Synthesis of conformationally locked Carba-LNAs through intramolecular free-radical addition to C=N. Electrostatic and steric implication of the carba-LNA substituents in the modified oligos for nuclease and thermodynamic stabilities.
The syntheses of the hitherto unavailable parent fully unsubstituted carba-LNA and its C7'-amino and/or C6'-hydroxyl substituted derivatives, have been accomplished by the intramolecular 5-exo free-radical addition to a C4'-tethered C=N to give carba-LNAs with variable hydrophilic substituents at C6'/C7' (amino and/or hydroxyl). They have been introduced into isosequential antisense oligonucleotides (AONs) to examine how their relative electrostatic and steric effects in the minor groove of a putative AON-RNA duplex affect the target affinity, nuclease resistance, and RNase H elicitation. We show that 2'-oxygen in LNA is important in stabilizing the DNA/DNA and DNA/RNA duplexes vis-a-vis the unsubstituted carba-LNA and its other derivatives and that hydrophobic groups at C6'/C7' in both carba-LNA and carba-ENA relatively destabilize the AON/DNA duplex more profoundly than those in the AON/RNA duplexes. Two main factors affect the relative stabilities of AON/DNA versus AON/RNA duplexes: (i) hydration in the minor groove depending upon hydrophilicity vis-a-vis hydrophobicity of the substituents, and (ii) the relative size of the minor groove in the AON/DNA versus AON/RNA duplexes dictates the steric clash with the substituents depending upon their relative chiralities. We also show how the chirality and chemical nature of the C6'/C7' substituents affect the nuclease stability as well as the thermal stability and the RNase recruitment by AON/RNA duplexes.
Xu
Jianfeng
J
Department of Bioorganic Chemistry, Box 581, ICM, Biomedical Center, Uppsala University, SE-751 23 Uppsala, Sweden.
Liu
Yi
Y
Dupouy
Christelle
C
Chattopadhyaya
Jyoti
J
D002244:Carbon / Q000737:chemistry*; D002625:Chemistry, Organic / Q000379:methods*; D005609:Free Radicals; D008956:Models, Chemical; D008968:Molecular Conformation; D009584:Nitrogen / Q000737:chemistry*; D009696:Nucleic Acids / Q000737:chemistry*; D009841:Oligonucleotides / Q000737:chemistry; D016376:Oligonucleotides, Antisense / Q000737:chemistry; D010100:Oxygen / Q000737:chemistry; D016914:Ribonuclease H / Q000737:chemistry; D055672:Static Electricity; D013237:Stereoisomerism; D013816:Thermodynamics
D005609:Free Radicals; D009696:Nucleic Acids; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense; D002244:Carbon; D016914:Ribonuclease H; D009584:Nitrogen; D010100:Oxygen