![[Feedback]](/icons/banner/feedbackACT.gif){kind=icon}
![[For Subscribers]](/icons/banner/subscriberACT.gif){kind=icon}
![[Archive]](/icons/banner/archiveACT.gif){kind=icon}
![[Search]](/icons/banner/searchACT.gif){kind=icon}
![[Contents]](/icons/banner/tocACT.gif){kind=icon}
|
|
|
|
|
||
Vol. 19, Issue 8, 3283-3289, August 2008
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
*Institute of Legal Medicine, University of Ulm, 89075 Ulm, Germany; University Children's Hospital, 89075 Ulm, Germany; and Department of Transfusion Medicine, Institute for Clinical Transfusion Medicine, and Institute for Transfusion Medicine and Immunogenetics, University Hospital of Ulm, 89081 Ulm, Germany
Submitted March 20, 2008;
Revised May 14, 2008;
Accepted May 19, 2008
Monitoring Editor: William P. Tansey
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
; Kaina, 2003; Willmore et al., 2004; Deriano et al., 2005).
DNA repair mechanisms such as mismatch repair, base excision repair, nucleotide excision repair, direct damage reversal, and DNA double-strand break repair (DNA-DSB-repair) are involved in the integrity of DNA. Defects in DNA repair give rise to hypersensitivity to DNA-damaging agents, accumulation of mutations in the genome, and finally to the development of cancer and various metabolic disorders (Christmann et al., 2003).
Doxorubicin, cisplatin, cyclophosphamide, and etoposide induce DNA damage, which generates a complex cascade of events leading to cell cycle arrest, transcriptional and posttranscriptional activation of a subset of genes including those associated with DNA repair, and triggering of apoptosis (Friesen et al., 1996, 1999a; Christmann et al., 2003; Kaina, 2003).
The genotoxic effects of DNA double-strand breaks (DNA-DSBs) are highly potent inducers of cell death (Christmann et al., 2003). In higher eukaryotes a single nonrepaired DNA-DSB inactivating an essential gene can be sufficient for the indention of cell death via apoptosis (Rich et al., 2000). The repair of DNA-DSBs is critical for the survival of cells exposed to DNA-damaging agents (Christmann et al., 2003). Homologous recombination (HR) and nonhomologous end joining (NHEJ) are the two principal pathways repairing DNA-DSBs (Christmann et al., 2003). In simple eukaryotes, HR is the main pathway, whereas in mammalian cells, NHEJ predominantly repairs DNA-DSBs, resulting from DNA-damaging agents (Jeggo, 1998a,b; Haber, 2000; Cromie et al., 2001). NHEJ does not depend on the presence of homologous DNA sequences and requires the DNA-dependent protein kinase (DNA-PK) complex and the XRCC4/DNA-ligase IV complex (Jeggo, 1998a,b; Lieber, 1999). DNA-PK is a member of the phosphatidylinositol-3 kinase family that includes ataxia-telangiectasia-mutated (ATM) and ATM-Rad3-related (ATR) (Durocher and Jackson, 2001). It consists of a heterotrimeric complex consisting of proteins Ku70 and Ku80 (Ku70/80) and the catalytic DNA-PK subunit (Christmann et al., 2003). DNA-PK exhibits protein kinase activity only when bound to DNA (Christmann et al., 2003). The first step in DNA-DSB-repair by NHEJ is the binding of Ku70/80 to the damaged DNA followed by the recruitment of the catalytic subunit DNA-PK, thereby forming the active DNA-PK holoenzyme (Smith and Jackson, 1999; Christmann et al., 2003). One of the targets of DNA-PK is XRCC4, which forms a stable complex with DNA-ligase IV, joining the ends of broken DNA strands (Grawunder et al., 1998a,b).
Doxorubicin, cisplatin, cyclophosphamide, and etoposide not only induce DNA damage but also induce apoptosis and activate caspases (Friesen et al., 1996, 1999a,b, 2004; Kaufmann and Earnshaw, 2000; Christmann et al., 2003; Deriano et al., 2005). Caspases play a critical role in apoptosis induction (Kaufmann and Earnshaw, 2000). The apoptotic caspases are divided into two classes, effector caspases and initiator caspases. Effector caspases are responsible for the cleavage that disassemble the cell, and initiator caspases initiate a proteolytic caspase-activating cascade (Kaufmann and Earnshaw, 2000). Caspase-3, -6, and -7 are the effector caspases, and caspase-8 and -9 are the major initiator caspases (Kaufmann and Earnshaw, 2000). Caspase-8 or -9 is activated in a multimeric complex. Caspase-8 is activated in the death-inducing signaling complex, and caspase-9 is within the apoptosome (Kaufmann and Earnshaw, 2000). On activation of initiator caspases, caspase-8 and -9 acquire the ability to cleave and activate effector caspases such as caspase-3 (Kaufmann and Earnshaw, 2000). Caspases activation can be triggered by two different mechanisms, the death receptor pathway and the mitochondrial pathway, and it is negatively regulated at the receptor level by Flice-inhibitory protein that blocks caspase-8 activation, at the mitochondria level by Bcl-2 family proteins, and by inhibitor of apoptosis proteins (Kroemer, 1997; Deveraux et al., 1999; Srinivasula et al., 2001).
In the present study, we investigate the role of NHEJ-DNA-DSB-repair, DNA-ligase IV, and DNA-PK in deficient caspases activation by doxorubicin. A greater understanding of the links between deficient caspases activation and DNA repair in apoptosis-, chemo-, as well as radioresistance will lead to the development of more effective cancer prevention and treatment strategies.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
DNA-PK–deficient (–/–) and Proficient (+/+) Cells
The human glioblastoma cell lines MO59K [DNA-PK (+/+)] and MO59J [DNA-PK (–/–)] were obtained from American Type Culture Collection (Manassas, VA) and grown in complete DMEM (Invitrogen), which contained 10% heat-inactivated fetal bovine serum (Biochrom), 5 mM HEPES (Biochrom), 100 U/ml penicillin (Invitrogen), 100 µg/ml streptomycin (Invitrogen), and 2 mM L-glutamine (Biochrom) at 37°C in an atmosphere of 5% CO2.
Treatment of Adherent Cells
DNA-PK (+/+) and DNA-PK (–/–) glioblastoma cells were trypsinized and replated on 75-cm2 tissue culture flask at a concentration of 8000 cells/cm2. Cells were allowed to attached to the bottom of the wells, and then cells were treated with different concentrations of doxorubicin. At different time points, the Comet assay, the Nicoletti analysis, flow cytometry analysis, and Western Blot analysis were performed.
Treatment of Nonadherent Cells
Cells (2 x 105 cells/ml; Nalm6, Nalm6DoxoR, Nalm6 [DNA-ligase IV +/+, +/–, –/–)], were treated with different concentrations of doxorubicin. At different time points, the Comet assay, the Nicoletti analysis, flow cytometry analysis, and Western Blot analysis were performed.
Determination of Induction of Apoptosis and Cell Cycle
Apoptosis was determined by the Nicoletti method (Nicoletti et al., 1991) or forward light scatter/ side scatter (FSC/SSC) method (Carbonari et al., 1994). The percentage of apoptotic cells was measured by hypodiploid DNA (sub-G1) by the Nicoletti method or FSC/SSC analysis by flow cytometry (FACSCalibur; BD Biosciences, Heidelberg, Germany).
Inhibition of Caspases Activation by Benzoylcarbonyl-Val-Ala-Asp-fluoromethyl Ketone (z-VAD-fmk)
The broad-spectrum tripeptide inhibitor of caspases z-VAD-fmk (Enzyme Systems Products, Dublin, CA) was used at a concentration of 50 µM. Cells were treated 1 h before doxorubicin treatment.
Western Blot Analysis
Western blot analysis was done as described previously (Friesen et al., 2003). Immunodetection of poly(ADP-ribose) polymerase (PARP), caspase-3, active caspase-3, caspase-9, active caspase-9, and β-actin was done using rabbit anti-PARP polyclonal-antibody (1:5000; Enzyme Systems Products), mouse anti-caspase-3 monoclonal antibody (1:500; Cell Signaling Technology, Danvers, MA), rabbit anti-active-caspase-3 polyclonal-antibody (1:200; Millipore Bioscience Research Reagents, Temecula, CA), rabbit anti-caspase-9 polyclonal-antibody (1:5000; BD Biosciences Transduction Laboratories, Lexington, KY), rabbit anti-active caspase-9 polyclonal antibody (1:200; Millipore Bioscience Research Reagents), and mouse anti-β-actin monoclonal-antibody (Sigma Chemie, Deisenhofen, Germany). Peroxidase-conjugated goat anti-mouse immunoglobulin G (IgG) or peroxidase-conjugated goat anti-rabbit IgG (1:5000; Santa Cruz Biotechnology, Santa Cruz, CA) as secondary antibody was used for the enhanced chemiluminescence system (GE Healthcare, Little Chalfont, Buckinghamshire, United Kingdom). Equal protein loading was controlled by β-actin analysis.
Measurement of DNA Damage (Comet Assay)
DNA damage (DNA breaks) were measured by the alkaline Comet assay (Hartmann et al., 2003). Analysis was performed with a fluorescence microscope using a charge-coupled device camera connected to a personal computer and analysis software. Relative DNA breakage was expressed as olive tail moment (OTM), which was determined by measuring the fluorescence intensity of tail and nucleus using Kinetic Imgaging Komet 5.0 Software (BFI Optilas, Puchheim, Germany).
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
|
). We found that wortmannin enabled doxorubicin-induced apoptosis (Figure 1E) and DNA damage (Figure 1F) in doxorubicin-resistant Nalm6 cells.
DNA-Ligase IV and DNA-PK Play a Critical Role in Deficient Caspases Activation after Doxorubicin Treatment in Cancer Cells
NHEJ repairs predominant DNA-DSBs in mammalian cells (Jeggo, 1998a). DNA-PK as well as DNA-ligase IV play crucial roles in DNA-DSB-repair by NHEJ (Collis et al., 2005).
To analyze whether NHEJ plays an important role in deficient caspases activation in cancer cells, we measured first the role of DNA-PK in induction of DNA damage (DNA-DSBs and DNA single-strand breaks) and apoptosis after doxorubicin treatment in DNA-DSB-repair–proficient DNA-PK (+/+) and DNA-DSB-repair–deficient DNA-PK (–/–) glioblastoma cells lines (Figure 2A). After 6, 11, and 17 h, we found a strong induction of DNA damage in a DNA-PK (–/–) cells after treatment with doxorubicin (Figure 2A). In DNA-PK (+/+) cells, less DNA damage was detected after doxorubicin treatment, indicating that doxorubicin-induced DNA damage were repaired in DNA-PK (+/+) cells (Figure 2A). In parallel, we measured doxorubicin-induced apoptosis (Figure 2A). At 6, 11, and 17 h, we could not find an induction of apoptosis in DNA-PK (–/–) cells in contrast to induction of DNA damage. Induction of apoptosis was found after doxorubicin treatment in DNA-PK (–/–) cells at later time points (48 and 72 h) (Figure 2A), indicating that DNA damage, left unrepaired by DNA-PK, occurred before apoptosis was detected. In addition, in contrast to DNA-PK (–/–) cells, apoptosis could not be detected after doxorubicin treatment from 3 h until 24 h (Figure 2A) in DNA-PK (+/+) cells, suggesting that DNA-PK is involved in doxorubicin and apoptosis resistance. Minimal apoptosis induction was detected at 48 and 72 h. We next investigated whether DNA-PK is involved in deficient caspases activation in cancer cell (Figure 2B). We treated DNA-DSB-repair–proficient DNA-PK (+/+) and DNA-DSB-repair–deficient DNA-PK (–/–) cells with 0.1 and 0.3 µg/ml doxorubicin (Figure 2B). After 24 and 48 h, Western blot analysis was performed. We found caspase-3 activation, caspase-8 activation and PARP cleavage in DNA-PK (–/–) cells lines after doxorubicin treatment (Figure 2B). In DNA-PK (+/+) cells lines, caspases activation could not be detected after doxorubicin treatment (Figure 2B). In addition, inhibition of DNA-PK by wortmannin, a specific inhibitor of the phosphatidylinositol-3-kinase, including DNA-PK, ATM, and ATR (Durocher and Jackson, 2001), reversed apoptosis resistance (Figure 2C) and deficient activation of caspase-3, and caspase-8 after doxorubicin treatment in DNA-PK (+/+) cells (unpublished data). These findings indicate that DNA-PK plays an important role in doxorubicin-induced caspases activation and apoptosis.
|
). DNA-PK as well as DNA-ligase IV play crucial roles in DNA-DSB-repair by NHEJ (Collis et al., 2005). We next examined whether DNA-PK exclusively or DNA-ligase IV as a key enzyme for NHEJ also plays an important role in deficient caspases activation in cancer cells. Therefore, we incubated DNA-ligase IV (+/+), DNA-ligase IV (+/–), and DNA-ligase IV (–/–) Nalm6 cells with different concentrations of doxorubicin (Figure 3A). After 24, 48, and 72 h, we found a strong induction of apoptosis in DNA-ligase IV (–/–) cells in contrast to DNA-ligase IV (+/–) and DNA-ligase IV (+/+) cells (Figure 3A). In addition, we measured caspases activation after doxorubicin treatment in DNA-ligase IV (+/+), DNA-ligase IV (+/–), and DNA-ligase IV (–/–) Nalm6 cells by Western blot analysis (Figure 3B). After 48 h, we could detect activation of caspase-3, caspase-9, caspase-8, and PARP cleavage in DNA-ligase IV (–/–) Nalm6 cells lines (Figure 3B). In DNA-ligase IV (+/–) and DNA-ligase IV (+/+) Nalm6 cells, caspases activation could not be found at 48 h (Figure 3B). Furthermore, we measured induction of DNA damage (DNA-DSBs and DNA single-strand breaks) (Figure 3C). After 12, 16, 20, 24 h, we found a strong induction of DNA damage in DNA-ligase IV (–/–) Nalm6 cells after treatment with doxorubicin. Induction of DNA damage was barely detected in DNA-ligase IV (+/+) and DNA-ligase IV (+/–) Nalm6 cells at identical doxorubicin concentrations. This demonstrates that DNA-ligase IV as well as DNA-PK play a critical role in caspases activation, suggesting that DNA damage left unrepaired by NHEJ is crucial for caspases activation in cancer cells.
|
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
; Kaina, 2003; Willmore et al., 2004; Deriano et al., 2005). If left unrepaired, DNA damage can result in permanent cell cycle arrest, induction of apoptosis, and mitotic cell death caused by a loss of genomic material (Olive, 1998). Caspases such as caspase-3, caspase-8, and caspase-9 play a critical role in anticancer drug-induced apoptosis, in apoptosis-resistance and anticancer drug resistance (Friesen et al., 1999a; Christmann et al., 2003; Kaina, 2003). Anticancer drugs interact with DNA-activate caspases and induce DNA damage in anticancer drug-sensitive tumor cells (Friesen et al., 1999a; Kaufmann and Earnshaw, 2000; Christmann et al., 2003; Kaina, 2003). In anticancer drug-resistant tumor cells, caspases activation is blocked and DNA repair mechanisms are up-regulated (Friesen et al., 1997, 1999a; Los et al., 1997; Christmann et al., 2003; Kaina, 2003). In our study, we provide the evidence that DNA damage left unrepaired by NHEJ-DNA-DSB-repair initiates activation of caspase-8, caspase-9, and caspase-3 and that DNA-DSB-repair by NHEJ is critical for deficient caspases activation in cancer cells after doxorubicin treatment.
Doxorubicin activates caspases and induces DNA damage in cancer cells (Friesen et al., 1999a; Kaufmann and Earnshaw, 2000; Willmore et al., 2004). In chemosensitive Nalm6 cells, we found a strong activation of caspase-3, caspase-8, and caspase-9, cleavage of PARP, induction of apoptosis, as well as induction of DNA damage (DNA-DSBs and DNA single-strand breaks) after doxorubicin treatment in a time course. Induction of DNA damage was found before caspases activation and induction of apoptosis. This suggests that DNA damage was induced first and caspases activation and apoptosis was followed at later times after doxorubicin treatment. Doxorubicin-resistant cell lines are apoptosis-resistant, and they show a defect in caspases activation (Friesen et al., 1996, 1997, 1999a,b, 2004; Los et al., 1997; Posovszky et al., 1999). Consistent with these data, we found that doxorubicin could not induce apoptosis and activation of caspase-3, caspase-9, and caspase-8 in doxorubicin-resistant Nalm6 cells. DNA repair is critical for cell survival and plays an important role in chemo- and radioresistance (Christmann et al., 2003). Induction of DNA damage was not measurable after treatment with identical concentrations of doxorubicin in doxorubicin-resistant Nalm6 cells, in contrast to doxorubicin-sensitive Nalm6 cells. This suggests that caspases activation is blocked and DNA damage was repaired after doxorubicin treatment in doxorubicin-resistant Nalm6 cells. In doxorubicin-sensitive as well as in doxorubicin-resistant HL-60 and CEM leukemia cell lines, we found comparable results (unpublished data).
p53 accumulates in response to DNA damage and p53 plays an important role in the cellular choice between life and death (Friesen et al., 2003, 2004). p53 can transactivate numerous genes coding for proteins acting either at the receptor level of the apoptotic signals or at downstream stages of the apoptotic process. However, p53-deficient HL-60 cells and p53-mutant CEM cells harbor nonfunctional p53, suggesting that caspases activation and DNA damage may be independent of p53 in these cells lines (Friesen et al., 2003, 2004).
Antracyclines such as doxorubicin could be subject to a multidrug-resistant (MDR) phenotype with decreasing drug uptake in anthracycline-resistant tumor cells (Slapak et al., 1990). Doxorubicin resistance in Nalm6 as well as CEM and HL-60 cells is not due to an MDR phenotype, because we found that doxorubicin resistance is independent of MDR in leukemia cell lines (Friesen et al., 1997, 1999b).
DNA-PK is involved in DNA repair, and it is a molecular sensor for DNA damage that enhances the signal via phosphorylation of many downstream targets (Collis et al., 2005). Inhibition of DNA-PK seems to be a valid approach to enhance tumor cell-killing effects of treatments such as irradiation (Collis et al., 2005). Inhibition of DNA-PK sensitized doxorubicin-resistant Nalm6 cells to doxorubicin and reversed deficient doxorubicin-triggered apoptosis, deficient induction of DNA damage, and deficient caspases activation after doxorubicin in doxorubicin-resistant Nalm6, suggesting that DNA-PK is involved in insufficient tumor cell kill and in deficient caspases activation in doxorubicin- and apoptosis-resistant leukemia cells.
DNA-PK interacts with NHEJ mechanism (Collis et al., 2005). NHEJ is the predominant pathway for DNA-DSB-repair in mammalian cells (Lieber, 1999; Smith and Jackson, 1999; Durocher and Jackson, 2001; Christmann et al., 2003). NHEJ involves the XRCC4-DNA-ligase IV complex and the DNA-PK holoenzyme, consisting of the DNA binding heterodimer KU70/KU80 and the catalytic subunit DNA-PK (Grawunder et al., 1998a,b; Lieber, 1999; Smith and Jackson, 1999; Durocher and Jackson, 2001; Christmann et al., 2003). Cell lines deficient in any of these genes are generally highly sensitive to ionizing radiation and have marked deficiencies in DNA-DSB-repair (Riballo et al., 1999; Belenkov et al., 2002; Adachi et al., 2003; Holgersson et al., 2003; Collis et al., 2005; Deriano et al., 2005). Doxorubicin induced DNA damage in the cell line deficient in DNA-PK. Induction of DNA damage was strongly reduced after doxorubicin treatment in DNA-DSB-repair–proficient DNA-PK (+/+) cells at identical concentrations and at same time points, suggesting that doxorubicin-induced DNA damage was repaired in cell lines with intact DNA-PK. DNA-PK (–/–) cells are highly apoptosis-sensitive and DNA-PK (+/+) cells are apoptosis and doxorubicin resistant. In addition, doxorubicin induced activation of caspase-3 and caspase-8 in DNA-PK (–/–)-deficient cells, and caspases activation is lacking after doxorubicin treatment in DNA-PK (+/+) cells. Inhibition of DNA-PK reversed deficient caspases activation in DNA-PK (+/+) cells. These data suggest that caspases activation by doxorubicin depends on DNA-PK in cancer cells.
DNA-ligase IV is an key enzyme in NHEJ-DNA-DSB-repair (Lieber, 1999). Doxorubicin strongly induced apoptosis and caspases activation in DNA-ligase IV (–/–) cells similar to DNA-PK (–/–) cells. DNA-ligase IV (+/+) cells are apoptosis resistant to doxorubicin, and caspases activation is lacking after doxorubicin treatment similar to DNA-PK (+/+) cells, suggesting that not only DNA-PK exclusively but also ligase IV as a key enzyme for NHEJ-DNA-DSB-repair plays an important role in deficient caspases activation in cancer cells. On the basis of these results, we have developed a radioactive antibody that breaks chemo- and radioresistance by overcoming NHEJ-DNA-DSB-repair (Friesen et al., 2007).
NHEJ is involved in surviving topoisomerase II-mediated DNA damage (Malik et al., 2006). Doxorubicin intercalates with DNA and interacts with DNA-topoisomerase-II (topo-II) (Bodley et al., 1998). Mutated topo-II plays a critical role in anthracycline resistance such as doxorubicin resistance. In doxorubicin-resistant Nalm6 cells, doxorubicin resistance, and deficient caspases activation are highly unlikely attribute to topo-II mutation because wortmannin, a specific inhibitor of the phosphatidylinositol 3-kinase including DNA-PK, ATM, and ATR (Durocher and Jackson, 2001), reversed the phenotype in doxorubicin-resistant Nalm6 cells. This suggests that NHEJ plays a critical role in deficient caspases activation by doxorubicin in doxorubicin- and apoptosis-resistant cancer cells independently of topo-II mutation.
Caspases activation was inhibited by z-VAD-fmk a known inhibitor for caspases activation (Friesen et al., 2003). z-VAD-fmk blocked doxorubicin-induced apoptosis and caspases activation in doxorubicin and apoptosis-sensitive Nalm6 cells. Caspases activation and induction of apoptosis by doxorubicin were also inhibited by z-VAD-fmk in DNA-PK (–/–) and DNA-ligase IV (–/–)-deficient cell lines (unpublished data). This demonstrates that doxorubicin-induced apoptosis depends on caspases activation in anticancer drug-sensitive as well as in repair-deficient cell lines. DNA damage was induced by doxorubicin in doxorubicin-sensitive cells as well as in DSB-repair deficient DNA-PK (–/–) cells and DNA-ligase IV (–/–) cells before caspases were activated and apoptosis was detected. In contrast to induction of apoptosis, induction of DNA damage was not inhibited by blockade of caspases activation with z-VAD-fmk in doxorubicin-sensitive Nalm6 cells as well as in DNA-DSB-repair–deficient DNA-PK (–/–) cells and DNA-ligase IV (–/–) cells, indicating that induction of DNA damage is independent of caspases activation, but it depends on the NHEJ-DNA-DSB-repair. In addition, this suggests that caspases activation depends on induction of DNA damage left unrepaired by NHEJ-DNA-DSB-repair, and DNA-DSBs might be the initiator for caspases activation.
Collectively, we found that caspases activation depends on induction of DNA damage. DNA damage, which is left unrepaired in the NHEJ-DNA-DSB-repair pathway, seems to be the initiator for caspases activation and induction of apoptosis. Increasing NHEJ-DNA-DSB-repair leads to failure of induction of DNA damage, and it is responsible for deficient caspases activation in apoptosis-resistant and doxorubicin-resistant cancer cells. Modulation of NHEJ-DNA-DSB-repair restores caspases activation by doxorubicin in previously doxorubicin-resistant cells. Overcoming NHEJ-DNA-DBS-repair might offer promising strategies to enhance caspases dependent apoptosis-mediated tumor cell kill.
|
ACKNOWLEDGMENTS |
|---|
|
Footnotes |
|---|
Address correspondence to: Claudia Friesen (claudia.friesen{at}uni-ulm.de)
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Belenkov, A. I., Paiement, J. P., Panasci, L. C., Monia, B. P., and Chow, T. Y. (2002). An antisense oligonucleotide targeted to human Ku86 messenger RNA sensitizes M059K malignant glioma cells to ionizing radiation, bleomycin, and etoposide but not DNA cross-linking agents. Cancer Res 62, 5888–5896.
Bodley, A., Liu, L., Israel, M., Seshadri, R., Koseki, Y., Giuliani, F., Kirschenbaum, S., Silber, R., and Potmesil, M. (1998). DNA topoisomerase II-mediated interaction of doxorubicin and daunorubicin congeners with DNA. Cancer Res 48, 5969–5978.
Carbonari, M., Cibati, M., Cherchi, M., Sbarigia, D., Pesce, A. M., Dell'Ann, L., Modica, A., and Fiorelli, M. (1994). Detection and characterization of apoptotic peripheral blood lymphocytes in human immunodeficiency virus-infection and cancer chemotherapy by a novel flow immunocytometric method. Blood 83, 1268–1277.
Christmann, M., Tomicic, M. T., Roos, W. P., and Kaina, B. (2003). Mechanisms of human DNA repair: an update. Toxicology 193, 3–34.[CrossRef][Medline]
Collis, S. J., DeWeese, T. L., Jeggo, P. A., and Parker, A. R. (2005). The life and death of DNA-PK. Oncogene 24, 949–961.[CrossRef][Medline]
Cromie, G. A., Connelly, J. C., and Leach, D. R. (2001). Recombination at double-strand breaks and DNA end: conserved mechanisms from phage to humans. Mol. Cell 8, 1163–1174.[CrossRef][Medline]
Deriano, L. et al. (2005). Human chronic lymphocytic leukemia B cells can escape DNA damage-induced apoptosis through the nonhomologous end-joining DNA repair pathway. Blood 105, 4776–4783.
Deveraux, Q. L., Leo, E., Stennicke, H. R., Welsh, K., Salvesen, G. S., and Reed, J. (1999). Cleavage of human inhibitors of apoptosis protein XIAP results in fragments with distinct specificities for caspases. EMBO J 18, 5242–5251.[CrossRef][Medline]
Durocher, D., and Jackson, S. P. (2001). DNA-PK, ATM and ATR as sensors of DNA-damage: variations on a theme? Curr. Opin. Cell Biol 13, 225–231.[CrossRef][Medline]
Friesen, C., Fulda, S., and Debatin, K. M. (1997). Deficient activation of the CD95 (APO-1/Fas) system in drug-resistant cells. Leukemia 11, 1833–1841.[CrossRef][Medline]
Friesen, C., Fulda, S., and Debatin, K. M. (1999a). Cytotoxic drugs and the CD95 pathway. Leukemia 13, 1854–1858.[CrossRef][Medline]
Friesen, C., Fulda, S., and Debatin, K. M. (1999b). Induction of CD95 ligand and apoptosis by doxorubicin is modulated by the redox state in chemosensitive- and drug-resistant tumor cells. Cell Death Differ 6, 471–480.[CrossRef][Medline]
Friesen, C., Glatting, G., Koop, B., Schwarz, K., Morgenstern, A., Apostolidis, C., Debatin, K. M., and Reske, S. N. (2007). Breaking chemo- and radioresistance with [213Bi]anti-CD45 antibodies in leukemia cells. Cancer Res 67, 1950–1958.
Friesen, C., Herr, I., Krammer, P. H., and Debatin, K. M. (1996). Involvement of the CD95 (APO-1/FAS) receptor/ligand system in drug-induced apoptosis in leukemia cells. Nat. Med 2, 574–577.[CrossRef][Medline]
Friesen, C., Kiess, Y., and Debatin, K. M. (2004). A critical role of glutathione in determining apoptosis sensitivity and resistance in leukemia cells. Cell Death Differ 11, 73–85.[CrossRef]
Friesen, C., Lubatschofski, A., Kotzerke, J., Buchmann, I., Reske, S., and Debatin, K. M. (2003). beta-Irradiation used for systemic radioimmunotherapy induces apoptosis and activates apoptosis pathways in leukemia cells. Eur. J. Nucl. Med 30, 1251–1261.[CrossRef]
Grawunder, U., Zimmer, D., Fugmann, S., Schwarz, K., and Lieber, M. R. (1998a). DNA ligase IV is essential for V(D)J recombination and DNA double-strand break repair in human precursor lymphocytes. Mol. Cell 2, 477–484.[CrossRef][Medline]
Grawunder, U., Zimmer, D., Kulesza, P., and Lieber, M. R. (1998b). Requirement for an interaction of XRCC4 with DNA ligase IV for wild-type V(D)J recombination and DNA double-strand break repair in vivo. J. Biol. Chem 273, 24708–24714.
Haber, J. E. (2000). Partners and pathways repairing double-strand break. Trends Genet 16, 259–264.[CrossRef][Medline]
Hartmann, A. et al. (2003). Recommendations for conducting the in vivo alkaline Comet assay. Mutagenesis 18, 45–51.
Holgersson, A., Jernberg, A. R., Persson, L. M., Edgren, M. R., Lewensohn, R., Nilsson, A., Brahme, A., and Meijer, A. E. (2003). Low and high LET radiation-induced apoptosis in M059J and M059K cells. Int. J. Radiat. Biol 79, 611–621.[CrossRef][Medline]
Jeggo, P. A. (1998a). DNA breakage and repair. Adv. Genet 38, 185–218.[Medline]
Jeggo, P. A. (1998b). Identification of genes involved in repair of DNA double-strand breaks in mammalian cells. Radiat. Res 150, S80–S91.[Medline]
Kaina, B. (2003). DNA damage-triggered apoptosis: critical role of DNA repair, double-strand breaks, cell proliferation and signaling. Biochem. Pharmacol 66, 1547–1554.[CrossRef][Medline]
Kaufmann, S. H., and Earnshaw, W. C. (2000). Induction of apoptosis by cancer chemotherapy. Exp. Cell Res 256, 42–49.[CrossRef][Medline]
Kroemer, G. (1997). The proto-oncogene Bcl-2 and its role in regulating apoptosis. Nat. Med 3, 614–620.[CrossRef][Medline]
Lieber, M. R. (1999). The biochemistry and biological significance of nonhomologous DNA end joining: an essential repair process in multicellular eukaryotes. Genes Cells 4, 77–85.[Abstract]
Los, M., Herr, I., Friesen, C., Fulda, S., Schulze-Osthoff, K., and Debatin, K.-M. (1997). Cross-resistance of CD95- and drug-induced apoptosis as a consequence of deficient activation of caspases (ICE/Ced-3 proteases). Blood 90, 3118–3129.
Malik, M., Nitiss, K., Enriquez-Rios, V., and Nitiss, J. (2006). Roles of nonhomologous end-joining pathways in surviving topoisomerase II-mediated DNA damage. Mol. Cancer Ther 5, 1405–1414.
Nicoletti, I., Migliorati, G., Pagliacci, M. C., Grignani, F., and Riccardi, C. (1991). A rapid and simple method for measuring thymocyte apoptosis by propidium iodide staining and flow cytometry. J. Immunol. Methods 139, 271–279.[CrossRef][Medline]
Olive, P. L. (1998). The role of DNA single- and double-strand breaks in cell killing by ionizing radiation. Radiat. Res 150, 42–51.[CrossRef]
Posovszky, C., Friesen, C., Herr, I., and Debatin, K. M. (1999). Chemotherapeutic drugs sensitize pre-B ALL cells for CD95- and cytotoxic T-lymphocyte-mediated apoptosis. Leukemia 13, 400–409.[CrossRef][Medline]
Riballo, E. et al. (1999). Identification of defect in DNA ligase IV in radiosensitive leukaemia patient. Curr. Biol 9, 699–702.[CrossRef][Medline]
Rich, T., Allen, R. L., and Wyllie, A. H. (2000). Defying death after DNA damage. Nature 407, 777–783.[CrossRef][Medline]
Slapak, C. A., Daniel, J. C., and Levy, S. B. (1990). Sequential emergence of distinct resistance Phenotypes in murine erythroleukemia cells under adriamycin selection: decreased anthracycline uptake precedes increased P-glycoprotein expression. Cancer Res 50, 7895–7901.
Smith, G. C., and Jackson, S. P. (1999). The DNA-dependent protein kinase. Genes Dev 13, 916–934.
Srinivasula, S. M. et al. (2001). A conserved XIAP-interaction in caspase-9 and Smac/DIABLO regulates caspase activity and apoptosis. Nature 410, 112–116.[CrossRef][Medline]
Willmore, E., de Caux, S., Sunter, N. J., Tilby, M. J., Jackson, G. H., Austin, C. A., and Durkacz, B. W. (2004). A novel DNA-dependent protein kinase inhibitor, NU7026, potentiates the cytotoxicity of topoisomerase II poisons used in treatment of leukemia. Blood 103, 4659–4665.
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
18 kDa, the active fragment of caspase-9 at 
