title: LY2606368 Causes Replication Catastrophe and Antitumor Effects through CHK1-Dependent Mechanisms.
abstract: CHK1 is a multifunctional protein kinase integral to both the cellular response to DNA damage and control of the number of active replication forks. CHK1 inhibitors are currently under investigation as chemopotentiating agents due to CHK1's role in establishing DNA damage checkpoints in the cell cycle. Here, we describe the characterization of a novel CHK1 inhibitor, LY2606368, which as a single agent causes double-stranded DNA breakage while simultaneously removing the protection of the DNA damage checkpoints. The action of LY2606368 is dependent upon inhibition of CHK1 and the corresponding increase in CDC25A activation of CDK2, which increases the number of replication forks while reducing their stability. Treatment of cells with LY2606368 results in the rapid appearance of TUNEL and pH2AX-positive double-stranded DNA breaks in the S-phase cell population. Loss of the CHK1-dependent DNA damage checkpoints permits cells with damaged DNA to proceed into early mitosis and die. The majority of treated mitotic nuclei consist of extensively fragmented chromosomes. Inhibition of apoptosis by the caspase inhibitor Z-VAD-FMK had no effect on chromosome fragmentation, indicating that LY2606368 causes replication catastrophe. Changes in the ratio of RPA2 to phosphorylated H2AX following LY2606368 treatment further support replication catastrophe as the mechanism of DNA damage. LY2606368 shows similar activity in xenograft tumor models, which results in significant tumor growth inhibition. LY2606368 is a potent representative of a novel class of drugs for the treatment of cancer that acts through replication catastrophe.
authors.affiliation: Oncology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Eli Lilly and Company, Indianapolis, Indiana.
authors.forename: Constance
authors.initials: C
authors.lastname: King
authors.affiliation: Oncology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Eli Lilly and Company, Indianapolis, Indiana.
authors.forename: H Bruce
authors.initials: HB
authors.lastname: Diaz
authors.affiliation: Oncology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Eli Lilly and Company, Indianapolis, Indiana.
authors.forename: Samuel
authors.initials: S
authors.lastname: McNeely
authors.affiliation: Oncology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Eli Lilly and Company, Indianapolis, Indiana.
authors.forename: Darlene
authors.initials: D
authors.lastname: Barnard
authors.affiliation: Oncology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Eli Lilly and Company, Indianapolis, Indiana.
authors.forename: Jack
authors.initials: J
authors.lastname: Dempsey
authors.affiliation: Oncology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Eli Lilly and Company, Indianapolis, Indiana.
authors.forename: Wayne
authors.initials: W
authors.lastname: Blosser
authors.affiliation: Oncology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Eli Lilly and Company, Indianapolis, Indiana.
authors.forename: Richard
authors.initials: R
authors.lastname: Beckmann
authors.affiliation: Chemistry Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Eli Lilly and Company, Indianapolis, Indiana.
authors.forename: David
authors.initials: D
authors.lastname: Barda
authors.affiliation: Oncology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Eli Lilly and Company, Indianapolis, Indiana. mmarshall@lilly.com.
authors.forename: Mark S
authors.initials: MS
authors.lastname: Marshall
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology*; D045744:Cell Line, Tumor; D000071877:Checkpoint Kinase 1; D051357:Cyclin-Dependent Kinase 2 / Q000378:metabolism; D004249:DNA Damage / Q000187:drug effects; D004261:DNA Replication / Q000187:drug effects*; D004195:Disease Models, Animal; D005260:Female; D006801:Humans; D051379:Mice; D009369:Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D011494:Protein Kinases / Q000378:metabolism*; D011719:Pyrazines / Q000494:pharmacology*; D011720:Pyrazoles / Q000494:pharmacology*; D016196:S Phase / Q000187:drug effects / Q000235:genetics; D023041:Xenograft Model Antitumor Assays; D020687:cdc25 Phosphatases / Q000378:metabolism
chemical_list: D000970:Antineoplastic Agents; D047428:Protein Kinase Inhibitors; D011719:Pyrazines; D011720:Pyrazoles; C000608121:prexasertib; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; C000605869:Chek1 protein, mouse; D051357:Cyclin-Dependent Kinase 2; C517004:CDC25A protein, human; D020687:cdc25 Phosphatases