Regulation of anoikis resistance by NADPH oxidase 4 and epidermal growth factor receptor.

BACKGROUND: Normal cells are sensitive to anoikis, which is a cell detachment-induced apoptosis. However, cancer cells acquire anoikis resistance that is essential for successful metastasis. This study aimed to demonstrate the function and potential mechanism of NADPH oxidase 4 (NOX4) and EGFR activation in regulating anoikis resistance in lung cancer. METHODS: Cells were cultured either in the attached or suspended condition. Cell viability was measured by cell counting and live and dead cell staining. Expression levels of NOX4 and EGFR were measured by PCR and immunoblotting. Reactive oxygen species (ROS) levels were measured by flow cytometry. Effects of NOX4 overexpression or NOX4 knockdown by si-NOX4 on anoikis sensitivity were explored. Levels of NOX4 and EGFR in lung cancer tissues were evaluated by IHC staining. RESULTS: NOX4 was upregulated but EGFR decreased in suspended cells compared with attached cells. Accordingly, ROS levels were increased in suspended cells, resulting in the activation of Src and EGFR. NOX4 knockdown decreased activation of Src and EGFR, and thus sensitised cells to anoikis. NOX4 overexpression increased EGFR levels and attenuated anoikis. NOX4 expression is upregulated and is positively correlated with EGFR levels in the lung cancer patient tissues. CONCLUSIONS: NOX4 upregulation confers anoikis resistance by ROS-mediated activation of EGFR and Src, and by maintaining EGFR levels, which is critical for cell survival.

Effects of low-dose irradiation on mice with Escherichia coli-induced sepsis.

Although favorable immune responses to low-dose irradiation (LDI) have been observed in normal mice, i.e., a hormesis effect, little is known about the effects of LDI in infectious diseases. In this study, we examined the effects of LDI on mice with sepsis, a severe and often lethal hyperinflammatory response to bacteria. Female C57BL/6 mice were whole-body irradiated with 10cGy 48h before Escherichia coli infection, and survival, bacterial clearance, cytokines, and antioxidants were quantified. LDI pretreatment significantly increased survival from 46.7% in control mice to 75% in mice with sepsis. The bacterial burden was significantly lower in the blood, spleen, and kidney of LDI-treated mice than in those of control septic mice. The levels of pro-inflammatory cytokines, e.g., IL-1ß and IL-6, as well as anti-inflammatory IL-10 were markedly reduced in pre-LDI septic mice. Nitric oxide production by peritoneal macrophages was also reduced in pre-LDI septic mice. Immune cells in the spleen increased and Nrf2 and HO-1 were induced in pre-LDI septic mice. LDI stimulates the immune response and minimizes lethality in septic mice via enhanced bacterial clearance and reduced initial proinflammatory responses.

Inhibition of cancer cell invasion by new ((3,4-dihydroxy benzylidene)hydrazinyl)pyridine-3-sulfonamide analogs.

Rab GTPases regulate various types of intracellular membrane trafficking in all eukaryotes. Since Rab27a and its multiple effectors are involved in exocytosis of lysosome-related organelles and play a major role in malignancy, compounds targeting Rab27a could be likely used to inhibit invasive growth and tumor metastasis. Thus, we designed and synthesized several compounds based on the previously reported Rab27a-targeting synthetic compounds identified by virtual screening, and investigated their anti-metastatic effects in MDA-MB231 and A375 cells. Among the synthesized compounds, (E)-N-(3-chlorophenyl)-6-(2-(3,4-dihydroxy benzylidene)hydrazinyl)pyridine-3-sulfonamide (3d) and (E)-N-benzyl-6-(2-(3,4-dihydroxy benzylidene)hydrazinyl)-N-methylpyridine-3-sulfonamide (3f) significantly inhibited the invasiveness of both tumor cell lines. Compounds 3d and 3f also decreased the levels of signature extracellular matrix marker proteins (fibronectin, collagen, and α-smooth muscle actin) and representative mesenchymal cell markers (N-cadherin and vimentin). Taken together, our results suggest that novel sulfonamide analogs have anti-metastatic activity in breast and melanoma cancer cell lines and may be used as therapeutic agents to treat malignant cancer.

Novel focal adhesion kinase 1 inhibitor sensitizes lung cancer cells to radiation in a p53-independent manner.

Focal adhesion kinase 1 (FAK1) is known to promote tumor progression and metastasis by controlling cell movement, invasion, survival and the epithelial-to-mesenchymal transition in the tumor microenvironment. As recent reports imply that FAK1 is highly associated with tumor cell development and malignancy, the inhibition of FAK1 activity could be an effective therapeutic approach for inhibiting the growth and metastasis of tumor cells. In this study, we aimed to determine the effect of a novel synthetic FAK1 inhibitor 2-[2-(2-methoxy-4-morpholin-4-yl-phenylamino)-5-trifluoromethyl-pyrimidin-4-ylamino]-N-methyl-benzamide, (MPAP) on lung cancer cells. MPAP suppressed cancer cell proliferation and the phosphorylation of FAK1. Combined treatment with MPAP and irradiation (IR) showed enhanced suppression of cancer cell proliferation in wild-type p53 cells and more intense suppression in p53-null cells. In addition, the combination treatment effectively induced G1 cell cycle arrest in a p53-independent manner. In an in vivo tumor xenograft mouse model, treatment with both MPAP and IR reduced tumor growth more than the treatment with IR or MPAP alone. Overall, these data demonstrate that the radiosensitizing effect of MPAP is mediated by the regulation of retinoblastoma protein (RB) phosphorylation in a p53-independent manner.

Crucial role of TSC-22 in preventing the proteasomal degradation of p53 in cervical cancer.

The p53 tumor suppressor function can be compromised in many tumors by the cellular antagonist HDM2 and human papillomavirus oncogene E6 that induce p53 degradation. Restoration of p53 activity has strong therapeutic potential. Here, we identified TSC-22 as a novel p53-interacting protein and show its novel function as a positive regulator of p53. We found that TSC-22 level was significantly down-regulated in cervical cancer tissues. Moreover, over-expression of TSC-22 was sufficient to inhibit cell proliferation, promote cellular apoptosis in cervical cancer cells and suppress growth of xenograft tumors in mice. Expression of also TSC-22 enhanced the protein level of p53 by protecting it from poly-ubiquitination. When bound to the motif between amino acids 100 and 200 of p53, TSC-22 inhibited the HDM2- and E6-mediated p53 poly-ubiquitination and degradation. Consequently, ectopic over-expression of TSC-22 activated the function of p53, followed by increased expression of p21(Waf1/Cip1) and PUMA in human cervical cancer cell lines. Interestingly, TSC-22 did not affect the interaction between p53 and HDM2. Knock-down of TSC-22 by small interfering RNA clearly enhanced the poly-ubiquitination of p53, leading to the degradation of p53. These results suggest that TSC-22 acts as a tumor suppressor by safeguarding p53 from poly-ubiquitination mediated-degradation.