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High-Throughput Screening Approach for Identifying Compounds That Inhibit Nonhomologous End Joining.
Bredemeyer, Andrea L; Edwards, Bruce S; Haynes, Mark K; Morales, Abigail J; Wang, Yinan; Ursu, Oleg; Waller, Anna; Sklar, Larry A; Sleckman, Barry P.
Afiliación
  • Bredemeyer AL; 1 Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO, USA.
  • Edwards BS; 2 University of New Mexico Health Sciences Center and Cancer Research and Treatment Center, Department of Pathology, Cytometry, New Mexico Molecular Libraries Screening Center, Albuquerque, NM, USA.
  • Haynes MK; 2 University of New Mexico Health Sciences Center and Cancer Research and Treatment Center, Department of Pathology, Cytometry, New Mexico Molecular Libraries Screening Center, Albuquerque, NM, USA.
  • Morales AJ; 3 Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, NY, USA.
  • Wang Y; 3 Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, NY, USA.
  • Ursu O; 2 University of New Mexico Health Sciences Center and Cancer Research and Treatment Center, Department of Pathology, Cytometry, New Mexico Molecular Libraries Screening Center, Albuquerque, NM, USA.
  • Waller A; 2 University of New Mexico Health Sciences Center and Cancer Research and Treatment Center, Department of Pathology, Cytometry, New Mexico Molecular Libraries Screening Center, Albuquerque, NM, USA.
  • Sklar LA; 2 University of New Mexico Health Sciences Center and Cancer Research and Treatment Center, Department of Pathology, Cytometry, New Mexico Molecular Libraries Screening Center, Albuquerque, NM, USA.
  • Sleckman BP; 3 Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, NY, USA.
SLAS Discov ; 23(7): 624-633, 2018 08.
Article en En | MEDLINE | ID: mdl-29232168
DNA double-strand breaks (DSBs) are repaired primarily by homologous recombination (HR) or nonhomologous end joining (NHEJ). Compounds that modulate HR have shown promise as cancer therapeutics. The V(D)J recombination reaction, which assembles antigen receptor genes in lymphocytes, is initiated by the introduction of DNA DSBs at two recombining gene segments by the RAG endonuclease, followed by the NHEJ-mediated repair of these DSBs. Here, using HyperCyt automated flow cytometry, we develop a robust high-throughput screening (HTS) assay for NHEJ that utilizes engineered pre-B-cell lines where the V(D)J recombination reaction can be induced and monitored at a single-cell level. This approach, novel in processing four 384-well plates at a time in parallel, was used to screen the National Cancer Institute NeXT library to identify compounds that inhibit V(D)J recombination and NHEJ. Assessment of cell light scattering characteristics at the primary HTS stage (83,536 compounds) enabled elimination of 60% of apparent hits as false positives. Although all the active compounds that we identified had an inhibitory effect on RAG cleavage, we have established this as an approach that could identify compounds that inhibit RAG cleavage or NHEJ using new chemical libraries.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Descubrimiento de Drogas / Ensayos Analíticos de Alto Rendimiento / Reparación del ADN por Unión de Extremidades Tipo de estudio: Diagnostic_studies / Prognostic_studies / Screening_studies Límite: Humans Idioma: En Revista: SLAS Discov Año: 2018 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Descubrimiento de Drogas / Ensayos Analíticos de Alto Rendimiento / Reparación del ADN por Unión de Extremidades Tipo de estudio: Diagnostic_studies / Prognostic_studies / Screening_studies Límite: Humans Idioma: En Revista: SLAS Discov Año: 2018 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos