ABSTRACT
Radiation therapy is frequently used to treat non-small cell lung cancers (NSCLCs). We have previously shown that a combination of ionizing radiation (IR) and the staurosporine analog PKC 412, but not Ro 31-8220, increases cell death in NSCLC cells. To identify genes involved in the enhancement of cell death, a total gene profiling in response to co-administration of (i) PKC 412 with IR, or (ii) Ro 31-8220 with IR was implemented. These combined treatments caused upregulation of 140 and 179 genes and downregulation of 253 and 425 genes, respectively. Certain genes were selected and verified by real-time quantitative PCR and, of these genes, robust suppression of Ephrin B3 expression was suggested as a possible cell death-inducing mechanism of combined treatment with IR and PKC 412. Indeed, silencing of Ephrin B3 using siRNA in NSCLC cells resulted in a major alteration of their morphology with an elongated phenotype, decreased proliferation and increased cell death signaling. Moreover, silencing of Ephrin B3 in combination with IR caused a decrease in IR-mediated G(2)-arrest, induced cellular senescence, inhibited MAPK ERK and p38 phosphorylation, and caused an upregulation of p27(kip1) expression. Finally, silencing of Ephrin B3 in combination with IR sensitized U-1810 cells to IR-induced apoptosis. In conclusion, we identify and describe Ephrin B3 as a putative signaling molecule involved in the response of NSCLC cells to combined treatment with PKC 412 and ionizing radiation.
Subject(s)
Apoptosis/drug effects , Apoptosis/radiation effects , Ephrin-B3/antagonists & inhibitors , Radiation, Ionizing , Staurosporine/analogs & derivatives , Carcinoma, Non-Small-Cell Lung/metabolism , Carcinoma, Non-Small-Cell Lung/pathology , Carcinoma, Non-Small-Cell Lung/therapy , Cell Line, Tumor , Combined Modality Therapy , Cyclin-Dependent Kinase Inhibitor p27/metabolism , Ephrin-B3/genetics , Ephrin-B3/metabolism , Extracellular Signal-Regulated MAP Kinases/metabolism , G2 Phase Cell Cycle Checkpoints/radiation effects , Humans , Phosphorylation/radiation effects , RNA Interference , RNA, Small Interfering/metabolism , Signal Transduction/drug effects , Staurosporine/pharmacology , Staurosporine/therapeutic use , Up-Regulation/radiation effects , p38 Mitogen-Activated Protein Kinases/metabolismABSTRACT
We have re-evaluated the selectivity of fumagillin against endothelial cell proliferation and compared it to the reported selectivity of its potent analog TNP-470. We showed that fumagillin does not inhibit endothelial cell proliferation in a specific manner, but on the contrary it inhibits the proliferation of other cell types at the same range of concentrations. Furthermore, the IC50 values of fumagillin for endothelial cells are two orders of magnitude higher than those values reported for TNP-470 on endothelial cells; on the contrary, the IC50 value of fumagillin for human breast cancer MDA-MB231 cells is four orders of magnitude lower than the value reported for TNP-470 on the same cell line.
Subject(s)
Angiogenesis Inhibitors/pharmacology , Endothelium, Vascular/drug effects , Fatty Acids, Unsaturated/pharmacology , 3T3 Cells/cytology , 3T3 Cells/drug effects , Animals , Aorta/cytology , Aorta/drug effects , Cattle , Cell Division/drug effects , Cells, Cultured , Cyclohexanes , Dose-Response Relationship, Drug , Endothelium, Vascular/cytology , Humans , Inhibitory Concentration 50 , Mice , O-(Chloroacetylcarbamoyl)fumagillol , Organ Specificity , Sesquiterpenes/pharmacology , Tumor Cells, CulturedABSTRACT
BACKGROUND: Ehrlich ascites tumor is an experimental tumor model very suitable for performing comparative studies relating its growth in vitro and in vivo. We used this tumor model to study the potential modulatory effects of genistein and 2-methoxyestradiol, two anti-angiogenic compounds, on the proteolytic balance MMP/TIMP. Ehrlich cells grown in vitro secreted MMP-9, MMP-2 and two TIMPs; the treatment with either of the anti-angiogenic compounds here tested stimulated all these activities, but the increase in TIMPs activities of genistein-treated cells were higher than those of MMPs, thus inducing a decrease in the proteolytic balance. On the other hand, Ehrlich cells growing in vivo did not produce any detectable TIMP activity, but accumulated MMP-9 and MMP-2 during tumor growth. Both compounds induced significant decrease of MMPs activity when tumor cells were actively proliferating. It was concluded that both genistein and 2-methoxyestradiol could shift the proteolytic balance MMP/TIMP towards antiproteolysis in media or ascitic fluid conditioned by actively growing Ehrlich cells.