Because signal transducer and activator of transcription 3 (STAT3) is constitutively activated in most human solid tumors and is involved in the proliferation, angiogenesis, immune evasion, and antiapoptosis of cancer cells, researchers have focused on STAT3 as a target for cancer therapy. We screened for natural compounds that inhibit the activity of STAT3 using a dual-luciferase assay. Cryptotanshinone was identified as a potent STAT3 inhibitor. Cryptotanshinone rapidly inhibited STAT3 Tyr705 phosphorylation in DU145 prostate cancer cells and the growth of the cells through 96 hours of the treatment. Inhibition of STAT3 Tyr705 phosphorylation in DU145 cells decreased the expression of STAT3 downstream target proteins such as cyclin D1, survivin, and Bcl-xL. To investigate the cryptotanshinone inhibitory mechanism in DU145 cells, we analyzed proteins upstream of STAT3. Although phosphorylation of Janus-activated kinase (JAK) 2 was inhibited by 7 Mmol/L cryptotanshinone at 24 hours, inhibition of STAT3 Tyr705 phosphorylation occurred within 30 minutes and the activity of the other proteins was not affected. These results suggest that inhibition of STAT3 phosphorylation is caused by a JAK2-independent mechanism, with suppression of JAK2 phosphorylation as a secondary effect of cryptotanshinone treatment. Continuing experiments revealed the possibility that cryptotanshinone might directly bind to STAT3 molecules. Cryptotanshinone was colocalized with STAT3 molecules in the cytoplasm and inhibited the formation of STAT3 dimers. Computational modeling showed that cryptotanshinone could bind to the SH2 domain of STAT3. These results suggest that cryptotanshinone is a potent anticancer agent targeting the activation STAT3 protein. It is the first report that cryptotanshinone has antitumor activity through the inhibition of STAT3.

Heat shock factor 1 (HSF1) is the master switch for heat shock protein (HSP) expression in eukaryotes. A synthetic chemical library was screened to identify inhibitors of HSF1 using a luciferase reporter under the control of a heat shock element. A compound named KRIBB11 (N 2 -(1H-indazole-5-yl)-N 6 -methyl-3-nitropyridine-2,6-diamine) was identified for its activity in abolishing the heat shock-induced luciferase activity with an IC 50 of 1.2 mol/liter. When the cells were exposed to heat shock in the presence of KRIBB11, the induction of HSF1 downstream target proteins such as HSP27 and HSP70 was blocked. In addition, treatment of HCT-116 cells with KRIBB11 induced growth arrest and apoptosis. Markers of apoptosis, such as cleaved poly(ADPribose) polymerase, were detected after KRIBB11 treatment. Biotinyl-KRIBB11 was synthesized as an affinity probe for the identification of KRIBB11 target proteins. Using affinity chromatography and competition assays, KRIBB11 was shown to associate with HSF1 in vitro. Chromatin immunoprecipitation analysis showed that KRIBB11 inhibited HSF1-dependent recruitment of p-TEFb (positive transcription elongation factor b) to the hsp70 promoter. Finally, intraperitoneal treatment of nude mice with KRIBB11 at 50 mg/kg resulted in a 47.4% (p < 0.05) inhibition of tumor growth without body weight loss. Immunoblotting assays showed that the expression of HSP70 was lower in KRIBB11-treated tumor tissue than in control tissues. Because HSPs are expressed at high levels in a wide range of tumors, these results strengthen the rationale for targeting HSF1 in cancer therapy.The heat shock response (HSR) 4 was first reported in 1962 by Ritossa (1). Since then, many investigators have reported that the HSR is an evolutionarily conserved protective mechanism against a wide range of stresses, including heat shock, heavy metal, oxidative stress, fever, or protein misfolding (reviewed in Refs. 2, 3). The HSR is mediated by the heat shock factor family, which in mammalian cells is composed of three heat shock factors (HSF1, HSF2, and HSF4) that control the transcription of heat shock proteins (4, 5). Although HSF2 and HSF4 play a role in the HSR, HSF1 is the master regulator of the heat shock response in eukaryotes.RNA polymerase II (pol II) transcribes all mRNAs and has an extended carboxyl-terminal domain (CTD) in its largest subunit. This CTD consists of multiple repeats of the heptapeptide 1 YSPTSPS 7 . Before heat shock induction, pol II associates with the heat shock promoters, transcribes 20 -50 bases downstream of the transcription site, and stays there in an arrested inactive state (6, 7). Releasing pol II requires the recruitment and activation of HSF1. However, HSF1 alone is not sufficient to release arrested RNA pol II (8).HSF1 is required to recruit a second factor, p-TEFb, a heterodimer of CDK9 and cyclin T (8). The artificial recruitment of p-TEFb to the hsp70 promoter is sufficient for the induction of the hsp70 gene in the absence of heat shock (8). Phosphorylation of pol II Ser-2 o...

Structures of the protein-chromophore complex and the apoprotein form of neocarzinostatin were determined at 1.8 angstrom resolution. Neocarzinostatin is composed of a labile chromophore with DNA-cleaving activity and a stabilizing protein. The chromophore displays marked nonlinearity of the triple bonds and is bound noncovalently in a pocket formed by the two protein domains. The chromophore pi-face interacts with the phenyl ring edges of Phe52 and Phe78. The amino sugar and carbonate groups of the chromophore are solvent exposed, whereas the epoxide, acetylene groups, and carbon C-12, the site of nucleophilic thiol addition during chromophore activation, are unexposed. The position of the amino group of the chromophore carbohydrate relative to C-12 supports the idea that the amino group plays a role in thiol activation.

No abstract

Five compounds were isolated from the chloroform-soluble fraction of the methanolic extract of the dried rhizomes of Zingiber officinale (Zingiberaceae) through repeated column chromatography. Their chemical structures were elucidated as 4-, 6-, 8-, and 10-gingerols, and 6-shogaol using spectroscopic analysis. Among the five isolated compounds, 6-shogaol exhibited the most potent cytotoxicity against human A549, SK-OV-3, SK-MEL-2, and HCT15 tumor cells. 6-shogaol inhibited proliferation of the transgenic mouse ovarian cancer cell lines, C1 (genotype: p53(-/-), c-myc, K-ras) and C2 (genotype: p53(-/-), c-myc, Akt), with ED(50) values of 0.58 microM (C1) and 10.7 microM (C2).

No abstract

Cancer cells, compared to normal cells, are under oxidative stress associated with an elevated level of reactive oxygen species (ROS) and are more vulnerable to oxidative stress induced by ROS generating agents. Thus, manipulation of the ROS level provides a logical approach to kill cancer cells preferentially, without significant toxicity to normal cells, and great efforts have been dedicated to the development of strategies to induce cytotoxic oxidative stress for cancer treatment. Fenton reaction is an important biological reaction in which irons convert hydrogen peroxide (H2O2) to highly toxic hydroxyl radicals that escalate ROS stress. Here, we report Fenton reaction-performing polymer (PolyCAFe) micelles as a new class of ROS-manipulating anticancer therapeutic agents. Amphiphilic PolyCAFe incorporates H2O2-generating benzoyloxycinnamaldehyde and iron-containing compounds in its backbone and self-assembles to form micelles that serve as Nano-Fenton reactors to generate cytotoxic hydroxyl radicals, killing cancer cells preferentially. When intravenously injected, PolyCAFe micelles could accumulate in tumors preferentially to remarkably suppress tumor growth, without toxicity to normal tissues. This study demonstrates the tremendous translatable potential of Nano-Fenton reactors as a new class of anticancer drugs.

Background and purpose: Obovatol isolated from the medicinal herb Magnolia obovata exhibits a variety of biological activities. Here, the effect of obovatol and its mechanism of action on microglial activation, neuroinflammation and neurodegeneration were investigated. Experimental approach: In microglial BV-2 cells stimulated with lipopolysaccharide (LPS), we measured nitric oxide (NO) and cytokine production, and activation of intracellular signalling pathways by reverse transcription-polymerase chain reaction and Western blots. Cell death was assayed in co-cultures of activated microglia (with bacterial LPS) and neurons and in LPS-induced neuroinflammation in mice in vivo. Key results: Obovatol inhibited microglial NO production with an IC50 value of 10 mM. Obovatol also inhibited microglial expression of proinflammatory cytokines and inducible nitric-oxide synthase, which was accompanied by the inhibition of multiple signalling pathways such as nuclear factor kappa B, signal transducers and activators of transcription 1, and mitogen-activated protein kinases. In addition, obovatol protected cultured neurons from microglial toxicity and inhibited neuroinflammation in mice in vivo. One molecular target of obovatol in microglia was peroxiredoxin 2 (Prx2), identified by affinity chromatography and mass spectrometry. Obovatol enhanced the reactive oxygen species (ROS)-scavenging activity of Prx2 in vitro, thereby suppressing proinflammatory signalling pathways of microglia where ROS plays an important role. Conclusions and implications:Obovatol is not only a useful chemical tool that can be used to investigate microglial signalling, but also a promising drug candidate against neuroinflammatory diseases. Furthermore, our results indicate that Prx2 is a novel drug target that can be exploited for the therapeutic modulation of neuroinflammatory signalling.