rate. Although the resazurin assay allows for comparatively improved detection of slow-killing compounds, it also has higher false-positive rates that are likely to arise from the assay experimental conditions. The compounds with the most potent antitrypanosomal activity were selected in both screens and grouped into 13 structural clusters, with 11 new scaffolds as antitrypanosomal agents. Several of the identified compounds had IC50 <1 µM coupled with high selectivity toward the parasite. The core structures of the scaffolds are shown, providing promising new starting points for drug discovery for HAT.
authors.affiliation: Center for Neglected Diseases Drug Discovery (CND3), Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, South Korea Parasite Disease Group, Instituto de Biologia Molecular e Celular, Universidade do Porto, Porto, Portugal.
authors.forename: Joana
authors.initials: J
authors.lastname: Faria
authors.affiliation: Center for Neglected Diseases Drug Discovery (CND3), Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, South Korea Laboratório Nacional de Biociências (LNBio), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Campinas-SP, Brazil.
authors.forename: Carolina B
authors.initials: CB
authors.lastname: Moraes
authors.affiliation: MedChem & Chemical Biology Group, Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, South Korea.
authors.forename: Rita
authors.initials: R
authors.lastname: Song
authors.affiliation: Laboratório Nacional de Biociências (LNBio), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Campinas-SP, Brazil.
authors.forename: Bruno S
authors.initials: BS
authors.lastname: Pascoalino
authors.affiliation: Center for Neglected Diseases Drug Discovery (CND3), Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, South Korea.
authors.forename: Nakyung
authors.initials: N
authors.lastname: Lee
authors.affiliation: Center for Neglected Diseases Drug Discovery (CND3), Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, South Korea Skaggs School of Pharmacy and Pharmaceutical Science, University of California San Diego, La Jolla, CA, USA.
authors.forename: Jair L
authors.initials: JL
authors.lastname: Siqueira-Neto
authors.affiliation: Center for Neglected Diseases Drug Discovery (CND3), Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, South Korea.
authors.forename: Deu John M
authors.initials: DJ
authors.lastname: Cruz
authors.affiliation: Drugs for Neglected Diseases initiative (DNDi), Geneva, Switzerland.
authors.forename: Tanya
authors.initials: T
authors.lastname: Parkinson
authors.affiliation: Drugs for Neglected Diseases initiative (DNDi), Geneva, Switzerland.
authors.forename: Jean-Robert
authors.initials: JR
authors.lastname: Ioset
authors.affiliation: Parasite Disease Group, Instituto de Biologia Molecular e Celular, Universidade do Porto, Porto, Portugal Departamento de Ciências Biológicas, Faculdade de Farmácia, Universidade do Porto, Porto, Portugal.
authors.forename: Anabela
authors.initials: A
authors.lastname: Cordeiro-da-Silva
authors.affiliation: Center for Neglected Diseases Drug Discovery (CND3), Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, South Korea Laboratório Nacional de Biociências (LNBio), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Campinas-SP, Brazil lucio.freitasjunior@lnbio.cnpem.br.
authors.forename: Lucio H
authors.initials: LH
authors.lastname: Freitas-Junior
mesh_terms: D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D055808:Drug Discovery / Q000379:methods*; D005456:Fluorescent Dyes; D005470:Fluorometry / Q000379:methods; D057166:High-Throughput Screening Assays; D006801:Humans; D058069:Neglected Diseases / Q000188:drug therapy*; D010078:Oxazines; D054852:Small Molecule Libraries; D014344:Trypanocidal Agents / Q000494:pharmacology* / Q000627:therapeutic use; D014346:Trypanosoma brucei brucei / Q000187:drug effects*; D014353:Trypanosomiasis, African / Q000188:drug therapy*; D014966:Xanthenes
chemical_list: D005456:Fluorescent Dyes; D010078:Oxazines; D054852:Small Molecule Libraries; D014344:Trypanocidal Agents; D014966:Xanthenes; C005843:resazurin
keywords: anti-infective drugs; cell-based assays; fluorescence methods; ultrahigh-throughput screening