Down-regulation of the global regulator SATB1 by statins in COLO205 colon cancer cells

Special AT-rich sequence binding protein 1 (SATB1) regulates the expression of more than 1,000 genes in tumor cells. SATB1 expression has been implicated in metastasis, and its silencing results in reduced cancer progression and the reversion of metastatic cells to normal appearance. Therefore, any compound causing down-regulation of SATB1 expression or activity may be exploited for its therapeutic potential in terms of cancer regression. Earlier studies showed that the 3-hydroxy-3-methylglutaryl coenzymeA (HMG-CoA) reductase inhibitors (statin drugs), which are widely used to treat hypercholesterolemia, possess other pleotropic activities. These are now increasingly gaining attention for their cancer prevention abilities. However, the downstream interplay of the molecular mechanisms of such anti-cancer activities is unclear. Here, we show that SATB1 is down-regulated by statins in a time- and dose-dependent manner in COLO205 cells. This effect was statin-specific as the down-regulation of SATB1 was brought about by hydrophobic statins, such as simvastatin and fluvastatin, but not by hydrophilic pravastatin. Notably, treatment with mevalonate, an intermediate in the cholesterol and isoprenoid biosynthetic pathways, led to the inhibition of SATB1 down-regulation and cytotoxicity mediated by statins. Treatment with the proteasome inhibitors lactacystine and MG-132 inhibited the statin-mediated down-regulation of SATB1, suggesting that regulation occurs at the post-translational level. Thus, our results demonstrate a novel molecular mechanism for the anti-cancer activity of statin drugs in colon cancer cells, without invoking significant cytotoxicity

Fluvastatin mediated breast cancer cell death: a proteomic approach to identify differentially regulated proteins in MDA-MB-231 cells.

Statins are increasingly being recognized as anti-cancer agents against various cancers including breast cancer. To understand the molecular pathways targeted by fluvastatin and its differential sensitivity against metastatic breast cancer cells, we analyzed protein alterations in MDA-MB-231 cells treated with fluvastatin using 2-DE in combination with LC-MS/MS. Results revealed dys-regulation of 39 protein spots corresponding to 35 different proteins. To determine the relevance of altered protein profiles with breast cancer cell death, we mapped these proteins to major pathways involved in the regulation of cell-to-cell signaling and interaction, cell cycle, Rho GDI and proteasomal pathways using IPA analysis. Highly interconnected sub networks showed that vimentin and ERK1/2 proteins play a central role in controlling the expression of altered proteins. Fluvastatin treatment caused proteolysis of vimentin, a marker of epithelial to mesenchymal transition. This effect of fluvastatin was reversed in the presence of mevalonate, a downstream product of HMG-CoA and caspase-3 inhibitor. Interestingly, fluvastatin neither caused an appreciable cell death nor did modulate vimentin expression in normal mammary epithelial cells. In conclusion, fluvastatin alters levels of cytoskeletal proteins, primarily targeting vimentin through increased caspase-3- mediated proteolysis, thereby suggesting a role for vimentin in statin-induced breast cancer cell death

Activation of nuclear factor-kappaB during doxorubicin-induced apoptosis in endothelial cells and myocytes is pro-apoptotic: the role of hydrogen peroxide.

Doxorubicin (DOX) is a widely used anti-tumour drug. Cardiotoxicity is a major toxic side effect of DOX therapy. Although recent studies implicated an apoptotic pathway in DOX-induced cardiotoxicity, the mechanism of DOX-induced apoptosis remains unclear. In the present study, we investigated the role of reactive oxygen species and the nuclear transcription factor nuclear factor kappaB (NF-kappaB) during apoptosis induced by DOX in bovine aortic endothelial cells (BAECs) and adult rat cardiomyocytes. DOX-induced NF-kappaB activation is both dose- and time-dependent, as demonstrated using electrophoretic mobility-shift assay and luciferase and p65 (Rel A) nuclear-translocation assays. Addition of a cell-permeant iron metalloporphyrin significantly suppressed NF-kappaB activation and apoptosis induced by DOX. Overexpression of glutathione peroxidase, which detoxifies cellular H(2)O(2), significantly decreased DOX-induced NF-kappaB activation and apoptosis. Inhibition of DOX-induced NF-kappaB activation by a cell-permeant peptide SN50 that blocks translocation of the NF-kappaB complex into the nucleus greatly diminished DOX-induced apoptosis. Apoptosis was inhibited when IkappaB mutant vector, another NF-kappaB inhibitor, was added to DOX-treated BAECs. These results suggest that NF-kappaB activation in DOX-treated endothelial cells and myocytes is pro-apoptotic, in contrast with DOX-treated cancer cells, where NF-kappaB activation is anti-apoptotic. Removal of intracellular H(2)O(2) protects endothelial cells and myocytes from DOX-induced apoptosis, possibly by inhibiting NF-kappaB activation. These findings suggest a novel mechanism for enhancing the therapeutic efficacy of DOX

High Affinity Neutral Bodipy Fluorophores for Mitochondrial Tracking

We report the first high affinity neutral Bodipy fluorophores for selective imaging of mitochondria with notable sensitivity (∼100 nM) and insignificant cytotoxicity even at very high concentration (∼100 μM), when tested against HeLa cells. Further, these fluorophores are chemically robust and require no special conditions for storage

1-Methyl-4-phenylpyridinium (MPP+)-induced apoptosis and mitochondrial oxidant generation: role of transferrin-receptor-dependent iron and hydrogen peroxide.

1-Methyl-4-phenylpyridinium (MPP(+)) is a neurotoxin used in cellular models of Parkinson's Disease. Although intracellular iron plays a crucial role in MPP(+)-induced apoptosis, the molecular signalling mechanisms linking iron, reactive oxygen species (ROS) and apoptosis are still unknown. We investigated these aspects using cerebellar granule neurons (CGNs) and human SH-SY5Y neuroblastoma cells. MPP(+) enhanced caspase 3 activity after 24 h with significant increases as early as 12 h after treatment of cells. Pre-treatment of CGNs and neuroblastoma cells with the metalloporphyrin antioxidant enzyme mimic, Fe(III)tetrakis(4-benzoic acid)porphyrin (FeTBAP), completely prevented the MPP(+)-induced caspase 3 activity as did overexpression of glutathione peroxidase (GPx1) and pre-treatment with a lipophilic, cell-permeable iron chelator [N, N '-bis-(2-hydroxybenzyl)ethylenediamine-N, N '-diacetic acid, HBED]. MPP(+) treatment increased the number of TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick-end-labelling)-positive cells which was completely blocked by pre-treatment with FeTBAP. MPP(+) treatment significantly decreased the aconitase and mitochondrial complex I activities; pre-treatment with FeTBAP, HBED and GPx1 overexpression reversed this effect. MPP(+) treatment increased the intracellular oxidative stress by 2-3-fold, as determined by oxidation of dichlorodihydrofluorescein and dihydroethidium (hydroethidine). These effects were reversed by pre-treatment of cells with FeTBAP and HBED and by GPx1 overexpression. MPP(+)-treatment enhanced the cell-surface transferrin receptor (TfR) expression, suggesting a role for TfR-induced iron uptake in MPP(+) toxicity. Treatment of cells with anti-TfR antibody (IgA class) inhibited MPP(+)-induced caspase activation. Inhibition of nitric oxide synthase activity did not affect caspase 3 activity, apoptotic cell death or ROS generation by MPP(+). Overall, these results suggest that MPP(+)-induced cell death in CGNs and neuroblastoma cells proceeds via apoptosis and involves mitochondrial release of ROS and TfR-dependent iron

Synthesis of novel 5-substituted isoxazole-3-carboxamide derivatives and cytotoxicity studies on lung cancer cell line

1369-1375A series of novel 5-substituted isoxazole-3-carboxamide derivatives 6 have been prepared by coupling of 5-substituted isoxazole-3-carboxylic acids 3 with substituted-3-benzyloxyaniline <b style="mso-bidi-font-weight: normal">5 using DCC/HOBT as coupling agent. The products 6 have been evaluated for cytotoxicity on A549 lung cancer cell line and compounds <b style="mso-bidi-font-weight: normal">6a, 6b, 6e, 6j are<b style="mso-bidi-font-weight: normal"> found to show moderate proliferative activity and low cytotoxicity

Nanomaterial-based electrochemical biosensors for cytochrome c using cytochrome c reductase

Emerging evidences have pointed out that the release of cytochrome c (cyt c) from mitochondria into cytosol is a critical step in the activation of apoptosis. This article presents a novel approach for the detection of mitochondrial cyt c release for the first time using cytochrome c reductase (CcR) immobilized on nanoparticles decorated electrodes. Two kinds of nanomaterial-based biosensor platforms were used: (a) carbon nanotubes (CNT) incorporated polypyrrole (PPy) matrix on Pt electrode and (b) self-assembled monolayer (SAM) functionalized gold nanoparticles (GNP) in PPy-Pt. Scanning electron microscope was used to characterize the surface morphologies of the nanomaterial modified electrodes. Cyclic voltammograms of both the biosensors showed reversible redox peaks at − 0.45 and − 0.34 V vs Ag/AgCl, characteristic of CcR. In comparison, the CcR-CNT biosensor gave a detection limit of 0.5 ± 0.03 μM cyt c, which was 4-fold better than the CcR-GNP biosensor (2 ± 0.03 μM). Moreover, the CcR-CNT biosensor achieved a much larger linear range (1–1000 μM) over the CcR-GNP biosensor (5–600 μM) with 2-fold better sensitivity. The CcR-CNT-PPy-Pt biosensor was further applied to quantify the mitochondrial cyt c released in cytosol of A549 cells upon induction of apoptosis with doxorubicin, the results agreed well with standard western blot analysis