![]() This showed that this cyclic phosphotriester modification destabilizes the AON/DNA and AON/RNA duplex by about -6 to -9 degrees C/modification. The effect of pure (S(p))-D(2)-CNA modification in the AONs was estimated by complexing to the complementary RNA and DNA strands by the thermal denaturation studies. ![]() Thus, after removal of the solid supports from the (S(p))-D(2)-CNA-modified AONs by BDU/MeCN, they were treated with hydrazine hydrate in pyridine/AcOH to give pure AONs in 35-40% yield, which was unequivocally characterized by MALDI-TOF to show that they have an intact six-membered dioxaphosphorinane ring. In contrast, the (S(p))-D(2)-CNA was about 2 times more stable than (R(p))-D(2)-CNA under hydrazine hydrate/pyridine/AcOH (pH = 5.6), which was exploited in the deprotection of pure (S(p))-D(2)-CNA-incorporated antisense oligodeoxynucleotides (AON). ![]() Thus, both (S(p))- and (R(p))-D(2)-CNA dimers (17a and 17b) were very labile toward nucleophile attack in concentrated aqueous ammonia to give carba-LNA-6',5'-phosphodiester (21) approximately 70-90%, carba-LNA-3',5'-phosphodiester (22) approximately 10%, and carba-LNA-6',3'-phosphodiester (23) <10%. The chemical reactivity of the six-membered dioxaphosphorinane ring in D(2)-CNA was found to be dependent on the internucleotidic phosphate stereochemistry. It turned out that at equilibrium concentration the (S(p))-D(2)-CNA isomer is preferred over the (R(p))-D(2)-CNA isomer by 0.39 kcal/mol. It has been found that F(-) ion can catalyze the isomerization of pure (S(p))-D(2)-CNA or (R(p))-D(2)-CNA to give an equilibrium mixture (K = 1.94). Structural studies by NMR as well as by ab initio calculations showed that in (S(p))- and (R(p))-D(2)-CNA the following occur: (i) the sugar is locked in extreme North-type conformation with P = 11 degrees and Phi(m) = 54 degrees (ii) the six-membered 1,3,2-dioxaphosphorinane ring adopts a half-chair conformation (iii) the fixed phosphate backbone delta, epsilon, and zeta torsions were found to be delta, epsilon (cis), zeta for (S(p))-D(2)-CNA, and delta, epsilon (cis), zeta for (R(p))-D(2)-CNA. Two diastereomerically pure carba-LNA dioxaphosphorinane nucleotides, simultaneously conformationally locked at the sugar and the phosphate backbone, have been designed and synthesized.
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