Drug Target In Prostate Cancer Research Articles

MiR-21 as an Independent Biochemical Recurrence Predictor and Potential Therapeutic Target for Prostate Cancer

Abnormal miRNA expression is associated with prostate cancer progression. However, the relationship between miRNA and biochemical recurrence after radical prostatectomy is not well established. Thus, we evaluated the miRNA miR-21 as a biomarker to predict the risk of biochemical failure, and as a potential drug target for prostate cancer therapy. miR-21 levels were assayed using locked nucleic acid in situ hybridization coupled with tissue microarray techniques in 169 radical prostatectomy tissue samples. The Cox proportional hazard model was used to analyze miR-21 expression as an independent predictor of biochemical recurrence. The association of miR-21 with recurrence was estimated using the Kaplan-Meier method. miR-21 was also evaluated as a potential drug target for prostate cancer therapy. miR-21 expression in prostate cancer tissue samples was significantly associated with pathological stage, lymph node metastasis, capsular invasion, organ confined disease, Gleason score, biochemical recurrence and patient followup. Multivariate analysis also indicated that miR-21 expression could be an independent predictor of biochemical recurrence. The 5-year recurrence-free probability for patients positive vs negative for miR-21 expression was 33.9% vs 44.5%. In vivo treatment with antagomir-21 also repressed the tumor growth of DU145 cells in nude mice. Positive miR-21 expression was associated with poor biochemical recurrence-free survival and predicted the risk of biochemical recurrence in patients with prostate cancer after radical prostatectomy. Accordingly gene therapy using miR-21 inhibition strategies may prove useful for prostate cancer therapy.

Abstract A9: Androgen-regulated miR-32 targets BTG2 and is overexpressed in castration-resistant prostate cancer

Abstract The androgen receptor (AR) signaling pathway is central to the emergence of castration-resistant prostate cancer (CRPC). We set out to identify androgen-regulated microRNAs (miRNAs) that may contribute to the development of CRPC. We found miR-32 to be an androgen regulated miRNA which is differentially expressed in CRPC compared to benign prostate hyperplasia (BPH) according to microarray analyses. qRT-PCR analyses confirmed that miR-32 expression increases in CRPC compared to BPH, and that in untreated prostate cancer (PC) miR-32 expression is higher in tumors of stage pT3 compared to pT2. Transfection of pre-miR-32 to prostate cancer cell line LNCaP provided a significant growth advantage to the cells, and miR-32 was further demonstrated to reduce apoptosis. To identify target genes for miR-32, an mRNA microarray analysis was performed with LNCaP cells transfected with pre-miR-32. In this assay, B cell translocation gene 2 (BTG2) was identified as a novel putative target for miR-32. BTG2 was reduced both at mRNA and protein level in the cells transfected with pre-miR-32, and BTG2 was confirmed to be a miR-32 target by 3′-UTR-luciferase assay. Furthermore, BTG2 was able to partially rescue the growth advantage of miR-32 in LNCaP cells. Immunostainings of prostate cancer patient material showed a statistically significant (p < 0.0001) reduction of BTG2 protein in CRPCs compared to PC. The lack of BTG2 staining was also associated (p=0.0021) with a short progression-free time in patients who underwent prostatectomy and inversely correlated to miR-32 expression levels. In conclusion, we find that miR-32 is an androgen-regulated miRNA targeting BTG2 and overexpressed in CRPC. Thus, miR-32 is a potential marker for aggressive disease and is a putative drug target in PC. Citation Format: Sanni E. Jalava, Leena Latonen, Mauro Scaravilli, Kati K. Waltering, Janne Seppälä, Harri Lähdesmäki, Teuvo L.J. Tammela, Tapio Visakorpi. Androgen-regulated miR-32 targets BTG2 and is overexpressed in castrationresistant prostate cancer [abstract]. In: Proceedings of the AACR Special Conference on Advances in Prostate Cancer Research; 2012 Feb 6-9; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(4 Suppl):Abstract nr A9.

Androgen-regulated miR-32 targets BTG2 and is overexpressed in castration-resistant prostate cancer

The androgen receptor (AR) signaling pathway is involved in the emergence of castration-resistant prostate cancer (CRPC). Here, we identified several androgen-regulated microRNAs (miRNAs) that may contribute to the development of CRPC. Seven miRNAs, miR-21, miR-32, miR-99a, miR-99b, miR-148a, miR-221 and miR-590-5p, were found to be differentially expressed in CRPC compared with benign prostate hyperplasia (BPH) according to microarray analyses. Significant growth advantage for LNCaP cells transfected with pre-miR-32 and pre-miR-148a was found. miR-32 was demonstrated to reduce apoptosis, whereas miR-148a enhanced proliferation. Androgen regulation of miR-32 and miR-148a was confirmed by androgen stimulation of the LNCaP cells followed by expression analyses. The AR-binding sites in proximity of these miRNAs were demonstrated with chromatin immunoprecipitation (ChIP). To identify target genes for the miRNAs, mRNA microarray analyses were performed with LNCaP cells transfected with pre-miR-32 and pre-miR-148a. Expression of BTG2 and PIK3IP1 was reduced in the cells transfected with pre-miR-32 and pre-miR-148a, respectively. Also, the protein expression was reduced according to western blot analysis. BTG2 and PIK3IP1 were confirmed to be targets by 3'UTR-luciferase assays. Finally, immunostainings showed a statistically significant (P<0.0001) reduction of BTG2 protein in CRPCs compared with untreated prostate cancer (PC). The lack of BTG2 staining was also associated (P<0.01) with a short progression-free time in patients who underwent prostatectomy. In conclusion, androgen-regulated miR-32 is overexpressed in CRPC, leading to reduced expression of BTG2. Thus, miR-32 is a potential marker for aggressive disease and is a putative drug target in PC.

Phospholipase PLA2G7, associated with aggressive prostate cancer, promotes prostate cancer cell migration and invasion and is inhibited by statins

Prostate cancer is the second leading cause of cancer mortality in men in developed countries. Due to the heterogeneous nature of the disease, design of novel personalized treatments is required to achieve efficient therapeutic responses. We have recently identified phospholipase 2 group VII (PLA2G7) as a potential drug target especially in ERG oncogene positive prostate cancers. Here, the expression profile of PLA2G7 was studied in 1137 prostate cancer and 409 adjacent non-malignant prostate tissues using immunohistochemistry to validate its biomarker potential and putative association with disease progression. In order to reveal the molecular alterations induced by PLA2G7 impairment, lipidomic and gene expression profiling was performed in response to PLA2G7 silencing in cultured prostate cancer cells. Moreover, the antineoplastic effect of statins combined with PLA2G7 impairment was studied in prostate cancer cells to evaluate the potential of repositioning of in vivo compatible drugs developed for other indications towards anti-cancer purposes. The results indicated that PLA2G7 is a cancer-selective biomarker in 50 % of prostate cancers and associates with aggressive disease. The alterations induced by PLA2G7 silencing highlighted the potential of PLA2G7 inhibition as an anti-proliferative, pro-apoptotic and anti-migratorial therapeutic approach in prostate cancer. Moreover, the anti-proliferative effect of PLA2G7 silencing was potentiated by lipid-lowering statins in prostate cancer cells. Taken together, our results support the potential of PLA2G7 as a biomarker and a drug target in prostate cancer and present a rationale for combining PLA2G7 inhibition with the use of statins in prostate cancer management.

Abstract 3046: Phosphodiesterase 4D (PDE4D) as a new drug target for prostate cancer

Abstract Our laboratory used an insertional somatic mutagenesis screen in mice to identify novel genes that may drive prostate cancer initiation and progression. Using this screen, we have identified candidate prostate cancer genes including the candidate oncogene phosphodiesterase 4D (PDE4D). Investigation of PDE4D showed that it was over-expressed in human prostate cancer cell lines and in human prostate cancers. Stable knockdown of PDE4D by shRNA reduced the growth and migration of prostate cancer cells in vitro and in vivo. PDE4D has been implicated in the pathogenesis of chronic obstructive pulmonary disease (COPD). Several small molecule PDE4D selective inhibitors have been developed and tested in clinical trials for COPD. Although these studies showed that the inhibitors had modest efficacy as a therapy for COPD, they confirmed that the inhibitors were safe for use in humans at doses that inhibited PDE4D in vivo. Based on our previous studies with PDE4D, we postulated that PDE4D small molecule inhibitors would be candidate anti-prostate cancer drugs. Daily oral administration of PDE4D inhibitors, NVP-ABE171 or cilomilast, for 6 weeks significantly reduced the growth of LNCaP-C4 prostate cancer xenografts in immune-compromised mice. Under in vitro conditions in which prostate cancer cell growth was inhibited by either NVP-ABE171 or cilomilast, we also observed changes in the activation of potential downstream molecular targets of PDE4D inhibition. Affected pathways included the sonic hedgehog and mitogen-activated protein kinase pathways. We also tested the potential role of the PDE4D pathway in prostate cancer stromal-epithelial interactions by modeling these interactions using novel microchannel co-culture devices. In this context, PDE4D inhibitors reduced the activation of Shh pathway target genes in mesenchymal cells and reduced the proliferation of epithelial cancer cell lines, supporting the potential importance of the PDE4D pathway for prostate cancer stromal-epithelial interactions. We are continuing in vitro and in vivo studies with these candidate drugs to further support the use of NVP-ABE171 and cilomilast as anti-prostate cancer drugs and to understand their mechanism of action. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3046. doi:10.1158/1538-7445.AM2011-3046

USP2a is an oncogenic isopeptidase and a potential target in prostate cancer

14522 Background: De-ubiquitinating enzymes (isopeptidases) remove ubiquitin side chains prior to degradation by the proteosome thus stabilizing their protein targets. We have identified a novel androgen regulated isopeptidase, USP2a, and demonstrated that it binds and prolongs the half-life of fatty acid synthase (FAS), a key enzyme in lipid metabolism of tumor cells. Methods: We determined whether USP2a has oncogenic properties in vitro and in vivo. Wild-type and catalytically inactive USP2a were introduced in immortalized normal prostate epithelial cells (AR-iPrECs). Clonogenicity assays and apoptosis induction by chemotherapeutic agents were performed on these cells. Anti-USP2a siRNA were transfected in normal and transformed (LNCaP, DU145 and PC-3) prostate cell lines. Oncogenicity in vivo was shown by s.c. injection of NIH3T3-USP2a cells in nude mice. Furthermore, USP2a mRNA expression and gene microarrays were tested in 52 human prostate adenocarcinomas. Results: Wild-type USP2a overexpression in AR-iPrEC cells resulted in a significant increase in number and size of colonies compared to those obtained in parental cells. Growth in soft agar was significantly enhanced as well. Silencing of USP2a in LNCAP and DU145 cells resulted in a strong apoptotic effect, evaluated by FACS analysis and cleaved-PARP expression. The role of this isopeptidase in apoptosis regulation was confirmed on AR-iPrEC-USP2a cells, that showed resistance to apoptosis induced by cisplatin and taxol. Importantly, USP2a overexpression was able to transform NIH3T3 cells, generating greater than or equal to 0.5 cm subcutaneous tumors in 12/12 nude mice within 3 weeks, while none of the negative controls grew. USP2a mRNA was overexpressed in 39% of human prostate cancers, showing 1.6–104 (median 5.48) fold induction relative to normal tissues by qRT-PCR. Gene expression profiling of the same tumors revealed specific signatures in USP2a-overexpressing tumors. Conclusions: Our results demonstrate that USP2a behaves as an oncogene in vitro and in vivo and is overexpressed in organ-confined prostate cancer. These data strongly suggest that this isopeptidase is a potential drug target in prostate cancer. [Table: see text]