ABSTRACT
We identified pyrvinium pamoate, an old anthelminthic medicine, which preferentially inhibits anchorage-independent growth of cancer cells over anchorage-dependent growth (approximately 10 fold). It was also reported by others to have anti-tumor activity in vivo and selective toxicity against cancer cells under glucose starvation in vitro, but with unknown mechanism. Here, we provide evidence that pyrvinium suppresses the transcriptional activation of GRP78 and GRP94 induced by glucose deprivation or 2-deoxyglucose (2DG, a glycolysis inhibitor), but not by tunicamycin or A23187. Other UPR pathways induced by glucose starvation, e.g. XBP-1, ATF4, were also found suppressed by pyrvinium. Constitutive expression of GRP78 via transgene partially protected cells from pyrvinium induced cell death under glucose starvation, suggesting that suppression of the UPR is involved in pyrvinium mediated cytotoxicity under glucose starvation. Xenograft experiments showed rather marginal overall anti-tumor activity for pyrvinium as a monotherapy. However, the combination of pyrvinium and Doxorubicin demonstrated significantly enhanced efficacy in vivo, supporting a mechanistic treatment concept based on tumor hypoglycemia and UPR.
Subject(s)
Antineoplastic Agents/pharmacology , Antineoplastic Combined Chemotherapy Protocols , Hypoglycemia/metabolism , Protein Folding/drug effects , Pyrvinium Compounds/pharmacology , Activating Transcription Factor 4/metabolism , Animals , Cell Adhesion/drug effects , Cell Death/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Down-Regulation/drug effects , Doxorubicin/pharmacology , Drug Synergism , Endoplasmic Reticulum Chaperone BiP , Female , Gene Expression Regulation, Neoplastic/drug effects , Glucose/deficiency , Glycolysis/drug effects , HSP70 Heat-Shock Proteins/genetics , HSP70 Heat-Shock Proteins/metabolism , Heat-Shock Proteins/genetics , Heat-Shock Proteins/metabolism , Humans , Membrane Proteins/genetics , Membrane Proteins/metabolism , Mice , Mice, Nude , Molecular Chaperones/genetics , Molecular Chaperones/metabolism , Regulatory Factor X Transcription Factors , Transcription Factors/genetics , Transcription Factors/metabolism , Transcriptional Activation/drug effects , Up-Regulation/drug effects , X-Box Binding Protein 1ABSTRACT
Ischemia-reperfusion (I/R) has critical consequences in the heart. Recent studies on the functions of I/R-activated kinases, such as p38 mitogen-activated protein kinase (MAPK), showed that I/R injury is reduced in the hearts of transgenic mice that overexpress the p38 MAPK activator MAPK kinase 6 (MKK6). This protection may be fostered by changes in the levels of many proteins not currently known to be regulated by p38. To examine this possibility, we employed the multidimensional protein identification technology MudPIT to characterize changes in levels of proteins in MKK6 transgenic mouse hearts, focusing on proteins in mitochondria, which play key roles in mediating I/R injury in the heart. Of the 386 mitochondrial proteins identified, the levels of 58 were decreased, while only 2 were increased in the MKK6 transgenic mouse hearts. Among those that were decreased were 21 mitochondrial oxidative phosphorylation complex proteins, which was unexpected because p38 is not known to mediate such decreases. Immunoblotting verified that proteins in each of the five oxidative phosphorylation complexes were reduced in MKK6 mouse hearts. On assessing functional consequences of these reductions, we found that MKK6 mouse heart mitochondria exhibited 50% lower oxidative respiration and I/R-mediated reactive oxygen species (ROS) generation, both of which are predicted consequences of decreased oxidative phosphorylation complex proteins. Thus the cardioprotection observed in MKK6 transgenic mouse hearts may be partly due to decreased electron transport, which is potentially beneficial, because damaging ROS are known to be generated by mitochondrial complexes I and III during reoxygenation.
