Irene L Ibañez

Publications (4)20.42 Total impact

• Article: Phosphorylation and Subcellular Localization of p27Kip1 Regulated by Hydrogen Peroxide Modulation in Cancer Cells.

  Irene L Ibañez, Candelaria Bracalente, Cintia Notcovich, Ivanna Tropper, Beatriz L Molinari, Lucía L Policastro, Hebe Durán
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  ABSTRACT: The Cyclin-dependent kinase inhibitor 1B (p27Kip1) is a key protein in the decision between proliferation and cell cycle exit. Quiescent cells show nuclear p27Kip1, but this protein is exported to the cytoplasm in response to proliferating signals. We recently reported that catalase treatment increases the levels of p27Kip1 in vitro and in vivo in a murine model. In order to characterize and broaden these findings, we evaluated the regulation of p27Kip1 by hydrogen peroxide (H(2)O(2)) in human melanoma cells and melanocytes. We observed a high percentage of p27Kip1 positive nuclei in melanoma cells overexpressing or treated with exogenous catalase, while non-treated controls showed a cytoplasmic localization of p27Kip1. Then we studied the levels of p27Kip1 phosphorylated (p27p) at serine 10 (S10) and at threonine 198 (T198) because phosphorylation at these sites enables nuclear exportation of this protein, leading to accumulation and stabilization of p27pT198 in the cytoplasm. We demonstrated by western blot a decrease in p27pS10 and p27pT198 levels in response to H(2)O(2) removal in melanoma cells, associated with nuclear p27Kip1. Melanocytes also exhibited nuclear p27Kip1 and lower levels of p27pS10 and p27pT198 than melanoma cells, which showed cytoplasmic p27Kip1. We also showed that the addition of H(2)O(2) (0.1 µM) to melanoma cells arrested in G1 by serum starvation induces proliferation and increases the levels of p27pS10 and p27pT198 leading to cytoplasmic localization of p27Kip1. Nuclear localization and post-translational modifications of p27Kip1 were also demonstrated by catalase treatment of colorectal carcinoma and neuroblastoma cells, extending our findings to these other human cancer types. In conclusion, we showed in the present work that H(2)O(2) scavenging prevents nuclear exportation of p27Kip1, allowing cell cycle arrest, suggesting that cancer cells take advantage of their intrinsic pro-oxidant state to favor cytoplasmic localization of p27Kip1.
  PLoS ONE 01/2012; 7(9):e44502. · 4.09 Impact Factor
• Article: H2O2 scavenging inhibits G1/S transition by increasing nuclear levels of p27KIP1.

  Irene L Ibañez, Lucía L Policastro, Ivanna Tropper, Candelaria Bracalente, Mónica A Palmieri, Paola A Rojas, Beatriz L Molinari, Hebe Durán
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  ABSTRACT: The aim of the present study was to evaluate cell cycle regulation by scavenging H(2)O(2) in tumor cells. A significant arrest in the G1 phase of the cell cycle was demonstrated in CH72-T4 carcinoma cells exposed to catalase, associated with a decrease in cyclin D1 and an increase in the CDK inhibitory protein p27(KIP1). Moreover, we found a differential intracellular distribution of p27(KIP1), which remained in the nucleus after catalase treatment. In vivo experiments showed an increase in nuclear levels of p27(KIP1) associated with the inhibition of tumor growth by H(2)O(2) scavenging, confirming in vitro results. To conclude, H(2)O(2) scavenging may induce cell cycle arrest through the modulation of cyclin D1 and p27(KIP1) levels and nuclear localization of p27(KIP1). To our knowledge, this is the first report that demonstrates that the modulation of ROS alters the intracellular localization of a key regulatory protein of G1/S transition.
  Cancer letters 03/2011; 305(1):58-68. · 4.86 Impact Factor
• Source

  Available from: Jorge Davidson

  Article: Induction and rejoining of DNA double strand breaks assessed by H2AX phosphorylation in melanoma cells irradiated with proton and lithium beams.

  Irene L Ibañez, Candelaria Bracalente, Beatriz L Molinari, Mónica A Palmieri, Lucía Policastro, Andrés J Kreiner, Alejandro A Burlón, Alejandro Valda, Daniela Navalesi, Jorge Davidson, Miguel Davidson, Mónica Vázquez, Mabel Ozafrán, Hebe Durán
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  ABSTRACT: The aim of this study was to evaluate the induction and rejoining of DNA double strand breaks (DSBs) in melanoma cells exposed to low and high linear energy transfer (LET) radiation. DSBs and survival were determined as a function of dose in melanoma cells (B16-F0) irradiated with monoenergetic proton and lithium beams and with a gamma source. Survival curves were obtained by clonogenic assay and fitted to the linear-quadratic model. DSBs were evaluated by the detection of phosphorylated histone H2AX (gammaH2AX) foci at 30 min and 6 h post-irradiation. Survival curves showed the increasing effectiveness of radiation as a function of LET. gammaH2AX labeling showed an increase in the number of foci vs. dose for all the radiations evaluated. A decrease in the number of foci was found at 6 h post-irradiation for low LET radiation, revealing the repair capacity of DSBs. An increase in the size of gammaH2AX foci in cells irradiated with lithium beams was found, as compared with gamma and proton irradiations, which could be attributed to the clusters of DSBs induced by high LET radiation. Foci size increased at 6 h post-irradiation for lithium and proton irradiations in relation with persistent DSBs, showing a correlation with surviving fraction. Our results showed the response of B16-F0 cells to charged particle beams evaluated by the detection of gammaH2AX foci. We conclude that gammaH2AX foci size is an accurate parameter to correlate the rejoining of DSBs induced by different LET radiations and radiosensitivity.
  International journal of radiation oncology, biology, physics 08/2009; 74(4):1226-35. · 4.59 Impact Factor
• Article: Suppression of cancer growth by nonviral gene therapy based on a novel reactive oxygen species-responsive promoter.

  Lucía L Policastro, Irene L Ibañez, Hebe A Durán, Gastón Soria, Vanesa Gottifredi, Osvaldo L Podhajcer
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  ABSTRACT: Increased reactive oxygen species (ROS) production has been reported as a distinctive feature of different pathologies including cancer. Therefore, we assessed whether increased ROS production in the cancer microenvironment could be selectively exploited to develop a selective anticancer therapy. For this purpose, we constructed a novel chimeric promoter, based on a ROS-response motif located in the VEGF gene promoter placed, in turn, downstream of a second ROS-response motif obtained from the early growth response 1 (Egr-1) gene promoter. The activity of the chimeric promoter was largely dependent on variations in intracellular ROS levels and showed a high inducible response to exogenous H(2)O(2). Transient expression of the thymidine kinase (TK) gene driven by the chimeric promoter, followed by gancyclovir (GCV) administration, inhibited human colorectal cancer and melanoma cell growth in vitro and in vivo. Moreover, electrotransfer of the TK gene followed by GCV administration exerted a potent therapeutic effect on established tumors. This response was improved when combined with chemotherapeutic drugs. Thus, we show for the first time that a distinctive pro-oxidant state can be used to develop new selective gene therapeutics for cancer.
  Molecular Therapy 06/2009; 17(8):1355-64. · 6.87 Impact Factor