A prostate-derived cDNA that is mapped to human chromosome 19 encodes a novel protein

The epithelial cells of prostate gland secrete various secretory products that play an important role in the growth and differentiation of prostate gland. These secretory products have also been implicated in neuroendocrine differentiation of benign prostatic hyperplasia and prostate malignancy. We have cloned a prostate-derived cDNA encoding a novel protein with a predicted molecular weight of 78 kDa (P(78)), and precisely mapped the cDNA sequence to chromosome 19. The P(78) gene has a complex genomic structure with 18 exons and 17 introns. The P(78) contains two conserved structural domains with limited similarity to domain D of synapsin I. The P(78) mRNA was expressed in various human cell lines. Western blot analysis using antibody specific for the P(78) revealed the presence of the P(78) protein in the prostate cancer cell lines with much lower level in metastatic prostate cancer cell lines compared to that in a primary prostate cancer cell line.     

Discovery of novel fatty acid synthase (FAS) inhibitors based on the structure of ketoaceyl synthase (KS) domain

Development of fatty acid synthase (FAS) inhibitors has increasingly attracted much attention in recent years due to their potential therapeutic use in obesity and cancers. In this investigation, pharmacophore modeling based on the first crystal structure of human KS domain of FAS was carried out. The established pharmacophore model was taken as a 3D query for retrieving potent FAS inhibitors from the chemical database Specs. Docking study was further carried out to refine the obtained hit compounds. Finally, a total of 28 compounds were selected based on the ranking order and visual examination, which were first evaluated by a cell line-based assay. Seven compounds that have good inhibition activity against two FAS overexpressing cancer cell lines were further evaluated by an enzyme-based assay. One compound with a new chemical scaffold was found to have low micromolar inhibition potency against FAS, which has been subjected to further chemical structural modification.     

Inflammatory Cytokines and Survival Factors from Serum Modulate Tweak-Induced Apoptosis in PC-3 Prostate Cancer Cells

Tumor necrosis factor-like weak inducer of apoptosis (TWEAK, TNFSF12) is a member of the tumor necrosis factor superfamily. TWEAK activates the Fn14 receptor, and may regulate cell death, survival and proliferation in tumor cells. However, there is little information on the function and regulation of this system in prostate cancer. Fn14 expression and TWEAK actions were studied in two human prostate cancer cell lines, the androgen-independent PC-3 cell line and androgen-sensitive LNCaP cells. Additionally, the expression of Fn14 was analyzed in human biopsies of prostate cancer. Fn14 expression is increased in histological sections of human prostate adenocarcinoma. Both prostate cancer cell lines express constitutively Fn14, but, the androgen-independent cell line PC-3 showed higher levels of Fn14 that the LNCaP cells. Fn14 expression was up-regulated in PC-3 human prostate cancer cells in presence of inflammatory cytokines (TNFα/IFNγ) as well as in presence of bovine fetal serum. TWEAK induced apoptotic cell death in PC-3 cells, but not in LNCaP cells. Moreover, in PC-3 cells, co-stimulation with TNFα/IFNγ/TWEAK induced a higher rate of apoptosis. However, TWEAK or TWEAK/TNFα/IFNγ did not induce apoptosis in presence of bovine fetal serum. TWEAK induced cell death through activation of the Fn14 receptor. Apoptosis was associated with activation of caspase-3, release of mitochondrial cytochrome C and an increased Bax/BclxL ratio. TWEAK/Fn14 pathway activation promotes apoptosis in androgen-independent PC-3 cells under certain culture conditions. Further characterization of the therapeutic target potential of TWEAK/Fn14 for human prostate cancer is warranted.     

Topotecan and methotrexate alter expression of the apoptosis-related genes BCL2, FAS and BCL2L12 in leukemic HL-60 cells

The BCL2 family of genes (B-cell CLL/lymphoma 2; Bcl-2) plays a pivotal role in the highly regulated process of apoptosis. We have recently cloned a newly identified member of this family, BCL2L12, which was found to be differentially expressed in many tumors. It is known that topotecan and methotrexate act through induction of apoptosis in cancer cells. In the present study we investigated the expression profile of the novel apoptotic gene BCL2L12 in relation to other apoptotic genes in the human leukemic cell line HL-60, after treatment with topotecan or methotrexate. The kinetics of apoptosis induction and cell toxicity were investigated by DNA laddering and the MTT method, respectively. Gene expression levels were analyzed by RT-PCR using gene-specific primers. Downregulation of BCL2L12, BCL2 and FAS was observed after treatment of HL-60 cells with topotecan, while treatment with methotrexate led to downregulation of BCL2 and FAS, with no change in BCL2L12 expression. Our results support the significance of mRNA modulations in the expression of apoptosis-related genes during treatment of human leukemic cells with anticancer drugs.     

Cisplatin induces production of reactive oxygen species via NADPH oxidase activation in human prostate cancer cells

This study aimed to examine the roles of reactive oxygen species (ROS) in cisplatin treatment of human prostate cancer cells; hormone-sensitive LNCaP and hormone-refractory PC3 and DU145 cells. Intracellular levels of ROS and H(2)O(2) were measured and visualized using specific fluorescent probes. NADPH oxidase (NOX) activity was detected by lucigenin chemiluminescence assay. Expression levels of NOX isoforms were determined by semi-quantitative RT-PCR. Cisplatin treatment increased the intracellular levels of ROS and H(2)O(2) in three prostate cancer cell lines. The increase was transient and robust in hormone-sensitive LNCaP cells compared with hormone-refractory PC3 and DU145 cells. Consistent with these findings, the NOX activity induced by cisplatin was higher in LNCaP cells than in PC3 and DU145 cells. Expression pattern of NOX isoforms varied among three cell lines and the NOX activity was independent of NOX expression. Taken together, we have shown that cisplatin induces production of ROS and H(2)O(2) via NOX activation in human prostate cancer cell lines, which is most prominent in hormone-sensitive LNCaP cells.