Pancreatic cancer is a disease with an extremely poor prognosis. Tumor protein 53-induced nuclear protein 1 ( TP53INP1 ) is a proapoptotic stress-induced p53 target gene. In this article, we show by immunohistochemical analysis that TP53INP1 expression is dramatically reduced in pancreatic ductal adenocarcinoma (PDAC) and this decrease occurs early during pancreatic cancer development. TP53INP1 reexpression in the pancreatic cancer-derived cell line MiaPaCa2 strongly reduced its capacity to form s.c., i.p., and intrapancreatic tumors in nude mice. This anti-tumoral capacity is, at least in part, due to the induction of caspase 3-mediated apoptosis. In addition, TP53INP1 −/− mouse embryonic fibroblasts (MEFs) transformed with a retrovirus expressing E1A/ras V12 oncoproteins developed bigger tumors than TP53INP1 +/+ transformed MEFs or TP53INP1 −/− transformed MEFs with restored TP53INP1 expression. Finally, TP53INP1 expression is repressed by the oncogenic micro RNA miR-155, which is overexpressed in PDAC cells. TP53INP1 is a previously unknown miR-155 target presenting anti-tumoral activity.
OBJECTIVE To determine whether a specifically designed bispecific ( Bcl‐2/Bcl‐xL ) antisense oligonucleotide (ASO) induces apoptosis and enhances chemosensitivity in human prostate cancer LNCaP cells, as Bcl‐2 and Bcl‐xL are both anti‐apoptotic genes associated with treatment resistance and tumour progression in many malignancies, including prostate cancer. MATERIALS AND METHODS Inhibition of Bcl‐2 and Bcl‐xL expression by the bispecific ASO was evaluated using real‐time reverse transcription‐polymerase chain reaction and Western blotting, while growth inhibition and induction of apoptosis were analysed by a crystal violet assay, flow cytometry and Western blotting of apoptosis‐relevant proteins. The effect of combined treatment with bispecific ASO and chemotherapy or small‐interference RNA (siRNA) targeting the clusterin gene was also investigated. RESULTS Bispecific ASO reduced Bcl‐2 and Bcl‐xL expression in LNCaP cells in a dose‐dependent manner. There was cell growth inhibition, increases in the sub‐G0–G1 fraction, and cleavage of caspase‐3 and poly(ADP‐Ribose) polymerase proteins in LNCaP cells after bispecific ASO treatment. Interestingly, Bcl‐2 / Bcl‐xL bispecific ASO treatment also resulted in the down‐regulation of Mcl‐1 and up‐regulation of Bax. The sensitivity of LNCaP cells to mitoxantrone, docetaxel or paclitaxel was significantly increased, reducing the 50% inhibitory concentration by 45%, 80% or 90%, respectively. Furthermore, the apoptotic induction by Bcl‐2 / Bcl‐xL bispecific ASO was synergistically enhanced by siRNA‐mediated inhibition of clusterin, a cytoprotective chaperone that interacts with and inhibits activated Bax. CONCLUSIONS These findings support the concept of the targeted suppression of Bcl‐2 anti‐apoptotic family members using multitarget inhibition strategies for prostate cancer, through the effective induction of apoptosis.
Berberis (Barberry) species contain an important amount of alkaloids and are commonly used in traditional medicine. Alkaloids are preponderantly characterized by their numerous properties including anticancer activity. The aims of the present study were to investigate the effect and the mechanism of action of the Algerian Berberis hispanica alkaloids extract (BHAE) on Hep-2 laryngeal cancer cells. In this purpose, total alkaloids were extracted from the roots bark of the plant, then, were submitted to HPLC analysis. In order to determine the cells inhibition concentration (IC50), we used the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cells were enumerated using Trypan blue exclusion assay and viability by cell cycle analysis. The morphology was assessed by coloration with May Grunwald Giemsa (MGG). The evaluation of such stress markers as malondialdehyde (MDA), cytochrome c, p38MAPK, NF-kB, Akt, Erk 1/ 2 and p53 was assessed by colorimetric assays and western blotting. Our results demonstrated that BHAE contains Berberine, an isoquinoline plant alkaloid. The IC50 was determined at 75 μg/mL and the treatment of Hep-2 cells with this dose showed anti-proliferative effect and increased SubG0 cell population (cells undergoing apoptosis) in treated cells (15.4%). Further, the coloration of fixed cells showed that BHAE induced cells density and morphological changes. Moreover, our results showed that the treatment induced elevation of MDA, cytochrome c, p38MAPK and NF-kB level, but did not affect Akt and Erk 1/2 rates. We have also observed that the treatment with BHAE enhanced the expression of p53. Taken together, our results suggest that the BHAE triggered cell death by activation of apoptosis through ROS induction.
Bcl-2 and Bcl-xL are associated with treatment resistance and progression in many cancers, including prostate cancer. The objective of this study was to determine whether a novel bispecific antisense oligonucleotide targeting both Bcl-2 and Bcl-xL induces apoptosis and enhances chemosensitivity in androgen-independent PC3 prostate cancer cells. An antisense oligonucleotide with complete sequence identity to Bcl-2 and three-base mismatches to Bcl-xL selected from five antisense oligonucleotides targeting various regions with high homology between Bcl-2 and Bcl-xL was found to be the most potent inhibitor of both Bcl-2 and Bcl-xL expression in PC3 cells. This selected Bcl-2/Bcl-xL bispecific antisense oligonucleotide reduced mRNA and protein levels in a dose-dependent manner, reducing Bcl-2 and Bcl-xL protein levels to 12% and 19%, respectively. Interestingly, Mcl-1 was down-regulated as well, although levels of Bax, Bad, or Bak were not altered after treatment with this bispecific antisense oligonucleotide. Indirect down-regulation of inhibitor of apoptosis (IAP) family, including XIAP, cIAP-1 and cIAP-2, via second mitochondria-derived activator of caspases was also observed after bispecific antisense oligonucleotide treatment. Executioner caspase-3, caspase-6, and caspase-7 were shown to be involved in apoptosis induced by bispecific antisense oligonucleotide. This Bcl-2/Bcl-xL bispecific antisense oligonucleotide also enhanced paclitaxel chemosensitivity in PC3 cells, reducing the IC50 of paclitaxel by >90%. These findings illustrate that combined suppression of antiapoptotic Bcl-2 family members using this antisense oligonucleotide could be an attractive strategy for inhibiting cancer progression through alteration of the apoptotic rheostat in androgen-independent prostate cancer.
A fluorinated bola-amphiphilic dendrimer, sensitive to the reactive oxygen species, and its application in specific delivery of small interfering RNA in cancer cells, is reported by L. Peng and co-workers on page 8594. The dendrimer combines both lipid and dendrimer delivery advantages along with fluorine tags enabling 19F NMR studies on the delivery process, constituting a promising vector for on-demand stimulus-responsive delivery.
Abstract siRNA delivery remains a major challenge in RNAi‐based therapy. Here, we report for the first time that an amphiphilic dendrimer is able to self‐assemble into adaptive supramolecular assemblies upon interaction with siRNA, and effectively delivers siRNAs to various cell lines, including human primary and stem cells, thereby outperforming the currently available nonviral vectors. In addition, this amphiphilic dendrimer is able to harness the advantageous features of both polymer and lipid vectors and hence promotes effective siRNA delivery. Our study demonstrates for the first time that dendrimer‐based adaptive supramolecular assemblies represent novel and versatile means for functional siRNA delivery, heralding a new age of dendrimer‐based self‐assembled drug delivery in biomedical applications.
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