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Identification of microRNA as possible risk marker for drug-induced liver injury using chimeric PXB-Mouse® with highly humanized liver
Abstracts / Toxicology Letters 258S (2016) S62–S324
genes related to the XM, response to oxidative stress and apoptosis. Experiments were conducted with third instar larvae of two strains of D. melanogaster, Oregon-flare3 and flare3, treated with ZEN (260 microM) and controls water (negative), phenobarbital (positive) and buffer (dissolvent). Oligonucleotides were designed and, through end-point PCR of genomic DNA, the amplification of a single product and its size (100–200 bp) was verified. The expression of Cyp6a2 and Cyp6g1, Hsp60, Hsp70, Dmp53, H99 was evaluated through semiquantitative PCR using cDNA and amplification conditions were normalized with the ˇ-actin transcript. We obtained Hsp70 and Cyp6g1 transcripts in treatment and controls. The presence of Hsp70 is attributed to the great variety of functions it fulfills in the cell. As for Cyp6g1, we propose that its presence could be due to the fact that in D. melanogaster it takes part in the metabolism of 20-hydroxyecdysone. Hsp60 and Cyp6a2 transcripts was also found in the phenobarbital treatments. Results will be validated through Real Time PCR. Grant PAPIIT 200814. http://dx.doi.org/10.1016/j.toxlet.2016.06.1357 P01-048 Modulation of the genotoxicity of the 4-nitroquinoline-1-oxide by cotreatments with lycopene, resveratrol, vitamin C or vitamin C + FESO4 using the [Drosophila melanogaster] wing spot test ˜ Garcia, M.R. Arellano Llamas, J. De M.E. Heres Pulido ∗ , I.E. Duenas ˜ La Cruz Núnez, V. Cisneros Carrillo, S. Palacios López, L. Acosta ˜ Gómez, L.F. Santos Cruz, L. Castaneda Partida, A. Duran Diaz, E. Piedra Ibarra, J.T. Torres Gregorio UNAM, Iztacala, Tlalnepantla, Mexico It has been suggested that the intake of antoxidants would help prevent oxidant stress effects. 4-Nitroquinoline-1-oxide (4NQO) is a pro-oxidant carcinogen and promutagen bioactivated by the Xenobiotic Metabolism (XM). We investigated if antioxidants lycopene [LYC, 0.45, 0.9 and 1.8 [M], resveratrol [RES, 11, 43 and 172 M] and vitamin C [VitC, 5.6 mM], added or not with FeSO4 [0.2 mM], modulate the genotoxicity produced by 4NQO [2 mM]. Treatments were performed with the Drosophila wing spot test standard (ST) and high bioactivation (HB) crosses, with basal and high levels of cytochromes P450 (Cyp450s), respectively. Genotoxicity of 4-NQO was higher in the HB cross and when dissolved in ethanol or acetone. The antioxidants did not show any protection against 4-NQO in any of both crosses. In the ST cross, RES [11 [M] and VitC resulted genotoxic. Also, FeSO4 and VitC + FeSO4 modulated the genotoxicity of 4-NQO in the following fashion: 4-NQO + VitC + FeSO4 > 4-NQO + FeSO4 > 4-NQO + VitC > 4NQO + LYC > 4-NQO + RES > 4-NQO. In the HB cross, none of the three antioxidants proved to be genotoxic. RES [172 M] + 4-NQO increased the genotoxic activity of 4-NQO in both crosses. We concluded that the effects of the three antioxidants, FeSO4 and the modulation of 4-NQO were the result of the difference of Cyp450s levels, regulated and high, in the ST and HB crosses, respectively. In order to prove this, the expression levels of some molecular markers related to the XM and oxidative stress are currently under investigation. Grant PAPIIT #200814. http://dx.doi.org/10.1016/j.toxlet.2016.06.1358
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P02 – Biomarkers P02-001 Downregulation of mir-144-451 cluster promotes esophageal carcinoma Z. Gao ∗ , R. Liu, L. Yin, Y. Pu Key Laboratory of Environmental Medicine Engineering, Ministry of Education, School of Public Health, Southeast University, China miRNA cluster is widely expressed across species, accumulating studies have illustrated that miRNA cluster functioned more efficiently than a single miRNA in some cancer oncogenesis. It is likely that miRNA cluster is more stable and reliable than individual miRNA to be biomarkers for diagnosis and therapy. However, the regulatory mechanism of miRNA cluster was largely unknown. In current study, we detected the expression of the mature miRNAs of miR-144-451 cluster from 102 patients pathologically diagnosed with esophageal carcinoma using stem-loop real-time reverse transcription PCR. The expression level of has-miR-451a, has-miR144-3p, has-miR-144-5p in tumor tissues were significantly lower compared with adjacent non-tumor tissues, the fold-change for these three miRNAs were 0.376, 0.463 and 0.443 respectively (P < 0.05). Principal component regression analysis indicated down regulation of has-miR-144-3p and has-miR-144-5p increased the risk for esophageal carcinoma, the OR were 0.850 and 0.840 respectively. Stable transgenic cells overexpressing miR-144-451 were obtained using lentivirus containing region of pri-miR-144-451. The overexpression of miR-144-451 cluster leads to the arrest of S and G2 phase, while the invasion ability of stable transgenic cells was seriously inhibited. However, no difference of apoptosis, migration, proliferation were found between stable transgenic and negative control cells. We further found c-Myc, p-ERK were downregulated in stable transgenic cells, while p53 was upregulated. Furthermore, the downstream effectors of c-Myc, MMP9 and pcdc2 were downregulated in stable transgenic cells. miR-144-451 may or partly inhibited cell cycles and invasion of EC9706 through inhibiting ERK/c-Myc signaling pathway. http://dx.doi.org/10.1016/j.toxlet.2016.06.1359
P02-002 Identification of microRNA as possible risk marker for drug-induced liver injury using chimeric PXB-Mouse® with highly humanized liver T. Kitahara 1,∗ , T. Shimada 2 , S. Nagatsuka 3 , H. Enomoto 4 , H. Tachiki 4 , H. Sasaki 1 1 Department of Hospital Pharmacy, Nagasaki University Hospital, Nagasaki, Japan 2 PhoenixBio Co., Ltd., Hiroshima, Japan 3 ADME & Tox. Research Institute, Sekisui Medical Co., Ltd., Ibaraki, Japan 4 Research and Development Division, Towa Pharmaceutical Co., Ltd., Kyoto, Japan
Purpose: Recently, microRNAs (miRNAs) have emerged as risk marker of drug-induced liver injury (DILI) in clinical. We developed a novel method to identify miRNAs as possible risk markers for DILI. We analyzed expression of miRNAs which fluctuated under the treatment of drugs with relatively higher rate of hepatotoxic
S78
Abstracts / Toxicology Letters 258S (2016) S62–S324
incidents (hTOX) and those with negligible hepatotoxic incidents (non-hTOX) using chimeric mice with highly humanized liver. Methods: We selected 8 hTOX (Amiodarone, Acetaminophen, Ticlopidine, Terbinafine, Methotrexate, Valproate, Diclofenac, Benzbromarone) drugs and 8 non-hTOX (Dipyridamole, Cefalexin, Eperisone, Propranolol, Pirenzepine, Trihexyphenidyl, Dapagliflozin, Bumetanide) drugs based on information shown in the package insert. The drugs were administered to chimeric mice with humanized liver (PXB-mice® : PhoenixBio Co., Ltd.) threetimes daily. Hepatic miRNAs were analyzed using GeneChip® miRNA 3.0 array (Affymetrix Inc.), in order to select miRNAs that increased by 1.5 times or more in hTOX drugs and not increased in the non-hTOX drugs. Results and conclusion: We found 7 miRNAs (miR-10a, miR183, miR-557, miR-1202, miR-3065-5p, miR-3195 and miR-4535) that increased only in hTOX treatments and not in non-hTOX treatments. Previous studies revealed the role of miR-10a, miR-183 and miR-1202 in non-hepatic tissues. This is the first report showing these 7 miRNAs might be involved in DILI. The present method is useful for discovery of new miRNAs as risk marker of DILI without clinical trials. http://dx.doi.org/10.1016/j.toxlet.2016.06.1360 P02-003 Endocrine disruptor compound’s levels in hair of Greek children M.N. Tzatzarakis 1 , V. Karzi 1 , E. Vakonaki 1 , I. Katsikantami 1 , M. Kiriakakis 1 , I. Tsakiris 2 , M. Kavvalakis 1 , A.K. Rizos 3 , A.M. Tsatsakis 1,∗ 1 Laboratory of Toxicology, Medical School, University of Crete, Heraklion, Crete, Greece 2 Department of Agricultural Technology, School of Agriculture Technology, Food Technology and Nutrition, TEI of Western Macedonia, Greece 3 Department of Chemistry, University of Crete and Foundation for Research and Technology-Hellas (FORTH-IESL), Heraklion, Crete, Greece
Purpose: Bisphenol A (BPA), triclosan (TCS) and perfluorooctanoic acid (PFOA) are endocrine disrupting chemicals that interfere with the body’s endocrine system. They are widely used in personal care products, toys, plastic containers and metal food cans. The aim of this research is to evaluate the exposure of children to these chemicals by analyzing head hair samples using a liquid chromatography–mass spectrometry based protocol. Methods: Head hair samples (1–55 cm) were collected from 72 children (2–11 years old), residents of Crete, Greece. A consent form was obtained from the children’s parents. The extraction of the analytes was performed by ultrasonic assisted methanolic extraction for 4 h at 50 ◦ C. Results: The method was linear (r2 > 0.99). The mean, inter day precision (%RSD), % accuracy and % recovery of the method for BPA were 14.9, 99.7 and 98.0, for PFOA were 16.3, 98.2 and 81.3, for TCS were 18.3, 94.1 and 103.3, respectively. The 71.2% of the samples were positive for BPA, 6.8% for PFOA and 89.0% for TCS. The median concentration of the positive samples was found to be 12.2 pg/mg (mean 20.4 pg/mg, range 2.6–205.5 pg/mg) for BPA, 21.0 pg/mg (mean 21.1 pg/mg, range 20.8–21.5 pg/mg) for PFOA and 48.8 pg/mg (mean 275.2 pg/mg, range 3.6–7656.6 pg/mg) for TCS. Conclusion: This is the first reported methodology concerning simultaneous detection of BPA, TCS and PFOA in hair. High frequencies of positive samples were found both for triclosan and BPA.
Moreover, a wide range of the detected levels, mostly for TCS, was observed. http://dx.doi.org/10.1016/j.toxlet.2016.06.1361 P02-004 Estrogen and androgen receptor transactivation by actual serum mixtures of lipophilic persistent organic pollutants extracted from Greenlandic pregnant women M. Ghisari ∗ , E.C. Bonefeld Jørgensen Centre for Arctic Health & Molecular Epidemiology, Department of Public Health, Aarhus University, Aarhus, Denmark Greenlandic Inuit have high blood levels of environmental persistent organic pollutants (POPs) as a result of high consumption of contaminated marine mammals. Since the human exposure is not limited to a single compound, the assessment of the combined hormone disrupting potential of the actual serum POP mixture is important. We aimed to analyze the potential interference of the actual serum lipophilic POP mixture with estrogen receptor (ER) and androgen receptor (AR) function. The lipophilic POPs were extracted by SPE-HPLC from 215 pregnant Greenlandic women serum samples collected 2013–2015. The extracts were analyzed for xenoestrogenic and xenoandrogenic activities using luciferase reporter gene assays. Using Spearman’s analyses the correlations between the POP-induced ER and AR transactivities and the actual serum levels of POPs [14 polychlorinated biphenyls (PCBs), one polybrominated biphenyl, four polybrominated diphenyl ethers and 11 organochlorine pesticides] were assessed. The preliminary results show that serum lipophilic POPs have the potential to transactivate the ER and AR, suggesting an overall estrogenic and androgenic activity. When the serum extracts were analyzed in the presence of the natural AR and ER ligands, dihydrotestosterone (DHT) and 17-estradiol (E2), most of the serum extracts antagonized the DHT-induced AR transactivity, suggesting an overall AR inhibiting potential of the serum lipophilic POP. A positive correlation between ER transactivities and the sum of the PCB concentrations in the blood was observed. In conclusion, our data shows that the actual serum mixture of lipophilic POPs in Greenlandic pregnant women have hormone disruptive potential. http://dx.doi.org/10.1016/j.toxlet.2016.06.1362 P02-005 Allele and genotype frequencies of CYP2B6 gene *5 polymorphism in a Turkish population S. Süzen ∗ , N.Y. Artun Ankara University, Faculty of Pharmacy, Department of Toxicology, Tandogan, Ankara, Turkey The cytochrome P450 (CYP450) proteins are monooxygenases, which catalyze many reactions involved in drug metabolism and synthesis of cholesterol, steroids and other lipids (1). CYP2B6 plays a major role in the metabolism of several therapeutically important drugs, including the anticancer agents, the antiretrovirals, the narcotics, the antidepressants, the antiestrogen tamoxifen; the synthetic opioid methadone and many more (2,3). The enzyme also metabolizes certain endogenous compounds such as testosterone, as well as recreational drugs, including nicotine and ecstasy/MDMA
genes related to the XM, response to oxidative stress and apoptosis. Experiments were conducted with third instar larvae of two strains of D. melanogaster, Oregon-flare3 and flare3, treated with ZEN (260 microM) and controls water (negative), phenobarbital (positive) and buffer (dissolvent). Oligonucleotides were designed and, through end-point PCR of genomic DNA, the amplification of a single product and its size (100–200 bp) was verified. The expression of Cyp6a2 and Cyp6g1, Hsp60, Hsp70, Dmp53, H99 was evaluated through semiquantitative PCR using cDNA and amplification conditions were normalized with the ˇ-actin transcript. We obtained Hsp70 and Cyp6g1 transcripts in treatment and controls. The presence of Hsp70 is attributed to the great variety of functions it fulfills in the cell. As for Cyp6g1, we propose that its presence could be due to the fact that in D. melanogaster it takes part in the metabolism of 20-hydroxyecdysone. Hsp60 and Cyp6a2 transcripts was also found in the phenobarbital treatments. Results will be validated through Real Time PCR. Grant PAPIIT 200814. http://dx.doi.org/10.1016/j.toxlet.2016.06.1357 P01-048 Modulation of the genotoxicity of the 4-nitroquinoline-1-oxide by cotreatments with lycopene, resveratrol, vitamin C or vitamin C + FESO4 using the [Drosophila melanogaster] wing spot test ˜ Garcia, M.R. Arellano Llamas, J. De M.E. Heres Pulido ∗ , I.E. Duenas ˜ La Cruz Núnez, V. Cisneros Carrillo, S. Palacios López, L. Acosta ˜ Gómez, L.F. Santos Cruz, L. Castaneda Partida, A. Duran Diaz, E. Piedra Ibarra, J.T. Torres Gregorio UNAM, Iztacala, Tlalnepantla, Mexico It has been suggested that the intake of antoxidants would help prevent oxidant stress effects. 4-Nitroquinoline-1-oxide (4NQO) is a pro-oxidant carcinogen and promutagen bioactivated by the Xenobiotic Metabolism (XM). We investigated if antioxidants lycopene [LYC, 0.45, 0.9 and 1.8 [M], resveratrol [RES, 11, 43 and 172 M] and vitamin C [VitC, 5.6 mM], added or not with FeSO4 [0.2 mM], modulate the genotoxicity produced by 4NQO [2 mM]. Treatments were performed with the Drosophila wing spot test standard (ST) and high bioactivation (HB) crosses, with basal and high levels of cytochromes P450 (Cyp450s), respectively. Genotoxicity of 4-NQO was higher in the HB cross and when dissolved in ethanol or acetone. The antioxidants did not show any protection against 4-NQO in any of both crosses. In the ST cross, RES [11 [M] and VitC resulted genotoxic. Also, FeSO4 and VitC + FeSO4 modulated the genotoxicity of 4-NQO in the following fashion: 4-NQO + VitC + FeSO4 > 4-NQO + FeSO4 > 4-NQO + VitC > 4NQO + LYC > 4-NQO + RES > 4-NQO. In the HB cross, none of the three antioxidants proved to be genotoxic. RES [172 M] + 4-NQO increased the genotoxic activity of 4-NQO in both crosses. We concluded that the effects of the three antioxidants, FeSO4 and the modulation of 4-NQO were the result of the difference of Cyp450s levels, regulated and high, in the ST and HB crosses, respectively. In order to prove this, the expression levels of some molecular markers related to the XM and oxidative stress are currently under investigation. Grant PAPIIT #200814. http://dx.doi.org/10.1016/j.toxlet.2016.06.1358
S77
P02 – Biomarkers P02-001 Downregulation of mir-144-451 cluster promotes esophageal carcinoma Z. Gao ∗ , R. Liu, L. Yin, Y. Pu Key Laboratory of Environmental Medicine Engineering, Ministry of Education, School of Public Health, Southeast University, China miRNA cluster is widely expressed across species, accumulating studies have illustrated that miRNA cluster functioned more efficiently than a single miRNA in some cancer oncogenesis. It is likely that miRNA cluster is more stable and reliable than individual miRNA to be biomarkers for diagnosis and therapy. However, the regulatory mechanism of miRNA cluster was largely unknown. In current study, we detected the expression of the mature miRNAs of miR-144-451 cluster from 102 patients pathologically diagnosed with esophageal carcinoma using stem-loop real-time reverse transcription PCR. The expression level of has-miR-451a, has-miR144-3p, has-miR-144-5p in tumor tissues were significantly lower compared with adjacent non-tumor tissues, the fold-change for these three miRNAs were 0.376, 0.463 and 0.443 respectively (P < 0.05). Principal component regression analysis indicated down regulation of has-miR-144-3p and has-miR-144-5p increased the risk for esophageal carcinoma, the OR were 0.850 and 0.840 respectively. Stable transgenic cells overexpressing miR-144-451 were obtained using lentivirus containing region of pri-miR-144-451. The overexpression of miR-144-451 cluster leads to the arrest of S and G2 phase, while the invasion ability of stable transgenic cells was seriously inhibited. However, no difference of apoptosis, migration, proliferation were found between stable transgenic and negative control cells. We further found c-Myc, p-ERK were downregulated in stable transgenic cells, while p53 was upregulated. Furthermore, the downstream effectors of c-Myc, MMP9 and pcdc2 were downregulated in stable transgenic cells. miR-144-451 may or partly inhibited cell cycles and invasion of EC9706 through inhibiting ERK/c-Myc signaling pathway. http://dx.doi.org/10.1016/j.toxlet.2016.06.1359
P02-002 Identification of microRNA as possible risk marker for drug-induced liver injury using chimeric PXB-Mouse® with highly humanized liver T. Kitahara 1,∗ , T. Shimada 2 , S. Nagatsuka 3 , H. Enomoto 4 , H. Tachiki 4 , H. Sasaki 1 1 Department of Hospital Pharmacy, Nagasaki University Hospital, Nagasaki, Japan 2 PhoenixBio Co., Ltd., Hiroshima, Japan 3 ADME & Tox. Research Institute, Sekisui Medical Co., Ltd., Ibaraki, Japan 4 Research and Development Division, Towa Pharmaceutical Co., Ltd., Kyoto, Japan
Purpose: Recently, microRNAs (miRNAs) have emerged as risk marker of drug-induced liver injury (DILI) in clinical. We developed a novel method to identify miRNAs as possible risk markers for DILI. We analyzed expression of miRNAs which fluctuated under the treatment of drugs with relatively higher rate of hepatotoxic
S78
Abstracts / Toxicology Letters 258S (2016) S62–S324
incidents (hTOX) and those with negligible hepatotoxic incidents (non-hTOX) using chimeric mice with highly humanized liver. Methods: We selected 8 hTOX (Amiodarone, Acetaminophen, Ticlopidine, Terbinafine, Methotrexate, Valproate, Diclofenac, Benzbromarone) drugs and 8 non-hTOX (Dipyridamole, Cefalexin, Eperisone, Propranolol, Pirenzepine, Trihexyphenidyl, Dapagliflozin, Bumetanide) drugs based on information shown in the package insert. The drugs were administered to chimeric mice with humanized liver (PXB-mice® : PhoenixBio Co., Ltd.) threetimes daily. Hepatic miRNAs were analyzed using GeneChip® miRNA 3.0 array (Affymetrix Inc.), in order to select miRNAs that increased by 1.5 times or more in hTOX drugs and not increased in the non-hTOX drugs. Results and conclusion: We found 7 miRNAs (miR-10a, miR183, miR-557, miR-1202, miR-3065-5p, miR-3195 and miR-4535) that increased only in hTOX treatments and not in non-hTOX treatments. Previous studies revealed the role of miR-10a, miR-183 and miR-1202 in non-hepatic tissues. This is the first report showing these 7 miRNAs might be involved in DILI. The present method is useful for discovery of new miRNAs as risk marker of DILI without clinical trials. http://dx.doi.org/10.1016/j.toxlet.2016.06.1360 P02-003 Endocrine disruptor compound’s levels in hair of Greek children M.N. Tzatzarakis 1 , V. Karzi 1 , E. Vakonaki 1 , I. Katsikantami 1 , M. Kiriakakis 1 , I. Tsakiris 2 , M. Kavvalakis 1 , A.K. Rizos 3 , A.M. Tsatsakis 1,∗ 1 Laboratory of Toxicology, Medical School, University of Crete, Heraklion, Crete, Greece 2 Department of Agricultural Technology, School of Agriculture Technology, Food Technology and Nutrition, TEI of Western Macedonia, Greece 3 Department of Chemistry, University of Crete and Foundation for Research and Technology-Hellas (FORTH-IESL), Heraklion, Crete, Greece
Purpose: Bisphenol A (BPA), triclosan (TCS) and perfluorooctanoic acid (PFOA) are endocrine disrupting chemicals that interfere with the body’s endocrine system. They are widely used in personal care products, toys, plastic containers and metal food cans. The aim of this research is to evaluate the exposure of children to these chemicals by analyzing head hair samples using a liquid chromatography–mass spectrometry based protocol. Methods: Head hair samples (1–55 cm) were collected from 72 children (2–11 years old), residents of Crete, Greece. A consent form was obtained from the children’s parents. The extraction of the analytes was performed by ultrasonic assisted methanolic extraction for 4 h at 50 ◦ C. Results: The method was linear (r2 > 0.99). The mean, inter day precision (%RSD), % accuracy and % recovery of the method for BPA were 14.9, 99.7 and 98.0, for PFOA were 16.3, 98.2 and 81.3, for TCS were 18.3, 94.1 and 103.3, respectively. The 71.2% of the samples were positive for BPA, 6.8% for PFOA and 89.0% for TCS. The median concentration of the positive samples was found to be 12.2 pg/mg (mean 20.4 pg/mg, range 2.6–205.5 pg/mg) for BPA, 21.0 pg/mg (mean 21.1 pg/mg, range 20.8–21.5 pg/mg) for PFOA and 48.8 pg/mg (mean 275.2 pg/mg, range 3.6–7656.6 pg/mg) for TCS. Conclusion: This is the first reported methodology concerning simultaneous detection of BPA, TCS and PFOA in hair. High frequencies of positive samples were found both for triclosan and BPA.
Moreover, a wide range of the detected levels, mostly for TCS, was observed. http://dx.doi.org/10.1016/j.toxlet.2016.06.1361 P02-004 Estrogen and androgen receptor transactivation by actual serum mixtures of lipophilic persistent organic pollutants extracted from Greenlandic pregnant women M. Ghisari ∗ , E.C. Bonefeld Jørgensen Centre for Arctic Health & Molecular Epidemiology, Department of Public Health, Aarhus University, Aarhus, Denmark Greenlandic Inuit have high blood levels of environmental persistent organic pollutants (POPs) as a result of high consumption of contaminated marine mammals. Since the human exposure is not limited to a single compound, the assessment of the combined hormone disrupting potential of the actual serum POP mixture is important. We aimed to analyze the potential interference of the actual serum lipophilic POP mixture with estrogen receptor (ER) and androgen receptor (AR) function. The lipophilic POPs were extracted by SPE-HPLC from 215 pregnant Greenlandic women serum samples collected 2013–2015. The extracts were analyzed for xenoestrogenic and xenoandrogenic activities using luciferase reporter gene assays. Using Spearman’s analyses the correlations between the POP-induced ER and AR transactivities and the actual serum levels of POPs [14 polychlorinated biphenyls (PCBs), one polybrominated biphenyl, four polybrominated diphenyl ethers and 11 organochlorine pesticides] were assessed. The preliminary results show that serum lipophilic POPs have the potential to transactivate the ER and AR, suggesting an overall estrogenic and androgenic activity. When the serum extracts were analyzed in the presence of the natural AR and ER ligands, dihydrotestosterone (DHT) and 17-estradiol (E2), most of the serum extracts antagonized the DHT-induced AR transactivity, suggesting an overall AR inhibiting potential of the serum lipophilic POP. A positive correlation between ER transactivities and the sum of the PCB concentrations in the blood was observed. In conclusion, our data shows that the actual serum mixture of lipophilic POPs in Greenlandic pregnant women have hormone disruptive potential. http://dx.doi.org/10.1016/j.toxlet.2016.06.1362 P02-005 Allele and genotype frequencies of CYP2B6 gene *5 polymorphism in a Turkish population S. Süzen ∗ , N.Y. Artun Ankara University, Faculty of Pharmacy, Department of Toxicology, Tandogan, Ankara, Turkey The cytochrome P450 (CYP450) proteins are monooxygenases, which catalyze many reactions involved in drug metabolism and synthesis of cholesterol, steroids and other lipids (1). CYP2B6 plays a major role in the metabolism of several therapeutically important drugs, including the anticancer agents, the antiretrovirals, the narcotics, the antidepressants, the antiestrogen tamoxifen; the synthetic opioid methadone and many more (2,3). The enzyme also metabolizes certain endogenous compounds such as testosterone, as well as recreational drugs, including nicotine and ecstasy/MDMA