title: Chk1 signaling pathways that mediated G(2)M checkpoint in relation to the cellular resistance to the novel topoisomerase I poison BNP1350.
abstract: A novel karenitecin, BNP1350, is a topoisomerase I-targeting anticancer agent with significant antitumor activity in vitro and in vivo. A BNP1350-resistant human head and neck carcinoma A253 cell line, denoted A253/BNPR, was developed. The A253/BNPR cell line was approximately 9-fold resistant to BNP1350 and 4-fold cross-resistant to another topoisomerase I inhibitor SN-38, the active metabolite of irinotecan. After drug treatment with equimolar concentrations of BNP1350 (0.7 microM) for 2h, activation of the DNA double-strand break repair protein complexes was similar in the two cell lines, suggesting that DNA dsb repair is not attributable to resistance to BNP1350 in the A253/BNPR cells. Cell cycle analysis indicates that the A253 cell line accumulated primarily in S phase, but G(2) phase accumulation was observed in the A253/BNPR cell line at 48 h after drug removal. Elevated chk1 phosphorylation at Ser(345) following DNA damage induced by BNP1350 was accompanied by G(2) accumulation in the A253/BNPR cell line, while exposure to equimolar concentrations of BNP1350 (0.7 microM) induced S-phase arrest and no increased phosphorylation of chk1 at Ser(345) in the A253 cell line. Under the same conditions, increased chk1 activity was observed in the A253/BNPR cell line, but not in the A253 cell line. Moreover, stimulated binding of 14-3-3 proteins to chk1 was observed in BNP1350-treated A253/BNPR cells. To confirm relationship between chk1 expression/phosphorylation and drug resistance to topo I poisons, we examined the effects of chk1 or chk2 antisense oligonucleotides on the cellular growth inhibition. Chk1 antisense oligonucleotide can sensitize the A253/BNPR cells to killing by topo I inhibitor BNP1350, but no significant sensitization of BNP1350-induced growth inhibition was observed in the drug-sensitive cell line. Chk2 antisense oligonucleotide has only a small sensitization effect on BNP1350-induced growth inhibition in both cell lines. The data indicate that the chk1 signaling pathways that mediate cell cycle checkpoint are associated with cellular resistance to BNP1350 in the A253/BNPR cell line.
authors.affiliation: Department of Pharmacology and Therapeutics, Grace Cancer Drug Center, Roswell Park Cancer Institute, Buffalo, NY 14263, USA.
authors.forename: Ming-biao
authors.initials: MB
authors.lastname: Yin
authors.forename: Gunnar
authors.initials: G
authors.lastname: Hapke
authors.forename: Jiaxi
authors.initials: J
authors.lastname: Wu
authors.forename: Rami G
authors.initials: RG
authors.lastname: Azrak
authors.forename: Cheryl
authors.initials: C
authors.lastname: Frank
authors.forename: Carol
authors.initials: C
authors.lastname: Wrzosek
authors.forename: Youcef M
authors.initials: YM
authors.lastname: Rustum
mesh_terms: D048948:14-3-3 Proteins; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000494:pharmacology*; D002455:Cell Division / Q000187:drug effects; D000071877:Checkpoint Kinase 1; D004249:DNA Damage; D004260:DNA Repair; D019008:Drug Resistance, Neoplasm; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D016195:G2 Phase; D006801:Humans; D008938:Mitosis; D016376:Oligonucleotides, Antisense / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D015398:Signal Transduction; D059004:Topoisomerase I Inhibitors; D014407:Tumor Cells, Cultured; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism
chemical_list: D048948:14-3-3 Proteins; D004791:Enzyme Inhibitors; D016376:Oligonucleotides, Antisense; D059004:Topoisomerase I Inhibitors; C406143:cositecan; D014446:Tyrosine 3-Monooxygenase; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D002166:Camptothecin