Exploring site‐specific activation of bis‐N,N’‐dialkylaminophosphordiamidites and the synthesis of morpholinophosphoramidate oligonucleotides

Morpholinos are six‐membered rings that may provide higher conformational rigidity when incorporated into an oligonucleotide (ODN) backbone. Phosphorodiamidate morpholinos are chemically modified ODNs containing morpholinos in place of 2’‐deoxyribose moieties throughout their backbone and have garnered much interest in recent years due to their ability to function as highly effective steric blockers in exon skipping therapy. To further explore the biophysical and biological properties of ODNs derived from morpholino nucleosides, we have replaced the 2’‐deoxyribonucleotides of phosphodiester DNA with morpholinonucleotides to generate phosphoramidate ODNs. Here, we evaluate the mechanistic pathways observed during the solution‐phase synthesis of morpholinonucleoside phosphoramidites, solid‐phase synthesis of morpholinonucleotide phosphoramidates of mA, mG, mC and mT (prefix ‘m’ represents morpholino) and our first attempts directed at the solid‐phase synthesis of chimeric DNA‐phosphoramidate ODNs, as well as fully modified 22‐mer phosphoramidate ODNs.