Project description:Our previous studies of proteomic-coupled-network analysis of AR protein interaction complexes (Paliouras et al., Integrative Biology, 2011) identified a number of proteins involved in RNA metabolism, specifically alternative RNA splicing. We selected two interacting RNA splicing proteins, SAM68 and DDX5 to examine RNA splicing events in prostate cancer (PCa). This analysis suggests a much more robust effect on RNA splicing with AR dictating either an exon-inclusion or -exclusion pathway. To establish the true physiological roles of AR in alternative RNA splicing, we opted to further examine the changes in global splicing profiles of LNCaP PCa cells, stimulated with and without androgens in conjunction with overexpression studies of SAM68 and DDX5.
Project description:Rapid immunoprecipitation mass spectrometry of endogenous protein (RIME) was conducted to examine interactome of androgen receptor (AR) in LNCaP cells.
Project description:G-1 is an agonist to GPR30. Activation of GPR30 by G-1 inhibited prostate cancer cell growth in LNCaP xenografts regrown after catration of the host (nude mice), but not in the androgen-sensitive LNCaP xenograft grown in an intact host. Results provide insights into the molecular basis of G-1 action in castration-resistant prostate cancer.
Project description:G-1 is an agonist to GPR30. Activation of GPR30 by G-1 inhibited prostate cancer cell growth in LNCaP xenografts regrown after catration of the host (nude mice), but not in the androgen-sensitive LNCaP xenograft grown in an intact host. Results provide insights into the molecular basis of G-1 action in castration-resistant prostate cancer. Male nude mice were injected with LNCaP cells. When the LNCaP tumors reached 150–300 mm3, mice were divided into two groups: intact (androgen-sensitive tumor) and castrated. For the intact group, mice were subcutaneously injected with vehicle alone (95% PBS, 2.5% DMSO, 2.5% ethanol) or G-1 (4 mg/kg/day in vehicle) daily for 16 days. For the castrated group, tumors regressed and then regrew to ~300-400mm3. Mice were treated daily with vehicle or G-1 as described for 16 days. Tumors were harvested for RNA extraction and microarray experiments.
Project description:Aberrant androgen receptor (AR)-mediated transcription is a critical driver in progression of human prostate cancer. It's known that different doses of androgens can elicit differential transcriptional and proliferative responses in prostate-tumor cells. Here, we set out to examine the androgenic regulation of glycoprotein expression in the membrane fraction of prostate-tumor cells that could serve as mediators or markers of androgen-induced proliferative responses observed in prostate-tumor cells. A bioanalytical workflow involving lectin-affinity chromatography and label-free quantitative mass spectrometry was used to identify androgen-sensitive glycomembrane protein expression associated with androgen-mediated proliferation. This study would facilitate the identification of surface membrane proteins involved in androgen-mediated proliferation and provide potential therapeutic targets in the detection treatment of proliferation prostate-tumors.
Project description:Androgens are required for the development of normal prostate, and they are also linked to the development of prostate cancer. We used microarrays to understand the role of androgen in an androgen dependent, androgen receptor (AR) positive human metastatic cell line, LNCaP.
Project description:We generated and characterized an androgen-independent LNCaP-AI cell line by long-term culture of androgen-dependent LNCaP cells in RPMI-1640 medium containing charcoal-stripped serum. This approach used to generate the line mimics the castration resistant condition for treating prostate cancer, supporting the relevance of the LNCAP-AI cell line to Castration Resistant Prostate Cancer.
Project description:Androgens are required for the development of normal prostate, and they are also linked to the development of prostate cancer. We used microarrays to understand the role of androgen in an androgen dependent, androgen receptor (AR) positive human metastatic cell line, LNCaP. LNCaP cells were grown in RPMI medium and they were subjected to stay in phenol-red free, RPMI with charcoal stripped serum for 48h. Synthetic androgen R1881 was added and the cells were allowed to grow for 48h. Control cells were given with corresponding amount of ethanol as vehicle which is used for the solubilization of R1881. Cells were harvested and RNA was isolated for microarray analysis.
Project description:Gedunin is a natural product that affects LNCaP androgen-signalling by 24h; We used microarrays to detail the androgen-responsive program of gene expression affected by gedunin treatment at 24h Experiment Overall Design: LNCaP cells were grown to 50% confluency and deprived of androgen, and subsequently treated with gedunin plus androgen, androgen alone, or vehicle alone for 24h prior to direct Trizol lysis and RNA isolation
Project description:celastrol is a natural product that affects LNCaP androgen-signalling by 24h; We used microarrays to detail the androgen-responsive program of gene expression affected by celastrol treatment at 24h Experiment Overall Design: LNCaP cells were grown to 50% confluency and deprived of androgen, and subsequently treated with celastrol plus androgen, androgen alone, or vehicle alone for 24h prior to direct Trizol lysis and RNA isolation