Neurosteroid Analogues. 9. Conformationally Constrained Pregnanes: Structure-Activity Studies of 13,24-Cyclo-18,21-dinorcholane Analogues of the GABA Modulatory and Anesthetic Steroids (3α,5α)- and (3α,5β)-3-Hydroxypregnan-20-one

The hydrogen-bond-acceptor properties of the carbonyl moiety in the 17β-acetyl group on the D-ring of the anesthetic steroids (3α,5α)- and (3α5α)-3-hydroxypregan-20-one form an important part of the anesthetic steroid pharmacophore. 13,24-Cyclo-18,21-dinorcholanes containing a ketone or conjugated ketone group at C-20, C-22, C-23, or C-24 were prepared as conformationally constrained analogues of these anesthetic steroids and were used to probe for alternate locations for the D-ring hydrogen-bond-accepting carbonyl group. The analogues were evaluated (1) in [³⁵S]-tert-butylbicyclophosphorothionate binding experiments, (2) in electrophysiological experiments using rat α₁β₂γ_2L GABA_A receptors expressed in Xenopus laevis oocytes, and (3) as tadpole anesthetics. In the binding assay, the relative order of potencies for the analogues in the 5α- and 5β-series is identical. For the ketones, the order is 24-one ≥ 23-one > 20-one > 22-one. Likewise, for the enones, the order is Δ²²-24-one > Δ²⁰⁽²²⁾-23-one > Δ²²-20-one > Δ²³-22-one. Similar relative orders of potencies are also found in the other two bioassays. The activities of the 24-one and Δ²²-24-one compounds were expected to be very low, because the carbonyl group in these compounds is located over the steroid C-ring and oriented toward C-8. Instead, these compounds have the highest activities in their respective series, with the Δ ²²-24-one compounds having activities comparable to those of the reference anesthetic steroids. The electrophysiology results obtained with the 24-oxo-cyclosteroids suggest that rat α₁β ₂γ_2L GABA_A receptors contain more than one donor for the hydrogen-bond-acceptor group of anesthetic steroids. The family of cyclosteroids should be useful for future structure-activity relationship studies of steroid modulation of other GABA_A receptor subtypes.