title: Chlorambucil cytotoxicity in malignant B lymphocytes is synergistically increased by 2-(morpholin-4-yl)-benzo[h]chomen-4-one (NU7026)-mediated inhibition of DNA double-strand break repair via inhibition of DNA-dependent protein kinase.
abstract: Chlorambucil (CLB) treatment is used in chronic lymphocytic leukemia (CLL) but resistance to CLB develops in association with accelerated repair of CLB-induced DNA damage. Phosphorylated histone H2AX (gammaH2AX) is located at DNA double-strand break (DSB) sites; furthermore, it recruits and retains damage-responsive proteins. This damage can be repaired by nonhomologous DNA end-joining (NHEJ) and/or homologous recombinational repair (HR) pathways. A key component of NHEJ is the DNA-dependent protein kinase (DNA-PK) complex. Increased DNA-PK activity is associated with resistance to CLB in CLL. We used the specific DNA-PK inhibitor 2-(morpholin-4-yl)-benzo[h]chomen-4-one (NU7026) to sensitize CLL cells to chlorambucil. Our results indicate that in a CLL cell line (I83) and in primary CLL-lymphocytes, chlorambucil plus NU7026 has synergistic cytotoxic activity at nontoxic doses of NU7026. CLB treatment results in G(2)/M phase arrest, and NU7026 increases this CLB-induced G(2)/M arrest. Moreover, a kinetic time course demonstrates that CLB-induced DNA-PK activity was inhibited by NU7026, providing direct evidence of the ability of NU7026 to inhibit DNA-PK function. DSBs, visualized as gammaH2AX, were enhanced 24 to 48 h after CLB and further increased by CLB plus NU7026, suggesting that the synergy of the combination is mediated by NU7026 inhibition of DNA-PK with subsequent inhibition of DSB repair.
authors.affiliation: Montreal Centre for Experimental Therapeutics in Cancer-Lady Davis Institute for Medical Research, Sir Mortimer B Davis-Jewish General Hospital, McGill University, Montreal, Quebec, Canada.
authors.forename: Lilian
authors.initials: L
authors.lastname: Amrein
authors.forename: Martin
authors.initials: M
authors.lastname: Loignon
authors.forename: Anne-Christine
authors.initials: AC
authors.lastname: Goulet
authors.forename: Michael
authors.initials: M
authors.lastname: Dunn
authors.forename: Bertrand
authors.initials: B
authors.lastname: Jean-Claude
authors.forename: Raquel
authors.initials: R
authors.lastname: Aloyz
authors.forename: Lawrence
authors.initials: L
authors.lastname: Panasci
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D001402:B-Lymphocytes / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002699:Chlorambucil / Q000494:pharmacology*; D002867:Chromones / Q000494:pharmacology*; D004260:DNA Repair / Q000187:drug effects*; D051747:DNA-Activated Protein Kinase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D019008:Drug Resistance, Neoplasm; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006657:Histones / Q000378:metabolism; D006801:Humans; D020128:Inhibitory Concentration 50; D009025:Morpholines / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects
chemical_list: C479235:2-(morpholin-4-yl)benzo(h)chromen-4-one; D002867:Chromones; D004791:Enzyme Inhibitors; C468783:H2AX protein, human; D006657:Histones; D009025:Morpholines; D002699:Chlorambucil; D051747:DNA-Activated Protein Kinase