L15 Endocrine disrupting compounds.pdf

6/14/2021 L15 Endocrine disrupting chemicals (EDCs) Timo Hamers Make dilution plan for two mixtures Test compound 1B Own Alternative combination combination 1 CuSO4 CuSO4 + TBT TBT + T...

6/14/2021 L15 Endocrine disrupting chemicals (EDCs) Timo Hamers Make dilution plan for two mixtures Test compound 1B Own Alternative combination combination 1 CuSO4 CuSO4 + TBT TBT + TCS 2 TBT TBT + TCS CuSO4 + TBT 3 TCS TCS + CuSO4 TBT + TCS 1  6/14/2021 Set-up  Why Endocrine Disrupting Chemicals [EDCs]?  Endocrine system and hormones  Sex hormones  Definition and examples of EDCs  Mode of action of EDCs  Examples of EDC effects in humans and wildlife  Bioassay: ER-CALUX to measure (anti-)estrogenic EDCs 3 4 2  6/14/2021 Why EDCs? Theo Colborn (1927-2014) Our Stolen Future (1996) Are We Threatening Our Fertility, Intelligence, and Survival? A Scientific Detective Story Popular press publication With a foreword from vice-president Al Gore Development of the “endocrine disruption” hypothesis 5 5 WHO reports 2002 and 2012 The effects are endocrine system This work related and not concluded that necessarily species scientific knowledge dependent. Effects at that time provided shown in wildlife or evidence that certain experimental animals effects observed in wildlife may also occur in humans can be attributed to chemicals if they are exposed to EDCs at a that function as endocrine vulnerable time and at concentrations disrupting chemicals (EDCs); that the leading to alterations of endocrine evidence of a causal link was weak in most regulation. Of special concern are effects cases and that most effects had been observed on early development of both humans and in areas where chemical contamination was wildlife, as these effects are often irreversible high; and that experimental data supported this and may not become evident until later in life. conclusion. The document further concluded that there was only weak evidence for endocrine-related effects in humans. https://www.who.int/ipcs/publications/new_issues/endocrine_disruptors/en/ http://www.who.int/ceh/publications/endocrine/en/ 6 3  6/14/2021 Endocrine system system of glands producing and transporting chemical signals (hormones) throughout the body Biological response is taking place in the target cell (receptor) 8 Endocrine system adipose tissue! leptin adiponectin estrogen 9 9 4  6/14/2021 Hormones Signal compounds Essential (vital) functions Development Growth Reproduction Behavior Metabolism 10 Chemical classification of hormones  Peptide hormones Synthesis via gene transcription, e.g. thyroid hormone, insulin  Steroid hormones Synthesis from cholesterol, e.g. > sex hormones like androgens, estrogens and progestagens > corticoids involved in glucose and mineral regulation 11 5  6/14/2021 Thyroid hormone synthesis By Mikael Häggström, used with permission. [CC0], from Wikimedia Commons 12 CYPs and biosynthesis of steroid hormones P450 enzymes  important for biotransformation, e.g. CYP1A1: planar aromatic compounds CYP 11B1/2, CYP17, CYP19, CYP21A2: steroidogenesis  Tissue specific 13 6  6/14/2021 Biological classification of hormones - Role  “Releasing hormones” Hypothalamus → Pituitary  “Stimulating (or tropic) hormones“ Pituitary → endocrine tissue (gland)  “Non-tropic hormones” Endocrine tissue → target cell 14 Biological classification of hormones: receptors Membrane receptors Nuclear receptors Non-steroid hormones Steroid and thyroid hormones Extracellular Intracellular Hormone-receptor binding  Hormone-receptor complex  activates second messenger system activates cell itself (signaling cascade) in target cell 15 7  6/14/2021 Characteristics of sex hormones  Not stored, but immediately released  Steroid hormones, derived from cholesterol  Lipophilic: diffuse through membranes  Transport through blood: bound to proteins like “Steroid-binding Globulins” (SBGs)  Release: controlled by negative feedback E.g. hypothalamus-pituitary-gonad (HPG) axis 16 Negative feedback HPG axis: hypothalamus- pituitary-gonad axis  GnRH: Gonadotropin- releasing hormone  Gonadotropic hormones FSH: follicle stimulating hormone LH; luteinizing hormone  Sex hormones E2: estradiol T: testosterone DHT: dihydrotestosterone 17 8  6/14/2021 Sex hormones estrogens and progestogens  Produced mainly in the ovary (testis) and adrenal gland  Ovulation, menstruation cycle  Embryo implantation and maturation  Birth and lactation  Secondary sex characteristics Estradiol 18 Sex hormone testosterone  Produced in the testis (ovary) and adrenal gland  In embryonic development: causes differentiation into man (“default” is woman)  Sperm production  Libido  Secondary sex characteristics 19 9  6/14/2021 Mechanism of action steroid hormones  Intracellular  Nuclear receptors  Hormone-receptor complex  activates cell itself 20 Set-up  Why Endocrine Disrupting Chemicals [EDCs]?  Endocrine system and hormones  Sex hormones  Definition and examples of EDCs  Mode of action of EDCs  Examples of EDC effects in humans and wildlife  Bioassay: ER-CALUX to measure (anti-)estrogenic EDCs 21 10  6/14/2021 Endocrine disrupting chemicals – Definition (WHO, 2002)  “An endocrine disruptor is an exogenous substance or mixture that alters function(s) of the endocrine system and consequently causes adverse health effects in an intact organism, or its progeny, or (sub) populations.”  “A potential endocrine disruptor is an exogenous substance or mixture that possesses properties that might be expected to lead to endocrine disruption in an intact organism, or its progeny, or (sub)populations.” 22 Endocrine disrupting chemicals 23 11  6/14/2021 EDC examples  Pharmaceuticals and Personal Care Products (PCPP)  Not in the picture: Handsoaps Toothpaste 24 Most focus on EATS modalities Compounds acting through mediation of  Estrogen  Androgen  Thyroid  Steroidogenic systems 25 12  6/14/2021 EDCs Over 400 chemicals on EU priority list 26 Mode of action EDCs Agonism: binding and activation of hormone receptors (“hormone mimic”) Antagonism: binding and inactivation of hormone receptors (“hormone blocking”) 27 13  6/14/2021 Promiscuous character of the estrogen receptor 28 Mode of action EDCs Indirect by disrupting the normal hormonal balance Disrupting the synthesis, release, metabolism and/or excretion of hormones Disrupting hormone binding to transporter proteins Changing the sensitivity of the target cell > Up or down regulation of gene expression > Epigenetic regulation of promotor regions 29 14  6/14/2021 Possible sites of action of environmental contaminants on HPT axis Gilbert et al. 2011 30 Thyroid hormone disruption Woodruff et al. 2008  Associated effects of increased/decreased thyroid hormone activity: Developmental neurotoxicity Cardiovascular disorders Metabolic disorders Carcinogenicity 31 15  6/14/2021 Set-up  Why Endocrine Disrupting Chemicals [EDCs]?  Endocrine system and hormones  Sex hormones  Definition and examples of EDCs  Mode of action of EDCs  Examples of EDC effects in humans and wildlife  Bioassay: ER-CALUX to measure (anti-)estrogenic EDCs 32 Examples of EDC effects in wildlife  Adverse effects on reproduction due to egg shell thinning and feminization of males in birds by DDT and other pesticides (North America, Europe)  Impairment of reproduction and immune function in seals by PCB/DDT metabolites (e.g. DDE), dioxins (Baltic Sea, Wadden Sea)  Distorted sex organ development in alligators linked to a DDT spill (Florida)  Feminization of male fish populations by natural estrogens and xeno-estrogens (world- wide)  Masculinization of female marine snails by TBT from antifouling paints (world-wide) Vos et al. 2000 34 16  6/14/2021 Intersex in Roach oc sd po t po t See lecture L11 on Effect-based Monitoring 35 Sexual disruption (intersex) in wild fish populations (UK) See lecture L11 on Effect-based Monitoring A BC D E F GH I J F G H I J KLM Jobling et al. 1998 36 17  6/14/2021 Relationship between VTG biomarker and intersex See lecture L11 on Effect-based Monitoring Jobling et al. 1998 37 Signal transduction pathway for steroid hormones OH Steroids, e.g. estrogens 17ß-estradiol CELL OH HO NUCLEUS 17ß-ethynyl-estradiol ERE gene Binding to HO Receptors mRNA bisphenol-A HO OH 4-nonylphenol OH Protein O O di(2-ethylhexyl)phthalate O O Feminizing effects See lecture L11 on Effect-based Monitoring 38 18  6/14/2021 CYPs and biosynthesis of steroid hormones P450 enzymes  important for biotransformation, e.g. CYP1A1: planar aromatic compounds CYP 11B1/2, CYP17, CYP19, CYP21A2: steroidogenesis  CYP19 (aromatase): androgen estrogen  Tissue specific 39 Hypospadias in USA Effects of EDCs in humans: Epidemiological studies Breast cancer Testicular cancer http://www.unep.org/pdf/WHO_HSE_PHE_IHE_2013.1_eng.pdf 40 19  6/14/2021 Effects of EDCs in humans: epidemiological studies Sperm density Fertility 41 Adapted from State of the Science of Endocrine Disrupting Chemicals - 2012 41 Effects of EDCs in humans  Effects depend on dose, age, sex, and timing of exposure  In utero exposure vs exposure at later age 42 20  6/14/2021 Development is sensitive 43 EDCs and human health: endocrine diseases associated with exposure to EDCs  Evidence for a role for developmental exposure to EDCs and these diseases has been shown in both animal and epidemiological studies 44 21  6/14/2021 Diethylstilbestrol (DES): endocrine disruption in humans  Synthetic estrogen  “wonder drug”  Prevention of miscarriages and complications during pregnancy  About 2-8 million pregnancies worldwide between 1946 and 1977 45 DES: endocrine disruption in humans  DES daughters: Morphological malformation of gonads Abnormal pregnancies Disturbed menstrual cycle Increased incidence of carcinoma in cervix or vagina  DES sons: Cryptorchidism and hypospadias Decreased sperm quality 46 22  6/14/2021 Set-up  Why Endocrine Disrupting Chemicals [EDCs]?  Endocrine system and hormones  Sex hormones  Definition and examples of EDCs  Mode of action of EDCs  Examples of EDC effects in humans and wildlife  Bioassay: ER-CALUX to measure (anti-)estrogenic EDCs 47 Bioassays to measure (anti-)estrogenic activity  ER-LUC reporter gene bioassay CELL (xeno-)estrogen NUCLEUS ERE gene Estrogen Receptors mRNA pERE -tata-Luc Luciferase Protein In stably transfected T47D human breast cancer cell line (Legler et al, 1999) 48 23  6/14/2021 Estrogenic potency 49 Relative Potency (REP) = EC50E2/EC50X Relative potency (REP) 1.0 0.74 0.00005 0.00003 0.0000036 0.0000011 0.0000004 0.0062 0.01 0.00005 0.000004 50 24  6/14/2021 Intersex in roach oc sd po t po t 51 Monitoring of estrogenic compounds in NL Results from ER-LUC assay LOES project water bile sediment Legler et al, 2002 52 25  6/14/2021 Monitoring of estrogenic compounds in humans Results from ER-LUC assay Using fractionation techniques to separate endogenous hormones from xeno-estrogens 53 Goals: After this lecture, you are able to  Explain what hormones are, what they do, where they are formed, and what their mechanism of action is  Classify hormones Chemically Biologically based on role and receptor type  Explain what endocrine disrupting chemicals (EDCs) are  Explain the most important modes of action of EDCs Receptor agonism/antagonism Affecting the hormonal balance  Name examples of endocrine disruption in human and wildlife  Explain the principle of the ER-LUC bioassay and how to use it for environmental monitoring 54 26