title: Involvement of ATM-mediated Chk1/2 and JNK kinase signaling activation in HKH40A-induced cell growth inhibition.
abstract: HKH40A (RTA 502), or 5-nitro-2-(3-{4-[3-(8-methoxy-6-oxo-6H-2,10b-diaza-aceanthrylen-5-ylamino)propyl]piperazin-1-yl}-propyl)-2-aza-phenalene-1,3-dione, has been shown to be a potent cell growth inhibitor. To investigate HKH40A actions, we focused on exploring the signal transduction pathways that are involved in its cell growth inhibitory mechanisms. We found that HKH40A activated ataxia telangiectasia mutated (ATM) kinase, which then triggered activation of the Chk1/2 signaling pathway, evidenced by Chk1/2 mediated inhibitory phosphorylation of Cdc25C protein phosphatase. This resulted in Cdk1 tyrosine phosphorylation at Tyr-15, leading to cell cycle block at G2/M phase. HKH40A also activated the c-Jun N-terminal kinase (JNK) pathway, most likely regulated by ATM kinase, since pre-treatment of Hep3B cells with ATM inhibitor KU-55933 or ATM siRNA transfection antagonized HKH40A-induced c-Jun phosphorylation. HKH40A-induced apoptosis was probably mediated by JNK-H2A.X interaction, since phospho-c-Jun and phospho-H2A.X were able to co-localize in the nucleus and to co-immuno-precipitate. Furthermore, inhibition of JNK kinase activity by JNK inhibitor SP600125 abolished both HKH40A-induced H2A.X phosphorylation and apoptosis. Our data support the hypothesis that binding of HKH40A to cellular DNA likely activates ATM kinase, which then induces parallel Chk 1/2 and JNK signaling pathways, leading to G2/M cell cycle block and apoptosis.
authors.affiliation: Department of Medical Oncology, Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, Pennsylvania, USA. ziqiu.wang@jefferson.edu
authors.forename: Ziqiu
authors.initials: Z
authors.lastname: Wang
authors.forename: Meifang
authors.initials: M
authors.lastname: Wang
authors.forename: Sid
authors.initials: S
authors.lastname: Kar
authors.forename: Brian I
authors.initials: BI
authors.lastname: Carr
mesh_terms: D054831:Acridones / Q000494:pharmacology*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D064007:Ataxia Telangiectasia Mutated Proteins; D006528:Carcinoma, Hepatocellular / Q000201:enzymology / Q000473:pathology; D018797:Cell Cycle Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D000071877:Checkpoint Kinase 1; D064447:Checkpoint Kinase 2; D004247:DNA / Q000096:biosynthesis; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D006657:Histones / Q000378:metabolism; D006801:Humans; D008113:Liver Neoplasms / Q000201:enzymology / Q000473:pathology; D048670:MAP Kinase Kinase 4 / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D008297:Male; D053644:Naphthalimides / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats; D011916:Rats, Inbred F344; D025521:Tumor Suppressor Proteins / Q000378:metabolism*
chemical_list: D054831:Acridones; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D006657:Histones; D053644:Naphthalimides; C525457:RTA 502; D025521:Tumor Suppressor Proteins; D004247:DNA; D011494:Protein Kinases; D064447:Checkpoint Kinase 2; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C000605867:CHEK1 protein, human; C578634:CHEK2 protein, human; D000071877:Checkpoint Kinase 1; C000605871:Chek1 protein, rat; C578632:Chek2 protein, rat; D017346:Protein Serine-Threonine Kinases; D048670:MAP Kinase Kinase 4