title: Celastrol synergistically enhances temozolomide cytotoxicity in melanoma cells.
abstract: Efforts to improve melanoma response rates to temozolomide (TMZ) have thus far been unsuccessful. We screened a library of 2,000 marketed drugs and natural products to identify agents with the potential to sensitize melanoma cells to the effects of TMZ. Celastrol (CEL), a natural compound found in the Thunder of God vine, was identified based on its ability to enhance cell death in TMZ-resistant melanoma cells. A cell proliferation assay was used to compare the growth-inhibitory effects of TMZ alone versus TMZ/CEL combination treatment. Cytotoxic synergy was assessed using combination-index methods. The expression of nuclear factor-kappaB (NF-kappaB), IkappaB, mitogen-activated protein kinase, and ubiquitinated proteins were examined using Western blotting, and the localization of NF-kappaB in CEL-treated melanoma cells was evaluated using immunofluorescence microscopy. The CEL/TMZ combination synergistically inhibited cell proliferation in melanoma cells. CEL treatment increased the levels of ubiquitinated proteins, reduced the levels of tumor necrosis factor-alpha-induced IkappaB phosphorylation, and blocked NF-kappaB translocation to the nucleus. Inhibition of NF-kappaB with small interfering RNA mimicked the ability of CEL to sensitize melanoma cells to TMZ, suggesting that inhibition of NF-kappaB may play a role in TMZ/CEL-induced cytotoxicity. The TMZ/CEL combination induced the phosphorylation of c-Jun NH(2)-terminal kinase, implicating the mitogen-activated protein kinase pathway in the treatment effects. Our data suggest that CEL may be effective in sensitizing resistant melanoma cells to the effects of TMZ.
authors.affiliation: The Cutaneous Biology Program, The Ronald O. Perelman Department of Dermatology, NYU School of Medicine, New York, NY 10016, USA.
authors.forename: Ming
authors.initials: M
authors.lastname: Chen
authors.forename: Amy E
authors.initials: AE
authors.lastname: Rose
authors.forename: Nicole
authors.initials: N
authors.lastname: Doudican
authors.forename: Iman
authors.initials: I
authors.lastname: Osman
authors.forename: Seth J
authors.initials: SJ
authors.lastname: Orlow
mesh_terms: D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D003606:Dacarbazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D004354:Drug Screening Assays, Antitumor; D004357:Drug Synergism; D055785:Gene Knockdown Techniques; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism; D020128:Inhibitory Concentration 50; D008545:Melanoma / Q000201:enzymology / Q000473:pathology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism; D053978:Pentacyclic Triterpenes; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D034741:RNA, Small Interfering; D000077204:Temozolomide; D014315:Triterpenes / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D057149:Ubiquitinated Proteins
chemical_list: D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D053978:Pentacyclic Triterpenes; D034741:RNA, Small Interfering; D014315:Triterpenes; D014409:Tumor Necrosis Factor-alpha; D057149:Ubiquitinated Proteins; D000072000:NF-KappaB Inhibitor alpha; D003606:Dacarbazine; D020928:Mitogen-Activated Protein Kinases; D046988:Proteasome Endopeptidase Complex; C050414:celastrol; D000077204:Temozolomide