Which information identifies a chemical as endocrine disrupting? Poul Bjerregaard Institute of Biology University of Southern Denmark Odense and Danish Centre on Endocrine Disrupters UNEP EDC Advisory Group meeting, Geneva , September 25-26, 2015

WHO EDC definition Endocrine mechanism Adversity Plausible link between the two

Adversity according to WHO/IPCS 2004 A change in morphology, physiology, growth, reproduction, development or lifespan of an organism which results in impairment of functional capacity or impairment of capacity to compensate for additional stress or increased susceptibility to the harmful effects of other environmental influences

Adversity? Risk assessment of chemicals What do we want to protect? Human risk assessment: Every individual Environmental risk assessment: The structure and function of the ecosystem

Environmental risk assessment Protection of ecosystem structure and function The aim is to protect 95% of the species ‘Adversity ’defined from population effects Not effects on the individual

Environmental risk assessment Protection of ecosystem structure and function The aim is to protect 95% of the species ‘Adversity’ defined from population effects Not effects on the individual

Relevant OECD test guidelines Human risk assessment TG 416 or 433 (mammals) E.g. malformed male genitals, altered AGD Environmental risk assessment TG 234 Fish Sexual Development Test

Fish Sexual Development Test Altered content of yolk proteins Controlled by oestrogen Indicates endocrine mechanism - not adversity Altered sex ratio Indicates endocrine mechanism and adversity OECD Guidance Document 150

Danish suggestion for EDC-criteria Category 1 – Endocrine Disrupter Adverse in vivo effects ED mode of action in vivo clearly linked to adverse in vivo effects Category 2a – Suspected Endocrine Disrupter Adverse effects in vivo where an ED MoA is suspected ED MoA in vitro combined with toxicokinetic in vivo data ED MoA in vivo suspected to be linked to adverse effects in vivo Category 2b – Indicated Endocrine Disrupter in vitro/in silico evidence indicating potential for ED in vivo

Exemplified by chemical UV filters Category 2a Category 1 Danish 2012-assessment BP-3: Indications of oestrogenic and anti-androgenic effects. Not consistent Category 2a Category 2a TG234 needed

Sexual development in zebrafish ♀ ♂ Hatch 20 days 40 days 60 days

Sex ratio altered – more ♀ Kinnberg et al. 2015. Environ.Toxicol.Chem. In press.

No consistent effect on yolk proteins 12 d exposure

Conclusion Exposure to benzophenone 3 skews sex ratio so Benzophenone 3 is an EDC

Is BP-3 oestrogenic or anti-androgenic? Does it matter ? Depends on the requirement for detailed knowledge about the mechanism of action Detailed knowledge about ‘Adverse Outcome Pathways’ is desirable But not always obtainable

A chemical may have multiple AOPs Prochloraz Fungicide

Prochloraz: More male zebrafish * 100 Female Male Intersex Undifferentiated 80 60 % 40 20 73 72 72 69 Control 16 m g/l 64 m g/l 202 m g/l Prochloraz Prochloraz Prochloraz Kinnberg et al. 2007. Comp. Biochem. Physiol.145C, 165-170

Prochloraz is anti-androgenic in rats Laier et al. 2006. Toxicol.Appl.Pharmacol. 213, 160-171

Effect of prochloraz Cholesterol Prochloraz Anti-androgenic in rats Testosterone Prochloraz Inhibits aromatase in zebrafish (?) Oestrogen

Conclusion Detailed knowledge on ‘Adverse Outcome Pathways’ is not necessarily needed to reach conclusions on endocrine disrupting activity