High-affinity DNA targeting using readily accessible mimics of N2′-functionalized 2′-amino-α-L-LNA

Abstract

N2′-Pyrene-functionalized 2′-amino-α-L-LNAs (locked nucleic acids) display extraordinary affinity toward complementary DNA targets due to favorable preorganization of the pyrene moieties for hybridization- induced intercalation. Unfortunately, the synthesis of these monomers is challenging (∼20 steps, <3% overall yield), which has precluded full characterization of DNA-targeting applications based on these materials. Access to more readily accessible functional mimics would be highly desirable. Here we describe short synthetic routes to a series of O2′-intercalator- functionalized uridine and N2′-intercalator-functionalized 2′-N-methyl-2′-aminouridine monomers and demonstrate, via thermal denaturation, UV-vis absorption and fluorescence spectroscopy experiments, that several of them mimic the DNA-hybridization properties of N2′-pyrene- functionalized 2′-amino-α-L-LNAs. For example, oligodeoxyribonucleotides (ONs) modified with 2′-O-(coronen-1-yl) methyluridine monomer Z, 2′-O-(pyren-1-yl)methyluridine monomer Y, or 2′-N-(pyren-1-ylmethyl)-2′-N-methylaminouridine monomer Q display prominent increases in thermal affinity toward complementary DNA relative to reference strands (average ΔT m/mod up to +12 °C), pronounced DNA-selectivity, and higher target specificity than 2′-amino-?-L-LNA benchmark probes. In contrast, ONs modified with 2′-O-(2-napthyl)uridine monomer W, 2′-O-(pyren-1-yl)uridine monomer X or 2′-N-(pyren-1-ylcarbonyl)-2′-N-methylaminouridine monomer S display very low affinity toward DNA targets. This demonstrates that even conservative alterations in linker chemistry, linker length, and surface area of the appended intercalators have marked impact on DNA-hybridization characteristics. Straightforward access to high-affinity building blocks such as Q, Y, and Z is likely to accelerate their use in DNA-targeting applications within nucleic acid based diagnostics, therapeutics, and material science. © 2011 American Chemical Society.