Abstract 4995: HO1 induction interrupts the glucocorticoid receptor signaling in prostate cancer cells
Authors: Leonardi Daiana B. , Paez Alejandra V. , Schuster Federico , Anselmino Nicolas , Brandani Javier N. , Abbate Mercedes , Gueron Geraldine , Cotignola Javier H. , Vazquez Elba S. 
Published: August 2015

Although initially treatable, prostate cancer can recur in a hormone refractory form that is not responsive to current available therapies. The mortality rate associated with hormone refractory prostate cancer is high, and there is an urgent need for new therapeutic agents to treat prostate cancer. In order to overcome the toxicity of chemotherapy, glucocorticoids have long been used in the treatment of hormone-resistant prostate cancer. Nevertheless, many authors reported that the glucocorticoid receptor (GR) can substitute the androgen receptor and activate a similar but distinguishable set of genes. A common feature of prostate cancer is the dependence on activated signal transducer and activator of transcription 3 (STAT3), a transcription factor for survival and a coactivator of GR signaling. We have previously demonstrated that the induction of HO1, an antioxidant and anti-inflammatory protein, increases STAT3 cytoplasmic retention, interfering with its signaling and promotes HO1 nuclear localization. Our aim is to study the effect of HO1 induction on GR signaling in prostate cancer. PC3 cells were treated with Dexamethasone (Dex) at different concentrations (10−7, 10−8 and 10−9 M) during 24 h in 10% charcoaled-serum media and were exposed to the pharmacological inducer of HO1 (hemin, 80 uM, 24 h). Control cells were treated with vehicles. No significant differences in cell viability were detected by MTT assay. In order to investigate whether HO1 interrupts the GR transcriptional activity, PC3 cells were transfected with a luciferase reporter plamid containing three glucocorticoid response elements. Transfected cells were treated either with hemin (80 uM, 24 h), Dex (10−8 M, 6 h) or the combination of both drugs. The GR transcriptional activation was confirmed when cells were treated with the agonist Dex (p&lt;0.01). HO1 induction significantly reduced either the GR basal transcriptional activity (p&lt;0.01) as well as the Dex induced (p&lt;0.01). The expression of Ikb, a GR regulated gene, was screened by RT-qPCR and was confirmed its inhibition in hemin exposed cells treated or not with dexamethasone. The repressive activity of HO1 was also demonstrated using an NFkB luciferase reporter assay. Using confocal fluorescence microscopy, GR nuclear translocation upon Dex treatment was confirmed, and a decrease of GR nuclear localization was demonstrated under hemin+Dex treatment (p&lt;0.01). These results correlate with the diminished luciferase activity of the MMTV-luc plasmid in the presence of hemin+Dex vs Dex alone. Strikingly this is the first report showing the immunoprecipitation of GR and HO1, showing physical interaction between these proteins, both in the nucleus and in the cytoplasm. In conclusion, these results provide a novel function for HO1 downmodulating GR transcriptional activity, repressing the expression of its target genes and interfering with the receptor's nuclear localization in prostate cancer cells.

Citation Format: Daiana B. Leonardi, Alejandra V. Paez, Federico Schuster, Nicolas Anselmino, Javier N. Brandani, Mercedes Abbate, Geraldine Gueron, Javier H. Cotignola, Elba S. Vazquez. HO1 induction interrupts the glucocorticoid receptor signaling in prostate cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4995. doi:10.1158/1538-7445.AM2015-4995