Synthetic progestogens also known as synthetic gestagens

Synthetic progestogens (also known as ‘synthetic gestagens’, ‘synthetic progestins’ or simply ‘progestins’) are designed to mimic the effects of progesterone (P4) and are the active pharmaceutical ingredient in many contraceptives and hormone replacement therapies. Since most progestins are derived from steroid hormones, they can be classified according to the structure of the parent compound, and include P4 derivatives (dydrogestrone, medrogestone), pregnane derivatives (chlormadinone acetate, cyproterone acetate, medroxyprogesterone acetate), norpregnane derivatives (progmegestone, nomegestrol acetate), the spironolactone derivative, drospirenone (DPN), and 19-nortestosterone derivatives, which are subdivided into estranes (19-norethindrone [19-NE], lynestrenol, tibolone), ethinylated gonanes (levonorgestrel [LNG], norgestimate [NGM], etonogestrel [ETG], gestodene [GSD]) and non-ethinylated gonanes (dienogest). Like other pharmaceuticals with endocrine receptor-mediated modes of action, progestins are considered as aquatic contaminants of increasing concern (, ) and occur widely in treated sewage and effluent-receiving waters (, , , ). Off-target receptor modulation is relatively common for synthetic progestogens. Most progestins structurally related to progesterone, pregnane or norpregnane are antagonists of the human androgen receptor (hAR), similarly to P4 itself (), while 19-nortestosterone-derived progestins are generally androgenic (). In the present study, we focused on 19-nortestosterone-derived compounds since progestins of this class can elicit sub-acute effects in fish at environmentally relevant concentrations. In developing zebrafish embryos, changes in the Protease Inhibitor Cocktail (EDTA-Free, 200X in DMSO) sale of selected nuclear hormone receptors occur in response to 2ng/L 19-NE or LNG (). A significant reduction in fecundity (eggs produced per pair) in fathead minnow breeding pairs was observed in response to LNG at concentrations as low as 0.8ng/L, with 100ng/L nominal LNG causing the development of male secondary sex characteristics in females after 21 days (). Exposure of fish to LNG, 19-NE and GSD at or above 25ng/L caused a significant reduction in fecundity in medaka breeding pairs and induced male coloration in female fathead minnow (). Egg production in fathead minnow was significantly reduced in response to 1ng/L or 100ng/L GSD, although a clear concentration–response relationship was not observed (). Perhaps the most compelling evidence for strong androgenic activity comes from two studies conducted using the three-spined stickleback (), currently the leading indicator species for studying (anti) androgenic effects. In female stickleback, LNG exposure at or above 40ng/L induced the expression of spiggin in kidney tissue and repressed vitellogenin transcript levels in liver (). A follow-up study showed that exposure of male sticklebacks to ≥65ng/L LNG resulted in the prolonged maintenance of breeding condition, which the authors proposed could have adverse population-level effects in the wild due to increased energy expenditure and susceptibility to predation (). In vitro reporter gene transactivation assays driven by the fathead minnow androgen receptor (fmAR) revealed potent activation by 19-nortestosterone-derived progestins (). The same study showed that DPN was an agonist of fathead minnow progestogen receptor (fmPR), albeit weaker than the endogenous ligand 17α,20β-dihydroxy-4-pregnen-3-one (DHP), but that none of the 19-nortestosterone-derived progestins tested activate fmPR. This suggests that unlike the human (hPR), which is more potently activated by most synthetic progestins than P4 itself, 19-nortestosterone-derived progestins do not act as strong agonists of fish PR. While cyprinids such as fathead minnow express a single AR, some percomorphes are known to express two AR subtypes (, ) that respond differently to environmentally relevant (anti) androgens (), raising the question of whether 19-nortestosterone-derived progestins preferentially activate either AR subtype.