title: Functional evaluation of a fluorescent schweinfurthin: mechanism of cytotoxicity and intracellular quantification.
abstract: Schweinfurthins are potent inhibitors of cancer cell growth, especially against human central nervous system tumor lines such as SF-295 cells. However, the mechanisms through which these compounds impede cell growth are not fully understood. In an effort to understand the basis for the effects of schweinfurthins, we present a fluorescent schweinfurthin, 3-deoxyschweinfurthin B-like p-nitro-bis-stilbene (3dSB-PNBS), which displays biological activity similar to that of 3-deoxyschweinfurthin B (3dSB). These two schweinfurthins retain the unique differential activity of the natural schweinfurthins, as evidenced by the spindle-like morphological changes induced in SF-295 cells and the unaltered appearance of human lung carcinoma A549 cells. We demonstrate that incubation with 3dSB or 3dSB-PNBS results in cleavage of poly-ADP-ribose polymerase (PARP) and caspase-9, both markers of apoptosis. Coincubation of 3dSB or 3dSB-PNBS with the caspase-9 inhibitor (Z)-Leu-Glu(O-methyl)-His-Asp(O-methyl)-fluoromethylketone prevents PARP cleavage. Therapeutic agents that induce apoptosis often activate cellular stress pathways. A marker for multiple stress pathways is the phosphorylation of eukaryotic initiation factor 2α, which is phosphorylated in response to 3dSB and 3dSB-PNBS treatment. Glucose-regulated protein 78 and protein disulfide isomerase, both endoplasmic reticulum chaperones, are up-regulated with schweinfurthin exposure. Using the fluorescent properties of 3dSB-PNBS and dimethoxyphenyl-p-nitro-bis-stilbene (DMP-PNBS), a control compound, we show that the intracellular levels of 3dSB-PNBS are higher than those of Rhodamine 123 or DMP-PNBS in SF-295 and A549 cells.
authors.affiliation: Department of Internal Medicine, University of Iowa, Iowa City, Iowa 52242, USA.
authors.forename: Craig H
authors.initials: CH
authors.lastname: Kuder
authors.forename: Ryan M
authors.initials: RM
authors.lastname: Sheehy
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Neighbors
authors.forename: David F
authors.initials: DF
authors.lastname: Wiemer
authors.forename: Raymond J
authors.initials: RJ
authors.lastname: Hohl
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D053453:Caspase 9 / Q000378:metabolism; D045744:Cell Line, Tumor; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D000091342:Endoplasmic Reticulum Chaperone BiP; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism; D005456:Fluorescent Dyes / Q000494:pharmacology*; D005909:Glioblastoma / Q000188:drug therapy / Q000473:pathology; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D019704:Protein Disulfide-Isomerases / Q000378:metabolism; D013267:Stilbenes / Q000494:pharmacology*; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; D015852:Eukaryotic Initiation Factor-2; D005456:Fluorescent Dyes; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D008565:Membrane Proteins; D013267:Stilbenes; C018551:glucose-regulated proteins; C117233:schweinfurthin B; D011065:Poly(ADP-ribose) Polymerases; D053453:Caspase 9; D019704:Protein Disulfide-Isomerases