The ratios of phosphoprotein to total protein are shown for pAkt ser473, pS6 and p4EBP1 (bottom). Three xenografts from each treatment group are shown. E, Western blot analyses of protein lysates from xenografts harvested 1hr after the last treatment (top). D, Body weight change over the duration of the experiment. C, LNCaP95 tumor growth in castrated mice administered vehicle, a half-dose of EPI (100mg/kg body weight), BEZ (5mg/kg body weight) or combination daily by oral gavage for two weeks. Data for “A” and “B” are shown as ratio to vehicle control. Proliferation was assessed by BrdU incorporation. B, LNCaP95 cells were also treated with everolimus (EVE, 10 nM) instead with BEZ235. Proliferation was measured by BrdU incorporation. not statistically significant *p < 0.05 ** p < 0.01 *** p < 0.001 ****p < 0.0001 # p < 0.01 # p < 0.001 # p < 0.0001.Ī, LNCaP95 cells were treated with DMSO, EPI, ENZ, BEZ or combination of EPI and BEZ for 1hr prior to the addition of R1881 (0.1 nM) for 48 hr. One-Way ANOVA, post-hoc Turkey’s multiple comparisons test. Data is presented as the mean ± SEM from three independent experiments. Luciferase activities are shown as percentage of vehicle control. LNCaP95 cells were harvested 24hr after the treatment of indicated compound (right). EPI, BEZ, or combination of EPI and BEZ were added 1hr before addition of IL-6 (50 ng/ml) or FSK (50 uM) in LNCaP cells and harvested after 24hr. D, Transactivation assays of the AR NTD were performed in LNCaP (Left and middle) and LNCaP95 cells (right) cotransfected with p5xGal4UAS-TATA-luciferase and AR NTD-Gal4DBD. Ectopic protein levels of AR-V7 and AR-V567es in Cos-1 cells are shown relative to endogenous levels of FL-AR in LNCaP cells (right). C, Cos-1 cells transfected with PB-luciferase and expression vectors for AR-V567es or AR-V7 were treated with DMSO, EPI, BEZ or combination of EPI and BEZ (left). LNCaP95 cells transfected with PSA-luciferase reporter were also treated with everolimus (10 nM) or combination with ENZ or EPI (A, right panel). LNCaP95 (A) and LNCaP (B) cells transfected with PSA-, ARR3- or PB-luciferase reporters were treated with DMSO, EPI, ENZ, BEZ or combination for 1hr prior to the addition of R1881 for 48hr. ©2015 American Association for Cancer Research. A combination of EPI-002 and BEZ235 decreased the growth of LNCaP95 cells in vitro and in vivoĬotargeting mTOR and AR-NTD to block transcriptional activities of FL-AR and AR-Vs provided maximum antitumor efficacy in PTEN-null, enzalutamide-resistant CRPC. Inhibition of mTOR provided additional blockade of UBE2C expression. Gene expression and efficacy were examined in vitro and in vivoĮPI-002 had antitumor activity in enzalutamide-resistant LNCaP95 cells that was associated with decreased expression of AR-V target genes (e.g., UBE2C). To determine the functional roles of FL-AR, AR-Vs, and PI3K/Akt/mTOR pathways, we employed EPI-002 or enzalutamide and BEZ235 (low dose) or everolimus in human prostate cancer cells that express FL-AR or FL-AR and AR-Vs (LNCaP95). Here, we evaluated the efficacy and mechanism of combination therapy to block mTOR activity together with EPI-002, an AR N-terminal domain (NTD) antagonist that blocks the transcriptional activities of FL-AR and AR-Vs in models of CRPC. It is currently unknown how blocking the PI3K/Akt/mTOR pathway impacts prostate cancer driven by AR-Vs. However, truncated AR-splice variants (AR-V) that are constitutively active and dominant over FL-AR are associated with tumor progression and resistance mechanisms in CRPC. Therefore, cotargeting full-length (FL) AR and PI3K/Akt/mTOR signaling has been proposed as a possible, more effective therapeutic approach for CRPC.

Transcriptionally active androgen receptor (AR) plays a role in the majority of CRPCs. Retrieved 5 February 2006.The PI3K/Akt/mTOR pathway is activated in most castration-resistant prostate cancers (CRPC). Archived from the original on 10 January 2008. Archived from the original on 10 February 2011. Archived from the original on 5 January 2006. Archived from the original on 31 December 2005. 'A Ukrainian child porn ring exposed'.Archived from the original on 10 August 2004. 'Police Shut Ukraine Model Agency in Porn Crackdown'.Archived from the original on 18 October 2007. ^ 'Criminal group involves 1,500 underage Ukrainian girls in porn business with parents' knowledge'.Archived from the original on 7 April 2006.

^ '2005 Country Report on Human Rights Practices in Ukraine'.

^ a b 'Police Shut Ukraine Model Agency in Porn Crackdown'.

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