Figure 2.

Modifications of anti-sense nucleotides. Anti-sense nucleotides are delineated into two major types, anti-sense oligonucleotides (ASOs), which maintain a ribose/deoxyribose backbone, and Morpholinos, which replace the sugar backbone with a morpholine group. ASOs have been modified in several ways to increase their resistance to endogenous nucleases, either by changing the phosphodiester bond or adding molecules onto the sugar backbone itself. Initial modifications altered the phosphodiester bond to a phosphorothioate bond by replacing an O⁻ with an S⁻, resulting in robust nuclease resistance. However, a disadvantage of this modification is the reduced affinity for target mRNA, which has been addressed by combining phosophothioate with modifications to the 2′ C that replace the H with an O-alkyl group, either an O-methyl or O-methyoxy-ethyl (MOE) or an LNA configuration. These modifications further enhance nuclease resistance and increase the affinity of the ASO for mRNA. However, all of these modifications require the addition of oxygen (colored in green) to the 2′ C in the deoxyribose base, thus changing the sugar from deoxyribose into ribose and preventing the recruitment of RNase H to degrade transcripts. Further, modifications designed to recruit RNase H have been achieved by forming “gapmers” that consist of RNA with “gaps” filled by DNA, allowing for successful recruitment of RNase H. Morpholinos reduce nuclease degradation by completely replacing the sugar backbone with a morpholine group. Morpholinos are unable to recruit RNase H, and importantly, they cannot cross the cell membrane without the addition of an octaguanidine dendrimer (vivo-Morpholino). The traits of each modification are summed up in the bottom of the panel.