title: Development and validation of high-throughput screening assays for poly(ADP-ribose) polymerase-2 inhibitors.
abstract: Poly(ADP-ribose) polymerase-1 and -2 (PARP1/2) are two key facilitators of DNA repair and are implicated in the pathogenesis of cancers and several chronic diseases. Inhibitors of PARP1/2 have shown powerful therapeutic effects in the treatment of cancer, cerebral ischemia, and inflammation. In addition, evidence from several studies suggests unique functions for PARP2 in genome surveillance, spermatogenesis, adipogenesis, and T cell development, and PARP2-specific inhibitors might have many other applications. To acquire PARP1/2 inhibitors, many high-throughput screening (HTS) assays for PARP1 inhibitors have been developed. However, detailed screening assays for PARP2 inhibitors have not been reported. Herein, three HTS assays for PARP2 inhibitors were developed and validated with reference inhibitors in each case. The results suggest that the HTS assays for PARP2 inhibitors using chemical quantification of NAD(+), biotin-based quantification of PAR, and ELISA quantification of PAR are sensitive, robust, and cost effective.
authors.affiliation: Department of Pharmacology, State Key Laboratory of Bioactive Substances and Functions of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, People's Republic of China.
authors.forename: Zhixiang
authors.initials: Z
authors.lastname: Zhu
authors.affiliation: Department of Pharmacology, State Key Laboratory of Bioactive Substances and Functions of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, People's Republic of China.
authors.forename: Jing
authors.initials: J
authors.lastname: Jin
authors.affiliation: Department of Pharmacology, State Key Laboratory of Bioactive Substances and Functions of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, People's Republic of China.
authors.forename: Nina
authors.initials: N
authors.lastname: Xue
authors.affiliation: Department of Pharmacology, State Key Laboratory of Bioactive Substances and Functions of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, People's Republic of China.
authors.forename: Xiuyun
authors.initials: X
authors.lastname: Song
authors.affiliation: Department of Pharmacology, State Key Laboratory of Bioactive Substances and Functions of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, People's Republic of China. Electronic address: chxg@imm.ac.cn.
authors.forename: Xiaoguang
authors.initials: X
authors.lastname: Chen
mesh_terms: D019921:Biotinylation; D004353:Drug Evaluation, Preclinical / Q000379:methods; D057075:Enzyme Assays / Q000379:methods*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004797:Enzyme-Linked Immunosorbent Assay; D006367:HeLa Cells; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D009243:NAD / Q000737:chemistry / Q000378:metabolism; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D009243:NAD; C483041:PARP2 protein, human; D011065:Poly(ADP-ribose) Polymerases
keywords: Biotinylated NAD(+); Chemical quantification; ELISA; High-throughput screening; Inhibitor; PARP2