riccardin-d and Prostatic-Neoplasms

RD-N, an aminomethylated derivative of riccardin D, is a lysosomotropic agent that can trigger lysosomal membrane permeabilization followed by cathepsin B (CTSB)-dependent apoptosis in prostate cancer (PCa) cells, but the underlying mechanisms remain unknown. Here we show that RD-N treatment drives CTSB translocation from the lysosomes to the nucleus where it promotes DNA damage by suppression of the breast cancer 1 protein (BRCA1). Inhibition of CTSB activity with its specific inhibitors, or by CTSB-targeting siRNA or CTSB with enzyme-negative domain attenuated activation of BRCA1 and DNA damage induced by RD-N. Conversely, CTSB overexpression resulted in inhibition of BRCA1 and sensitized PCa cells to RD-N-induced cell death. Furthermore, RD-N-induced cell death was exacerbated in BRCA1-deficient cancer cells. We also demonstrated that CTSB/BRCA1-dependent DNA damage was critical for RD-N, but not for etoposide, reinforcing the importance of CTSB/BRCA1 in RD-N-mediated cell death. In addition, RD-N synergistically increased cell sensitivity to cisplatin, and this effect was more evidenced in BRCA1-deficient cancer cells. This study reveals a novel molecular mechanism that RD-N promotes CTSB-dependent DNA damage by the suppression of BRCA1 in PCa cells, leading to the identification of a potential compound that target lysosomes for cancer treatment.

Topics: Amines; Apoptosis; BRCA1 Protein; Cathepsin B; Cell Membrane Permeability; DNA Damage; Humans; Lysosomes; Male; Methylation; Phenyl Ethers; Prostatic Neoplasms; Proteolysis; Stilbenes; Tumor Cells, Cultured

Bisbibenzyl compounds have gained our interests for their potential antitumor activity in malignant cell-types.. The objective of this study is to investigate the effect of bisbibenzyl compounds riccardin C (RC), marchantin M (MM), and riccardin D (RD) on androgen receptor (AR) in prostate cancer (PCa) cells.. After exposure to 10 μM of the compounds for 24 h, cell cycle and cell survival analyses were performed using FACS and MTT assay to confirm the effect of these bisbibenzyls on PCa LNCaP cells. Changes in the AR expression and function, as the result of exposure to the compounds, were investigated using real-time PCR, ELISA, transient transfection, western blotting (WB), immunoprecipitation, and immunofluorescence staining (IF). Chemical-induced autophagy was examined by WB, IF, and RNAi.. RC, MM, and RD reduced the viability of LNCaP cells accompanied with arrested cell cycle in the G0/G1 phase and induction of apoptosis. Further investigation revealed that these compounds significantly inhibited AR expression at mRNA and protein levels, leading to the suppression of AR transcriptional activity. Moreover, inhibition of proteasome activity by bisbibenzyls, which in turn caused the induction of autophagy, as noted by induction of LC3B expression, conversion, and accumulation of punctate dots in treated cells. Co-localization of AR/LC3B and AR/Ub suggested that autophagy contributed to the degradation of polyubiquitinated-AR when proteasome activity was suppressed by the bisbibenzyls.. Suppression of proteasome activity and induction of autophagy were involved in bisbibenzyl-mediated modulation of AR activities and apoptosis, suggesting their potential in treating PCa.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Autophagy; Bibenzyls; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Survival; Ethers, Cyclic; Gene Expression; Hepatophyta; Humans; Male; Phenyl Ethers; Prostatic Neoplasms; Proteasome Inhibitors; Protein Transport; Receptors, Androgen; Stilbenes; Transcription, Genetic

Our previous studies had shown that Riccardin D (RD) exhibited cytotoxic effects by induction of apoptosis and inhibition of angiogenesis and topoisomerase II. Here, we reported that apoptosis is not the sole mechanism by which RD inhibits tumor cell growth because low concentrations of RD caused cellular senescence in prostate cancer (PCa) cells.. Low concentrations of RD were used to treat PCa cells in vitro and in vivo, and senescence-associated β-galactosidase activity, DNA damage response markers, and/or colony-forming ability, cell cycle were analyzed, respectively. We then used siRNA knockdown to identify key factor in RD-triggered cellular senescence.. RD treatment caused growth arrest at G0/G1 phase with features of cellular senescence phenotype such as enlarged and flattened morphology, increased senescence-associated-beta-galactosidase staining cells, and decreased cell proliferation in PCa cells. Induction of cellular senescence by RD occurred through activation of DNA damage response including increases in the phosphor-H2AX, inactivation of Chk1/2, and suppression of repair-related Ku70/86 and phosphor-BRCA1 in PCa cells in vitro and in vivo. Analysis of expression levels of p53, p21(CIP1), p16(INK4a), p27(KIP1), pRb and E2F1 and genetic knockdown of p21(CIP1) demonstrated an important role of p21(CIP1) in RD-triggered cellular senescence.. Involvement of the DNA damage response and p21(CIP1) defines a novel mechanism of RD action and indicates that RD could be further developed as a promising anticancer agent for cancer therapy.

Topics: Animals; Apoptosis; Cell Division; Cell Line, Tumor; Cell Proliferation; Cellular Senescence; Cyclin-Dependent Kinase Inhibitor p21; DNA Damage; G1 Phase Cell Cycle Checkpoints; Gene Knockdown Techniques; Humans; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Phenyl Ethers; Prostatic Neoplasms; Random Allocation; Resting Phase, Cell Cycle; Stilbenes; Transfection; Xenograft Model Antitumor Assays

We recently reported that Riccardin D (RD) was able to induce apoptosis by targeting Topo II. Here, we found that RD induced cell cycle arrest in G2/M phase in PC-3 cells, and caused remarkable DNA damage as evidenced by induction of γH2AX foci, micronuclei, and DNA fragmentation in Comet assay. Time kinetic and dose-dependent studies showed that ATM/Chk2 and ATR/Chk1 signaling pathways were sequentially activated in response to RD. Blockage of ATM/ATR signaling led to the attenuation of RD-induced γH2AX, and to the partial recovery of cell proliferation. Furthermore, RD exposure resulted in the inactivation of BRCA1, suppression of HR and NHEJ repair activity, and downregulation of the expressions and DNA-end binding activities of Ku70/86. Consistent with the observations, microarray data displayed that RD triggered the changes in genes responsible for cell proliferation, cell cycle, DNA damage and repair, and apoptosis. Administration of RD to xenograft mice reduced tumor growth, and coordinately caused alterations in the expression of genes involved in DNA damage and repair, along with cell apoptosis. Thus, this finding identified a novel mechanism by which RD affects DNA repair and acts as a DNA damage agent in prostate cancer.

Topics: Animals; Antigens, Nuclear; Antineoplastic Agents, Phytogenic; Apoptosis; Ataxia Telangiectasia Mutated Proteins; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Checkpoint Kinase 1; Checkpoint Kinase 2; Disease Models, Animal; DNA Damage; DNA End-Joining Repair; DNA-Binding Proteins; Gene Expression Regulation, Neoplastic; Humans; Ku Autoantigen; Male; Mice; Phenyl Ethers; Prostatic Neoplasms; Protein Kinases; Recombinational DNA Repair; Signal Transduction; Stilbenes; Transcriptome; Xenograft Model Antitumor Assays