Syntheses of 19-[ O-(carboxymethyl)oxime] haptens of epipregnanolone and pregnanolone

Abstract

O-(Carboxymethyl)oximes 1 and 2 derived from two epimeric 5β-pregnanolones (3β-hydroxy-5β-pregnan-20-one and 3α-hydroxy-5β-pregnan-20-one) in position 19 were prepared. Two synthetic routes were employed, both using protection of the 20-keto group after reduction into the (20R)-alcohol in the form of acetate. In the first route, (20R)-19-hydroxy-5β-pregnan-3β,20-diyl diacetate ( 3) was transformed into the corresponding 19-[ O-(carboxymethyl)oxime] methyl ester 6, then deacetylated by acid and partially silylated with tert-butyldimethylsilyl chloride. The desired 3- O-silylated derivative 8 was separated, oxidized to the 20-ketone and protecting groups were sequentially removed to give the first title hapten 1. The second route started from (20R)-19-hydroxy-3-oxopregn-4-en-20-yl acetate ( 11), which was hydrogenated in the presence of base to the 5β-pregnan-3-one derivative 12, protected in position 19 with tert-butyldimethylsilyl group and reduced with borohydride. The prevailing 3α-alcohol 15 was separated, protected in position 3 with a methoxymethyl group, deprotected in position 19 and transformed into the 19-[ O-(carboxymethyl)oxime] 19. After deacetylation, esterification with diazomethane and oxidation in position 20, the pregnanolone skeleton was regenerated. Final deprotection steps gave the second title hapten 2. Both haptens, i.e., (19E)-3β- and -3α-hydroxy-20-oxo-5β-pregnan-19-al 19-[ O-(carboxymethyl)oxime], were designed for the development of immunoassays of the corresponding parent neuroactive steroids.