Solution phase synthesis of short oligoribonucleotides on a precipitative tetrapodal support

An effective method for the synthesis of short oligoribonucleotides in solution has been elaborated. Novel 2'-O-(2-cyanoethyl)-5'-O-(1-methoxy-1-methylethyl) protected ribonucleoside 3'-phosphoramidites have been prepared and their usefulness as building blocks in RNA synthesis on a soluble support has been demonstrated. As a proof of concept, a pentameric oligoribonucleotide, 3'-UUGCA-5', has been prepared on a precipitative tetrapodal tetrakis(4-azidomethylphenyl)pentaerythritol support. The 3'-terminal nucleoside was coupled to the support as a 3'-O-(4-pentynoyl) derivative by Cu(I) promoted 1,3-dipolar cycloaddition. Couplings were carried out with 1.5 equiv of the building block. In each coupling cycle, the small molecular reagents and byproducts were removed by two quantitative precipitations from MeOH, one after oxidation and the second after the 5'-deprotection. After completion of the chain assembly, treatment with triethylamine, ammonia and TBAF released the pentamer in high yields.

Compound 1a (2.35g, 3.65mmol) was dissolved in THF (47 mL), 1:1 mixture (v/v) of TFA and water (23.5 mL) was added dropwise on an ice-bath and the mixture was left with stirring for 3.5 h at 0 °C.
The progress of acetalization was monitored by TLC and p-toluensulfonic acid monohydrate was added portionwise until the starting material had disappeared. The crude mixture was extracted with EtOAc (100 mL). The organic phase was washed with aq. NaHCO3 (200 mL) and dried over Na2SO4.
Compound 3a was obtained in 96% yield as white foam (
The mixture was agitated for 96 h at room temperature. The solvent was removed under reduced pressure and the residue was sujected to column chromatography on silica gel using a gradient of 1-
4-Pentynoic anhydride obtained was then added to a solution of 4a (0.49 g, 1.03 mmol) in pyridine (20 mL) on an ice-bath. A catalytic amount of DMAP was added and the mixture was stirred for 2 h at room temperature. After completion, the solvent was removed by evaporation and the yellowish oil was subjected to chromatographic purification on a silica gel column using a gradient of 1-2% MeOH in DCM containing 1% TEA as an eluent. 6a was obtained in 95% yield (0.544 g, 0.983 mmol