title: Silibinin inhibits prostate cancer cells- and RANKL-induced osteoclastogenesis by targeting NFATc1, NF-κB, and AP-1 activation in RAW264.7 cells.
abstract: Currently, there are limited therapeutic options against bone metastatic prostate cancer (PCA), which is primarily responsible for high mortality and morbidity in PCA patients. Enhanced osteoclastogenesis is an essential feature associated with metastatic PCA in the bone microenvironment. Silibinin, an effective chemopreventive agent, is in phase II clinical trials in PCA patients but its efficacy against PCA cells-induced osteoclastogenesis is largely unknown. Accordingly, here we examined silibinin effect on PCA cells-induced osteoclastogenesis employing human PCA (PC3MM2, PC3, and C4-2B) and murine macrophage RAW264.7 cells. We also assessed silibinin effect on receptor activator of nuclear factor κB ligand (RANKL)-induced signaling associated with osteoclast differentiation in RAW264.7 cells. Further, we analyzed silibinin effect on osteomimicry biomarkers in PCA cells. Results revealed that silibinin (30-90 μM) inhibits PCA cells-induced osteoclast activity and differentiation in RAW264.7 cells via modulating expression of several cytokines (IGF-1, TGF-β, TNF-α, I-TAC, M-CSF, G-CSF, GM-CSF, etc.) that are important in osteoclastogenesis. Additionally, in RAW264.7 cells, silibinin decreased the RANKL-induced expression and nuclear localization of NFATc1, which is considered the master regulator of osteoclastogenesis. Furthermore, silibinin decreased the RANKL-induced DNA binding activity of NFATc1 and its regulators NF-κB and AP1, and the protein expression of osteoclast specific markers (TRAP, OSCAR, and cathepsin K). Importantly, silibinin also decreased the expression of osteomimicry biomarkers (RANKL, Runx2, osteocalcin, and PTHrP) in cell culture (PC3 and C4-2B cells) and/or in PC3 tumors. Together, our findings showing that silibinin inhibits PCA cells-induced osteoclastogenesis, suggest that silibinin could be useful clinically against bone metastatic PCA.
authors.affiliation: Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Denver, Aurora, Colorado.
authors.forename: Chandagirikoppal V
authors.initials: CV
authors.lastname: Kavitha
authors.forename: Gagan
authors.initials: G
authors.lastname: Deep
authors.forename: Subhash C
authors.initials: SC
authors.lastname: Gangar
authors.forename: Anil K
authors.initials: AK
authors.lastname: Jain
authors.forename: Chapla
authors.initials: C
authors.lastname: Agarwal
authors.forename: Rajesh
authors.initials: R
authors.lastname: Agarwal
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D007124:Immunoenzyme Techniques; D008264:Macrophages / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D050778:NFATC Transcription Factors / Q000378:metabolism*; D010010:Osteoclasts / Q000187:drug effects* / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D053245:RANK Ligand / Q000378:metabolism*; D000077385:Silybin; D012838:Silymarin / Q000494:pharmacology*; D018808:Transcription Factor AP-1 / Q000378:metabolism*
chemical_list: D016207:Cytokines; D016328:NF-kappa B; D050778:NFATC Transcription Factors; D053245:RANK Ligand; D012838:Silymarin; D018808:Transcription Factor AP-1; D000077385:Silybin
keywords: NFATc1; RANK ligand; osteoclastogenesis; prostate cancer; silibinin