Abstract
Background
Replication-dependent radiosensitization of tumors ranks among the most promising
tools for future improvements in tumor therapy. However, cell cycle checkpoint signaling
during S phase is a key for maintaining genomic stability after ionizing irradiation
allowing DNA damage repair by stabilizing replication forks, inhibiting new origin
firing and recruiting DNA repair proteins. As the impact of the different types of
DNA damage induced by ionizing radiation on replication fork functionality has not
been investigated, this study was performed in tumor cells treated with various agents
that induce specific DNA lesions.
Methods
U2OS cells were exposed to methyl methanesulfonate (MMS) to induce base damage, low
or high concentrations of hydrogen peroxide for the induction of SSBs, Topotecan to
induce DSBs at replication, Mitomycin C (MMC) to induce interstrand cross-links or
ionizing irradiation to analyze all damages. Chk1 phosphorylation, origin firing and
replication fork progression, and cell cycle distribution were analyzed.
Results
In our system, the extent of Chk1 phosphorylation was dependent on the type of damage
induced and prolonged Chk1 phosphorylation correlated with the inhibition of replication
initiation. Ionizing radiation, high concentrations of hydrogen peroxide, and Topotecan
affected replication elongation much more strongly that the other agents. Almost all
agents induced a slight increase in the S phase population but subsequent G2 arrest
was only observed in response to those agents that strongly inhibited replication
elongation and caused prolonged Chk1 phosphorylation.
Conclusions
Our data suggest that to improve radiotherapy, radiosensitivity in S phase could be
increased by combining irradiation with agents that induce secondary DSB or inhibit
checkpoint signaling, such as inhibitors of PARP or Chk1.
Abbreviations:
BD (base damage), DDR (DNA damage response), DSB (double strand break), ICL (interstrand crosslink), MMC (Mitomycin C), MMS (methyl methanesulfonate), SSB (single strand break)Keywords
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Article info
Publication history
Published online: February 13, 2012
Accepted:
January 2,
2012
Received in revised form:
December 8,
2011
Received:
August 10,
2011
Identification
Copyright
© 2012 Elsevier Ireland Ltd. Published by Elsevier Inc. All rights reserved.
