Identification of metabolites, clinical chemistry markers and transcripts associated with hepatotoxicity.

Early and accurate pre-clinical and clinical biomarkers of hepatotoxicity facilitate the drug development process and the safety monitoring in clinical studies. We selected eight known model compounds to be administered to male Wistar rats to identify biomarkers of drug induced liver injury (DILI) using transcriptomics, metabolite profiling (metabolomics) and conventional endpoints. We specifically explored early biomarkers in serum and liver tissue associated with histopathologically evident acute hepatotoxicity.

A real-time impedance-based screening assay for drug-induced vascular leakage.

Vascular leakage is a serious side effect of therapies based on monoclonal antibodies or cytokines which may lead to life-threatening situations. With the steady increase of new drug development programs for large molecules, there is an urgent need for reliable tools to assess this potential liability of new medicines in a rapid and cost-effective manner. Using human umbilical vein endothelial cells (HUVECs) as a model for endothelium, we established an impedance-based assay measuring the integrity of the endothelial cell monolayer in real time.

Increased levels of urinary phenylacetylglycine associated with mitochondrial toxicity in a model of drug-induced phospholipidosis.

Background: Phospholipidosis (PLD) is a lysosomal storage disorder induced by a class of cationic amphiphilic drugs. However, drug-induced PLD is reversible. Evidence of PLD from animal studies with some compounds has led to discontinuation of development.

Decreased translation of Dio3 mRNA is associated with drug-induced hepatotoxicity.

Recent work has demonstrated the importance of post-transcriptional gene regulation in toxic responses. In the present study, we used two rat models to investigate mRNA translation in the liver following xenobiotic-induced toxicity. By combining polysome profiling with genomic methodologies, we were able to assess global changes in hepatic mRNA translation.

Bioengineered 3D human kidney tissue, a platform for the determination of nephrotoxicity.

The staggering cost of bringing a drug to market coupled with the extremely high failure rate of prospective compounds in early phase clinical trials due to unexpected human toxicity makes it imperative that more relevant human models be developed to better predict drug toxicity. Drug-induced nephrotoxicity remains especially difficult to predict in both pre-clinical and clinical settings and is often undetected until patient hospitalization. Current pre-clinical methods of determining renal toxicity include 2D cell cultures and animal models, both of which are incapable of fully recapitulating the in vivo human response to drugs, contributing to the high failure rate upon clinical trials.

Effect of GLP1R agonists taspoglutide and liraglutide on primary thyroid C-cells from rodent and man.

Glucagon-like peptide 1 (GLP1) analogs have been associated with an increased incidence of thyroid C-cell hyperplasia and tumors in rodents. This effect may be due to a GLP1 receptor (GLP1R)-dependent mechanism. As the expression of GLP1R is much lower in primates than in rodents, the described C-cell proliferative lesions may not be relevant to man.

A long-term three dimensional liver co-culture system for improved prediction of clinically relevant drug-induced hepatotoxicity.

Drug-induced liver injury (DILI) is the major cause for liver failure and post-marketing drug withdrawals. Due to species-specific differences in hepatocellular function, animal experiments to assess potential liabilities of drug candidates can predict hepatotoxicity in humans only to a certain extent. In addition to animal experimentation, primary hepatocytes from rat or human are widely used for pre-clinical safety assessment.

A simple whole blood bioassay detects cytokine responses to anti-CD28SA and anti-CD52 antibodies.

Introduction: In 2006 the anti-CD28 superagonistic IgG4 TGN1412, having passed pre-clinical safety screens, caused a severe 'cytokine storm' in 6 healthy volunteers. Others have shown that for TGN1412 to induce an inflammatory signal in human peripheral blood mononuclear cells (PBMCs) or in human diluted blood, endothelial cells or bound monoclonal antibody (mAb) is required as part of a bioassay complex. These types of protocols rely on different donor cells and therefore have limitations as bioassays for pre-clinical testing.

Current challenges and controversies in drug-induced liver injury.

Current key challenges and controversies encountered in the identification of potentially hepatotoxic drugs and the assessment of drug-induced liver injury (DILI) are covered in this article. There is substantial debate over the classification of DILI itself, including the definition and validity of terms such as 'intrinsic' and 'idiosyncratic'. So-called idiosyncratic DILI is typically rare and requires one or more susceptibility factors in individuals.

Gene expression-based in vivo and in vitro prediction of liver toxicity allows compound selection at an early stage of drug development.

We have analyzed gene expression and histopathology of rat liver treated with a histamine-3 receptor inverse agonist under development for the treatment of obesity 24 h after a single acute administration. While histopathology did not identify a clear liver toxicity, analysis of gene changes strongly suggested the development of toxicity. This prediction was confirmed in a 2-week repeat-dose rat study where prominent liver pathology occurred, while gene changes that lead to the prediction persisted.

A comparative integrated transcript analysis and functional characterization of differential mechanisms for induction of liver hypertrophy in the rat.

The main goal of the present work was to better understand the molecular mechanisms underlying liver hypertrophy (LH), a recurrent finding observed following acute or repeated drug administration to animals, using transcriptomic technologies together with the results from conventional toxicology methods. Administration of 5 terminated proprietary drug candidates from participating companies involved in the EU Innomed PredTox Project or the reference hepatotoxicant troglitazone to rats for up to a 14-day duration induced LH as the main liver phenotypic toxicity outcome. The integrated analysis of transcriptomic liver expression data across studies turned out to be the most informative approach for the generation of mechanistic models of LH.

Cross-study and cross-omics comparisons of three nephrotoxic compounds reveal mechanistic insights and new candidate biomarkers.

The European InnoMed-PredTox project was a collaborative effort between 15 pharmaceutical companies, 2 small and mid-sized enterprises, and 3 universities with the goal of delivering deeper insights into the molecular mechanisms of kidney and liver toxicity and to identify mechanism-linked diagnostic or prognostic safety biomarker candidates by combining conventional toxicological parameters with "omics" data. Mechanistic toxicity studies with 16 different compounds, 2 dose levels, and 3 time points were performed in male Crl: WI(Han) rats. Three of the 16 investigated compounds, BI-3 (FP007SE), Gentamicin (FP009SF), and IMM125 (FP013NO), induced kidney proximal tubule damage (PTD).

EU framework 6 project: predictive toxicology (PredTox)--overview and outcome.

In this publication, we report the outcome of the integrated EU Framework 6 PROJECT: Predictive Toxicology (PredTox), including methodological aspects and overall conclusions. Specific details including data analysis and interpretation are reported in separate articles in this issue. The project, partly funded by the EU, was carried out by a consortium of 15 pharmaceutical companies, 2 SMEs, and 3 universities.

The enhanced value of combining conventional and "omics" analyses in early assessment of drug-induced hepatobiliary injury.

The InnoMed PredTox consortium was formed to evaluate whether conventional preclinical safety assessment can be significantly enhanced by incorporation of molecular profiling ("omics") technologies. In short-term toxicological studies in rats, transcriptomics, proteomics and metabolomics data were collected and analyzed in relation to routine clinical chemistry and histopathology. Four of the sixteen hepato- and/or nephrotoxicants given to rats for 1, 3, or 14days at two dose levels induced similar histopathological effects.

Renal biomarker qualification submission: a dialog between the FDA-EMEA and Predictive Safety Testing Consortium.

The first formal qualification of safety biomarkers for regulatory decision making marks a milestone in the application of biomarkers to drug development. Following submission of drug toxicity studies and analyses of biomarker performance to the Food and Drug Administration (FDA) and European Medicines Agency (EMEA) by the Predictive Safety Testing Consortium's (PSTC) Nephrotoxicity Working Group, seven renal safety biomarkers have been qualified for limited use in nonclinical and clinical drug development to help guide safety assessments. This was a pilot process, and the experience gained will both facilitate better understanding of how the qualification process will probably evolve and clarify the minimal requirements necessary to evaluate the performance of biomarkers of organ injury within specific contexts.

Preclinical evidence for nitrogen-containing bisphosphonate inhibition of farnesyl diphosphate (FPP) synthase in the kidney: implications for renal safety.

Bisphosphonates are potent inhibitors of osteoclast-mediated bone resorption and play an important role in the treatment of osteoporosis, metastatic bone disease, and Paget disease. However, nephrotoxicity has been reported with some bisphosphonates. Nitrogen-containing bisphosphonates directly inhibit farnesyl diphosphate (FPP) synthase activity (mevalonate pathway) and reduce protein prenylation leading to osteoclast cell death.

Application of metabonomics in a comparative profiling study reveals N-acetylfelinine excretion as a biomarker for inhibition of the farnesyl pathway by bisphosphonates.

In this work, the results of metabolic profiling of urine from a preclinical comparative profiling study with the two biphosphonates ibandronate and zoledronate are reported. Toxicological assessment showed very different effects for the two compounds. Ibandronate did not cause major signs of toxicity, whereas zoledronate elicited hepatotoxicity and nephrotoxicity.

Biomarker discovery by imaging mass spectrometry: transthyretin is a biomarker for gentamicin-induced nephrotoxicity in rat.

Adverse drug effects are often associated with pathological changes in tissue. An accurate depiction of the undesired affected area, possibly supported by mechanistic data, is important to classify the effects with regard to relevance for human patients. MALDI imaging MS represents a new analytical tool to directly provide the spatial distribution and the relative abundance of proteins in tissue.

Assessment of hepatotoxic liabilities by transcript profiling.

Male Wistar rats were treated with various model compounds or the appropriate vehicle controls in order to create a reference database for toxicogenomics assessment of novel compounds. Hepatotoxic compounds in the database were either known hepatotoxicants or showed hepatotoxicity during preclinical testing. Histopathology and clinical chemistry data were used to anchor the transcript profiles to an established endpoint (steatosis, cholestasis, direct acting, peroxisomal proliferation or nontoxic/control).

Toxicogenomics in the pharmaceutical industry: hollow promises or real benefit?

Almost 10 years ago, microarray technology was established as a new powerful tool for large-scale analysis of gene expression. Soon thereafter the new technology was discovered by toxicologists for the purpose of deciphering the molecular events underlying toxicity, and the term "Toxicogenomics" appeared in scientific literature. Ever since, the toxicology community was fascinated by the multiplicity of sophisticated possibilities toxicogenomics seems to offer: genome-wide analysis of toxicant-induced expression profiles may provide a means for prediction of toxicity prior to classical toxicological endpoints such as histopathology or clinical chemistry.

Discriminating different classes of toxicants by transcript profiling.

Male rats were treated with various model compounds or the appropriate vehicle controls. Most substances were either well-known hepatotoxicants or showed hepatotoxicity during preclinical testing. The aim of the present study was to determine if biological samples from rats treated with various compounds can be classified based on gene expression profiles.

Toxicogenomics in predictive toxicology in drug development.

The goal of toxicology is the assessment of possible risk to man. An emerging technology with the potential to have a major impact on risk assessment is toxicogenomics. In this review, we provide an overview of the many possibilities for toxicogenomics including technology platforms, data interpretation, and regulatory perspective and we give examples of toxicogenomics investigations.

Interlaboratory evaluation of rat hepatic gene expression changes induced by methapyrilene.

Several studies using microarrays have shown that changes in gene expression provide information about the mechanism of toxicity induced by xenobiotic agents. Nevertheless, the issue of whether gene expression profiles are reproducible across different laboratories remains to be determined. To address this question, several members of the Hepatotoxicity Working Group of the International Life Sciences Institute Health and Environmental Sciences Institute evaluated the liver gene expression profiles of rats treated with methapyrilene (MP).

Gene expression in two hepatic cell lines, cultured primary hepatocytes, and liver slices compared to the in vivo liver gene expression in rats: possible implications for toxicogenomics use of in vitro systems.

Microarray technology allows the simultaneous analysis of mRNA expression levels of thousands of genes. In the field of toxicogenomics, this technology could help to identify potentially unsafe compounds based on the changes in mRNA expression patterns they induce. Rodent in vivo and in vitro systems are currently the experimental models of choice for predictive toxicology, especially in early phases of development.