Abstract

The use of oligonucleotides as therapeutic and diagnostic tools has grown significantly in recent decades. However, native oligonucleotides face numerous challenges such as poor cellular uptake and low resistance to nucleases hence necessitating the incorporation of modifications to their scaffold. Small circular oligonucleotides have shown promise in these fields; however, their formation is limited by their length and the proximity of the ligating ends. To address this challenge, we investigated a methodology to synthesize circular constructs containing a chemically stable butylene or base labile sulfonylethane interstrand cross-link to pre-organize the system to aid in the circularization via CNBr-assisted ligation or click chemistry. Moreover, nucleic acid crystallography has proven to be an asset in determining nucleic acid structures and complexes. Incorporation of a selenium atom to the scaffold of DNA has helped with phasing issues. Thus, we investigated a novel approach to introduce a 5’-methylphosphoroselenoate handle by the use of a methylphosphoramidite followed by treatment with potassium selenocyanate. Lastly, the O6-position of guanine bases can be methylated by various exo- and endogenous agents. This adduct can be repaired by the protein O6-Alkylguanine DNA Alkyltransferase (AGT). The activity of AGT can reduce the efficiency of some alkylating chemotherapeutic drugs. Thus, the discovery and evaluation of AGT inhibitors is desirable. Hence, we explored development of a method to evaluate the repair proficiency of AGT in the presence of inhibitors towards DNA duplexes which contain an O6-methyl-2’-deoxyguanosine adduct.