Chemical synthesis and biological characterization of phosphorothioate analogs of 2’, 5’-3’-deoxyadenylate trimer

In continued studies to elucidate the requirements for binding to and activation of the 2’,5’-oligoadenylate (2-5A) dependent endorlbonuclease (RNase L), four 2-5A trimer analogs were examined to evaluate the effect of chlrality of phosphorothioate substitution on biological activity. The chemical syntheses and purification of the four Isomers of P-thlo-3’-deoxy-adenylyl-(2′-5′)-P-thlo-3′-deoxyadenylyl-(2′-5′)-3′-deoxyadenosine, by the phosphoramldlte approach, is described. The isolated intermediates were characterized by elemental and spectral analyses. The fully deblocked compounds were characterized by 1H and 31P NMR and HPLC analyses. The 2′,5′-(3′dA)3 cores with either Rp or Sp chlrality in the 2′,5′-internucleotlde linkages will bind to but will not activate RNase L. This is in contrast to 2′,5′-A3 core analogs with either RpRp or SpRp phosphorothioate substitution in the 2′,5′-internucleotide linkages which can bind to and activate RNase L. There are also marked differences In the ability of the 2′,5′-A3 analogs to activate RNase L following introduction of the 5′-monophosphate. For example, the 5′mono-phosphates of 2′,5′-(3′dA)3-RpRp and 2′,5′-(3′dA)3-SpRp can bind to and activate RNase L, whereas the 5’-monophosphates of 2′,5′-(3′dA)3-RpSp and 2′,5′-(3′dA)3rSpSp can bind to but can not activate RNase L.