Hormonal properties of norethisterone, 7alpha-methyl-norethisterone and their derivatives.

Norethisterone (NET) is a progestagenic compound with very weak androgenicity and estrogenicity. These low androgenic and estrogenic activities may be attributed to NET itself or induced by metabolites of NET. In order to improve the bioactivity of NET, the effects of a 7alpha-methyl substitution were studied. Thus this study has two objectives: first the comparison between biological activities of NET and 7alpha-methyl-NET (MeNET), and second the biological activity of tentative metabolites of NET and those of MeNET. The metabolites consist of a 3-keto-, 3alpha- or 3beta-hydroxy-group located next to a carbon 4 to 5 double bond (Delta(4)) or a 5alpha-hydrogen atom. The 7alpha-methyl substitution was of special interest as it prevents 5alpha-reduction. The biological activities of NET, MeNET and their potential metabolites were assessed by in vitro binding, transactivation and proliferation assays on progesterone (PR), androgen (AR), estrogen (ER) and glucocorticoid (GR) receptors and by in vivo progestagenic McPhail, androgenic Hershberger, estrogenic Allen-Doisy tests and combined estrogenic and progestagenic ovulation inhibition tests. NET is a compound with five- to eight-fold weaker PR binding and transactivation activities than the reference compound Org 2058 (100%) and two-fold stronger than progesterone. Binding and transactivation activities of NET for AR (DHT=100%) are 3.2 and 1.1%, respectively, for ER none (E2=100%) and for GR below 1% (DEX=100%). MeNET is 1.5- to two-fold less progestagenic and ten- to 20-fold more androgenic than NET, while it does not show activity for ER and GR. The relative binding affinity of 5alpha-NET was seven-fold lower for PR and 1.5-fold higher for AR than for NET, while in transactivation assays 5alpha-NET was only active at levels below 1% for all tested receptors. 3beta-Hydroxy-(5alpha-reduced)-metabolites showed clear ER binding and transactivation activities, while 3alpha-hydroxy-(5alpha-reduced)-metabolites did hardly possess these characteristics. These hydroxy metabolites did not bind or activate other receptors. Substitution of 7alpha-methyl to NET metabolites led to similar characteristics, but with higher activities for AR and ER and weaker activity for PR. The outcome of in vivo tests showed a remarkable effect for MeNET. Progestagenic activity in rabbits appeared for NET equipotent to or eight-fold higher than for MeNET, after subcutaneous or oral treatment, respectively. On the other hand, MeNET showed in rats a ten-fold higher androgenicity and eight-fold higher estrogenicity than NET. Ovulation inhibition was induced at very low oral or subcutaneous dose levels, being 120- or ten-fold lower than for NET, respectively. The estrogenicity can also be induced by 3alpha- or 3beta-hydroxy metabolites of MeNET, which are 15 or even more than 40-fold stronger than those of NET, respectively. In conclusion, after the introduction of a 7alpha-methyl substituent to NET an increased estrogenicity and androgenicity and a reduced progestagenic activity was found. The in vivo estrogenicity is mainly due to 3beta-hydroxy-MeNET and to a lesser extent to 3alpha-hydroxy-MeNET, while the androgenicity and progestagenicity are most likely caused by MeNET itself. Since the 7alpha-methyl substituent inhibits 5alpha-reductase, 5alpha-reduced MeNET metabolites can be excluded from biological activities. As MeNET is a very effective ovulation inhibitor, due to its mixed progestagenic and estrogenic profile, a further reduction of androgenicity of MeNET may yield new contraceptives with an attractive profile for contraception.