Phosphodiester Phosphonate method

Phosphorothioates. All three synthetic approaches appHcable to unmodified oligonucleotides can be adapted for synthesis of phosphorothioates (11) (33,46). If all of the phosphodiester linkages in an oligonucleotide are to be replaced with phosphorothioates, the ff-phosphonate method for coupling, followed by oxidation with Sg in carbon disulfide and triethylamine in the final step, is the most straightforward method. 

We reported earlier that the presence in a molecule of phosphodiester units does not interfere with the next 0-phosphoryIation step by means of the H-phosphonate method (i.e., P-protection for phosphodiesters is not required). Indeed, compound 40 (containing a phosphodiester at 0-6" ) was successfully phospholipidated (Scheme 6) by pivaloyl chloride assisted reaction with the acylalkylglyceiyl hydrogenphosphonate 7 followed by oxidation (in situ) with iodine to provide the fully protected oleic ester GPI [Sp = -1.4 and -1.2 (0.5 P each),... 

Scheme 19.2 Methods for polymer-supported oligonucleotide synthesis. Dinudeoside phosphate formation in the common methods for oligonucleotide synthesis phosphodiester (I), phosphotriester (II), phosphite triester (III) and H-phosphonate (IV).

The only exception was an attempt at the 5 -to-3 solid-phase oligonucleotide synthesis using nucleoside 3 -H-phosphonate monoesters, polymer-bound through a trityl-type linker, and oxidative coupling conditions (HgCl2) [123], The H-phosphonates were produced by phosphitylation of the polymer-bound N-unpro-tected nucleoside with phosphorus trichloride followed by hydrolysis by aqueous pyridine [61c]. However, despite some advanced features [124] the stepwise yields were comparable with those in the contemporary phosphodiester method-that is they were too low for the method to be truly competitive. 

Triesters of type 182 (n = 2-5) have been prepared from the nucleoside 3 -H-phosphonate, via the cyclic phosphites. Methods were also developed for making H-phosphonates and phosphodiesters involving just one of the hydroxy-groups of the diols. ... 

Workers at Isis Pharmaceuticals have developed methods for the synthesis of chimeric oligonucleotides containing blocks of phosphodiester, phos-phorothioate and phosphoramidate intemucleosidic links. H-Phosphonate... 

Dinucleoside phosphorothioates are chiral analogues of phosphodiesters in which one of the nonbridging oxygens has been replaced by sulfur. These compounds are important research tools in stereochemical investigations, in mechanistic studies of various enzymatic reactions, and other biochemical stndies [210-212]. There have been only a few methods for the preparation of ribonncleoside phosphorothioates [213-216]. The H-phosphonate approach is the best method for the preparation of phosphorothioate diesters [217-219]. 

Phosphorylation methods that are used in the synthesis of GPIs are adopted from the weU-estabhshed protocols for the synthesis of oligonucleotides. The two commonly used methods are both based on phosphitylation that is followed by oxidation. Phosphitylation of an alcohol with H-phosphonates activated via transient formation of mixed anhydride with pivaloyl chloride followed by oxidation with iodine in wet pyridine is commonly used to prepare the phosphate salts (Scheme 12.3, introduction of the phosphoHpid phosphodiester). The reagents... 

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