title: Small-molecule inhibitors identify the RAD52-ssDNA interaction as critical for recovery from replication stress and for survival of BRCA2 deficient cells.
abstract: The DNA repair protein RAD52 is an emerging therapeutic target of high importance for BRCA-deficient tumors. Depletion of RAD52 is synthetically lethal with defects in tumor suppressors BRCA1, BRCA2 and PALB2. RAD52 also participates in the recovery of the stalled replication forks. Anticipating that ssDNA binding activity underlies the RAD52 cellular functions, we carried out a high throughput screening campaign to identify compounds that disrupt the RAD52-ssDNA interaction. Lead compounds were confirmed as RAD52 inhibitors in biochemical assays. Computational analysis predicted that these inhibitors bind within the ssDNA-binding groove of the RAD52 oligomeric ring. The nature of the inhibitor-RAD52 complex was validated through an in silico screening campaign, culminating in the discovery of an additional RAD52 inhibitor. Cellular studies with our inhibitors showed that the RAD52-ssDNA interaction enables its function at stalled replication forks, and that the inhibition of RAD52-ssDNA binding acts additively with BRCA2 or MUS81 depletion in cell killing.
authors.affiliation: Department of Biochemistry, University of Iowa, Iowa City, United States.
authors.forename: Sarah R
authors.initials: SR
authors.lastname: Hengel
authors.affiliation: Department of Environment and Health, Section of Experimental and Computational Carcinogenesis, Istituto Superiore di Sanita, Rome, Italy.
authors.forename: Eva
authors.initials: E
authors.lastname: Malacaria
authors.affiliation: Division of Medicinal and Natural Products Chemistry, Department of Pharmaceutical Sciences and Experimental Therapeutics, University of Iowa, Iowa City, United States.
authors.forename: Laura
authors.initials: L
authors.lastname: Folly da Silva Constantino
authors.affiliation: Department of Biochemistry, University of Iowa, Iowa City, United States.
authors.forename: Fletcher E
authors.initials: FE
authors.lastname: Bain
authors.affiliation: Department of Biochemistry, University of Iowa, Iowa City, United States.
authors.forename: Andrea
authors.initials: A
authors.lastname: Diaz
authors.affiliation: Department of Biochemistry, University of Iowa, Iowa City, United States.
authors.forename: Brandon G
authors.initials: BG
authors.lastname: Koch
authors.affiliation: Department of Biochemistry, University of Iowa, Iowa City, United States.|NMR Core Facility, Carver College of Medicine, University of Iowa, Iowa City, United States.
authors.forename: Liping
authors.initials: L
authors.lastname: Yu
authors.affiliation: Department of Biochemistry, University of Iowa, Iowa City, United States.|Division of Medicinal and Natural Products Chemistry, Department of Pharmaceutical Sciences and Experimental Therapeutics, University of Iowa, Iowa City, United States.|High Throughput Screening Facility, University of Iowa, Iowa City, United States.
authors.forename: Meng
authors.initials: M
authors.lastname: Wu
authors.affiliation: Department of Environment and Health, Section of Experimental and Computational Carcinogenesis, Istituto Superiore di Sanita, Rome, Italy.
authors.forename: Pietro
authors.initials: P
authors.lastname: Pichierri
authors.affiliation: Department of Biochemistry, University of Iowa, Iowa City, United States.|Division of Medicinal and Natural Products Chemistry, Department of Pharmaceutical Sciences and Experimental Therapeutics, University of Iowa, Iowa City, United States.
authors.forename: M Ashley
authors.initials: MA
authors.lastname: Spies
authors.affiliation: Department of Biochemistry, University of Iowa, Iowa City, United States.
authors.forename: Maria
authors.identifier: 0000-0002-7375-8037
authors.initials: M
authors.lastname: Spies
mesh_terms: D024682:BRCA2 Protein / Q000172:deficiency*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D005347:Fibroblasts / Q000187:drug effects / Q000502:physiology; D006801:Humans; D011485:Protein Binding / Q000187:drug effects; D051721:Rad52 DNA Repair and Recombination Protein / Q000378:metabolism*
chemical_list: D024682:BRCA2 Protein; C551750:BRCA2 protein, human; D004277:DNA, Single-Stranded; D004791:Enzyme Inhibitors; D051721:Rad52 DNA Repair and Recombination Protein
keywords: BRCA2; DNA repair; MUS81; RAD52; biochemistry; chromosomes; genes; high throughput screening; human; small-molecule inhibitors