Hubert Chassaigne

Mechanism of cisplatin proximal tubule toxicity revealed by integrating transcriptomics, proteomics, metabolomics and biokinetics.

Cisplatin is one of the most widely used chemotherapeutic agents for the treatment of solid tumours. The major dose-limiting factor is nephrotoxicity, in particular in the proximal tubule. Here, we use an integrated omics approach, including transcriptomics, proteomics and metabolomics coupled to biokinetics to identify cell stress response pathways induced by cisplatin. The human renal proximal tubular cell line RPTEC/TERT1 was treated with sub-cytotoxic concentrations of cisplatin (0.5 and 2 μM) in a daily repeat dose treating regime for up to 14 days. Biokinetic analysis showed that cisplatin was taken up from the basolateral compartment, transported to the apical compartment, and accumulated in cells over time. This is in line with basolateral uptake of cisplatin via organic cation transporter 2 and bioactivation via gamma-glutamyl transpeptidase located on the apical side of proximal tubular cells. Cisplatin affected several pathways including, p53 signalling, Nrf2 mediated oxidative stress response, mitochondrial processes, mTOR and AMPK signalling. In addition, we identified novel pathways changed by cisplatin, including eIF2 signalling, actin nucleation via the ARP/WASP complex and regulation of cell polarization. In conclusion, using an integrated omic approach together with biokinetics we have identified both novel and established mechanisms of cisplatin toxicity.

A combined proteomics and metabolomics approach to assess the effects of gold nanoparticles in vitro

Abstract Omics technologies, such as proteomics or metabolomics, have to date been applied in the field of nanomaterial safety assessment to a limited extent. To address this dearth, we developed an integrated approach combining the two techniques to study the effects of two sizes, 5 and 30 nm, of gold nanoparticles (AuNPs) in Caco-2 cells. We observed differences in cells exposed for 72 h to each size of AuNPs: 61 responsive (up/down-regulated) proteins were identified and 35 metabolites in the cell extract were tentatively annotated. Several altered biological pathways were highlighted by integrating the obtained multi-omics data with bioinformatic tools. This provided a unique set of molecular information on the effects of nanomaterials at cellular level. This information was supported by complementary data obtained by immunochemistry, microscopic analysis, and multiplexed assays. A part from increasing our knowledge on how the cellular processes and pathways are affected by nanomaterials (NMs), these findings could be used to identify specific biomarkers of toxicity or to support the safe-by-design concept in the development of new nanomedicines.

Proteomics study of silver nanoparticles on Caco-2 cells

Silver nanoparticles (AgNPs) have been incorporated into several consumer products. While these advances in technology are promising and exciting, the effects of these nanoparticles have not equally been studied. Due to the size, AgNPs can penetrate the body through oral exposure and reach the gastrointestinal tract. The present study was designed as a comparative proteomic analysis of Caco-2 cells, used as an in vitro model of the small intestine, exposed to 30 nm citrate stabilized-silver nanoparticles (AgNPs) for 24 or 72 h. Using two complementary proteomic approaches, 2D gel-based and label-free mass spectrometry, we present insight into the effects of AgNPs at proteins level. Exposure of 1 or 10 μg/mL AgNPs to Caco-2 cells resulted in 56 and 88 altered proteins at 24 h and 72 h respectively, by 2D gel-based technique. Ten of these proteins were found to be common between the two time-points. Using label-free mass spectrometry technique, 291 and 179 altered proteins were found at 24 h and 72 h, of which 24 were in common. Analysis of the proteomes showed several major biological processes altered, from which, cell cycle, cell morphology, cellular function and maintenance were the most affected.

Collaboration of the Joint Research Centre and European Customs Laboratories for the Identification of New Psychoactive Substances

Background: The emergence of psychoactive designer drugs has significantly increased over the last few years. Customs officials are responsible for the control of products entering the European Union (EU) market. This control applies to chemicals in general, pharmaceutical products and medicines. Numerous products imported from non-EU countries, often declared as ‘bath salts’ or ‘fertilizers’, contain new psychoactive substance (NPS). Review: These are not necessarily controlled under international law, but may be subject to monitoring in agreement with EU legislation. This situation imposes substantial challenges, for example, for the maintenance of spectral libraries used for their detection by designated laboratories. The chemical identification of new substances, with the use of powerful instrumentation, and the time needed for detailed analysis and interpretation of the results, demands considerable commitment. The EU Joint Research Centre endeavors to provide scientific support to EU Customs laboratories to facilitate rapid identification and characterisation of seized samples. In addition to analysing known NPS, several new chemical entities have also been identified. Frequently, these belong to NPS classes already notified to the European Monitoring Centre for Drugs and Drug Addiction (EMCDDA) by the European Early-Warning System (EWS). Conclusion: The aim of this paper is to discuss the implementation of workflow mechanisms that are in place in order to facilitate the monitoring, communication and management of analytical data. The rapid dissemination of this information between control authorities strives to help protect EU citizens against the health risks posed by harmful substances.