Inhibition of ATR leads to increased sensitivity to hypoxia/reoxygenation.
The transient opening and closing of tumor vasculature result in periods of severe oxygen deprivation (hypoxia) followed by reoxygenation. This exerts a positive selective pressure for cells that have lost their sensitivity to killing by reduced oxygen levels. These cells are effectively resistant to hypoxia-induced apoptosis and conventional therapeutic approaches. Here we show hypoxia-induced S-phase arrest results in regions of single-stranded DNA in stalled replication forks and signals the activation of ATR. S-phase cells represent the most sensitive phase of the cell cycle to the stress of hypoxia/reoxygenation. Loss of ATR or inhibition of ATR kinase activity results in a further loss of reproductive viability in S-phase cells when exposed to hypoxic conditions followed by reoxygenation but has little effect on the inhibition of DNA synthesis. This is, at least in part, mediated via Chk1 signaling because loss of Chk1 also results in increased sensitivity to hypoxia/reoxygenation. The observed decrease in reproductive survival is in part because of the accumulation of DNA damage in S-phase cells during hypoxia exposure in the absence of full ATR activity. Therefore, ATR acts to protect stalled replication forks during hypoxia exposure. In conclusion, ATR and Chk1 play critical roles in the cellular response to hypoxia/reoxygenation, and inhibitors of ATR and Chk1 represent new hypoxic cell cytotoxins.
Hammond
Ester M
EM
Center for Clinical Sciences Research, Department of Radiation Oncology, Stanford University, Stanford, California 94305, USA.
Dorie
Mary Jo
MJ
Giaccia
Amato J
AJ
D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000502:physiology; D015687:Cell Hypoxia / Q000502:physiology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000502:physiology; D000071877:Checkpoint Kinase 1; D004249:DNA Damage; D004261:DNA Replication / Q000502:physiology; D004277:DNA, Single-Stranded / Q000378:metabolism; D045325:HCT116 Cells; D006801:Humans; D010100:Oxygen / Q000378:metabolism*; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000502:physiology; D034741:RNA, Small Interfering / Q000235:genetics / Q000494:pharmacology; D015398:Signal Transduction / Q000502:physiology
D018797:Cell Cycle Proteins; D004277:DNA, Single-Stranded; D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; D011494:Protein Kinases; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D017346:Protein Serine-Threonine Kinases; D010100:Oxygen