Hydrophobic alkyl-linked base-modified pyrimidine and 7-deazapurine 2′-deoxyribonucleoside phosphoramidites: synthesis and application in solid-phase synthesis of modified and hypermodified oligonucleotides

Abstract

A series of 2′-deoxyribonucleoside 3′-phosphoramidites bearing hydrophobic alkyl-linked modifications at nucleobases was synthesized, namely 5-phenylethyluracil, 5-pentylcytosine, 7-(indol-3-yl)ethyl-7-deazaadenine, and 7-isopentyl-7-deazaguanine derivatives. These nucleoside phosphoramidites were used for solid-phase synthesis of modified and hypermodified oligonucleotides containing up to fifteen modified nucleotides in a row. Their hybridization with complementary non-modified or modified oligonucleotides and thermal stability of the resulting DNA duplexes was studied using UV-vis denaturing experiments and CD spectroscopy. The results indicate that the partially modified hydrophobic DNA can still retain B-conformation, although with lower thermal stability. On the other hand, the hypermodified ONs containing all four modified nucleotides did not hybridize to duplexes likely due to formation of aggregates as indicated by dynamic light scattering measurement. This work expands the toolkit of chemically modified nucleotides for applications in functional nucleic acids or nucleic acid therapeutics, but also shows the scope and limitations of the use of hydrophobic nucleotides in hypermodified oligonucleotides and DNA.