Synthesis of O(6)-alkylated preQ(1) derivatives

A naturally occurring riboswitch can utilize 7-aminomethyl-O(6)-methyl-7-deazaguanine (m(6)preQ(1)) as cofactor for methyl group transfer resulting in cytosine methylation. This recently discovered riboswitch-ribozyme activity opens new avenues for the development of RNA labeling tools based on tailored O(6)-alkylated preQ(1) derivatives. Here, we report a robust synthesis for this class of pyrrolo[2,3-d]pyrimidines starting from readily accessible N(2)-pivaloyl-protected 6-chloro-7-cyano-7-deazaguanine. Substitution of the 6-chloro atom with the alcoholate of interest proceeds straightforward. The transformation of the 7-cyano substituent into the required aminomethyl group turned out to be challenging and was solved by a hydration reaction sequence on a well-soluble dimethoxytritylated precursor via in situ oxime formation. The synthetic path now provides a solid foundation to access O(6)-alkylated 7-aminomethyl-7-deazaguanines for the development of RNA labeling tools based on the preQ(1) class-I riboswitch scaffold.