BackgroundEmodin has been showed to induce apoptosis of pancreatic cancer cells and inhibit tumor growth in our previous studies. This study was designed to investigate whether emodin could inhibit the angiogenesis of pancreatic cancer tissues and its mechanism.Methodology/Principal FindingIn accordance with our previous study, emodin inhibited pancreatic cancer cell growth, induced apoptosis, and enhanced the anti-tumor effect of gemcitabine on pancreatic caner cells in vitro and in vivo by inhibiting the activity of NF-κB. Here, for the first time, we demonstrated that emodin inhibited tumor angiogenesis in vitro and in implanted pancreatic cancer tissues, decreased the expression of angiogenesis-associated factors (NF-κB and its regulated factors VEGF, MMP-2, MMP-9, and eNOS), and reduced eNOS phosphorylation, as evidenced by both immunohistochemistry and western blot analysis of implanted tumors. In addition, we found that emodin had no effect on VEGFR expression in vivo.Conclusions/SignificanceOur results suggested that emodin has potential anti-tumor effect on pancreatic cancer via its dual role in the promotion of apoptosis and suppression of angiogenesis, probably through regulating the expression of NF-κB and NF-κB-regulated angiogenesis-associated factors.

Evodiamine has therapeutic potential against cancers. This study was designed to investigate whether combination therapy with gemcitabine and evodiamine enhanced antitumor efficacy in pancreatic cancer. In vitro application of the combination therapy triggered significantly higher frequency of pancreatic cancer cells apoptosis, inhibited the activities of PI3K, Akt, PKA, mTOR and PTEN, and decreased the activation of NF-κB and expression of NF-κB-regulated products. In vivo application of the combination therapy induced significant enhancement of tumor cell apoptosis, reductions in tumor volume, and inhibited activation of mTOR and PTEN. In conclusion, evodiamine can augment the therapeutic effect of gemcitabine in pancreatic cancer through direct or indirect negative regulation of the PI3K/Akt pathway.

As a critical apoptosis executioner, caspase-3 becomes activated and then enters into the nucleus to exert its function. However, the molecular mechanism of this nuclear entry of active caspase-3 is still unknown. In this study, we revealed that caspase-3 harbors a crm-1-independent nuclear export signal (NES) in its small subunit. Using reversecaspase-3 as the study model, we found that the function of the NES in caspase-3 was not disturbed by the conformational changes during induced caspase-3 activation. Mutations disrupting the cleavage activity or p3-recognition site resulted in a defect in the nuclear entry of active caspase-3. We provide evidence that the p3-mediated specific cleavage activity of active caspase-3 abrogated the function of the NES. In conclusion, our results demonstrate that during caspase-3 activation, NES is constitutively present. p3-mediated specific cleavage activity abrogates the NES function in caspase-3, thus facilitating the nuclear entry of active caspase-3.

Gemcitabine resistance is a common problem of pancreatic cancer chemotherapy, and how to reverse it plays an important role in the treatment of pancreatic cancer. This study investigated the effect of emodin on the gemcitabine-resistant pancreatic cancer cell line SW1990/Gem, and explored the potential mechanism of its action. SW1990/Gem was obtained by culture of the pancreatic cancer cell line SW1990 in vitro by intermittently increasing the concentration of gemcitabine in the culture medium for 10 months, observing the morphology using inverted microscopy. SW1990/Gem cells were pretreated with emodin (10 μM) for different periods followed by treatment with gemcitabine (20 μM) for 48 h; cell proliferation was tested by MTT assay. SW1990/Gem cells were treated by emodin with different concentrations for 48 h, cell apoptosis was detected by flow cytometry (FCM). The expression of gene and protein, such as MDR-1 (P-gp), NF-κB, Bcl-2, Bax, cytochrome-C (cytosol), caspase-9 and -3 were measured by RT-PCR and Western blotting. The function of P-gp in SW1990/Gem cells was checked by FCM. The results showed that the SW1990/Gem cells changed greatly in morphology and the resistance index was 48.63. Emodin promoted cell apoptosis of the gemcitabine-resistant cell line SW1990/Gem in a dose-dependent manner. Emodin enhanced the SW1990/Gem cell sensitivity to gemcitabine in a time-dependent manner. Emodin monotherapy or combination with gemcitabine both decreased the gene and protein expression levels of MDR-1 (P-gp), NF-κB and Bcl-2 and inhibited the function of P-gp, but increased the expression levels of Bax, cytochrome-C (cytosol), caspase-9 and -3, and promoted cell apoptosis. This demonstrated that emodin had a reversing effect on the gemcitabine-resistant cell line SW1990/Gem, possibly via decreasing the function of P-gp and activating the mitochondrial apoptosis pathway in vitro.

Gemcitabine is currently the best treatment available for pancreatic cancer (PaCa); however, patients with the disease develop resistance to the drug over time. Agents that can either enhance the effects of gemcitabine or overcome chemoresistance to the drug are required for the treatment of PaCa. Oridonin is one such agent which is safe and multitargeted, and has been linked with the suppression of survival, proliferation, invasion and angiogenesis of cancer. In this study, we investigated whether oridonin could sensitize PaCa to gemcitabine in vitro and in vivo. In vitro, oridonin inhibited the proliferation of the PaCa cell line, BxPC-3, potentiated the apoptosis induced by gemcitabine, induced G1 cell cycle arrest and activated p38 and p53; these results were significant when oridonin was combined with gemcitabine. In vivo, we found that oridonin significantly suppressed tumor growth and this effect was further enhanced by gemcitabine (P<0.05). Tumors from nude mice injected with BxPC-3 PaCa cells and treated with a combination of oridonin and gemcitabine showed a significant upregulation in p38 and p53 activation (P<0.05 vs. control, P<0.05 vs. gemcitabine or oridonin alone). Taken together, our results demonstrate that oridonin can potentiate the effects of gemcitabine in PaCa through the mitogen-activated protein kinase (MAPK)-p38 signaling pathway, which is dependent on p53 activation.