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The role of reactive oxygen species and subsequent DNA-damage response in the emergence of resistance towards resveratrol in colon cancer models.

Colin DJ, Limagne E, Ragot K, Lizard G, Ghiringhelli F, Solary É, Chauffert B, Latruffe N, Delmas D - Cell Death Dis (2014)

Bottom Line: Ultimately, stable resistant populations towards RSV displaying higher degrees of ploidy and macronucleation as compared to parental cells emerged.We linked these transient effects and resistance emergence to the abilities of these cells to progressively escape RSV-induced DNA damage.This study provides a pre-clinical analysis of the long-term effects of RSV and highlights ROS as main agents in RSV's indirect DNA-damaging properties and consequences in terms of anticancer response and potent resistance emergence.

View Article: PubMed Central - PubMed

Affiliation: 1] Université de Bourgogne, Dijon F-21000, France [2] Centre de Recherche Inserm U866-Lipids, Nutrition, Cancers, Dijon F-21000, France [3] EA7270-Bio-PeroxIL Biochimie du peroxysome, inflammation et métabolisme lipidique, Dijon F-21000, France.

ABSTRACT
In spite of the novel strategies to treat colon cancer, mortality rates associated with this disease remain consistently high. Tumour recurrence has been linked to the induction of resistance towards chemotherapy that involves cellular events that enable cancer cells to escape cell death. Treatment of colon cancer mainly implicates direct or indirect DNA-damaging agents and increased repair or tolerances towards subsequent lesions contribute to generate resistant populations. Resveratrol (RSV), a potent chemosensitising polyphenol, might share common properties with chemotherapeutic drugs through its indirect DNA-damaging effects reported in vitro. In this study, we investigated how RSV exerts its anticancer effects in models of colon cancer with a particular emphasis on the DNA-damage response (DDR; PIKKs-Chks-p53 signalling cascade) and its cellular consequences. We showed in vitro and in vivo that colon cancer models could progressively escape the repeated pharmacological treatments with RSV. We observed for the first time that this response was correlated with transient activation of the DDR, of apoptosis and senescence. In vitro, a single treatment with RSV induced a DDR correlated with S-phase delay and apoptosis, but prolonged treatments led to transient micronucleations and senescence phenotypes associated with polyploidisation. Ultimately, stable resistant populations towards RSV displaying higher degrees of ploidy and macronucleation as compared to parental cells emerged. We linked these transient effects and resistance emergence to the abilities of these cells to progressively escape RSV-induced DNA damage. Finally, we demonstrated that this DNA damage was triggered by an overproduction of reactive oxygen species (ROS) against which cancer cells could adapt under prolonged exposure to RSV. This study provides a pre-clinical analysis of the long-term effects of RSV and highlights ROS as main agents in RSV's indirect DNA-damaging properties and consequences in terms of anticancer response and potent resistance emergence.

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Resveratrol induces a transient activation of the DNA-damage response invivo. (a) Effects of daily oral administrations of200 mg/kg of RSV on the growth of SW620 tumours in vivo. SW620cells were subcutaneously injected at day 0 to nude mice and the treatments withRSV or with the vehicle (Ctl) began at day 2. Data are medians of tumour volumesof 9–12 mice per group±the first and third quartile. Statisticalanalyses were performed by the Mann–Whitney's test and are shown assignificant with P<0.05 (*) and P<0.01 (**).(b, c) Levels of expression and phosphorylation of the componentsof the DNA-damage response were analysed by immunoblotting using the specifiedantibodies. Analyses were performed on SW620 tumours extracts collected from fivemice per group after 11 (b) and 15 days (c) of daily treatments with200 mg/kg of RSV (R200) and compared to control tumours (Ctl).Quantifications by densitometry are plotted as medians±the first and thirdquartile. Statistical analyses were performed by the Mann–Whitney'stest and are shown as significant with P<0.05 (*) andP<0.01 (**)
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fig3: Resveratrol induces a transient activation of the DNA-damage response invivo. (a) Effects of daily oral administrations of200 mg/kg of RSV on the growth of SW620 tumours in vivo. SW620cells were subcutaneously injected at day 0 to nude mice and the treatments withRSV or with the vehicle (Ctl) began at day 2. Data are medians of tumour volumesof 9–12 mice per group±the first and third quartile. Statisticalanalyses were performed by the Mann–Whitney's test and are shown assignificant with P<0.05 (*) and P<0.01 (**).(b, c) Levels of expression and phosphorylation of the componentsof the DNA-damage response were analysed by immunoblotting using the specifiedantibodies. Analyses were performed on SW620 tumours extracts collected from fivemice per group after 11 (b) and 15 days (c) of daily treatments with200 mg/kg of RSV (R200) and compared to control tumours (Ctl).Quantifications by densitometry are plotted as medians±the first and thirdquartile. Statistical analyses were performed by the Mann–Whitney'stest and are shown as significant with P<0.05 (*) andP<0.01 (**)

Mentions: These results correlating transient DDR, apoptosis and senescence activations byRSV led us to explore the response of a more standard model of human CCR xenograftin Nude mice. We used the SW620 model which has already exhibited sensitivitytowards RSV in vitro (Supplementary TableS1). In contrast to the PROb model, RSV treatment led to a growthdelay of SW620 tumours (Figure 3a), showing themodel-dependent macroscopic effects of RSV. Interestingly, the extent of thisdelay appeared to decrease with time according to the statistical analysis. Webiochemically characterised the response of this model and found a similar trendin terms of DDR induction as compared with the PROb model (Figures 3b and c). Indeed, all the markers described earlier wereinduced in RSV-treated tumours, including the senescence p16 and apoptosiscaspase-3 markers. Importantly, after 15 days of treatment, most of theseinductions were compromised, which agreed with what we found in the PRObmodel.


The role of reactive oxygen species and subsequent DNA-damage response in the emergence of resistance towards resveratrol in colon cancer models.

Colin DJ, Limagne E, Ragot K, Lizard G, Ghiringhelli F, Solary É, Chauffert B, Latruffe N, Delmas D - Cell Death Dis (2014)

Resveratrol induces a transient activation of the DNA-damage response invivo. (a) Effects of daily oral administrations of200 mg/kg of RSV on the growth of SW620 tumours in vivo. SW620cells were subcutaneously injected at day 0 to nude mice and the treatments withRSV or with the vehicle (Ctl) began at day 2. Data are medians of tumour volumesof 9–12 mice per group±the first and third quartile. Statisticalanalyses were performed by the Mann–Whitney's test and are shown assignificant with P<0.05 (*) and P<0.01 (**).(b, c) Levels of expression and phosphorylation of the componentsof the DNA-damage response were analysed by immunoblotting using the specifiedantibodies. Analyses were performed on SW620 tumours extracts collected from fivemice per group after 11 (b) and 15 days (c) of daily treatments with200 mg/kg of RSV (R200) and compared to control tumours (Ctl).Quantifications by densitometry are plotted as medians±the first and thirdquartile. Statistical analyses were performed by the Mann–Whitney'stest and are shown as significant with P<0.05 (*) andP<0.01 (**)
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4260744&req=5

fig3: Resveratrol induces a transient activation of the DNA-damage response invivo. (a) Effects of daily oral administrations of200 mg/kg of RSV on the growth of SW620 tumours in vivo. SW620cells were subcutaneously injected at day 0 to nude mice and the treatments withRSV or with the vehicle (Ctl) began at day 2. Data are medians of tumour volumesof 9–12 mice per group±the first and third quartile. Statisticalanalyses were performed by the Mann–Whitney's test and are shown assignificant with P<0.05 (*) and P<0.01 (**).(b, c) Levels of expression and phosphorylation of the componentsof the DNA-damage response were analysed by immunoblotting using the specifiedantibodies. Analyses were performed on SW620 tumours extracts collected from fivemice per group after 11 (b) and 15 days (c) of daily treatments with200 mg/kg of RSV (R200) and compared to control tumours (Ctl).Quantifications by densitometry are plotted as medians±the first and thirdquartile. Statistical analyses were performed by the Mann–Whitney'stest and are shown as significant with P<0.05 (*) andP<0.01 (**)
Mentions: These results correlating transient DDR, apoptosis and senescence activations byRSV led us to explore the response of a more standard model of human CCR xenograftin Nude mice. We used the SW620 model which has already exhibited sensitivitytowards RSV in vitro (Supplementary TableS1). In contrast to the PROb model, RSV treatment led to a growthdelay of SW620 tumours (Figure 3a), showing themodel-dependent macroscopic effects of RSV. Interestingly, the extent of thisdelay appeared to decrease with time according to the statistical analysis. Webiochemically characterised the response of this model and found a similar trendin terms of DDR induction as compared with the PROb model (Figures 3b and c). Indeed, all the markers described earlier wereinduced in RSV-treated tumours, including the senescence p16 and apoptosiscaspase-3 markers. Importantly, after 15 days of treatment, most of theseinductions were compromised, which agreed with what we found in the PRObmodel.

Bottom Line: Ultimately, stable resistant populations towards RSV displaying higher degrees of ploidy and macronucleation as compared to parental cells emerged.We linked these transient effects and resistance emergence to the abilities of these cells to progressively escape RSV-induced DNA damage.This study provides a pre-clinical analysis of the long-term effects of RSV and highlights ROS as main agents in RSV's indirect DNA-damaging properties and consequences in terms of anticancer response and potent resistance emergence.

View Article: PubMed Central - PubMed

Affiliation: 1] Université de Bourgogne, Dijon F-21000, France [2] Centre de Recherche Inserm U866-Lipids, Nutrition, Cancers, Dijon F-21000, France [3] EA7270-Bio-PeroxIL Biochimie du peroxysome, inflammation et métabolisme lipidique, Dijon F-21000, France.

ABSTRACT
In spite of the novel strategies to treat colon cancer, mortality rates associated with this disease remain consistently high. Tumour recurrence has been linked to the induction of resistance towards chemotherapy that involves cellular events that enable cancer cells to escape cell death. Treatment of colon cancer mainly implicates direct or indirect DNA-damaging agents and increased repair or tolerances towards subsequent lesions contribute to generate resistant populations. Resveratrol (RSV), a potent chemosensitising polyphenol, might share common properties with chemotherapeutic drugs through its indirect DNA-damaging effects reported in vitro. In this study, we investigated how RSV exerts its anticancer effects in models of colon cancer with a particular emphasis on the DNA-damage response (DDR; PIKKs-Chks-p53 signalling cascade) and its cellular consequences. We showed in vitro and in vivo that colon cancer models could progressively escape the repeated pharmacological treatments with RSV. We observed for the first time that this response was correlated with transient activation of the DDR, of apoptosis and senescence. In vitro, a single treatment with RSV induced a DDR correlated with S-phase delay and apoptosis, but prolonged treatments led to transient micronucleations and senescence phenotypes associated with polyploidisation. Ultimately, stable resistant populations towards RSV displaying higher degrees of ploidy and macronucleation as compared to parental cells emerged. We linked these transient effects and resistance emergence to the abilities of these cells to progressively escape RSV-induced DNA damage. Finally, we demonstrated that this DNA damage was triggered by an overproduction of reactive oxygen species (ROS) against which cancer cells could adapt under prolonged exposure to RSV. This study provides a pre-clinical analysis of the long-term effects of RSV and highlights ROS as main agents in RSV's indirect DNA-damaging properties and consequences in terms of anticancer response and potent resistance emergence.

Show MeSH
Related in: MedlinePlus