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BackgroundIt was well known that the clinical use of chemotherapeutic drugs is restricted by severe adverse reactions and drug resistances. Thus it is necessary to figure out a strategy to increase the specific anti-tumor efficiency of chemotherapeutic drugs. Apigenin, a kind of flavonoids, has been reported to possess anticancer activities with very low cytotoxicity to normal tissue.Methodology/Principal FindingsOur results from cell viability assay, western-blots and TdT-mediated dUTP-biotin nick end labeling (TUNEL) assay demonstrated the synergistic pro-apoptotic effects of a low dose of apigenin and paclitaxel in human cancer cell lines. To analyze the underlying mechanism, we examined reactive oxygen species (ROS) staining after cells were treated with a combination of apigenin and paclitaxel, or each of them alone. Data from flow-cytometry showed that superoxides but not reduction of peroxides accumulated in HeLa cells treated with apigenin or a combination of apigenin and paclitaxel. Apigenin and paclitaxel-induced HeLa cell apoptosis was related to the level of ROS in cells. We further evaluated activity and protein level of superoxide dismutase (SOD). Apigenin significantly inhibited SOD activity but did not alter the SOD protein level suggesting that apigenin promoted ROS accumulation through suppressing enzyme activity of SOD. Addition of Zn2+, Cu2+ and Mn2+ to cell lysates inhibited apigenin's effects on SOD activity. At the same time, data from caspase-2 over-expression and knocked-down experiments demonstrated that caspase-2 participated in apigenin and paclitaxel-induced HeLa cell apoptosis.Conclusions/SignificanceTaken together, our study demonstrated that apigenin can sensitize cancer cells to paclitaxel induced apoptosis through suppressing SOD activity, which then led to accumulation of ROS and cleavage of caspase-2, suggesting that the combined use of apigenin and paclitaxel was an effective way to decrease the dose of paclitaxel taken.

Inducible heat shock protein70 (HSP70) is one of the most important HSPs for maintenance of cell integrity during normal cellular growth as well as pathophysiological conditions. Apoptosis signal-regulating kinase (ASK) 1, a mammalian MAPKKK, activates the JNK and p38 pathways. Here we report a novel function of HSP70 in regulating TNF-alpha-induced cell apoptosis. Our study demonstrated that HSP70 physically interacted with ASK1 and promoted the ubiquitin-dependent proteasomal degradation of ASK1. CHIP (carboxyl terminus of the HSC70-interacting protein) which acted as a co-chaperone of HSP70 cooperated with HSP70 in regulating ASK1. We also found that TNF-alpha stimulated HSP70/CHIP/ASK1 association and through cooperating with CHIP, HSP70 inhibits TNF-alpha-induced cell apoptosis both in over-expression and RNAi conditions. Structural analysis indicated that C-terminal domain of HSP70 was necessary for ASK1 degradation, and N- terminal domain of ASK1 was essential for its binding to HSP70. All these findings indicated that HSP70 and CHIP association is important for HSP70 in interacting with ASK1. Through forming the complex of HSP70/CHIP/ASK1, HSP70 promotes ASK1 proteasomal degradation and prevents TNF-alpha-induced cell apoptosis.

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BackgroundAbnormal activity of STAT3 is associated with a number of human malignancies. Hsp90 plays a central role in stabilizing newly synthesized proteins and participates in maintaining the functional competency of a number of signaling transducers involved in cell growth, survival and oncogenesis, such as STAT3. Hsp90 interacts with STAT3 and stabilizes Tyr-phosphorylated STAT3. It has been reported that luteolin possesses anticancer activity through degradation of Tyr705-phosphorylated STAT3.Methodology/Principal FindingsWe found that overexpression of Hsp90 inhibited luteolin-induced degradation of Tyr705-phosphorylated STAT3 and luteolin also reduced the levels of some other Hsp90 interacting proteins. Results from co-immunoprecipitation and immunoblot analysis demonstrated that luteolin prevented the association between Hsp90 and STAT3 and induced both Tyr705- and Ser727-phosphorylated STAT3 degradation through proteasome-dependent pathway. The molecular modeling analysis with CHARMm–Discovery Studio 2.1(DS 2.1) indicated that luteolin could bind to the ATP-binding pocket of Hsp90. SPR technology-based binding assay confirmed the association between luteolin and Hsp90. ATP-sepharose binding assay displayed that luteolin inhibited Hsp90-ATP binding.Conclusions/SignificanceLuteolin promoted the degradation of Tyr705- and Ser727-phosphorylated STAT3 through interacting with Hsp90 and induced apoptosis of cancer cells. This study indicated that luteolin may act as a potent HSP90 inhibitor in antitumor strategies.