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@INPROCEEDINGS{Dahmen:818233,
author = {Dahmen, Volker and Kriehuber, Ralf},
title = {{S}pecificity, damaging potential and cell killing of
{I}-125 labeled {T}riplex-forming oligonucleotides in a
transgenic cell line},
reportid = {FZJ-2016-04716},
year = {2016},
abstract = {Introduction: Triplex-forming oligonucleotides (TFOs) are
known for their ability to bind DNA in a sequence specific
manner and are therefore a promising tool to manipulate
genes or gene regulatory units in a cellular environment.
Auger-Electron-Emitter-labeled TFOs can induce complex but
localized damage to the DNA. Using radionuclide-labeled TFO
the specificity of the target binding and the subsequent
damage in the cellular environment were analysed by
comparing the effects in two cell lines of which only one
possessed the proper TFO target sequence. Methods: The human
carcinoma cell line SCL-II wildtype (WT) was stably
transfected with a vector system (p2RT) producing a
transgenic strain, SCL-II p2RT, containing the TFO-p2RT
target sequence. TFO labeling with I-125 or P-32 was
performed using the primer extension method. Efficient
delivery of labeled TFO into SCL-II p2RT as well as SCL-II
WT cells was ensured by electroporation with the
Nucleofector I system (Lonza GmbH, Basel). Subsequently,
specific target binding was visualized by restriction digest
of isolated genomic DNA and autoradiographic analysis. DNA
damage and survival rate were determined with the 53BP1-foci
and the colony forming assay (CFA).Results: Autoradiographic
analysis revealed a specific triplex formation at the p2RT
vector target sequence, confirming the specific binding.
However, the 53BP1-foci assay and the CFA detected a similar
increased level of DNA damage and a pronounced reduction in
cell survival in both cell strains per cumulated decays.
Thus, the different target status in the two cell strains
was not reflected on damage level and in survival rate.
Conclusions: Even though a high degree of specific TFO-p2RT
binding to its target site could be verified, the 53BP1 and
CFA data indicate that also a considerable amount of
alternative DNA damaging is occurring.},
month = {Sep},
date = {2016-09-26},
organization = {19th Annual Meeting of the Society for
Biological Radiation Research, Erlangen
(Germany), 26 Sep 2016 - 28 Sep 2016},
subtyp = {After Call},
cin = {S-US},
cid = {I:(DE-Juel1)S-US-20090406},
pnm = {899 - ohne Topic (POF3-899)},
pid = {G:(DE-HGF)POF3-899},
typ = {PUB:(DE-HGF)24},
url = {https://juser.fz-juelich.de/record/818233},
}