title: Irinotecan and DNA-PKcs inhibitors synergize in killing of colon cancer cells.
abstract: This study sought to measure the degree of synergy induced by specific small molecule inhibitors of DNA-PK [NU7026 and IC486241 (ICC)], a major component of the non-homologous end-joining (NHEJ) pathway, with SN38 or oxaliplatin. Synergy between the DNA damaging drugs and the DNA-PK inhibitors was assessed using the sulforhodamine-B assay (SRB). Effects of drug combinations on cell cycle and DNA-PK activity were determined using flow cytometry and western blot analysis. DNA damage was assessed via comet assay and quantification of γH2AX. The role of homologous recombination repair (HRR) was determined by nuclear Rad51 protein levels and a GFP reporter recombination assay. Significant reductions in the IC(50) values of SN38 were observed at 5 and 10 μM of DNA-PK inhibitors. Moreover, at 1-2 μM (attainable concentrations with ICC in mice) these DNA-PKcs inhibitors demonstrated synergistic reductions in the IC(50) of SN38. Flow cytometric data indicated that SN38 and SN38 in combination with DNA-PKcs inhibitors showed dramatic G2/M arrest at 24 h. Furthermore, reduced phosphorylation of DNA-PKcs and increased DNA damage were observed at this time point with SN38 in combination with DNA-PKcs inhibitors as compared to cells treated with SN38 alone. SN38 alone and in the presence of ICC increased nuclear Rad51 protein levels. Furthermore, inhibition of DNA-PKcs increased HRR suggesting that NHEJ is a negative regulator of HRR. These data indicate that small molecule inhibitors of DNA-PKcs dramatically enhance the efficacy of SN38 in colon cancer cell lines.
authors.affiliation: Montreal Centre for Experimental Therapeutics in Cancer-Lady Davis Institute-Jewish General Hospital, McGill University, 3755, Côte Sainte Catherine Road, Montréal, Québec, H3T 1E2, Canada.
authors.forename: David
authors.initials: D
authors.lastname: Davidson
authors.forename: Yannick
authors.initials: Y
authors.lastname: Coulombe
authors.forename: Veronica L
authors.initials: VL
authors.lastname: Martinez-Marignac
authors.forename: Lilian
authors.initials: L
authors.lastname: Amrein
authors.forename: Jeremy
authors.initials: J
authors.lastname: Grenier
authors.forename: Keira
authors.initials: K
authors.lastname: Hodkinson
authors.forename: Jean-Yves
authors.initials: JY
authors.lastname: Masson
authors.forename: Raquel
authors.initials: R
authors.lastname: Aloyz
authors.forename: Lawrence
authors.initials: L
authors.lastname: Panasci
mesh_terms: D054831:Acridones / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000494:pharmacology; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002867:Chromones / Q000494:pharmacology*; D002945:Cisplatin / Q000494:pharmacology; D003110:Colonic Neoplasms / Q000473:pathology; D020552:Comet Assay; D051747:DNA-Activated Protein Kinase / Q000037:antagonists & inhibitors*; D004357:Drug Synergism; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D000077146:Irinotecan; D009025:Morpholines / Q000494:pharmacology*; D009944:Organoplatinum Compounds / Q000494:pharmacology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011725:Pyridines / Q000494:pharmacology; D059004:Topoisomerase I Inhibitors / Q000494:pharmacology
chemical_list: C479235:2-(morpholin-4-yl)benzo(h)chromen-4-one; D054831:Acridones; D000970:Antineoplastic Agents; D002867:Chromones; C575866:IC486241; D009025:Morpholines; D009944:Organoplatinum Compounds; D047428:Protein Kinase Inhibitors; D011725:Pyridines; D059004:Topoisomerase I Inhibitors; C431073:oxiplatin; D000077146:Irinotecan; D051747:DNA-Activated Protein Kinase; D002945:Cisplatin; D002166:Camptothecin