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Assessment of candidate biomarkers of drug induced liver injury in preclinical toxicity studies
S152
Abstracts / Toxicology Letters 189S (2009) S57–S273
sistence and their high potential to bioaccumulate in organisms they are of toxicological and public concern. The toxicity of PFOS and PFOA has been extensively studied in rodents and hepatotoxicity, developmental toxicity, and a carcinogenic potency are the effects of main concern for both compounds. In the present study, perfluorooctane sulphonate (PFOS), perfluorooctanoic acid (PFOA) and perfluorohexane sulfonate (PFHxS) were quantified in 304 breast milk samples collected during the Bavarian Monitoring of Breast Milk (BAMBI) program in 2007 and 2008. PFOS could be quantified in 273 (90%) of all samples. The concentration ranged between 0.03 and 0.263 g/l (median: 0.057 g/l). In only seven samples (2%) PFOA reached the LOQ of 0.150 g/l with a highest concentration of 0.289 g/l. PFHxS was detected in only five samples (2%) above the LOQ of 0.020 g/l. Based on these results, an intake of 0.009 g/kg b.w. PFOS per day (using median) respectively 0.042 g/kg b.w. (using maximum value) via breast milk was obtained for an infant. The data suggest that fully breastfed infants are unlikely to exceed the recommended tolerable daily intake (TDI) of PFOS of 0.150 g/kg b.w. per day. Due to similar or lower concentrations of PFOA in breast milk but higher TDI of PFOA (1.50 g/kg b.w. per day) this is also true for PFOA. For PFHxS no TDI exists so far. doi:10.1016/j.toxlet.2009.06.748
Y06 Contaminant effects in shore crabs (Carcinus maenas) from Ria Formosa Lagoon Vera Maria 1,∗ , Maria Ana Santos 2 , Maria João Bebianno 1 1
Algarve University, 2 CIMA & Faculty of Marine Environmental Sciences, Faro, Portugal, 2 Aveiro University, CESAM & Biology Department, Aveiro, Portugal
In the current work an overview regarding defence and damage biomarkers signals in relation to contamination in two sites at Ria Formosa Lagoon (South of Portugal) is given using female and male shore crabs, Carcinus maenas. The cross transplantation occurred during 6 days at a hypothetical reference site (Ramalhete (R)), and a contaminated site (Olhão (O)). General enzymatic antioxidant responses (gills and hepatopancreas (Hp) catalase (CAT) and gills glutathione peroxidase (GPx) activities increase) showed that transplanted crabs (female and male) are suffering pro-oxidant challenges at O site. Gills and Hp GST were reduced in both gender crabs transplanted from R to O. MT induction in crabs transplanted from site R to O (contaminated by metals and PAHs) was found, however differences were also observed towards gender and organ specificities. High gills LPO exist in male crabs transplanted from R to O, while in females it was the opposite. In both gender crabs from O site gills DNA integrity decreased compared to R feral crabs. Moreover, Hp DNA integrity decreases in male crabs transplanted from O to R site which may be related to environmental conditions (lower contamination levels) revealing the difficulty of selection of reference sites in field studies. Data demonstrated that female and male C. maenas antioxidant defences and damage biomarkers were sensitive to the mixture of contaminants present in these sites as well as good indicators of general stress. doi:10.1016/j.toxlet.2009.06.749
Y07 Assessment of candidate biomarkers of drug induced liver injury in preclinical toxicity studies Melanie Adler 1,∗ , Dana Hoffmann 1 , Heidrun Ellinger 2 , Phil Hewitt 3 , Katja Matheis 4 , Laoighse Mulrane 5 , William M. Gallagher 5 , Laura Suter-Dick 6 , Michael M. Fountoulakis 6 , Wolfgang Dekant 1 , Angela Mally 1 University of Würzburg, Toxicology, Würzburg, Germany, 2 Bayer Schering Pharma AG, Berlin, Germany, 3 Merck KGaA, Darmstadt, Germany, 4 Boehringer Ingelheim Pharma GmbH & Co.KG, Biberach, Germany, 5 University College Dublin, Dublin, Ireland, Ireland, 6 Hoffmann-La Roche AG, Basel, Switzerland, Switzerland
1
This study, which was conducted as part of the Innomed PredTox project (www.innomed-predtox.com), was designed to assess the value of a set of potential markers for improved detection of liver injury in preclinical toxicity studies. Male Wistar rats were treated with drug candidates (BAY16, EMD335823, BI-3) that previously failed during development, in part due to hepatotoxicity, at two dose levels for 1, 3 and 14 days. Concentrations of lipocalin-2 and clusterin, which are frequently overexpressed and released from damaged tissues, and thiostatin, recently identified within PredTox as being elevated in urine in response to liver injury, were determined in rat urine and serum by ELISA. This was supplemented by confirmatory qRT-PCR and immunohistochemical analyses in the target organ. Serum paraoxonase-1 activity, which has been suggested as a marker of hepatotoxicity, was determined using a fluorometric assay. Clusterin and paraoxonase-1 were not consistently altered in response to liver injury. In contrast, thiostatin and lipocalin-2 were increased in serum and urine of treated animals in a time- and dosedependent manner. These changes correlated well with mRNA expression in the target organ and generally reflected the onset and degree of liver injury, which was characterized by hepatocyte and bile duct damage, inflammation and regeneration in all studies. Although lipocalin-2 and thiostatin are both acute phase proteins expressed in a range of tissues and may thus not be specific for liver injury, our results indicate that both markers may serve as sensitive, minimally invasive diagnostic markers of inflammation and tissue damage. doi:10.1016/j.toxlet.2009.06.750
Y08 Evaluation of urinary kidney biomarkers in rat models of acute and subchronic nephrotoxicity Dana Hoffmann 1,∗ , Tanja Henzler 1 , Thomas Herget 2 , John Callanan 3 , Wolfgang Dekant 1 , Philip Hewitt 2 , Angela Mally 1 1
University of Wuerzburg, Toxicology, Wuerzburg, Germany, 2 Merck KGaA, Darmstadt, Germany, 3 University College Dublin, Dublin, Ireland For improved detection of nephrotoxicity, a panel of novel urinary kidney biomarkers was recently submitted to and approved by the US-FDA and EMEA. However, few data regarding the performance of these markers are publicly available so far. In this study, we investigated the potential of some of the newly accepted makers (Kim-1, b2m, cystatin C, clusterin) along with six additional urinary key proteins of kidney injury (GST-a, Timp-1, VEGF, calbindin, NGAL/lipocalin-2, osteopontin) to detect kidney injury in
Abstracts / Toxicology Letters 189S (2009) S57–S273
sistence and their high potential to bioaccumulate in organisms they are of toxicological and public concern. The toxicity of PFOS and PFOA has been extensively studied in rodents and hepatotoxicity, developmental toxicity, and a carcinogenic potency are the effects of main concern for both compounds. In the present study, perfluorooctane sulphonate (PFOS), perfluorooctanoic acid (PFOA) and perfluorohexane sulfonate (PFHxS) were quantified in 304 breast milk samples collected during the Bavarian Monitoring of Breast Milk (BAMBI) program in 2007 and 2008. PFOS could be quantified in 273 (90%) of all samples. The concentration ranged between 0.03 and 0.263 g/l (median: 0.057 g/l). In only seven samples (2%) PFOA reached the LOQ of 0.150 g/l with a highest concentration of 0.289 g/l. PFHxS was detected in only five samples (2%) above the LOQ of 0.020 g/l. Based on these results, an intake of 0.009 g/kg b.w. PFOS per day (using median) respectively 0.042 g/kg b.w. (using maximum value) via breast milk was obtained for an infant. The data suggest that fully breastfed infants are unlikely to exceed the recommended tolerable daily intake (TDI) of PFOS of 0.150 g/kg b.w. per day. Due to similar or lower concentrations of PFOA in breast milk but higher TDI of PFOA (1.50 g/kg b.w. per day) this is also true for PFOA. For PFHxS no TDI exists so far. doi:10.1016/j.toxlet.2009.06.748
Y06 Contaminant effects in shore crabs (Carcinus maenas) from Ria Formosa Lagoon Vera Maria 1,∗ , Maria Ana Santos 2 , Maria João Bebianno 1 1
Algarve University, 2 CIMA & Faculty of Marine Environmental Sciences, Faro, Portugal, 2 Aveiro University, CESAM & Biology Department, Aveiro, Portugal
In the current work an overview regarding defence and damage biomarkers signals in relation to contamination in two sites at Ria Formosa Lagoon (South of Portugal) is given using female and male shore crabs, Carcinus maenas. The cross transplantation occurred during 6 days at a hypothetical reference site (Ramalhete (R)), and a contaminated site (Olhão (O)). General enzymatic antioxidant responses (gills and hepatopancreas (Hp) catalase (CAT) and gills glutathione peroxidase (GPx) activities increase) showed that transplanted crabs (female and male) are suffering pro-oxidant challenges at O site. Gills and Hp GST were reduced in both gender crabs transplanted from R to O. MT induction in crabs transplanted from site R to O (contaminated by metals and PAHs) was found, however differences were also observed towards gender and organ specificities. High gills LPO exist in male crabs transplanted from R to O, while in females it was the opposite. In both gender crabs from O site gills DNA integrity decreased compared to R feral crabs. Moreover, Hp DNA integrity decreases in male crabs transplanted from O to R site which may be related to environmental conditions (lower contamination levels) revealing the difficulty of selection of reference sites in field studies. Data demonstrated that female and male C. maenas antioxidant defences and damage biomarkers were sensitive to the mixture of contaminants present in these sites as well as good indicators of general stress. doi:10.1016/j.toxlet.2009.06.749
Y07 Assessment of candidate biomarkers of drug induced liver injury in preclinical toxicity studies Melanie Adler 1,∗ , Dana Hoffmann 1 , Heidrun Ellinger 2 , Phil Hewitt 3 , Katja Matheis 4 , Laoighse Mulrane 5 , William M. Gallagher 5 , Laura Suter-Dick 6 , Michael M. Fountoulakis 6 , Wolfgang Dekant 1 , Angela Mally 1 University of Würzburg, Toxicology, Würzburg, Germany, 2 Bayer Schering Pharma AG, Berlin, Germany, 3 Merck KGaA, Darmstadt, Germany, 4 Boehringer Ingelheim Pharma GmbH & Co.KG, Biberach, Germany, 5 University College Dublin, Dublin, Ireland, Ireland, 6 Hoffmann-La Roche AG, Basel, Switzerland, Switzerland
1
This study, which was conducted as part of the Innomed PredTox project (www.innomed-predtox.com), was designed to assess the value of a set of potential markers for improved detection of liver injury in preclinical toxicity studies. Male Wistar rats were treated with drug candidates (BAY16, EMD335823, BI-3) that previously failed during development, in part due to hepatotoxicity, at two dose levels for 1, 3 and 14 days. Concentrations of lipocalin-2 and clusterin, which are frequently overexpressed and released from damaged tissues, and thiostatin, recently identified within PredTox as being elevated in urine in response to liver injury, were determined in rat urine and serum by ELISA. This was supplemented by confirmatory qRT-PCR and immunohistochemical analyses in the target organ. Serum paraoxonase-1 activity, which has been suggested as a marker of hepatotoxicity, was determined using a fluorometric assay. Clusterin and paraoxonase-1 were not consistently altered in response to liver injury. In contrast, thiostatin and lipocalin-2 were increased in serum and urine of treated animals in a time- and dosedependent manner. These changes correlated well with mRNA expression in the target organ and generally reflected the onset and degree of liver injury, which was characterized by hepatocyte and bile duct damage, inflammation and regeneration in all studies. Although lipocalin-2 and thiostatin are both acute phase proteins expressed in a range of tissues and may thus not be specific for liver injury, our results indicate that both markers may serve as sensitive, minimally invasive diagnostic markers of inflammation and tissue damage. doi:10.1016/j.toxlet.2009.06.750
Y08 Evaluation of urinary kidney biomarkers in rat models of acute and subchronic nephrotoxicity Dana Hoffmann 1,∗ , Tanja Henzler 1 , Thomas Herget 2 , John Callanan 3 , Wolfgang Dekant 1 , Philip Hewitt 2 , Angela Mally 1 1
University of Wuerzburg, Toxicology, Wuerzburg, Germany, 2 Merck KGaA, Darmstadt, Germany, 3 University College Dublin, Dublin, Ireland For improved detection of nephrotoxicity, a panel of novel urinary kidney biomarkers was recently submitted to and approved by the US-FDA and EMEA. However, few data regarding the performance of these markers are publicly available so far. In this study, we investigated the potential of some of the newly accepted makers (Kim-1, b2m, cystatin C, clusterin) along with six additional urinary key proteins of kidney injury (GST-a, Timp-1, VEGF, calbindin, NGAL/lipocalin-2, osteopontin) to detect kidney injury in