Attenuation of nucleoside and anti-cancer nucleoside analog drug uptake in prostate cancer cells by Cimicifuga racemosa extract BNO-1055.

This study aimed to investigate the mechanisms underlying the anti-proliferative effects of the ethanolic Cimicifuga racemosa extract BNO-1055 on prostate cells and evaluate its therapeutic potential. BNO-1055 dose-dependently attenuated cellular uptake and incorporation of thymidine and BrdU and significantly inhibited cell growth after long-time exposure. Similar results were obtained using saponin-enriched sub-fractions of BNO-1055. These inhibitory effects of BNO-1055 could be mimicked using pharmacological inhibitors and isoform-specific siRNAs targeting the equilibrative nucleoside transporters ENT1 and ENT2. Moreover, BNO-1055 attenuated the uptake of clinically relevant nucleoside analogs, e.g. the anti-cancer drugs gemcitabine and fludarabine. Consistent with inhibition of the salvage nucleoside uptake pathway BNO-1055 potentiated the cytotoxicity of the de novo nucleotide synthesis inhibitor 5-FU without significantly altering its uptake. Collectively, these data show for the first time that the anti-proliferative effects of BNO-1055 result from hindered nucleoside uptake due to impaired ENT activity and demonstrate the potential therapeutic use of BNO-1055 for modulation of nucleoside transport.

ROS signaling by NOX4 drives fibroblast-to-myofibroblast differentiation in the diseased prostatic stroma.

Stromal remodeling, in particular fibroblast-to-myofibroblast differentiation, is a hallmark of benign prostatic hyperplasia (BPH) and solid tumors, including prostate cancer (PCa). Increased local production of TGFß1 is considered the inducing stimulus. Given that stromal remodeling actively promotes BPH/PCa development, there is considerable interest in developing stromal-targeted therapies. Microarray and quantitative PCR analysis of primary human prostatic stromal cells induced to undergo fibroblast-to-myofibroblast differentiation with TGFß1 revealed up-regulation of the reactive oxygen species (ROS) producer reduced nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4) and down-regulation of the selenium-containing ROS-scavenging enzymes glutathione peroxidase 3, thioredoxin reductase 1 (TXNRD1), and the selenium transporter selenoprotein P plasma 1. Consistently, NOX4 expression correlated specifically with the myofibroblast phenotype in vivo, and loss of selenoprotein P plasma 1 was observed in tumor-associated stroma of human PCa biopsies. Using lentiviral NOX4 short hairpin RNA-mediated knockdown, pharmacological inhibitors, antioxidants, and selenium, we demonstrate that TGFß1 induction of NOX4-derived ROS is required for TGFß1-mediated phosphorylation of c-jun N-terminal kinase, which in turn is essential for subsequent downstream cytoskeletal remodeling. Significantly, selenium supplementation inhibited differentiation by increasing ROS-scavenging selenoenzyme biosynthesis because glutathione peroxidase 3 and TXNRD1 expression and TXNRD1 enzyme activity were restored. Consistently, selenium depleted ROS levels downstream of NOX4 induction. Collectively, this work demonstrates that dysregulated redox homeostasis driven by elevated NOX4-derived ROS signaling underlies fibroblast-to-myofibroblast differentiation in the diseased prostatic stroma. Further, these data indicate the potential clinical value of selenium and/or NOX4 inhibitors in preventing the functional pathogenic changes of stromal cells in BPH and PCa.

Attenuated proliferation and trans-differentiation of prostatic stromal cells indicate suitability of phosphodiesterase type 5 inhibitors for prevention and treatment of benign prostatic hyperplasia.

Benign prostatic hyperplasia (BPH) is characterized by tissue overgrowth and stromal reorganization primarily due to cellular proliferation and fibroblast-to-myofibroblast trans-differentiation. To evaluate the potential of phosphodiesterase type 5 (PDE5) inhibitors like tadalafil for prevention and treatment of BPH, we analyzed the role of the nitric oxide/cyclic GMP (cGMP)/PDE5 pathway for cellular proliferation and TGFbeta1-induced fibroblast-to-myofibroblast trans-differentiation in primary prostate stromal cells. Inhibition by tadalafil of PDE5, which is mainly expressed in the stromal compartment of the prostate, reduced proliferation of primary prostate stromal cells and to a lesser extent of primary prostate basal epithelial cells. Attenuated proliferation due to elevated intracellular cGMP levels was confirmed by inhibition of the cGMP-dependent protein kinase G by its inhibitor KT2358. Moreover, tadalafil strongly attenuated TGFbeta1-induced fibroblast-to-myofibroblast trans-differentiation. The inhibitory effect on trans-differentiation was also observed after small interfering RNA-mediated PDE5 knockdown. As confirmed by the MAPK kinase 1 inhibitor PD98059, this effect was mediated via MAPK kinase 1 signaling. We conclude that BPH patients might benefit from adjuvant therapies with PDE5 inhibitors that inhibit stromal enlargement due to cell proliferation, as well as TGFbeta1-induced trans-differentiation processes.