title: Chk1 inhibition in p53-deficient cell lines drives rapid chromosome fragmentation followed by caspase-independent cell death.
abstract: Activation of Checkpoint kinase 1 (Chk1) following DNA damage mediates cell cycle arrest to prevent cells with damaged DNA from entering mitosis. Here we provide a high-resolution analysis of cells as they undergo S- and G₂-checkpoint bypass in response to Chk1 inhibition with the selective Chk1 inhibitor GNE-783. Within 4-8 h of Chk1 inhibition following gemcitabine induced DNA damage, cells with both sub-4N and 4N DNA content prematurely enter mitosis. Coincident with premature transition into mitosis, levels of DNA damage dramatically increase and chromosomes condense and attempt to align along the metaphase plate. Despite an attempt to congress at the metaphase plate, chromosomes rapidly fragment and lose connection to the spindle microtubules. Gemcitabine mediated DNA damage promotes the formation of Rad51 foci; however, while Chk1 inhibition does not disrupt Rad51 foci that are formed in response to gemcitabine, these foci are lost as cells progress into mitosis. Premature entry into mitosis requires the Aurora, Cdk1/2 and Plk1 kinases and even though caspase-2 and -3 are activated upon mitotic exit, they are not required for cell death. Interestingly, p53, but not p21, deficiency enables checkpoint bypass and chemo-potentiation. Finally, we uncover a differential role for the Wee-1 checkpoint kinase in response to DNA damage, as Wee-1, but not Chk1, plays a more prominent role in the maintenance of S- and G₂-checkpoints in p53 proficient cells.
authors.affiliation: Discovery Oncology; Genentech; San Francisco, CA.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Del Nagro
authors.affiliation: Discovery Oncology; Genentech; San Francisco, CA.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Choi
authors.affiliation: Discovery Oncology; Genentech; San Francisco, CA.
authors.forename: Yang
authors.initials: Y
authors.lastname: Xiao
authors.affiliation: Department of Pathology; Genentech; San Francisco, CA.
authors.forename: Linda
authors.initials: L
authors.lastname: Rangell
authors.affiliation: Department of Research Diagnostics; Genentech; San Francisco, CA.
authors.forename: Sankar
authors.initials: S
authors.lastname: Mohan
authors.affiliation: Department of Research Diagnostics; Genentech; San Francisco, CA.
authors.forename: Ajay
authors.initials: A
authors.lastname: Pandita
authors.affiliation: Department of Pathology; Genentech; San Francisco, CA.
authors.forename: Jiping
authors.initials: J
authors.lastname: Zha
authors.affiliation: Discovery Oncology; Genentech; San Francisco, CA.
authors.forename: Peter K
authors.initials: PK
authors.lastname: Jackson
authors.affiliation: Discovery Oncology; Genentech; San Francisco, CA.
authors.forename: Thomas
authors.initials: T
authors.lastname: O'Brien
mesh_terms: D002243:Carbolines / Q000494:pharmacology; D020169:Caspases / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D000071877:Checkpoint Kinase 1; D002877:Chromosomes, Human / Q000235:genetics*; D053938:DNA Fragmentation / Q000187:drug effects*; D003841:Deoxycytidine / Q000031:analogs & derivatives / Q000494:pharmacology; D004789:Enzyme Activation; D059565:G2 Phase Cell Cycle Checkpoints / Q000187:drug effects; D006801:Humans; D008938:Mitosis / Q000187:drug effects; D009687:Nuclear Proteins / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011494:Protein Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D051135:Rad51 Recombinase / Q000378:metabolism; D059807:S Phase Cell Cycle Checkpoints / Q000187:drug effects; D016159:Tumor Suppressor Protein p53 / Q000235:genetics* / Q000378:metabolism; D000093542:Gemcitabine
chemical_list: D002243:Carbolines; D018797:Cell Cycle Proteins; C000589252:GNE-783; D009687:Nuclear Proteins; D047428:Protein Kinase Inhibitors; D016159:Tumor Suppressor Protein p53; D003841:Deoxycytidine; D011494:Protein Kinases; D011505:Protein-Tyrosine Kinases; C499349:WEE1 protein, human; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D051135:Rad51 Recombinase; D020169:Caspases; D000093542:Gemcitabine
keywords: Chk1; GNE-783; checkpoint-bypass; chemo-potentiation; gemcitabine; p53