Synthesis of Cyclohexyl Carbocyclic Puromycin and Its Inhibition of Protein Synthesis

In a continuation of our studies with puromycin analogues, four cyclohexyl carbocyclic puromycins, 6-dimethylamino-9-[(R)-[2(R)-hydroxy-3(R)-(p-methoxyphenyl-L-alanylamino)]cyclohexyl]purine (1a), 6-dimethylamino-9-[(S)-[2(S)-hydroxy-3(S)-(p-methoxyphenyl-L-alanylamino)]cyclohexyl]purine (1b), 6-dimethylamino-9-[(R)-[2-CR)-hydroxy-3(S)-(p-methoxyphenyl-L-alanylamino)]cyclohexyl]purine (2a), and 6-dimethylamino-9-[(S)-[2(S)-hydroxy-3(R)-(p-methoxyphenyl-L-alanylamino)]cyclohexyl]purine (2b), have been synthesized. trans-3-Amino-2-hydroxycyclohexanone ethylene ketal (4) was easily obtained by opening of epoxide 3 with liquid ammonia. Condensation of 4 with 5-amino-4,6-dichloropyrimidine and subsequent ring closure gave the 9-substituted 6-chloropurine 5 which was converted to the O-acetyloxime 8. Reduction of 8 with diborane gave a mixture of cisand trans-amino alcohols separated as their acetamides, 9 (7%) and 10 (33%), respectively. The amino alcohols were converted to 1a,b and 2a,b by a general method previously reported from our laboratory. In addition, the trans-cyclopentyl carbocyclic puromycins, 6-dimethylamino-9-[(R)-[2(R)-hydroxy-3(S)-(p-methoxyphenyl-L-alanylamino)] cyclopentyl]purine and 6-dimethylamino-9-[(S)-[2(S)-hydroxy-3(R)-(p-methoxyphenyl-L-alanylamino)] cyclopentyl]purine (19a and 19b), were prepared from the previously described 2a-acetamido-5a-(6-dimethylamino-9-purinyl)cyclopentan-1μ-ol. The puromycin analogues were evaluated as inhibitors of protein synthesis by their ability to inhibit the rate of poly(U,C)-directed L-[¹⁴C] poly (phenylalanine) formation in an Escherichia coli cell-free ribosome system. The results of this study, in conjunction with our previous reports, suggest that maximum activity of puromycin analogues is obtained when the purine moiety, the amino acid, and the hydroxyl group are oriented about a five-membered ring. In addition, the amino acid and hydroxyl group must be in a cis orientation, and the absolute stereochemistry of the parent antibiotic must be conserved.