Design of Gene Therapeutic Agent Possessing Cooperative DNA Alkylating Ability

具有协同DNA烷基化能力的基因治疗剂的设计

• 批准号: 09555283
• 负责人: SUGIYAMA Hiroshi
• 金额: $ 7.1万
• 依托单位: Tokyo Medical and Dental University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 1998
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09555283/
• 关键词: Antitumor agent; Heterodimer; Duocarmycin; Distamycin; DNA sequence; Molecular recognition; DNAアルキル化; 配列特異性; 遺伝子治療; 協同性

Sequence-specific DNA alkylations have been a topic of much current interest due to their significant potential in molecular biology and human medicine. We developed general method to synthesize novel conjugates between segment A of Duocarmycin A (Du) and N-methylimidazole (lm)-N-methylpyrrole (Py) hairpin polyamides and their DNA alkylation. The conjugates PyPyPygammalmPyPyDu (8a) and JmPyPygammalmPyPyDu (8b) were designed to alkylate the target sequences (A/T)G(A/T)_2N(A/G) and (A/T)G(A/T)CN(A/G), respectively, according to Dervan's ring-pairing rule. High resolution denaturing gel electrophoresis indicated that 8a exclusively alkylated the A of the 5'-TGTAAAA-3' within a -400 bp DNA fragment Similarly, alkylation by 8b occurred exclusively at the G of the 5'-AGTCAGA-3' sequence with efficiency at nanomolar concentration. In order to better understand the structure of the alkylated DNA by these conjugates, the alkylation of non-self complementary duplex decanucleotides, ODNl and ODN2, were investigated. HPLC and ESMS analyses of the reaction of these ODNs with 8a and 8b demonstrated that both conjugates efficiently and selectively alkylate N3 of the purine bases of their target sequence.

序列特异性DNA烷基化反应由于其在分子生物学和人类医学中的巨大潜力而成为当前研究的热点。我们发展了一种合成多卡霉素A（Du）A片段与N-甲基咪唑（Im）-N-甲基吡咯（Py）发夹状聚酰胺的新型偶联物及其DNA烷基化的通用方法。根据Dervan的环配对规则，设计了分别使靶序列（A/T）G（A/T）_2N（A/G）和（A/T）G（A/T）CN（A/G）烷基化的缀合物PyPyPyPygammalmPyPyDu（8a）和JmPygammalmPyDu（8b）。高分辨率变性凝胶电泳表明，8a仅烷基化~ 400 bp DNA片段内的5 ′-TGTAAAA-3 ′的A。类似地，8b的烷基化仅发生在5 ′-AGTCAGA-3 ′序列的G处，在纳摩尔浓度下有效。为了更好地理解这些缀合物的烷基化DNA的结构，研究了非自身互补双链体十核苷酸ODN 1和ODN 2的烷基化。这些ODN与8a和8b反应的HPLC和ESMS分析表明，这两种缀合物都有效地和选择性地烷基化其靶序列的嘌呤碱基的N3。

项目成果

Sugiyama,H: "Highly Efficient DNA Interstrand Crosslinking Induced by an Antitumor Antibiotic,Carzinophilin" Tetrahedron Letters. 40. 315-318 (1999)

Sugiyama,H：“抗肿瘤抗生素亲肉素诱导的高效 DNA 链间交联”四面体快报。

Sugiyama,H: "Preferential C1'Hydrogen Abstraction by a Uracilyl Radical in DNA-RNA Hybrid." Tetrahedron Letters. 38. 8057-8060 (1997)

Sugiyama,H：“DNA-RNA 杂合体中尿嘧啶自由基优先夺取 C1 氢。”

Sugiyama, H.: "Product Analysis of GG-SpecificPhotooxidation of DNA via Electron Transfer : 2-Aminoimidazolone as a Major Guanine Oxidation Product." J.Am.Chem.Soc.120. 7373-7374 (1998)

Sugiyama, H.：“通过电子转移对 DNA 进行 GG 特异性光氧化的产物分析：2-氨基咪唑酮作为主要鸟嘌呤氧化产物。”

Zhang, Q.-M.: "Replication of DNA Templates Containing 5-Fomyluracil,a Major Oxidative Lesion of Thymine in DNA." Ncleic Acids Res.25. 3969-3973 (1997)

张，Q.-M.：“含有 5-甲酰尿嘧啶的 DNA 模板的复制，这是 DNA 中胸腺嘧啶的主要氧化损伤。”

Sugiyama, H.: "Highly Efficient DNA Interstrand Crosslinking Induced by an Antitumor Antibiotic, Carzinophilin" Tetrahedron Letters. 40. 315-318 (1999)

Sugiyama, H.：“抗肿瘤抗生素、嗜肉素诱导的高效 DNA 链间交联”四面体快报。