Protection 2-cyanoethyl esters

Protection of carboxyl groups as 2-cyanoethyl esters OCH2CH2CN - OH... 

The dimethoxytrityl ester protecting group is now removed by treatment with mild acid (CCI3CO2H), which is insufficiently reactive to hydrolyse the amide protection of bases, or the cyanoethyl protection of the phosphate. The coupling cycle can now be repeated using a phosphoramidite derivative of the next appropriate nucleoside. The sequences will be continued as necessary until the desired oligonucleotide is obtained. 

It then remains to remove protecting groups and release the product from the support. All of these tasks, except for the removal of the dimethoxytrityl group, are achieved by use of a single deprotection reagent, aqueous base (ammonia). The cyanoethyl groups are lost from the phosphates by base-catalysed elimination, and amide protection of the bases is removed by base-catalysed hydrolysis. The latter process also achieves hydrolysis of the succinate ester link to the support. 

Solid-phase synthesis is usually done on a silica support with a covalently attached succinamide as shown in Eq. 5-25. The first nucleotide at the 3 end of the chain to be synthesized is attached by an ester linkage to the bound succinamide (step a, Eq. 5-25). The 5 -protecting group is removed in step b and the 5 -OH reacts with the activated phosphine of the second nucleotide (step c, Eq. 5-25). Steps b and c are then repeated as often as necessary to complete the chain. The finished polynucleotide can be removed from the solid support, the cyanoethyl groups removed... 

Introduction of a phosphate monoester and removal of its protecting groups was a serious impediment in a synthesis of the serine-threonine phosphatase inhibitor Calyculin A by Evans and co-workers [Scheme 7,22].38 The four phosphate protecting groups evaluated were 2-cyanoethyl and 2-(trimethylsi]y])ethyl (see below), benzyl and p-methoxybenzyl. Of these, the p-methoxybenzyl phosphate ester offered the best compromise between base stability (it survived a Wittig reaction in the presence of a metal amide base) and acid lability required for the final deprotection step in which three secondary terf-butyldimethylsiiyl... 

Protective group for nucleosides. Stepwise synthesis of oligonucleotides utilizing 2-cyanoethyl phosphate (1, 172-173) requires protection of the 3 -hydroxyl group which can be cleaved under essentially neutral conditions, since a methoxy-trityl ether is sensitive to acid and a /3-cyanoethyl phosphoric ester is sensitive to alkali. The /3-benzoylpropionyl group meets these requirements, since it is quantitatively cleaved by dilute solutions of hydrazine hydrate in pyridine-acetic acid. The esters are prepared by condensation with DCC (dicyclohexylcarbodiimide). Definitive paper R. L. Letsinger and P. S. Miller, Am. Soc., 91, 3356 (1969)... 

The deprotection of crude oligodeoxynucleotides traditionally requires a basic treatment that allows the concomitant removal of the exocyclic acyl protecting groups, -elimination of the 2-cyanoethyl phosphate protecting group, and cleavage of the succinic ester bond that links the oligonucleotide to the solid support (Fig. 9). 

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