title: Silymarin inhibits growth and causes regression of established skin tumors in SENCAR mice via modulation of mitogen-activated protein kinases and induction of apoptosis.
abstract: This study reports in vivo therapeutic efficacy of silymarin against skin tumors with mechanistic rationale. 7,12-Dimethylbenz[a]anthracene-12-O-tetradecanoyl-phorbol-13-acetate (DMBA-TPA)-induced established skin papilloma (tumor)-bearing SENCAR mice were fed with 0.5% silymarin in AIN-93M-purified diet (w/w), and both tumor growth and regression were monitored during 5 weeks of feeding regimen. Silymarin feeding significantly inhibited (74%, P < 0.01) tumor growth and also caused regression (43%, P < 0.01) of established tumors. Proliferating cell nuclear antigen and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling immunohistochemical staining of tumors showed that silymarin decreases proliferation index by 48% (P < 0.001) and increases apoptotic index by 2.5-fold (P < 0.001), respectively. Skin tumor growth inhibition and regression by silymarin were also accompanied by a strong decrease (P < 0.001) in phospho-ERK1/2 levels in tumors from silymarin-fed mice compared with controls. In the studies evaluating bioavailability and physiologically achievable level of silymarin (as silibinin) in plasma, skin tumor, skin, liver, lung, mammary gland and spleen, we found 10, 6.5, 3.1, 13.7, 7.7, 5.9 and 4.4 microg silibinin/ml plasma or per gram tissue, respectively. In an attempt to translate these findings to human skin cancer and to establish biological significance of physiologically achievable level, effect of plasma concentration of silibinin was next examined in human epidermoid carcinoma A431 cells. Silibinin treatment of cells in culture at 12.5, 25 (plasma level) and 50 microM doses resulted in 30-74% (P < 0.01-0.001) growth inhibition and 7-42% death of A431 cells in a dose- and time-dependent manner; apoptosis was identified as a cell death response by silibinin. Similar silibinin treatments also resulted in a significant decrease in phospho-mitogen-activated protein kinase/extracellular signal-regulated protein kinase 1/2 (MAPK/ERK1/2) levels, but an up-regulation of stress-activated protein kinase/jun NH(2)-terminal kinase (SAPK/JNK1/2) and p38 mitogen-activated protein kinase (p38 MAPK) activation in A431 cells. The use of MEK1 inhibitor, PD98059, showed that inhibition of ERK1/2 signaling, in part, contributes to silibinin-caused cell growth inhibition. Together, the data suggest that an inhibition of ERK1/2 activation and an increased activation of JNK1/2 and p38 by silibinin could be possible underlying molecular events involved in inhibition of proliferation and induction of apoptosis in A431 cells. These data suggest that silymarin and/or its major active constituent silibinin could be an effective agent for both prevention and intervention of human skin cancer.
authors.affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, University of Colorado Health Sciences Center, Denver, CO 80262, USA.
authors.forename: Rana P
authors.initials: RP
authors.lastname: Singh
authors.forename: Anil K
authors.initials: AK
authors.lastname: Tyagi
authors.forename: Jifu
authors.initials: J
authors.lastname: Zhao
authors.forename: Rajesh
authors.initials: R
authors.lastname: Agarwal
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; D051379:Mice; D018632:Mice, Inbred SENCAR; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009928:Organ Specificity; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D012838:Silymarin / Q000378:metabolism / Q000494:pharmacology* / Q000627:therapeutic use*; D012878:Skin Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology*; D013997:Time Factors; D014407:Tumor Cells, Cultured
chemical_list: D012838:Silymarin; D017346:Protein Serine-Threonine Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; C482125:MAP2K1 protein, human; C482127:Map2k1 protein, mouse; D020929:Mitogen-Activated Protein Kinase Kinases