Ionizing radiation activates AMP-activated kinase (AMPK): a target for radiosensitization of human cancer cells.
OBJECTIVE
Adenosine monophosphate (AMP)-activated kinase (AMPK) is a molecular energy sensor regulated by the tumor suppressor LKB1. Starvation and growth factors activate AMPK through the DNA damage sensor ataxia-telangiectasia mutated (ATM). We explored the regulation of AMPK by ionizing radiation (IR) and its role as a target for radiosensitization of human cancer cells.


METHODS
Lung, prostate, and breast cancer cells were treated with IR (2-8 Gy) after incubation with either ATM or AMPK inhibitors or the AMPK activator metformin. Then, cells were subjected to either lysis and immunoblotting, immunofluorescence microscopy, clonogenic survival assays, or cell cycle analysis.


RESULTS
IR induced a robust phosphorylation and activation of AMPK in all tumor cells, independent of LKB1. IR activated AMPK first in the nucleus, and this extended later into cytoplasm. The ATM inhibitor KU-55933 blocked IR activation of AMPK. AMPK inhibition with Compound C or anti-AMPK alpha subunit small interfering RNA (siRNA) blocked IR induction of the cell cycle regulators p53 and p21(waf/cip) as well as the IR-induced G2/M arrest. Compound C caused resistance to IR, increasing the surviving fraction after 2 Gy, but the anti-diabetic drug metformin enhanced IR activation of AMPK and lowered the surviving fraction after 2 Gy further.


CONCLUSIONS
We provide evidence that IR activates AMPK in human cancer cells in an LKB1-independent manner, leading to induction of p21(waf/cip) and regulation of the cell cycle and survival. AMPK appears to (1) participate in an ATM-AMPK-p21(waf/cip) pathway, (2) be involved in regulation of the IR-induced G2/M checkpoint, and (3) may be targeted by metformin to enhance IR responses.
Sanli
Toran
T
Department of Oncology, Juravinski Cancer Center and McMaster University, Hamilton, Ontario, Canada.
Rashid
Ayesha
A
Liu
Caiqiong
C
Harding
Shane
S
Bristow
Robert G
RG
Cutz
Jean-Claude
JC
Singh
Gurmit
G
Wright
James
J
Tsakiridis
Theodoros
T
D055372:AMP-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D064007:Ataxia Telangiectasia Mutated Proteins; D001943:Breast Neoplasms / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000528:radiation effects; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000037:antagonists & inhibitors / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects / Q000528:radiation effects; D005260:Female; D016195:G2 Phase / Q000187:drug effects; D006801:Humans; D008175:Lung Neoplasms / Q000378:metabolism; D008297:Male; D008687:Metformin / Q000494:pharmacology; D009025:Morpholines / Q000494:pharmacology; D010766:Phosphorylation / Q000528:radiation effects; D011471:Prostatic Neoplasms / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011720:Pyrazoles / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D011753:Pyrones / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000494:pharmacology; D011836:Radiation Tolerance / Q000187:drug effects; D016159:Tumor Suppressor Protein p53 / Q000037:antagonists & inhibitors / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000378:metabolism
C495818:2-morpholin-4-yl-6-thianthren-1-yl-pyran-4-one; C493956:CDKN1A protein, human; D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D004268:DNA-Binding Proteins; D009025:Morpholines; D011720:Pyrazoles; D011743:Pyrimidines; D011753:Pyrones; D034741:RNA, Small Interfering; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; C516138:dorsomorphin; D008687:Metformin; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases