New approach methodologies in human regulatory toxicology - Not if, but how and when!

The predominantly animal-centric approach of chemical safety assessment has increasingly come under pressure. Society is questioning overall performance, sustainability, continued relevance for human health risk assessment and ethics of this system, demanding a change of paradigm. At the same time, the scientific toolbox used for risk assessment is continuously enriched by the development of "New Approach Methodologies" (NAMs).

Gauging innovation and health impact from biomedical research: survey results and interviews with recipients of EU-funding in the fields of Alzheimer's disease, breast cancer and prostate cancer.

Biomedical research on Alzheimer's disease (AD), breast cancer (BC) and prostate cancer (PC) has globally improved our understanding of the etiopathological mechanisms underlying the onset of these diseases, often with the goal to identify associated genetic and environmental risk factors and develop new medicines. However, the prevalence of these diseases and failure rate in drug development remain high. Being able to retrospectively monitor the major scientific breakthroughs and impact of such investment endeavors is important to re-address funding strategies if and when needed.

Alzheimer's Disease, and Breast and Prostate Cancer Research: Translational Failures and the Importance to Monitor Outputs and Impact of Funded Research.

Dementia and cancer are becoming increasingly prevalent in Western countries. In the last two decades, research focused on Alzheimer's disease (AD) and cancer, in particular, breast cancer (BC) and prostate cancer (PC), has been substantially funded both in Europe and worldwide. While scientific research outcomes have contributed to increase our understanding of the disease etiopathology, still the prevalence of these chronic degenerative conditions remains very high across the globe.

Finding synergies for the 3Rs - Repeated Dose Toxicity testing: Report from an EPAA Partners' Forum.

The European Partnership for Alternative Approaches to Animal Testing (EPAA) convened a Partners' Forum on repeated dose toxicity (RDT) testing to identify synergies between industrial sectors and stakeholders along with opportunities to progress these in existing research frameworks. Although RTD testing is not performed across all industrial sectors, the OECD accepted tests can provide a rich source of information and play a pivotal role for safety decisions relating to the use of chemicals. Currently there are no validated alternatives to repeated dose testing and a direct one-to-one replacement is not appropriate.

Finding synergies for 3Rs - Toxicokinetics and read-across: Report from an EPAA partners' Forum.

The European Partnership for Alternative Approaches to Animal Testing (EPAA) convened a Partners' Forum Toxicokinetics and Read-Across to provide an overview on research activities to develop in vitro toxicokinetics methods and physiologically-based kinetic (PBK) models and to find synergies to enhance use of toxicokinetic data to strengthen read-across. Currently, lacking toxicokinetic data often prevent the application of read-across. Preferably, toxicokinetic data should be generated using in vitro and in silico tools and anchored towards human relevance.

The mammalian gene function resource: the International Knockout Mouse Consortium.

In 2007, the International Knockout Mouse Consortium (IKMC) made the ambitious promise to generate mutations in virtually every protein-coding gene of the mouse genome in a concerted worldwide action. Now, 5 years later, the IKMC members have developed high-throughput gene trapping and, in particular, gene-targeting pipelines and generated more than 17,400 mutant murine embryonic stem (ES) cell clones and more than 1,700 mutant mouse strains, most of them conditional. A common IKMC web portal (www.

Animal models for arthritis: innovative tools for prevention and treatment.

The development of novel treatments for rheumatoid arthritis (RA) requires the interplay between clinical observations and studies in animal models. Given the complex molecular pathogenesis and highly heterogeneous clinical picture of RA, there is an urgent need to dissect its multifactorial nature and to propose new strategies for preventive, early and curative treatments. Research on animal models has generated new knowledge on RA pathophysiology and aetiology and has provided highly successful paradigms for innovative drug development.

Murine Prkdc polymorphisms impact DNA-PKcs function.

Polymorphic variants of DNA repair genes can increase the carcinogenic potential of exposure to ionizing radiation. Two single nucleotide polymorphisms (SNPs) in Prkdc, the gene encoding the DNA-dependent protein kinase catalytic subunit (DNA-PKcs), have been identified in BALB/c mice and linked to reduced DNA-PKcs activity and mammary cancer susceptibility. We examined three additional mouse strains to better define the roles of the BALB/c Prkdc SNPs (R2140C and M3844V).

3rd US-EU workshop: systems level understanding of DNA damage responses.

The 3rd US-EU Workshop on systems level understanding of DNA damage responses was held from March 30 to April 1, 2009 in Egmond aan Zee, The Netherlands. Objectives of the workshop were (1) to assess the current science of the DDR, in particular network level responses to chemotherapeutic and environmentally induced DNA damage; and (2) to establish the basis for a reciprocal scientific exchange program between the EU and US in the relevant areas of DDR research. Here, we report the highlights of the meeting program and conclude that this third meeting in 2009 refined the role of DDR networks in human disease.

Research on animal model organisms funded by the European Commission's framework programmes.

Recognising the crucial role of model organisms in exploring the causes of human disease and in developing safe treatments, the European Commission has invested euro180 million in collaborative research projects on model organisms since 2002. Further financial support is planned for the future. Projects supported by the European Commission are playing an important role in structuring the research landscape in Europe and creating the knowledge base to understand health and disease.

DNA-PKcs and ATM influence generation of ionizing radiation-induced bystander signals.

The phenomenon by which irradiated cells influence non-irradiated neighboring cells, referred to as the bystander effect (BSE), is not well understood in terms of the underlying pathways involved. We sought to enlighten connections between DNA damage repair and the BSE. Utilizing sister chromatid exchange (SCE) frequencies as a marker of the BSE, we performed cell transfer strategies that enabled us to distinguish between generation versus reception of a bystander signal.

A new E6/P63 pathway, together with a strong E7/E2F mitotic pathway, modulates the transcriptome in cervical cancer cells.

Cervical carcinoma is associated with certain types of human papillomaviruses expressing the E6 and E7 oncogenes, which are involved in carcinogenesis through their interactions with the p53 and pRB pathways, respectively. A critical event on the path to malignant transformation is often manifested by the loss of expression of the viral E2 transcription factor due to the integration into the host genome of the viral DNA. Using microarrays, we have previously shown that reintroduction of a functional E2 in the HeLa cervical carcinoma cell line activates a cluster of p53 target genes while at the same time severely repressing a group of E2F target genes.

Genital warts in Burmeister's porpoises: characterization of Phocoena spinipinnis papillomavirus type 1 (PsPV-1) and evidence for a second, distantly related PsPV.

We identified sequences from two distantly related papillomaviruses in genital warts from two Burmeister's porpoises, including a PV antigen-positive specimen, and characterized Phocoena spinipinnis papillomavirus type 1 (PsPV-1). The PsPV-1 genome comprises 7879 nt and presents unusual features. It lacks an E7, an E8 and a bona fide E5 open reading frame (ORF) and has a large E6 ORF.

Transcriptional response to ionizing radiation in lymphocyte subsets.

Human lymphocyte subpopulations differ in their cellular responses to ionizing radiation. To shed light on the molecular basis of this effect, we characterized the transcriptional response to 1 Gy X-rays of CD4+ T lymphocytes. Of 18,433 genes tested, 102 were modulated more than 1.

Gene expression in response to ionizing radiation: an overview of molecular features in hematopoietic cells.

We review the molecular mechanisms involved in response to ionizing radiation in various hematopoietic cell types. First, a general overview of the radiation induced cell signaling molecules in mammals is given. The importance of highly conserved kinases, such as ATM and ATR, as well as the p53 protein for maintaining the genome stability is highlighted.

Effect of ionizing radiation on gene expression in CD4+ T lymphocytes and in Jurkat cells: unraveling novel pathways in radiation response.

To better understand at the molecular level the effect of ionizing radiation in leukocytes, the global transcriptional response to X-ray irradiation was studied in human CD4+ T lymphocytes and in Jurkat cells. Microarray analysis performed on freshly isolated human CD4+ lymphocytes 8 h after an LD50 irradiation dose of 1 Gy revealed that out of 13,825 genes, 1084 were modulated more than 1.5-fold.

Gene expression induced by X-ray irradiation in human blood cell lines.

The response to X-ray irradiation of three different human hematopoietic cell lines originating from T (Jurkat), B (Raji), and promyelocytic (HL60) leukemia was analyzed. The survival after irradiation differed among the three cell lines, with Jurkat cells being the most vulnerable and HL60 being the least sensitive. The profile of gene expression was studied with the microarray technique in both Jurkat and HL60 cell lines.

A genomic approach reveals a novel mitotic pathway in papillomavirus carcinogenesis.

More than 90% of cervical carcinomas are associated with human papillomavirus (HPV) infection. The two viral oncogenes E6 and E7 play a major role in transforming the cells by disrupting p53- and pRb-dependent cell cycle checkpoints. A hallmark of HPV-associated cervical carcinoma is loss of the expression of the viral E2 protein, often by disruption of E2-encoding gene.

Mouse one-cell embryos undergoing a radiation-induced G2 arrest may re-enter S-phase in the absence of cytokinesis.

PCC (premature chromosome condensation) can be used for visualizing and scoring damage induced by radiation in the chromatin of cells undergoing a G1 or G2 arrest. A method involving the fusion of irradiated single embryonic cells with single MI oocytes was used to induce PCC in mouse zygotes of the BALB/c strain, which suffer a drastic G2 arrest after X-irradiation (dose used 2.5 Gy).

Developmental abnormalities induced by X-irradiation in p53 deficient mice.

In order to assess the influence of p53 inactivation on radiation-induced developmental effects, male mice heterozygous for the wild-type p53 allele (mimicking the human Li-Fraumeni syndrome) were crossed with C57BL females, and their heterozygous p53+/- progeny were mated with each other to obtain p53+/-, p53-/- and p53+/+ embryos. Pregnant females were X-irradiated with 0.5 Gy on days 1 (pre-implantation period), 8 or 11 (organogenesis period) of gestation.

Radiation-induced chromosome aberrations in guinea-pig growing oocytes, and their relation to follicular atresia.

The female guinea-pig has been shown to represent a good model to investigate the genetic hazard of ionizing radiation in humans. The sensitivity of the guinea-pig oocytes to radiation-induced chromosome aberrations was, therefore, studied at different stages of oocyte and follicular growth. The sensitivity of oocytes enclosed in small follicles (15 weeks before ovulation) was found to be low and comparable to that of immature oocytes present at birth.

Histone H1 kinase activity in ovulated oocytes.

The level of kinase activity of cdkl is known to be high during metaphase of the two meioses. In this experiment, histone H1 kinase activity (which is known to reflect cdk1 activity) was assayed in BALB/c mouse ovulated oocytes at various timepoints after ovulation. Histone H1 kinase activity in ovulated oocytes was stable up to 37 hours after ovulation.

Delayed effects of prenatal low-dose irradiation in the white matter of the rat brain.

Purpose: This study is part of a general search for the anatomical bases of the severe mental retardation syndrome caused by prenatal irradiation. More specifically, it seeks reasons for the high radiosensitivity of a white matter area, the cingulum of the corpus callosum.

Materials And Methods: Pregnant primiparous Wistar rats were exposed to X-rays at 12, 13, 14 or 15 days of gestation (E12, E13, E14 or E15) with single low doses of 180 mGy.