Design of Novel Antisense Nucleic Acids with Resistance for Nuclease digestion

抗核酸酶消化的新型反义核酸的设计

• 批准号: 07457551
• 负责人: URATA Hidehito
• 金额: $ 3.52万
• 依托单位: Osaka University of Pharmaceutical Sciences
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07457551/
• 关键词: Antisense; Oligonucleotide; L-Nucleoside; Conformational fixation; Carbocyclic Nucleoside; オリゴヌクレチド

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

1.设计并外消旋合成了新型碳环尿苷、胞苷、腺苷和鸟苷类似物（1-4），其糖苷扭转角通过碱基和糖基之间的5元O-环结构固定在X=180 °附近。2.以环戊二烯为原料，通过（+）-二异松基硼烷的不对称硼氢化反应合成了光学活性环戊烷单元（5）。然后，根据用于合成外消旋1-4的合成方案，通过使用化合物5作为起始材料来进行光学活性1-4的合成。（-）-1的合成已经完成，（-）-2和（-）-3的合成将在不久的将来实现。3.将（-）-1转化为3 '-亚磷酰胺衍生物后，合成了含（-）-1残基的寡脱氧核苷酸（6-9），并对其进行了结构分析，发现（-）-4的合成途径存在缺陷，需要进一步改进。 ...更多信息 通过自动化DNA合成仪进行了合成。已经评估了这些寡聚脱氧核苷酸对3 '-毒液磷酸二酯酶消化的抗性以及对（dA）_<12>和（rA）_<12>的杂交特性。5'-TTTTTTTTTTTT-3'（6）5'-TTTTTcUcUTTTTT-3'（8）5'-TTTTTTcUTTTTT-3'（7）5'-cUcUcUcUcUcUcUcUcUcUcUcUcUcU-3'（9）cU=（-）-1（1）对毒液磷酸二酯酶消化的抗性在6在几分钟内被酶水解的条件下，9根本没有被水解。在7和8的情况下，cU残基和3 '端胸腺嘧啶残基之间的序列被快速水解，但在cU残基处水解速率大大降低。(2)与互补序列的杂交特性Tm值为7和8相比，每个取代降低了10 μ C以上，与盐浓度无关。9在低盐条件下不能与（dA）_<12>和（rA）_<12>杂交，而在高盐条件下只能与（rA）_形成稳定的双链体<12>。少

项目成果

Hidehito Urata et al.: "Design and Racemic Synthesis of Conformationally Restricted CarbocyclicPyrimidine Nucleoside Analogs Based on the Structure of the L-Nucleoside Residue in Heterochiral DNA" Chem.Pharm.Bull.46 (in press). (1998)

Hidehito Urata 等人：“基于异手性 DNA 中 L-核苷残基结构的构象限制性碳环嘧啶核苷类似物的设计和外消旋合成”Chem.Pharm.Bull.46（印刷中）。

Hidehito Urata et al.: "Sequence Dependence of Thermodynamic Stability of Heterochiral DNA" Tetrahedron Letters. 37. 5551-5554 (1996)

Hidehito Urata 等人：“异手性 DNA 热力学稳定性的序列依赖性”四面体快报。

Hirofumi Ohishi et al.: "The Crystal Structure of ((]SY.+-。[)) -Cyclocytidine Analogue Having a Low Anti Conformation Around Glycosyl Bond" Acta Crystallographica Section C. (in press). (1998)

Hirofumi Ohishi 等人：“糖基键周围具有低反构象的 ((]SY.+-.[)) -环胞苷类似物的晶体结构”《晶体学报》C 部分（出版中）。

Hidehito Urata, et al.: "Sequence Dependence of Thermodynamic Stability of Heterochirai DNA" Tetrahedron Letters. 37. 5551-5554 (1996)

Hidehito Urata 等人：“异手性 DNA 热力学稳定性的序列依赖性”四面体快报。