title: Potentiation of the novel topoisomerase I inhibitor indenoisoquinoline LMP-400 by the cell checkpoint and Chk1-Chk2 inhibitor AZD7762.
abstract: Novel topoisomerase I (Top1) inhibitors are in clinical development to circumvent the drawbacks of camptothecins (CPT). Here, we report molecular investigations into LMP-400, an indenoisoquinoline Top1 inhibitor in phase 1 clinical trial, by itself and in combination with the cell-cycle checkpoint inhibitor AZD7762. We examined drug effects on DNA replication and killing of cancer cells and found that LMP-400 showed synergistic antiproliferative activity when combined with AZD7762 in human colon carcinoma cells. Inhibition of S-phase progression and bromodeoxyuridine incorporation were similarly induced by LMP-400 and CPT and were abrogated by AZD7762. Replication studied by single DNA molecule analyses and immunofluorescence microscopy (molecular combing) showed rapid inhibition of fork progression in response to LMP-400 treatment with subsequent recapitulation after AZD7762 addition. AZD7762 inhibited both the activation/autophosphosphorylation of Chk1 and Chk2 at nanomolar concentrations in LMP-400-treated cells. This potent dual inhibition of Chk1 and Chk2 by AZD7762 was below the drug concentrations required to abrogate cell-cycle inhibition and produce synergism with LMP-400. Also, the synergism was independent of Chk2 both in Chk2-complemented cells and Chk2 knockout cells, suggesting additional mechanisms for cell-cycle abrogation by AZD7762. Together, our findings show a rationale for combining cell-cycle checkpoint inhibitors with the novel non-CPT indenoisoquinoline Top1 inhibitors.
authors.affiliation: Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, Bethesda, Maryland, USA.
authors.forename: Sheena M
authors.initials: SM
authors.lastname: Aris
authors.forename: Yves
authors.initials: Y
authors.lastname: Pommier
mesh_terms: D000971:Antineoplastic Combined Chemotherapy Protocols / Q000627:therapeutic use*; D052117:Benzodioxoles / Q000494:pharmacology*; D000071877:Checkpoint Kinase 1; D064447:Checkpoint Kinase 2; D003110:Colonic Neoplasms / Q000188:drug therapy*; D004261:DNA Replication / Q000187:drug effects; D004357:Drug Synergism; D019073:HT29 Cells; D006801:Humans; D007546:Isoquinolines / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011494:Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D013876:Thiophenes / Q000494:pharmacology*; D059004:Topoisomerase I Inhibitors / Q000494:pharmacology*; D014508:Urea / Q000031:analogs & derivatives* / Q000494:pharmacology
chemical_list: C532363:3-(carbamoylamino)-5-(3-fluorophenyl)-N-(3-piperidyl)thiophene-2-carboxamide; D052117:Benzodioxoles; D007546:Isoquinolines; C553898:NSC 724998; D047428:Protein Kinase Inhibitors; D013876:Thiophenes; D059004:Topoisomerase I Inhibitors; D014508:Urea; D011494:Protein Kinases; D064447:Checkpoint Kinase 2; C000605867:CHEK1 protein, human; C578634:CHEK2 protein, human; D000071877:Checkpoint Kinase 1; D017346:Protein Serine-Threonine Kinases