Efficient synthesis of novel 16-spiroisoxazolines in the androst-5-ene series was carried out by 1,3-dipolar cycloadditions of different aryl nitrile oxides to 3β-acetoxy-16-methylene-androst-5-en-17-one. During the intermolecular ring closures, the attack of the O terminus of the nitrile oxide dipole from the α side on C-16 predominated for steric reasons permitting the reactions to occur in a regio- and stereoselective manner. The minor isomers in which the angular methyl group on C-13 and the O atom of the isoxazoline heteroring were in the β, β-cis orientation were obtained in a yield of only ~10 %. Moreover, the conversions were influenced to a certain extent by the substituents on the aromatic moiety of the 1,3-dipoles. The stereostructures of the related diastereomers were confirmed by 2D NMR methods. Deacetylation of the primarily formed main products resulted in the corresponding 3β-OH analogs, which were further reduced to furnish 3β, 17β-diols. All of the synthetized compounds were subjected to in vitro pharmacological studies in order to investigate their antiproliferative effects on three malignant human adherent cell lines (HeLa, MCF7, and A431).