Design of Novel Antisense Nucleic Acids with Resistance for Nuclease digestion

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1.Design and racemic synthesis of novel carbocyclic uridine, cytidine, adenosine and guanosine analogs (1-4), whose glycosidic torsion angle is fixed at around X=180ﾟ by means of the 5-membered O-cyclic structure between the base and sugar moiety, has been achieved.2.The optically active cyclopentane unit (5) has been synthesized from cyclopentadiene via asymmetric hydroboration by using (+)-diisopinocampheylborane. Then, synthesis of optically active 1-4 was performed by using compound 5 as a starting material according to the synthetic protocol for synthesis of racemic 1-4. Synthesis of (-)-1 has been achieved, and synthesis of (-)-2 and (-)-3 will be achieved in near future. However, modification of the synthetic pathway of (-)-4 is now investigating, because low solubility of the synthetic intermediates causes difficulties for their purification.3.After conversion of (-)-1 to its 3'-phosphoramidite derivative, oligodeoxynucleotides containing the (-)-1 residues (6-9) have been synt … More hesized by means of automated DNA synthesizer. Resistance to 3'-venome phosphodiesterase digestion and hybridization properties towards (dA)_<12> and (rA)_<12> of these oligodeoxynucleotides have been evaluated.5'-TTTTTTTTTTTT-3' (6) 5'-TTTTTcUcUTTTTT-3' (8)5'-TTTTTTcUTTTTT-3' (7) 5'-cUcUcUcUcUcUcUcUcUcUcUcU-3' (9) cU= (-)-1(1) Resistance to venome phosphodiesterase digestionUnder the conditions 6 is hydrolyzed by the enzyme in a few minutes, 9 was not hydrolyzed at all. In the case of 7 and 8, the sequence between the cU residue and the 3'-end thymine residue was hydrolyzed repidly, but the hydrolyze rate was greatly decreased at the cU residue.(2) Hybridization properties with the complementary sequencesTm values of 7 and 8 were lowered by more than 10ﾟC per substitution compared to 6, independent of salt concentration. On the other hand, 9 was not be able to hybridize with (dA)_<12> and (rA)_<12> at low salt concentration, however, formed a stable duplex only with (rA)_<12> at high salt conditions. Less