Synthesis and characterization of oligonucleotides containing the C4′-oxidized abasic site produced by bleomycin and other DNA damaging agents
Synthesis and characterization of oligonucleotides containing the C4′-oxidized abasic site produced by bleomycin and other DNA damaging agents
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DOI：
10.1002/anie.200352102
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发表时间：
2003-01-01
影响因子：
16.6
通讯作者：
Greenberg, MM
中科院分区：
文献类型：
--
作者：
Kim, J;Gil, JM;Greenberg, MM
非环前体5应该导致非对映异构体的混合。[4C，d]这种期望与核磁共振对DNA中2的表征一致，这也使我们相信1的立体异构体应该相互平衡地存在，并存在少量的5。[10]由于我们没有理由期望1将作为单一的异构体存在于DNA中，我们认为没有理由试图从稳定的前体的单个立体异构体中产生它。在设计我们的方法时，我们还考虑到包含2和3的生物聚合物的化学合成能够利用寡核苷酸合成过程中通常使用的碱性去保护条件，在最后的光化学步骤中揭开不易碱的损伤。[7，8]这些考虑导致我们设计6作为自动合成过程中使用的亚磷酰胺，最终在DNA中合成1(方案2)。6的显著特征包括用作耐碱性光触发器的邻硝基藜芦基(ONV)和伯醇的硅氧基保护基团。后者允许我们在不使双缩醛(7)暴露在酸中的情况下合成受保护的寡核苷酸。[11]邻-硝基藜芦基是一类在合成，特别是寡核苷酸合成中普遍使用的邻硝基苄基耐光保护基的成员。[12]预期含有7的寡核苷酸将被提纯、储存，并根据需要用于生成1。亚磷酰胺6是由3‘，5’-O-硅氧基脱氧核糖缩醛8(图式3)合成的。甲缩醛在1，3-丙二硫醇存在下的裂解提供给二硫杂环9一个C4-羟基，然后被氧化。[13]随后在3，4-二甲氧基-6-硝基苯甲醇存在下氧化裂解二硫代酮10得到11作为四个立体异构体的混合物。[14]虽然在这一步分离异构体是不可行的，但相应的1H核磁共振谱表明两个形成的量要大得多。单个非对映异构体的分析样品在脱硅后获得(12)。用~1H核磁共振波谱确定了1R，4S-12和1S，4R-12占环化化合物混合物的80%。在实际应用中，主要的非对映异构体是在制备环十二氧基双三甲基硅氧基硅醚(1R，4S-1S，4R-1S，4R-1S)时分离出来的。
acyclic precursor 5 should result in a mixture of diastereomers.[4c, d] This expectation is consistent with NMR characterization of 2 in DNA, which also leads us to believe that the stereoisomers of 1 should exist in equilibrium with each other and small amounts of 5.[10] As there was no reason for us to expect that 1 will exist in DNA as a single isomer, we saw no reason to attempt to generate it from individual stereoisomers of a stable precursor. While devising our approach, we also took into account that chemical syntheses of biopolymers that contain 2 and 3 are able to utilize the alkaline deprotection conditions typically employed during oligonucleotide synthesis by unmasking the alkali-labile lesions in a final photochemical step.[7, 8] These considerations led us to design 6 as the phosphoramidite used during the automated synthesis to ultimately synthesize 1 in DNA (Scheme2). Notable features of 6 include the o-nitroveratryl moiety (ONV), which serves as the alkali-resistant phototrigger, and the silyloxy protecting group for the primary alcohol. The latter allows us to synthesize protected oligonucleotides without exposing the bisacetal (7) to acid.[11] The o-nitroveratryl group is a member of the class of o-nitrobenzyl photolabile protecting groups that are of general use in synthesis, and oligonucleotide synthesis in particular.[12] It was anticipated that oligonucleotides containing 7 would be purified, stored, and used to generate 1 as needed. Phosphoramidite 6 was prepared from the 3’, 5’-O-silyloxy deoxyribose acetal 8 (Scheme3). Cleavage of the methyl acetal in the presence of 1, 3-propane dithiol provided the dithiane 9 with a C4-hydroxy group, which was then oxidized.[13] Subsequent oxidative cleavage of the dithiane ketone, 10, in the presence of 3, 4-dimethoxy-6-nitrobenzyl alcohol provided 11 as a mixture of four stereoisomers.[14] Although separation of the isomers was not practical at this step, the corresponding 1H NMR spectrum indicated that two were formed in significantly greater amounts. Analytical samples of the individual diastereomers were obtained upon desilylation (12). 1H NMR spectroscopy was used to determine that 1R, 4S-12 and 1S, 4R-12 accounted for> 80% of the mixture of cyclized compounds. In practice, the major diastereomers were separated upon preparation of the cyclododecyloxy bis-trimethylsilyloxy silyl ethers (1R, 4S-1S, 4R-