Non-targeted effects of ionising radiation

Non-targeted effects of ionising radiation Sisko Salomaa STUK - Radiation and Nuclear Safety Authority Co-ordination Meeting on Low Dose Risk Research, Brussels, 20-21 June 2007 06/07/2007 NOTE, 2006-2010 1

General information Start date of the project: 1 September 2006 Duration: 48 months 19 partner organisations Coordinating organisation: STUK - Radiation and Nuclear Safety Authority Project coordinator: Prof. Sisko Salomaa EURATOM Specific Programme for Research and Training on Nuclear Energy, 6 th Framework Program Total eligible costs: 11.89 M EC contribution: 6.33 M 06/07/2007 NOTE, 2006-2010 2

No 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 List of NOTE partners Organisation name Acronym Country Radiation and Nuclear Safety Authority STUK Finland University of Dundee UNDUN UK Leipzig University UL Germany MRC Radiation and Genome Stability Unit MRC UK Imperial College ICFM UK Gray Cancer Institute GCI UK Belgian Nuclear Research Centre SCK-CEN Belgium Dublin Institute of Technology RESC Ireland National Institute of Health ISS Italy University of Leicester ULEICS UK McMaster University MAC Canada Atomic Energy of Canada Limited AECL Canada National Research Institute for Radiobiology and Radiohygiene NRIRR Hungary National Research Centre for Environment and Health GSF Germany University of Pavia UNIPV Italy University of Erlangen-Nuremberg FAU Germany University of Duisburg-Essen UDE Germany Norwegian Radium Hospital NRH Norway University of Ottawa Heart Institute OHIR Canada Scientific contact person Prof. Sisko Salomaa Prof. Eric G. Wright Dr. Guido Hildebrandt Dr. Munira Kadhim Dr. Mark Little Dr. Kevin M. Prise Dr. Paul Jacquet Dr. Fiona Lyng Dr. Maria Antonella Tabocchini Prof. Yuri Dubrova Prof. Carmel Mothersill Dr. Ron Mitchel/Dr. Nick Priest Dr. Géza Sáfrány Dr. Werner Friedland Prof. Andrea Ottolenghi Dr. Franz Roedel Prof. George Iliakis Dr. Jostein Dahle Prof. Stewart Whitman 06/07/2007 NOTE, 2006-2010 3

System of radiation protection Knowledge of radiation risk is based on direct epidemiological evidence, as well as scientific study of radiation biology The system is designed to protect against both deterministic and stochastic effects A linear, non-threshold (LNT) dose-response relationship is used for all long-term health effects (e.g. cancer, genetic effects) A dose and dose-rate correction factor is used to relate the effects of acute exposures to chronic exposures (DDREF) Radiation dose is used as a surrogate for risk The effects produced by different types of radiation are qualitatively the same Doses can be summed to predict overall risk 06/07/2007 NOTE, 2006-2010 4

The classical paradigm in radiobiology - the basis of radiation-induced health effects DNA damage induced at the time of exposure mutation fixed in the first cell division clonal proliferation cancer hereditary effects HEALTH EFFECTS 06/07/2007 NOTE, 2006-2010 5

mutation and cell killing seen in cells that were not directly hit by radiation but were nearby Bystander effect These effects occur in cells that have never been irradiated. 06/07/2007 NOTE, 2006-2010 6

The progeny of irradiated cells show occurrence of aberrations or new mutations and/or new chromosomal other genomic damage lethal mutations (delayed cell death/apoptosis or reproductive cell death) Genomic instability These non-clonal effects occur in cells that have never been irradiated. 06/07/2007 NOTE, 2006-2010 7

Concept of dose as surrogate of risk LNT and radiation protection a linear dose response means that every increment of dose and the associated risk can be assessed separately, irrespective of prior or future doses, as long as doses are below deterministic effects a fixed dose increment is always associated with the same additional risk doses received by an individual at different time points canbesummedup(cumulativedose) (collective dose can be used to predict risk at the population level) 06/07/2007 NOTE, 2006-2010 8

Low-dose effects Effects targeted effects classical effects, dominant at high doses non-targeted effects bystander genomic instability Dose 06/07/2007 NOTE, 2006-2010 9

Non-targeted effects may be important modifiers of risk at the low dose region 06/07/2007 NOTE, 2006-2010 10

A new paradigm of Radiation Biology Targeted effects Classical paradigm of radiation biology DNA damage occurs during or very shortly after irradiation of the nuclei in targeted cells The potential for biological consequences can be expressed within one or two cell generations Non-targeted effects New evidence Bystander effect Genomic instability Adaptive response Clastogenic factors Delayed reproductive death Induction of genes by radiation Premature differentation Low dose hypersensitivity Abscopal (out-of-field) effects 06/07/2007 NOTE, 2006-2010 11

Paradigm shift Classical paradigm New evidence DNA-targeted cells considered as isolated units clonal effects linear / linear quadratic dose response classical radiation biology (effects on DNA) non-targeted cells are communicating tissue responses non-clonal effects plateau-like dose response (on/off) need for new methodological approaches 06/07/2007 NOTE, 2006-2010 12

Key questions from the radiation protection policy point of view For cancer risk: is there a deviation from LNT at low doses? Can ionising radiation cause non-cancer diseases or modify their risk at low / intermediate doses? Are there differences in the radiation sensitivity between individuals? 06/07/2007 NOTE, 2006-2010 13

General objectives of the NOTE IP To investigate the mechanisms of non-targeted effects, in particular, bystander effects, genomic instability and adaptive response To investigate if and how non-targeted effects modulate the cancer risk in the low dose region To investigate if ionising radiation can cause noncancer diseases or beneficial effects at low and intermediate doses To investigate individual susceptibility and other factors modifying non-targeted responses 06/07/2007 NOTE, 2006-2010 14

General objectives of the NOTE IP To contribute to the conceptualisation of a new paradigm in radiation biology that would cover both the classical direct (DNA-targeted) and non-targeted effects. To assess the relevance of non-targeted effects for radiation protection and to set the scientific basis for a modern, more realistic, radiation safety system 06/07/2007 NOTE, 2006-2010 15

WP1 WP6 WP7 Management activities Management board Administrative coordinator Sisko Salomaa WP leaders WP2 WP3 WP4 Infrastructures, training and mobility Kevin Prise Mechanisms of non-targeted effects Eric Wright Non-cancer diseases Guido Hildebrandt Factors modifying non-targeted responses Munira Kadhim Dissemination and exploitation activities Oleg Belyakov Task 1 Communication with public Riikka Laitinen-Sorvari Task 2 Relevance for radiation protection Sisko Salomaa Advisory committee WP5 Modelling of non-targeted effects Mark Little Task 3 Conceptualisation of new paradigm Oleg Belyakov 06/07/2007 NOTE, 2006-2010 16

WP2: Mechanisms of non-targeted effects Eric Wright What is the target for non-targeted effects? What cells produce signals? What is the trigger for the production of signals? What cells respond to signals? What intercellular processes are involved in responding to signals? What are the messengers of bystander responses? What are the mechanisms of non-targeted effects in tissues? What are the mechanisms of genomic instability perpetuation? What are the mechanisms of transgenerational instability? 06/07/2007 NOTE, 2006-2010 17

WP2 Examples of specific hypotheses to be tested Production of signals is cell-type and genotype-dependent and involves targets other than nuclear DNA; Responses to signals are cell-type and genotype-dependent mediated by the stress response pathways that have evolved to deal with cellular injury; Both damaging and protective responses may be elicited in a context-dependent manner; Involvement of reactive oxygen/nitrogen species and cytokines are commonly involved; Perpetuation of genomic instability in both somatic and germ cells may be due to intrinsic epigenetic changes such as methylation; In vivo, an instability phenotype may not necessarily be a reflection of genomically unstable cells but responses to ongoing production of damaging signals; i.e. bystander-type processes; In vivo non-targeted effects may reflect inflammatory-type responses 06/07/2007 NOTE, 2006-2010 18

+ WP2 Signals from irradiated cells Target cells Instability Adaptive response Bystander effects Clonogenic cell survival Cytogenetic aberrations Apoptosis / death Damage responses Signal transduction pathways Gene and protein expression 06/07/2007 NOTE, 2006-2010 19

WP2 Non-targeted mechanisms in vivo T 1 T 2 Instability Adaptive responses Bystander mechanisms Transgenerational effects Cytogenetic aberrations Apoptosis / death Mutation Gene and protein expression Damage responses Inter-cellular signalling Signal transduction pathways 06/07/2007 NOTE, 2006-2010 20

WP 3: Non-cancer diseases Guido Hildebrandt Main question: Can ionising radiation cause non-cancer diseases or beneficial effects at low and intermediate doses? 06/07/2007 NOTE, 2006-2010 21

WP 3: Non-cancer diseases Guido Hildebrandt Tasks: 3.1 Mechanisms of induction of cardiovascular diseases in vitro and in vivo 3.2 Inflammatory vs. anti-inflammatory effects 3.3 Role of non-targeted effects in developmental defects 06/07/2007 NOTE, 2006-2010 22

3.1. Mechanisms of cardiovascular diseases induction -in vitro GCI To determine the responses of endothelial cells and tissue-based models to targeted low dose exposure as a surrogate for determining the role of radiation damage to the vasculature in non-cancer diseases in vitro. 06/07/2007 NOTE, 2006-2010 23

3.2. Mechanisms of cardiovascular diseases induction in vivo AECL, OHIRC, ULMED, ICFM To investigate radiation dose response on atherosclerosis progression (aortic lesions) after exposure at early stage disease in knockouts (ApoE-/-) and in ApoE-/- TP 53+/- as compared to "normals" (ApoE+/+) in vivo. Since inflammatory and thrombotic changes in endothelial cells have an important impact on the development of atherosclerotic lesions we further want to study whether low dose radiation exposure at low dose rate or high dose rate induces vascular changes in the heart of ApoE-/- mice 06/07/2007 NOTE, 2006-2010 24

3.3. Inflammatory or anti-inflammatory responses? ULMED, NRIRR, FAU To investigate the effect of acute and chronic low dose IR on the maturation of dentritic cells (DC) generated from peripheral blood of healthy volunteers as compared to untreated DC, and to analyze treated DC with respect to their protective or augmenting capacities in immunological reactions major immune parameters (lymphocyte number, proliferation rate and cytokine secretion of lymphocytes after stimulation with non-specific mitogens, changes in the proportion of different lymphocyte subpopulations) the inflammatory properties of endothelial cells, subgroups of leukocytes isolated from peripheral blood and macrophages 06/07/2007 NOTE, 2006-2010 25

3.4. Role of non-targeted effects in developmental defects SCK-CEN To investigate transgenerational radiation effects at the level of developmental effects and genomic instability, after irradiation of 1-cell mouse embryos with low doses of X-rays. Reproductive effects, congenital anomalies, cytokine levels and chromosomal instability 06/07/2007 NOTE, 2006-2010 26

WP4: Factors modifying non-targeted responses Munira Kadhim influence of genetic factors on the in vivo production of non-targeted effects impact of genetic predisposition on non-targeted effects response in vitro molecular mechanisms determination that contribute to the distinct radiation responses at low dose radiations in mice with different radiosensitivity role of DNA repair and cell cycle checkpoint responses 06/07/2007 NOTE, 2006-2010 27

WP4 - Factors modifying non-targeted responses Genomic instability Bystander effects Transgenerational instability In vitro & in vivo Genetic Factors : -Inter-individual differences - cell lines differences Mechanistic Studies : -Involvement of repair, checkpoints and cell cycle control. -Role of epigenetic factors. Radiation dose and quality Link with WP2 - The influence of genetic factors on the in vivo production of bystander signals producing non-targeted effects and the genotypedependency of response mechanisms. 06/07/2007 NOTE, 2006-2010 28

WP4 Factors modifying non-targeted responses Summary of Deliverables 4.1 The genotype-dependence of signal production at different times after in vivo irradiation (UDUN) 4.2 Inter-individual differences in bystander response, and genomic comparison between primary and permanent cultures pre- and post-irradiation (NRIRR) 4.3 Role of particular proteins in the genetic differences observed in CBA and C57 in response to low dose low LET radiations (MRC) 4.4 Capacity of targeted and non-targeted bystander cells to repair DNA damage using NHEJ or HR (UDE) 06/07/2007 NOTE, 2006-2010 29

WP5: Modelling of non-targeted responses Mark Little intracellular and local intercellular communication and damage mechanisms intercellular communication mechanisms in vitro extracellular signalling mechanims in vivo genomic instability mechanisms in vivo 06/07/2007 NOTE, 2006-2010 30

WP5 main questions 1. Quantitatively test working hypotheses on mechanisms underlying non-targeted effects (bystander effects, genomic instability, adaptive response and non-cancer diseases) 2. Assess role in determining deviations from linear nothreshold hypothesis 3. Address tissues in which effects might arise, also mechanisms (intracellular stress response, signalling molecules etc) that might explain what is observed. 06/07/2007 NOTE, 2006-2010 31

WP5 Tasks Task 5.0 Modeller- experimentalist interaction (all): devise experiments that allow modellers to discriminate between models, and ascertain model parameters Task 5.1 Intracellular and local intercellular communication and damage mechanisms (GSF, ISS): elucidate intracellular mechanisms that might account for non-targeted effects, and integrate into better understood mechanisms for targeted effects (DNA damage and repair) 06/07/2007 NOTE, 2006-2010 32

WP5 tasks Task 5.2 Intercellular communication mechanisms in vitro (UNIPV, RR HF): evaluate candidate intercellular signalling mechanisms that might account for non-targeted effects (in particular bystander effect), and way these can be modulated Task 5.3 Extracellular signalling mechanisms in vivo (ICFM): devise models for non-targeted effects at organ or tissue level, taking account of mechanisms proposed in subgroups. In particular, develop number of candidate models of radiation-induced cardiovascular disease 06/07/2007 NOTE, 2006-2010 33

WP 6: Integration activities Kevin Prise development and better use of existing radiobiological facilities strengthening collaborative RTD activity supporting training and mobility of scientists educating the next generation of European radiobiologists on non-targeted effects 06/07/2007 NOTE, 2006-2010 34

Infrastructures within NOTE Charged particle microbeams (Leipzig and Northwood) Soft X-ray microbeam (Belfast) Small animal low dose facility (Chalk River) Low level exposure facility (Rome) Proteomics and genomics facilities (Helsinki) 06/07/2007 NOTE, 2006-2010 35

NOTE Training Course Protocols and Pitfalls in the study of Non-targeted Effects of Radiation 15-16 th September, Crete General overview lectures on Non-targeted effects External lecturers including Bill Morgan / Christian Streffer Review talks on specific end-points with emphasis on protocols, data interpretation Break-out sessions on technical aspects with emphasis on pitfalls and problems Wrap up and general discussion session Targeted at research students and post-docs 06/07/2007 NOTE, 2006-2010 36

WP 7: Dissemination and exploitation activities Oleg Belyakov a public NOTE website, NOTE Newsletter, Press releases a workshop on the conceptualisation of the new paradigm (2008) a workshop on the relevance of non-targeted effects for radiation protection (2010) 06/07/2007 NOTE, 2006-2010 37

WP7 structure and links with other WPs WP6 Integration activities: Infrastructures, training and mobility WP2 Mechanisms of non-targeted effects WP3 Non-cancer diseases WP4 Factors modifying non-targeted responses WP5 Modelling of non-targeted effects WP7 Dissemination and exploitation activities Task 1 Communication with public Task 2 Conceptualisation of new paradigm Task 3 Relevance for radiation protection End-users General public Scientific community Radiation protection community Decision makers 06/07/2007 NOTE, 2006-2010 38

NOTE website A new NOTE IP website with public and secure areas was launched in April 2007 at http://www.note-ip.org/. 06/07/2007 NOTE, 2006-2010 39

NOTE-related news for scientific community, media and general public The first newsletter was released in May 2007; appears every half a year Subscribe the Newsletter at the NOTE web site! NOTE Newsletter 06/07/2007 NOTE, 2006-2010 40

Future directions... High versus low doses High versus low dose rate Cell cultures -> 3-D models -> tissues -> animals -> human data Tissue responses, effect of microstructure Non-targeted responses involve complex interactions, need for new holistic approaches Generation of hypothesis for mechanistic studies using systems biological approaches ( discovery science ) Mathematical modelling of genomics and proteomics data obtained during NOTE project 06/07/2007 NOTE, 2006-2010 41

Low-dose risk: Increasing the power of epidemiology High background in cancer incidence -> very large populations needed to detect small increases Confounding factors may mask risks Limitations in dosimetry, especially in environmental and protracted exposures -> hampers detection of risk Quantification of risk at low doses is very challenging Combine information from epidemiology and radiation biology (mechanisms, dose response, susceptibility)?? 06/07/2007 NOTE, 2006-2010 42