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The HESI inter-laboratory miRNA project
S48
Abstracts / Toxicology Letters 221S (2013) S31–S56
W13-3 MicroRNAs as markers of drug-induced tissue perturbation and adaptation
W13-5 The HESI inter-laboratory miRNA project
Chris Philip Daniel Jack Sharkey 1 , James Dear 3 , Jonathan Moggs 2 , Neil Kitteringham 1 , Derrick McWhirter 1 , Kevin Park 1
K.L. Thompson 1 , T. Chen 2 , P. Couttet 3 , H. Ellinger-Ziegelbauer 4 , M. Kanki 5 , J. Kelsall 6 , E. Boitier 7 , R. Nassirpour 8 , G. Searfoss 9 , T. Sharapova 10 , Catherine de la Moureyre-Spire 11 , P. Yuen 12 , R. O’Lone 13
1
1
Goldring 1 ,
Starkey-Lewis 1 ,
Antoine 1 ,
MRC Centre for Drug Safety Science, University of Liverpool, UK, Novartis Institutes for Biomedical Research, Basel, Switzerland, 3 Queen’s Medical Research Institute, Edinburgh, UK 2
MicroRNAs (miRNAs) potentially represent a new class of biomarkers which are stable and sensitive, and some of these molecules display a high degree of tissue-selective expression. Clearly they are likely to be useful as markers of injury in amenable biofluids. The challenge now is to understand some of the mechanisms involved in their release, whether this process is active or passive, which types of cellular perturbation are implicated in this process, and even if release serves a role as a signal. In this talk, data will be presented from studies in the MRC Centre for Drug Safety Science on miRNA markers of different forms of organ injury in pre-clinical models and clinical samples, with a particular focus on drug-induced liver injury. The potential of one particular miRNA (-122) as a marker of liver damage will be discussed, in a range of preclinical models (rudimentary hepatocyte-like cell lines, primary human hepatocytes and stem cell-derived hepatocyte-like cells, as well as animal models) and clinical studies. The role of such a marker to bridge between different in vitro and in vivo studies and real-life patient data will be considered, in the context of the need to make our experimental models relevant to human drug-induced tissue injury.Recent significant developments from other groups in this field will also be highlighted. http://dx.doi.org/10.1016/j.toxlet.2013.06.174
W13-4 The role of microRNAs in the response to toxic insult Martin Bushell MRC Toxicology Unit, Leicester, UK Compelling evidence has shown that microRNAs are critical downstream effectors of classic oncogene/tumour suppressor networks. The archetypal examples of oncogene and tumour suppressor microRNAs are the miR-17-92 (oncomir 1) polycistronmicroRNA cluster and the miR-34 family, respectively. Whilst the involvement of these two opposing families of microRNAs in oncogenesis has been known for some time, the mRNA targets through which they exert their phenotypes are only just beginning to be uncovered. Moreover, several recent reports have demonstrated that the relevant physiological targets of certain individual microRNAs are actually fairly limited, with the repression of just one or two major targets being sufficient to explain the observed phenotype. We will present our recent data examining the role of microRNAs in response to toxic insult and how they exert their effects, in particular, focusing on how these mechanisms are deregulated in human cancers and critically involved the in response to DNA damage. http://dx.doi.org/10.1016/j.toxlet.2013.06.175
CDER, Silver Spring, MD, 2 NCTR, Jefferson, AR, 3 Novartis Pharma AG, Basel, Switzerland, 4 Bayer Pharma AG, Wuppertal, Germany, 5 Astellas Pharma Inc., Osaka, Japan, 6 AstraZeneca, Macclesfield, Cheshire, UK, 7 Sanofi, Vitry-sur-Seine, France, 8 Pfizer, Andover, MA, 9 Lilly, Indianapolis, IN, 10 Abbott Laboratories, Abbott Park, IL, 11 Servier, Gidy, France, 12 NIDDK, NIH, Bethesda, MD, 13 ILSI Health and Environmental Sciences Institute, Washington, DC MicroRNAs are promising noninvasive biomarkers of druginduced toxicities due to their tissue-selective expression, rapid release post-injury, and stability in biofluids. However, low level expression of miRNAs in biofluids present challenges to their reliable quantification. Thepresentation will describe the aim and first results of this project developed by the HESI Application of Genomics to Mechanism-based Risk Assessment Technical Committee from 2011 (http://www.hesiglobal.org). A collaborative study was initiated by this HESI committee to assess best practices for measuring injury-related microRNAs in biofluids. A standard protocol for isolation and measurement of miRNA in serum, plasma, and urine to ultimately be used in preclinical toxicological studies were applied on the same samples by a number of participating laboratories (academics, pharma companies and regulators). Samples were derived from a model of acute cardiotoxicity in male Wistar rats induced by a single s.c. injection of 0.5 mg/kg isoproterenol. This study was run centrally in 3 stages and biofluidswere sent to 10 laboratories for analysis of 3 heart-enriched microRNAs miR-208, miR-499, and miR-1. Samples comprised of serum and plasma collected 4 h and 24 h post-injection, and urine collected overnight, each pooled together relatively to treated or control groups. At each site, same samples were assayed by qRTPCR using a standard protocol. Data were also normalized against levels of a spiked-in ath-miR159a control. The results from this interlaboratory analysis of multiple preanalytical and technical issues will provide guidelines for the accurate measurement and reporting of injury-related microRNAs in biofluids. http://dx.doi.org/10.1016/j.toxlet.2013.06.176 Workshop 14: Addressing safety in the immature organism: about children’s safety and risk-assessment
W14-1 Introduction to the immature organism Georg Schmitt Non-clinical Safety, F. Hoffmann-La Roche AG, Basel, Switzerland Developing children have morphological, biochemical, physiological and behavioral characteristics that contrast them from adults. Therefore, safety testing in adult animals may not be predictive for children. Some of the major factors responsible for sensitivity differences to chemical insult between the immature and adult organism are age-related changes in absorption, distribution, metabolism and excretion. Beyond mere quantitative
Abstracts / Toxicology Letters 221S (2013) S31–S56
W13-3 MicroRNAs as markers of drug-induced tissue perturbation and adaptation
W13-5 The HESI inter-laboratory miRNA project
Chris Philip Daniel Jack Sharkey 1 , James Dear 3 , Jonathan Moggs 2 , Neil Kitteringham 1 , Derrick McWhirter 1 , Kevin Park 1
K.L. Thompson 1 , T. Chen 2 , P. Couttet 3 , H. Ellinger-Ziegelbauer 4 , M. Kanki 5 , J. Kelsall 6 , E. Boitier 7 , R. Nassirpour 8 , G. Searfoss 9 , T. Sharapova 10 , Catherine de la Moureyre-Spire 11 , P. Yuen 12 , R. O’Lone 13
1
1
Goldring 1 ,
Starkey-Lewis 1 ,
Antoine 1 ,
MRC Centre for Drug Safety Science, University of Liverpool, UK, Novartis Institutes for Biomedical Research, Basel, Switzerland, 3 Queen’s Medical Research Institute, Edinburgh, UK 2
MicroRNAs (miRNAs) potentially represent a new class of biomarkers which are stable and sensitive, and some of these molecules display a high degree of tissue-selective expression. Clearly they are likely to be useful as markers of injury in amenable biofluids. The challenge now is to understand some of the mechanisms involved in their release, whether this process is active or passive, which types of cellular perturbation are implicated in this process, and even if release serves a role as a signal. In this talk, data will be presented from studies in the MRC Centre for Drug Safety Science on miRNA markers of different forms of organ injury in pre-clinical models and clinical samples, with a particular focus on drug-induced liver injury. The potential of one particular miRNA (-122) as a marker of liver damage will be discussed, in a range of preclinical models (rudimentary hepatocyte-like cell lines, primary human hepatocytes and stem cell-derived hepatocyte-like cells, as well as animal models) and clinical studies. The role of such a marker to bridge between different in vitro and in vivo studies and real-life patient data will be considered, in the context of the need to make our experimental models relevant to human drug-induced tissue injury.Recent significant developments from other groups in this field will also be highlighted. http://dx.doi.org/10.1016/j.toxlet.2013.06.174
W13-4 The role of microRNAs in the response to toxic insult Martin Bushell MRC Toxicology Unit, Leicester, UK Compelling evidence has shown that microRNAs are critical downstream effectors of classic oncogene/tumour suppressor networks. The archetypal examples of oncogene and tumour suppressor microRNAs are the miR-17-92 (oncomir 1) polycistronmicroRNA cluster and the miR-34 family, respectively. Whilst the involvement of these two opposing families of microRNAs in oncogenesis has been known for some time, the mRNA targets through which they exert their phenotypes are only just beginning to be uncovered. Moreover, several recent reports have demonstrated that the relevant physiological targets of certain individual microRNAs are actually fairly limited, with the repression of just one or two major targets being sufficient to explain the observed phenotype. We will present our recent data examining the role of microRNAs in response to toxic insult and how they exert their effects, in particular, focusing on how these mechanisms are deregulated in human cancers and critically involved the in response to DNA damage. http://dx.doi.org/10.1016/j.toxlet.2013.06.175
CDER, Silver Spring, MD, 2 NCTR, Jefferson, AR, 3 Novartis Pharma AG, Basel, Switzerland, 4 Bayer Pharma AG, Wuppertal, Germany, 5 Astellas Pharma Inc., Osaka, Japan, 6 AstraZeneca, Macclesfield, Cheshire, UK, 7 Sanofi, Vitry-sur-Seine, France, 8 Pfizer, Andover, MA, 9 Lilly, Indianapolis, IN, 10 Abbott Laboratories, Abbott Park, IL, 11 Servier, Gidy, France, 12 NIDDK, NIH, Bethesda, MD, 13 ILSI Health and Environmental Sciences Institute, Washington, DC MicroRNAs are promising noninvasive biomarkers of druginduced toxicities due to their tissue-selective expression, rapid release post-injury, and stability in biofluids. However, low level expression of miRNAs in biofluids present challenges to their reliable quantification. Thepresentation will describe the aim and first results of this project developed by the HESI Application of Genomics to Mechanism-based Risk Assessment Technical Committee from 2011 (http://www.hesiglobal.org). A collaborative study was initiated by this HESI committee to assess best practices for measuring injury-related microRNAs in biofluids. A standard protocol for isolation and measurement of miRNA in serum, plasma, and urine to ultimately be used in preclinical toxicological studies were applied on the same samples by a number of participating laboratories (academics, pharma companies and regulators). Samples were derived from a model of acute cardiotoxicity in male Wistar rats induced by a single s.c. injection of 0.5 mg/kg isoproterenol. This study was run centrally in 3 stages and biofluidswere sent to 10 laboratories for analysis of 3 heart-enriched microRNAs miR-208, miR-499, and miR-1. Samples comprised of serum and plasma collected 4 h and 24 h post-injection, and urine collected overnight, each pooled together relatively to treated or control groups. At each site, same samples were assayed by qRTPCR using a standard protocol. Data were also normalized against levels of a spiked-in ath-miR159a control. The results from this interlaboratory analysis of multiple preanalytical and technical issues will provide guidelines for the accurate measurement and reporting of injury-related microRNAs in biofluids. http://dx.doi.org/10.1016/j.toxlet.2013.06.176 Workshop 14: Addressing safety in the immature organism: about children’s safety and risk-assessment
W14-1 Introduction to the immature organism Georg Schmitt Non-clinical Safety, F. Hoffmann-La Roche AG, Basel, Switzerland Developing children have morphological, biochemical, physiological and behavioral characteristics that contrast them from adults. Therefore, safety testing in adult animals may not be predictive for children. Some of the major factors responsible for sensitivity differences to chemical insult between the immature and adult organism are age-related changes in absorption, distribution, metabolism and excretion. Beyond mere quantitative