Oligoribonucleotide Analogues Containing a Mixed Backbone of Phosphodiester and Formacetal Internucleoside Linkages, Together with Vicinal 2′‐O‐Methyl Groups

Abstract

Oligoribonucleotides containing formacetal internucleoside linkages have been prepared and studied by UV melting experiments. In RNA duplexes, the formacetal substitution is stabilizing (Deltat(m)=0 to +0.9 degrees C per modification) at physiological salt concentrations (0.1 M) but destabilizing (Deltat(m)=-0.4 to -0.8 degrees C per modification) at high salt concentrations (1 M); this suggests that reduction of electrostatic repulsion contributes substantially to the stabilization. The presence of 2'-O-Me substituents increases the stabilities of the duplexes (Deltat(m)=+0.5 to +1.1 degrees C per modification). The positive effects of formacetals and 2'-O-Me groups were independent and additive. (1)H NMR studies on monomeric model compounds containing 3'-(ethyl phosphate) or 3'-O-ethoxymethyl groups showed that the formacetal and 2'-O-Me substitutions shift the conformational equilibria of the ribose residues towards the North conformers by 5 to 12 %. Although the preference for the North conformers qualitatively correlates with increased duplex stabilities, changes in thermodynamic parameters (DeltaH degrees and TDeltaS degrees ) for formation of oligonucleotide duplexes and differences in dependence on concentrations of sodium acetate, sodium chloride and sodium perchlorate suggest that solvation effects are also important for the duplex stabilities. Overall the formacetal linkages fit well in A-type RNA duplexes, making them potentially interesting modifications for RNA-based gene-control strategies (e.g., antisense and RNA interference).

Interplay of Structure, Hydration and Thermal Stability in Formacetal Modified Oligonucleotides: RNA May Tolerate Nonionic Modifications Better than DNA

“…Matteucci found that formacetals in the DNA strand only slightly decreased the stability of DNA-RNA heteroduplexes 9. In contrast, our preliminary studies showed that formacetals were somewhat stabilizing in all RNA duplexes (+0.2 to +0.8 °C per modification) 4. However, each of these studies used different model sequences, which makes it difficult to compare and rationalize the results.…”

“…Several modifications, developed earlier to improve the properties of antisense oligonucleotides, have also shown promising results in RNAi 1,2. We are interested in developing RNA analogues having non-ionic internucleoside linkages 3,4. Among such, we have found that replacement of selected phosphates with amide linkages (CH 2 -CO-NH-5′) does not change the overall conformation and thermal stability of RNA 3.…”

Chemical Modification of Oligonucleotides for Therapeutic, Bioanalytical and other Applications

“…The formacetal modification on its own has a stabilising effect on RNA duplexes of 0 to +0.9 8C DT m /mod at physiological salt concentrations, but a destabilising effect (DT m /mod À0.4 to À0.8 8C) at high salt concentrations. The presence of the additional 2'-OMe modification combined with the form-A C H T U N G T R E N N U N G acetal modification had a stabilising effect of +0.5 to +1.9 8C DT m /mod compared to native RNA duplexes [5] (Scheme 2). The Sekine group used a 2'!5' squaryldiamide linkage to create the TpT dimer analogue U 2 -sq 5 -T (6), as replacement of the phosphodiester backbone of DNA.…”

“…Many moieties that are not chemically similar to phosphodiester, such as amide, guanidine and acetal have been used to replace the phosphodiester linkage. [1] More recently Strçmberg et al explored the combination of formacetal linkages and 2'-OMe modifications in oligoribonucleotides (5). The formacetal modification on its own has a stabilising effect on RNA duplexes of 0 to +0.9 8C DT m /mod at physiological salt concentrations, but a destabilising effect (DT m /mod À0.4 to À0.8 8C) at high salt concentrations.…”

Primitive Genetic Polymers

“…Phosphorus (V) esters and acetals are both tetrahedral and approximately isosteric, and it is known that aNAs can base pair with RNAs (Matteucci and Bischofberger 1991). Specifically, point substitutions of this linkage impart a modest destabilization to DNA-DNA and DNA-RNA duplexes and show saltdependent effects on RNA-RNA duplexes (stabilization at low salt, destabilization at high salt) (Jones et al 1993;Rice and Gao 1997;Rozners et al 2007;Kolarovic et al 2009). …”