title: Robust lanthanide-based assays for the detection of anti-apoptotic Bcl-2-family protein antagonists.
abstract: Anti-apoptotic Bcl-2-family proteins (Bcl-2, Bcl-x(L), Bfl-1, Mcl-1, Bcl-W and Bcl-B) have been recently validated as drug discovery targets for cancer, owed to their ability to confer tumor resistance to chemotherapy or radiation. The anti-apoptotic activity of Bcl-2 proteins is due to their ability to heterodimerize with their pro-apoptotic counterparts (proteins such as Bad, Bim or Bid) via a conserved peptide region termed BH3. Thus, molecules that mimic pro-apoptotic BH3 domains represent a direct approach to overcoming the protective effects of anti-apoptotic proteins such as Bcl-2 and Bcl-x(L). Here, we report on the development and evaluation of two novel Lanthanide-based assays that are formatted for high-throughput screening of small molecules capable of antagonizing BH3-Bcl-2 interactions. The assay conditions, robustness and reproducibility (Z' factors) are described. These assays represent useful tools to enable further studies in the search for novel, safe and effective anti-cancer agents targeting Bcl-2-family proteins.
authors.affiliation: Burnham Institute for Medical Research, 10901 North Torrey Pines Rd., La Jolla, CA 92037, USA.
authors.forename: Michele F
authors.initials: MF
authors.lastname: Rega
authors.forename: John C
authors.initials: JC
authors.lastname: Reed
authors.forename: Maurizio
authors.initials: M
authors.lastname: Pellecchia
mesh_terms: D000595:Amino Acid Sequence; D000970:Antineoplastic Agents / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D051023:BH3 Interacting Domain Death Agonist Protein / Q000037:antagonists & inhibitors*; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D031541:Fluorescence Resonance Energy Transfer; D015200:Fluoroimmunoassay; D006801:Humans; D028581:Lanthanoid Series Elements / Q000737:chemistry*; D008969:Molecular Sequence Data; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors*; D015203:Reproducibility of Results
chemical_list: D000970:Antineoplastic Agents; D051023:BH3 Interacting Domain Death Agonist Protein; D028581:Lanthanoid Series Elements; D019253:Proto-Oncogene Proteins c-bcl-2