Project description:An isogenic prostate cancer cell model was used to profile miRNA changes that contribute to the tumorigenic state of prostate cancer. P69 cells are an SV40 large T antigen immortalized cell line that is poorly tumorigenic and non-metastatic. The M12 derivative was derived from an in vivo selection process using nude, athymic mice. Immortalized cells were cultured and used to extract RNA and probed against the human miRNA panel from Exiqon. There were a total of 186 miRNAs that were at least two fold differentially regulated . Two human prostate cancer cell lines were used to evaluate miRNA expression differences contributing to oncogenesis. Two replicates were performed and the data presented as the mean fold difference between P69 and M12.
Project description:Purpose: Next-generation sequencing (NGS) of miRNA expression in cancer cell lines provides a path for analysis of dysregulated pathways and potential treatment of dysregulation. The goals of this study are to compare NGS-derived miRNA expression data in a non-tumorigenic prostate cell line (P69) to its metastatic derivative cell lines M12 and M2182.
Project description:EGR3 expression is upregulated in human prostate cancer compared to normal prostate tissue and is associated with absence of relapse, while low EGR3 expression in tumors is predicitive of disease relapse (Pio et al., PLOS One 2013; 8(1):e54096). However the function of EGR3 in prostate cancer is unknown. Human prostate cancer cells M12 containing high levels of EGR3 were used for shRNA-mediated silencing of EGR3. Gene expression analysis of EGR3 knockdown cells reveals a role in inflammation and the existence of a crosstalk with the NFkB pathway. The M12 prostate cancer cell line is a tumorigenic derivative of the P69 SV40-T immortalized epithelial cells obtained through serial passages of P69 tumor xenografts in mice (Bae et al., Prostate 1998; 34:275-82). M12 cells were transfected with shRNA scramble plasmid (shSCR) or shEGR3 plasmid and selected with puromycin (1 μg/ml). Single-cell clones were isolated using standard methods. Stably transfected cells (shSCR-M12 and shEGR3-M12) were then maintained in growth medium supplemented with puromycin. Two separate clones (termed cl 2 and cl 3) were studied. Knockdown of EGR3 (50% efficacy on average) was stable over many cell culture passages. Biological duplicates of control cells (scramble shRNA) and of two separate clones of shEGR3 cells (clone 2 and clone 3) were used for the gene expression arrays (i.e. 6 samples).
Project description:An isogenic prostate cancer cell model was used to profile miRNA changes that contribute to the tumorigenic state of prostate cancer. P69 cells are an SV40 large T antigen immortalized cell line that is poorly tumorigenic and non-metastatic. The M12 derivative was derived from an in vivo selection process using nude, athymic mice. Immortalized cells were cultured and used to extract RNA and probed against the human miRNA panel from Exiqon. There were a total of 186 miRNAs that were at least two fold differentially regulated .
Project description:The insulin-like growth factor-I (IGF-IR) and androgen (AR) receptors are important players in prostate cancer biology. Functional interactions between the IGF-I and androgen signaling pathways seem to have crucial roles in the progression of prostate cancer from early (benign) to advanced (metastatic) stages. DNA methylation is a major epigenetic alteration affecting gene expression. Hypermethylation of tumor suppressor promoters is a frequent event in human cancer, leading to inactivation and repression of specific genes. The aim of the present study was to identify the entire set of methylated genes (M-bM-^@M-^\methylomeM-bM-^@M-^]) in a cellular model that replicates prostate cancer progression. The methylation profiles of the P69 (early stage, benign) and M12 (advanced stage, metastatic) prostate cancer cell lines were established by treating cells with the demethylating agent 5-Aza-2M-bM-^@M-^Y-deoxycytidine (5-Aza) followed by DNA microarray analysis. Comparative genome-wide methylation analyses of 5-Aza-treated versus untreated cells identified 297 genes overexpressed in P69 and 191 genes overexpressed in M12 cells. One hundred and two genes were upregulated in both benign and metastatic cell lines. In addition, our analyses identified the PITX2 gene as a master regulator upstream of the AR and IGF-IR genes.
Project description:EGR3 expression is upregulated in human prostate cancer compared to normal prostate tissue and is associated with absence of relapse, while low EGR3 expression in tumors is predicitive of disease relapse (Pio et al., PLOS One 2013; 8(1):e54096). However the function of EGR3 in prostate cancer is unknown. Human prostate cancer cells M12 containing high levels of EGR3 were used for shRNA-mediated silencing of EGR3. Gene expression analysis of EGR3 knockdown cells reveals a role in inflammation and the existence of a crosstalk with the NFkB pathway.
