Anti-XIAP (G81) Antibody (A25633)

Cisplatin is a first-line chemotherapy drug against ovarian cancer. However, its strong toxic side effects and the development of cisplatin resistance in human cancer cells seriously influence the effects of chemotherapy and quality of life in patients. Noscapine (Nos), a non-toxic benzylisoquinoline alkaloid extracted from opium, has been recently reported to have anti-cancer activity, but the mechanism of that effect has not been clearly established. In the present study, we investigated cytotoxicity of Nos in combination with cisplatin (DDP) in drug-resistant human ovarian cancer cell line SKOV3/DDP in vitro and in vivo null mice xenograft model. Cell proliferation was measured by MTT assay, flow cytometry was used to analyze cell cycle and apoptosis, protein expression of several apoptotic factors was investigated by flow cytometry and immunohistochemical method, and their mRNA expression levels were determined by real-time PCR. In vitro experiments showed that Nos significantly inhibited proliferation of SKOV3/DDP cells. DDP/Nos-combined treatment notably enhanced DDP-induced inhibition of cell proliferation and increased the pro-apoptotic effect of DDP in SKOV3/DDP cells. DDP/Nos administration increased the proportion of G2/M cells, reduced both protein and mRNA expression of anti-apoptotic factors XIAP, surviving and NF-kB, and augmented protein and mRNA levels of pro-apoptotic caspase-3. In vivo experiments revealed that Nos/DDP treatment increased the apoptotic rate of xenograft tumors in null mice. Tumor volume decreased from 1.733 ± 0.155 g in mice treated with DDP alone to 1.191 ± 0.106 g in animals treated with Nos/DDP. These observations suggest that Nos increases the anti-cancer activity of DDP against the drug-resistant ovarian cancer cell line SKOV3/DDP by modulating the cell cycle and activating apoptotic pathways. The study provides a new chemotherapy strategy for the treatment of DDP-resistant human ovarian cancer.

Hepatocellular carcinoma remains one of the most prevalent malignancies worldwide. Curcuma aromatica and Polygonum cuspidatum are one of the commonly used paired-herbs for liver cancer treatment. Curcumin and resveratrol are the major anticancer constituents of Curcuma aromatica and Polygonum cuspidatum, respectively. Curcumin and resveratrol have been found to exhibit a synergistic anticancer effect in colon cancer. However, the combined effect of curcumin and resveratrol against hepatocellular carcinoma remains unknown. In the present study, we evaluated the combined effects of curcumin and resveratrol in hepatocellular carcinoma Hepa1-6 cells. The results showed that curcumin and resveratrol significantly inhibited the proliferation of Hepa1-6 cells in a dose- and time-dependent manner. The combination treatment of curcumin and resveratrol elicited a synergistic antiproliferative effect in Hepa1-6 cells. The apoptosis of Hepa1-6 cells induced by the combination treatment with curcumin and resveratrol was accompanied by caspase-3, -8 and -9 activation, which was completely abrogated by a pan caspase inhibitor, Z-VAD-FMK. Combination of curcumin and resveratrol upregulated intracellular reactive oxygen species (ROS) levels in Hepa1-6 cells. The ROS scavenger, NAC, partially attenuated the apoptosis and caspase activation induced by the combination treatment of curcumin and resveratrol. In addition, the combination of curcumin and resveratrol downregulated XIAP and survivin expression. These data suggest that the combination treatment of curcumin and resveratrol is a promising novel anticancer strategy for liver cancer. The present study also provides new insights into the effective mechanism of paired-herbs in traditional Chinese medicine.

Oxaliplatin is one of the agents used against colorectal cancer. Using PEG-liposome encapsulated oxaliplatin may enhance the accumulation of drugs in tumor cells, inducing apoptosis. However, the mechanism of action of PEG-liposome encapsulated oxaliplatin remains unclear. SW480 human colorectal cancer cells were treated with empty PEG-liposomes, free oxaliplatin or PEG-liposomal oxaliplatin. Cell cycle and apoptosis were assessed using fluorescence confocal microscopy and terminal deoxynucleotidyl transferase-mediated dUTP-fluorescein nick-end-labeling (TUNEL). Western blotting was used to analyze the expression of pro-apoptotic, anti-apoptotic and cyclin proteins. We found that PEG-liposomal oxaliplatin induced a stronger apoptotic response than empty PEG-liposomes or free oxaliplatin. Moreover, expression of Cyclin D1 increased, whereas expression of Cyclin A decreased after treatment with PEG-liposomal oxaliplatin. Furthermore, the cell cycle was arrested in the G1 phase. The results presented here indicate that PEG-liposome entrapment of oxaliplatin enhances the anticancer potency of the chemotherapeutic agent. The effect of PEG-liposomal oxaliplatin on apoptosis of SW480 human colorectal cancer cells may be through regulation of expression of Cyclin A or Cyclin D1, as well as pro-apoptotic and anti-apoptotic proteins.

MicroRNAs (miRNAs) have been shown to play critical roles in regulating the progress of leukemia. We performed miRNA expression profile in six Chinese patients with chronic lymphocytic leukemia (CLL), and in peripheral B cells from pooled 30 healthy donors, using a platform containing 866 human miRNAs. The most frequent changes in miRNAs in CLL cells included downregulation of miR-126, miR-572, miR-494, miR-923, miR-638, miR-130a, miR-181a and miR-181b and up-regulation of miR-29a, miR-660, miR-20a, miR-106b, miR-142-5p, miR-101, miR-30b, miR-34a, miR-let-7f, miR-21 and miR-155. Among the miRNAs down-regulated in CLL cells, we showed that miR-181a/b expression levels were significantly lower in poor prognostic subgroups defined by unmutated immunoglobulin heavy chain variable status and p53 aberrations. Furthermore, under-expression of miR-181a and miR-181b was associated with shorter overall survival and treatment-free survival in CLL patients. We further evaluated fludarabine-induced apoptosis after transfection of primary CLL cells from 40 patients with miR-15a, miR-16-1, miR-34a, miR-181a and miR-181b mimics. Transfection of miR-34a, miR-181a and miR-181b mimics into CLL cells from p53 wild-type patients led to significant increase in apoptosis compared with miRNA control. However, enforced expression of these miRNAs had no effect on B-CLL cells from p53-attenuated patients. We further demonstrated that miR-181a and miR-181b inhibiting BCL-2, MCL-1 and X-linked inhibitor of apoptosis protein by direct binding to 3'UTR. Thus, these results suggest that miR-181a/b may play important roles in the pathogenesis of CLL and may provide a possible therapeutic avenue and a sensitive indicator of the activity of the p53 axis in CLL.

Notch receptor signaling pathway (NRSP) is increasingly linked to carcinogenesis. Non-small cell lung cancer (NSCLC) appears to actively utilize this conserved developmental pathway. The aims of this study are to determine whether or not Notch 1-4 are overexpressed in NSCLC tissues compared with normal lung tissues and whether inhibiting NRSP could induce caspase-dependent or caspase-independent apoptosis. Immunohistochemistry was used to evaluate the expression of Notch 1-4 in 101 NSCLC tissue samples and 30 normal lung tissue samples. DAPT was used to repress NRSP in SK-MES-1 cells. Apoptosis was determined by Annexin V and PI staining. Cleaved poly ADP-ribose polymerase (PARP) was measured by Western blot; X-linked inhibitor of apoptosis protein (XIAP) and Survivin were assessed by qRT-PCR and Western blot; the release of second mitochondria-derived activator of caspase (Smac) from mitochondria to cytoplasm was evaluated by Western blot; the subcellular locations of endonuclease G (Endo G) and apoptosis inducing factor (AIF) were observed by Western blot and indirect immunofluorescence analysis. (Mech Dev, 98, 2000, 95) Notch 1-4 are up-regulated in NSCLC tissues and Notch 1, 2 are positively correlated with lymph node metastasis, (Proc Natl Acad Sci U S A, 106, 2009, 22293) DAPT treatment could inhibit NRSP and induce apoptosis, with a marked increase in cleaved PARP, decreases in XIAP and Survivin proteins and concomitant release of Smac, EndoG, and AIF from mitochondria, indicating that inhibiting NRSP by DAPT triggers caspase-dependent and caspase-independent apoptosis.

NF-κB plays a major role in the pathology of multiple myeloma. Here, we intended to investigate the regulating effect of cardamonin on NF-κB in myeloma cells. We found for the first time that cardamonin suppressed viability and induced apoptosis of myeloma cells. Cardamonin activated caspase-3 and PARP and suppressed the expression of various anti-apoptotic proteins. We discovered that NF-κB was repressed by cardamonin through suppression of IKK expression and IκBα phosphorylation. Furthermore, the expression of NF-κB-regulated gene products ICAM-1, COX-2 and VEGF was down-regulated by cardamonin. These results suggest that cardamonin blocks NF-κB pathway in human multiple myeloma cells.

OBJECTIVE:

To study the antitumor effects and associated mechanisms of extract of the Smilax china L. rhizome (SCR) on ovarian cancer cells.

METHODS:

Ovarian cancer cells A2780 were treated with different concentrations of SCR extract (SCRE), and compared with controls. Effects on cell growth were evaluated by cell counting kit-8 (CCK-8) assay; proliferation effects by EdU incorporation assay; cell cycle by propidium iodide staining; apoptosis by annexin V-fluorescein isothiocyanate/propidium iodide; cellular distribution of nuclear factor-κB (NF-κB) by immunofluorescence; protein levels of NF-κB, caspase-3, poly-adenosine diphosphate (ADP)-ribose polymerase (PARP), Bcl-2-associated X protein (Bax), cellular inhibitor of apoptosis (cIAP)-1, anti-X-linked inhibitor of apoptosis protein (XIAP), B-cell lymphoma-extra large (Bcl-XL), B-cell lymphoma-2 (Bcl-2) and AKT by Western blotting; and effects of SCRE combined with cisplatin or adriamycin on A2780 cells by CCK-8 assay.

RESULTS:

SCRE suppressed A2780 cell proliferation in a dose-dependent manner (P<0.05,P<0.01), arrested cells in G2/M phase and induced apoptosis by activating caspase-3, PARP and Bax. SCRE treatment also correlated with inhibition of NF-κB and downregulation of Bcl-2, Bcl-XL, cIAP-1, XIAP and AKT. SCRE can promote chemosensitivity to cisplatin and adriamycin in A2780 cells (P<0.01).

CONCLUSION:

SCR effectively inhibits NF-κB, induces apoptosis and reduces chemoresistance to cisplatin and adriamycin in ovarian cancer cells, which might be its molecular basis for treating ovarian cancer.

AIM:

To investigate the effects and underlying mechanisms of plumbagin, a naphthoquinone derived from medicinal plant Plumbago zeylanica, on human gastric cancer (GC) cells.

METHODS:

Human gastric cancer cell lines SGC-7901, MKN-28, and AGS were used. The cell viability was examined using CCK-8 viability assay. Cell proliferation rate was determined using both clonogenic assay and EdU incorporation assay. Apoptosis was detected via Annexin V/propidium iodide double-labeled flow cytometry. Western blotting was used to assess the expression of both NF-κB-regulated gene products and TNF-α-induced activation of p65, IκBα, and IKK. The intracellular location of NF-κB p65 was detected using confocal microscopy.

RESULTS:

Plumbagin (2.5-40 μmol/L) concentration-dependently reduced the viability of the GC cells. The IC(50) value of plumbagin in SGC-7901, MKN-28, and AGS cells was 19.12, 13.64, and 10.12 μmol/L, respectively. The compound (5-20 μmol/L) concentration-dependently induced apoptosis of SGC-7901 cells, and potentiated the sensitivity of SGC-7901 cells to chemotherapeutic agents TNF-αand cisplatin. The compound (10 μmol/L) downregulated the expression of NF-κB-regulated gene products, including IAP1, XIAP, Bcl-2, Bcl-xL, tumor factor (TF), and VEGF. In addition to inhibition of NF-κB p65 nuclear translocation, the compound also suppressed TNF-α-induced phosphorylation of p65 and IKK, and the degradation of IκBα.

CONCLUSION:

Plumbagin inhibits cell growth and potentiates apoptosis in human GC cells through the NF-κB pathway.

PURPOSE:

MicroRNAs (miRNAs) are short non-coding RNA molecules, which post-transcriptionally regulate genes expression and play crucial roles in diverse biological processes. Recent studies have shown that dysregulation of miRNAs might modulate the resistance of cancer cells to anti-cancer drugs, yet the modulation mechanism is not fully understood. We aimed to investigate the possible role of miRNAs in the development of multidrug resistance (MDR) in human gastric and lung cancer cell lines.

METHODS:

miRNA Quantitative real-time PCR was used to detect the different miRNA expression levels between drug resistant and parental cancer cells. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay was used to test the drug-resistant phenotype changes in cancer cells via over or downregulation of miRNAs. Dual-luciferase activity assay was used to verify the target genes of miRNAs. Western blot analysis and apoptosis assay were used to elucidate the mechanism of miRNAs on modulating drug resistance in cancer cells.

RESULTS:

miR-200bc/429 cluster was downregulated, while BCL2 and XIAP were upregulated in both MDR SGC7901/VCR (vincristine) and A549/CDDP (cisplatin) cells, compared with the parental SGC7901 and A549 cells, respectively. Overexpression of miR-200bc/429 cluster sensitized SGC7901/VCR and A549/CDDP cells to anti-cancer drugs, respectively. Both BCL2 and XIAP 3'-UTR reporters constructed in MDR cells suggested that BCL2 and XIAP were the common target genes of the miR-200bc/429 cluster. Enforced miR-200bc/429 cluster expression reduced BCL2 and XIAP protein level and sensitized both MDR cells to VCR-induced and CDDP-induced apoptosis, respectively.

CONCLUSIONS:

Our findings first suggest that miR-200bc/429 cluster could play a role in the development of MDR in both gastric and lung cancer cell lines, at least in part by modulation of apoptosis via targeting BCL2 and XIAP.