Targeting homeostatic mechanisms of endoplasmic reticulum stress to increase susceptibility of cancer cells to fenretinide-induced apoptosis: the role of stress proteins ERdj5 and ERp57

Endoplasmic reticulum (ER) malfunction, leading to ER stress, can be a consequence of genome instability and hypoxic tissue environments. Cancer cells survive by acquiring or enhancing survival mechanisms to counter the effects of ER stress and these homeostatic responses may be new therapeutic targets. Understanding the links between ER stress and apoptosis may be approached using drugs specifically to target ER stress responses in cancer cells. The retinoid analogue fenretinide [N-(4-hydroxyphenyl) retinamide] is a new cancer preventive and chemotherapeutic drug, that induces apoptosis of some cancer cell types via oxidative stress, accompanied by induction of an ER stress-related transcription factor, GADD153. The aim of this study was to test the hypothesis that fenretinide induces ER stress in neuroectodermal tumour cells, and to elucidate the role of ER stress responses in fenretinide-induced apoptosis. The ER stress genes ERdj5, ERp57, GRP78, calreticulin and calnexin were induced in neuroectodermal tumour cells by fenretinide. In contrast to the apoptosis-inducing chemotherapeutic drugs vincristine and temozolomide, fenretinide induced the phosphorylation of eIF2α, expression of ATF4 and splicing of XBP-1 mRNA, events that define ER stress. In these respects, fenretinide displayed properties similar to the ER stress inducer thapsigargin. ER stress responses were inhibited by antioxidant treatment. Knockdown of ERp57 or ERdj5 by RNA interference in these cells increased the apoptotic response to fenretinide. These data suggest that downregulating homeostatic ER stress responses may enhance apoptosis induced by oxidative stress-inducing drugs acting through the ER stress pathway. Therefore, ER-resident proteins such as ERdj5 and ERp57 may represent novel chemotherapeutic targets

An Intronic Variant in the GRP78, a Stress-Associated Gene, Improves Prediction for Liver Cirrhosis in Persistent HBV Carriers

Background: Our previous study indicated that a common variant (rs430397 G>A) in the intron 5 of glucose-regulated protein 78 (GRP78) gene was associated with risk and prognosis of primary hepatocellular carcinoma (HCC), including HBV- and cirrhosis-related HCC. rs430397 polymorphism may be a contributing factor or biomarker of HBV infection or HBV-related cirrhosis. Methodology/Principal Findings: 539 non-HBV-infected individuals, 205 self-limited infection and 496 persistent HBV infection were recruited between January 2001 and April 2005 from the hospitals in Southern China. Genomic DNA was genotyped for rs430397. The associations between the variation and susceptibility to liver cirrhosis (LC) in persistent HBV infection were examined. We observed that individuals carrying allele rs430397A were more likely to become HBV-related LC. When persistently infected patients were divided into four subgroups, patients with phase IV had an increased allele A and genotype AG compared with phase I and/or phase III. Decreased serum albumin and prolonged plasma prothrombin time (PT) were showed in LC patients carrying genotype AA. Furthermore, rs430397 genotype had an increased susceptibility to LC with dose-dependent manners (P-trend = 0.005), and the genotype did constitute a risk factor for the development of advanced LC (Child-Pugh classification C and B, P-trend = 0.021). Conclusions/Significance: rs430397 polymorphism may be a contributing factor to LC in persistent HBV carriers. © 2011 Zhu et al.published_or_final_versio

Toward Achieving Harmonization in a Nanocytotoxicity Assay Measurement Through an Interlaboratory Comparison Study

Development of reliable cell-based nanotoxicology assays is important for evaluation of potentially hazardous engineered nanomaterials. Challenges to producing a reliable assay protocol include working with nanoparticle dispersions and living cell lines, and the potential for nano-related interference effects. Here we demonstrate the use of a 96-well plate design with several measurement controls and an interlaboratory comparison study involving five laboratories to characterize the robustness of a nanocytotoxicity MTS cell viability assay based on the A549 cell line. The consensus EC50 values were 22.1 mg/L (95% confidence intervals 16.9 mg/L to 27.2 mg/L) and 52.6 mg/L (44.1 mg/L to 62.6 mg/L) for positively charged polystyrene nanoparticles for the serum-free and serum conditions, respectively, and 49.7 mu mol/L (47.5 mu mol/L to 51.5 mu mol/L) and 77.0 mu mol/L (54.3 mu mol/L to 99.4 mu mol/L) for positive chemical control cadmium sulfate for the serum-free and serum conditions, respectively. Results from the measurement controls can be used to evaluate the sources of variability and their relative magnitudes within and between laboratories. This information revealed steps of the protocol that may need to be modified to improve the overall robustness and precision. The results suggest that protocol details such as cell line ID, media exchange, cell handling, and nanoparticle dispersion are critical to ensure protocol robustness and comparability of nanocytotoxicity assay results. The combination of system control measurements and interlaboratory comparison data yielded insights that would not have been available by either approach by itself