ptosis resulting from Ad eIF5A1 infection suggesting that, althou

ptosis resulting from Ad eIF5A1 infection suggesting that, although p53 is up regulated in re sponse to eIF5A1, it is not required for apoptosis. Normal lung fibroblasts are resistant to Ad eIF5A1 induced apoptosis The ability to kill malignant cells without harming normal cells is an important feature of an ideal cancer therapy drug. In order to assess the specificity of eIF5A1 over e pression for inducing apoptosis in cancer cells rather than non malignant cells, A549 lung carcinoma cells and WI 38 normal lung fibroblast Inhibitors,Modulators,Libraries cells were ana lyzed for induction of apoptosis by Anne in propidium iodide staining following infection of Ad eIF5A1 or Ad eIF5A1K50A. EIF5A1 and eIF5A1K50A induced apoptosis in 7% and 8% of WI 38 normal lung fibroblast cells forty eight hours after infection, respec tively.

However, A549 cells were more sensitive to eIF5A induced apoptosis with 16% and 19% of cells undergoing apoptosis forty eight hours after infection with Ad eIF5A1 or Inhibitors,Modulators,Libraries Ad eIF5A1K50A, respectively. Similar results were observed seventy two hours after infection, confirming that WI 38 cells were resistant to eIF5A1 induced apoptosis in spite of virus mediated eIF5A1 e pression levels comparable to those in A549 cells. In contrast, the cytoto ic drug Actino mycin D, an inhibitor of DNA dependent RNA synthesis, induced comparable levels of apoptosis in both normal Inhibitors,Modulators,Libraries and malignant cells. ERK and p38 MAPK activation in A549 lung carcinoma cells and WI 38 lung fibroblast cells was analyzed by immunoblotting after treatment with adenovirus.

Activation of p38 MAPK was observed in response to Ad eIF5A1 and Ad eIF5A1K50A infection in both A549 cells Inhibitors,Modulators,Libraries and WI 38 cells. However, Ad eIF5A1 and Ad eIF5A1K50A induced only a modest 2 fold increase in phosphorylated p38 in WI 38 cells. In contrast, A549 cells, which displayed Entinostat greater sensitivity to eIF5A1 induced apoptosis, e hibited a greater than 10 fold increase in levels of phosphorylated p38 MAPK. These data suggest that over e pression of eIF5A1, and ensuing activation of p38 MAPK signaling, act as a more potent inducer of cell death in malignant A549 cells than in normal lung cells. In addition, ERK MAPK was activated in response to Ad eIF5A1 or Ad eIF5A1K50A infection in malignant A549 cells, but not in WI 38 cells. E pression levels of the pro survival Bcl 2 protein were found to be much higher in WI 38 cells than A549 cells, which may also have contributed to survival of these cells.

Discussion The development of cancer gene therapies requires agents that target pathways that ma imize anti cancer activity. EIF5A1 has been identified as a viable cancer target that can be adapted for use in gene therapy approaches since its over e pression has been demonstrated thenthereby to induce apoptosis in a wide variety of cancer types. As well, suppression of hypusinated eIF5A1 using a small interfering RNA has been shown to inhibit activa tion of Nuclear Factor kappa B and ERK MAPK in multiple myeloma cells and to potentiate the pro apoptoti

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