掲載種別：研究論文（学術雑誌）   出版者・発行元：Springer Science and Business Media LLC  

Abstract

Cell states are regulated by the response of signaling pathways to receptor ligand-binding and intercellular interactions. High-resolution imaging has been attempted to explore the dynamics of these processes and, recently, multiplexed imaging has profiled cell states by achieving a comprehensive acquisition of spatial protein information from cells. However, the specificity of antibodies is still compromised when visualizing activated signals. Here, we develop Precise Emission Canceling Antibodies (PECAbs) that have cleavable fluorescent labeling. PECAbs enable high-specificity sequential imaging using hundreds of antibodies, allowing for reconstruction of the spatiotemporal dynamics of signaling pathways. Additionally, combining this approach with seq-smFISH can effectively classify cells and identify their signal activation states in human tissue. Overall, the PECAb system can serve as a comprehensive platform for analyzing complex cell processes.

DOI： 10.1038/s41467-024-47989-9

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その他リンク： https://www.nature.com/articles/s41467-024-47989-9

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：MDPI  

DNA is constantly damaged by various external and internal factors. In particular, oxidative damage occurs in a steady state, and 8-oxo-2 '-deoxyguanosine (oxodG) is known as the main oxidative damage. OxodG is a strong genotoxic nucleoside and is thought to be involved in the pathogenesis of cancer and neurological diseases. However, a breakthrough method to detect the position of oxodG in DNA has not yet been developed. Therefore, we attempted to develop a novel method to detect oxodG in DNA using artificial nucleosides. Recently, we have succeeded in the recognition of oxodG in DNA by a single nucleotide elongation reaction using nucleoside derivatives based on a purine skeleton with a 1,3-diazaphenoxazine unit. In this study, we developed a new nucleoside derivative with a pyrimidine skeleton in order to further improve the recognition ability and enzymatic reaction efficiency. We, therefore, designed and synthesized 2 '-deoxycytidine-1,3-diazaphenoxazine (Cdap) and its triphosphate derivatives. The results showed that it was incorporated into the primer strand relative to the dG template because of its cytidine skeleton, but it was more effective at the complementary position of the oxodG template. These results indicate that the new nucleoside derivative can be considered as one of the new candidates for the detection of oxodG in DNA.

DOI： 10.3390/molecules29102270

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PHARMACEUTICAL SOC JAPAN  

Triplex DNA formation has generated much interest as a genomic targeting tool that directly targets duplex DNA. However, fundamental limitations in the base pairs of target duplex DNA sequences that can form stable triplex DNA have limited the application. Recently, we have reported on the recognition of CG and (5m)CG base pairs by artificial nucleic acid derivatives with a 2'-deoxynebularine skeleton. Therefore, we attempted to explore the basic skeleton that is important for the development of new artificial nucleic acids allowing for the recognition of TA base pairs. In this study, we focused on a benzimidazole skeleton and introduced a hydroxyl group to enable one -point hydrogen bonding. We have synthesized artificial nucleoside analogues with hydroxyl group on the benzimidazole and incorporated their amidite derivatives into triplex forming oligonucleotides (TFOs). The gel shift assay was performed to evaluate the triplex DNA formation ability of synthesized TFOs, and TFOs containing hydroxybenzimidazole were successfully recognized TA base pairs for all four different sequences. Moreover, compared to the results for the TFOs containing benzimidazole, which suggested hydrogen bonding formation at the hydroxyl group. Therefore, hydroxybenzimidazole would be an important artificial nucleic acid skeleton for TA base pair recognition.

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記述言語：英語   出版者・発行元：BENTHAM SCIENCE PUBL LTD  

Based on the sequence-specific recognition of target duplex DNA by triplex-forming oligonucleotides (TFOs) at the major groove side, the antigene strategy has been exploited as a gene-targeting tool with considerable attention. Triplex DNA is formed via the specific base triplets by the Hoogsteen or reverse Hoogsteen hydrogen bond interaction between TFOs and the homo-purine strand from the target duplex DNA, leading to the established sequence-specificity. However, the presence of inversion sites, which are known as non-natural nucleosides that can form satisfactory interactions with 2 '-deoxythymidine (dT) and 2 '-deoxycytidine (dC) in TA and CG base pairs in the target homo-purine DNA sequences, drastically restricts the formation of classically stable base triplets and even the triplex DNA. Therefore, the design of non-natural type nucleosides, which can effectively recognize CG or/and TA inversion sites with satisfactory selectivity, should be of great significance to expanding the triplex-forming sequence. Here, this review mainly provides a comprehensive review of the current development of novel non-natural nucleosides to recognize CG or/and TA inversion sites in triplex DNA formation against double-strand DNA (dsDNA).

DOI： 10.2174/0929867330666230512114130

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY  

This article describes the detailed synthetic protocol for the preparation of oligonucleotides containing 2-guanidinoethyl-2 '-deoxynebularine and 2-ureidoethyl-2 '-deoxynebularine nucleoside derivatives. These derivatives are obtained by a post-synthetic modification of triplex-forming oligonucleotides (TFOs) containing 2-aminoethyl-2 '-deoxynebularine, which is useful for forming stable triplex DNA with duplex DNA sequences containing 5mCG and CG interrupting sites. The hydroxyl groups of the sugar moiety of commercially available 2 '-deoxyguanosine are acetyl-protected, the 6-position is chlorinated and reduced to give a 2-substituted nebularine derivative, and then the sugar moiety is deprotected. The hydroxyl groups of the sugar moiety are silyl-protected and the amino group at the 2-position is iodinated before being coupled with diethyl malonate. The ethyl ester is reduced and the resulting alcohol converted to an amino group for protection. The compound is then converted to a phosphoramidite unit and incorporated into a TFO. Subsequent modification of the aminoethyl group on the TFO completes the synthesis of the oligonucleotides containing 2-guanidinoethyl-2 '-deoxynebularine and 2-ureidoethyl-2 '-deoxynebularine. (c) 2023 Wiley Periodicals LLC.Basic Protocol 1: Preparation of the phosphoramidite unit of the 2-aminoethyl-2 '-deoxynebularine derivative (14)Basic Protocol 2: Post-synthetic modification of oligonucleotides containing 2-aminoethyl-2 '-deoxynebularine derivativesBasic Protocol 3: Determination of the triplex-forming ability of oligonucleotides containing 2-aminoethyl-2 '-deoxynebularine derivatives

DOI： 10.1002/cpz1.893

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

Antigene methods are promising novel therapeutic approaches to suppress abnormal gene expression. One of these methods inhibits transcription by forming triplex DNA against duplex DNA. However, by using natural-type triplex-forming oligonucleotides (TFOs), stable triplex formation is limited to homopurine and homopyrimidine strands in targeted duplex DNA. We recently developed artificial nucleoside analogues with the ability to recognize CG and TA inversion sites. We successfully formed stable unnatural-type triplex DNA for duplex DNA containing a CG base pair and extended the target sequence using TFOs containing 2-amino-3-methylpyridinyl pseudo-dC (3MeAP-ΨdC). Therefore, this present study investigated triplex-forming regions and synthesized antigene TFOs containing 3MeAP-ΨdC. Some of the synthesized antigene TFOs reduced transcription products and inhibited cell proliferation in several types of cultured cancer cells. The antigene effects of antigene TFOs containing artificial nucleic acids were markedly stronger than those of natural-type TFOs, and these results clearly demonstrated the usefulness of incorporating artificial nucleic acids within TFOs.

DOI： 10.1039/d3md00139c

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掲載種別：研究論文（学術雑誌）   出版者・発行元：Pharmaceutical Society of Japan  

DOI： 10.1248/cpb.c22-00738

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

The formation of triplex DNA is a site-specific recognition method that directly targets duplex DNA. However, triplex DNA formation is generally formed for the GC and AT base pairs of duplex DNA, and there are no natural nucleotides that recognize the CG and TA base pairs, or even the 5-methyl-CG (5mCG) base pair. Moreover, duplex DNA, including 5mCG base pairs, epigenetically regulates gene expression in vivo, and thus targeting strategies are of biological importance. Therefore, the development of triplex-forming oligonucleotides (TFOs) with artificial nucleosides that selectively recognize these base pairs with high affinity is needed. We recently reported that 2'-deoxy-2-aminonebularine derivatives exhibited the ability to recognize 5mCG and CG base pairs in triplex formation; however, this ability was dependent on sequences. Therefore, we designed and synthesized new nucleoside derivatives based on the 2'-deoxy-nebularine (dN) skeleton to shorten the linker length connecting to the hydrogen-bonding unit in formation of the antiparallel motif triplex. We successfully demonstrated that TFOs with 2-guanidinoethyl-2'-deoxynebularine (guanidino-dN) recognized 5mCG and CG base pairs with very high affinity in all four DNA sequences with different adjacent nucleobases of guanidino-dN as well as in the promoter sequences of human genes containing 5mCG base pairs with a high DNA methylation frequency.

DOI： 10.1093/nar/gkac1110

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE  

Oxidative lesions, such as 8-oxo-dG and 8-oxo-dA, are continuously generated from exposure to reactive oxygen species. While 8-oxo-dG has been extensively studied, 8-oxo-dA has not received as much attention until recently. Herein, we report the synthesis of duplex DNAs incorporating dA, 8-oxo-dA, 7-deaza-dA, 8-Br-dA, and 8-Br-7-deaza-dA, which have different substitutions at 7-and 8-position, for the investigation into the implications of N7-hydrogen and C8-keto on the base pairing preference, mutagenic potential and repair of 8-oxo-dA. Base pairing study suggested that the polar N7-hydrogen and C8-keto of 8-oxo-dA, rather than the syn-preference, might be essential for 8-oxo-dA to form a stable base pair with dG. Insertion and extension studies using KF-exo? and human DNA polymerase beta indicated that the efficient dGTP insertion opposite 8-oxo-dA and extension past 8-oxo-dA:dG are contingent upon not only the stable base pair with dG, but also the flexibility of the active site in polymerase. The N7-hydrogen in 8-oxo-dA or C7-hydrogen in 7-deaza-dA and 8-Br-7-deaza-dA was suggested to be important for the recognition by hOGG1, although the excision efficiencies of 7-deaza-dA and 8-Br-7-deaza-dA were much lower than 8-oxo-dA. This study provides an insight into the structure-function relationship of 8-oxo-dA by nucleotide analogues.

DOI： 10.1016/j.bioorg.2022.106029

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER CHEMICAL SOC  

The formation of unnatural base pairs within duplex DNA would facilitate DNA nanotechnology and biotechnology. Iso-2'-deoxyguanosine (iso-dG) forms base pairs with iso-2'-deoxycytidine, and its use as an unnatural base pair was investigated. Iso-dG is one of the tautomers of 2-hydroxy-2'-deoxyadenosine (2-OH-dA), known as an oxidatively damaged nucleobase, and its selective recognition in DNA plays an important role in the diagnosis and pathogenesis of disease. Therefore, we focused on pseudo-dC (ψdC) as a suitable molecule that recognizes 2-OH-dA in DNA. Since 2-OH-dA shows tautomeric structures in DNA, we designed and used ψdC, which also has a tautomeric structure. We successfully synthesized a ψdC phosphoramidite compound for the synthesis of oligonucleotides (ODNs) as well as its triphosphate derivative (ψdCTP). Tm measurements revealed that ODNs including ψdC showed stable base pair formation with ODNs having 2-OH-dA. In contrast, low Tm values were observed for other bases (dG, dA, dC, and T). The results obtained for the single-nucleotide primer extension reaction revealed that ψdCTP was incorporated into the complementary position of 2-OH-dA in template DNA with high selectivity. In addition, the primer elongation reaction was confirmed to proceed in the presence of dNTPs. The present study reports an artificial nucleic acid that selectively and stably forms unnatural base pairs with 2-OH-dA in DNA.

DOI： 10.1021/jacs.2c07000

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PHARMACEUTICAL SOC JAPAN  

Due to the importance of the RNA chemical modifications, methods for the selective chemical modification at a predetermined site of the internal position of RNA have attracted much attention. We have developed functional artificial nucleic acids that modify a specific site of RNA in a site-and base-selective manner. In addition, the copper-catalyzed azide-alkyne cycloaddition (CuAAC) has been shown to introduce additional molecules on the alkynes attached to the pyridine ring. However, it was found that some azide compounds produced the cycloadduct in lower yields. Therefore, in this study, we synthesized the pyridinyl transfer group with the alkyne attached via a polyethylene glycol (PEG) linker with a different length and optimized its structure for both the transfer and CuAAC reaction. Three new transfer groups were synthesized by introducing an alkyne group at the end of the triethylene (11), tetraethylene (12) or pentaethylen glycol linker (13) at the 5-position of the pyridine ring of (E)-3-iodo-1-(pyridin-2-yl)prop-2-en-1-one. These transfer groups were introduced to the 6-thioguanine base in the oligodeoxynucleotide (ODN) in high yields. The transfer groups 11 and 12 more efficiently underwent the cytosine modification. For the CuAAC reaction, although 7 showed low adduct yields with the anionic azide compound, the new transfer groups, especially 12 and 13, significantly improved the yields. In conclusion, the transfer groups 12 and 13 were determined to be promising compounds for the modification of long RNAs.

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ROYAL SOC CHEMISTRY  

The identification of the position of 8-oxo-2'-deoxyguanosine (8-oxo-dG) in DNA is important to clarify the pathogenesis of many diseases. We herein developed a purine-1,3-diazaphenoxazine triphosphate (dPdapTP) and described the first example of detecting the presence of 8-oxo-dG by amplifying it several hundred times after the multiple-turnover single nucleotide primer extension reactions.

DOI： 10.1039/d2cc01372j

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記述言語：日本語   出版者・発行元：SOC SYNTHETIC ORGANIC CHEM JPN  

To determine of the position of oxidative nucleoside in DNA, the selective detection of 8-oxo-2'deoxyguanosine (8-oxo-dG) without chemical or enzymatic treatment is an attractive tool for genomic research. Here, I described the summary of our resent study which designed and synthesized the non -natural nucleoside analogue, the adenosine-1,3-diazaphenoxazine derivative (Adap), for selective recognition of 8-oxo-dG in DNA. This study has clearly shown that Adap has stabilizing effect of the duplex including the Adap-8-oxodG base pair with high selectivity. Furthermore, the fluorescent quenching property of Adap has been shown to detect of 8-oxo-dG in DNA. And also, the triphosphate of Adap (dAdapTP) was synthesized and tested for single nucleotide incorporation reaction to primer strand using DNA polymerase. The efficiency of dAdapTP incorporation into 8-oxo-dG-containing templates was better than with dG-containing templates. Then the detection of 8-oxo-dG in human telomeric DNA sequences extracted from H2O2-treated HeLa cells and in genome DNA extracted from small intestine of transgenic mice were successful. Furthermore, I will introduce the development of Adap derivatives, which are expected to be applied to sequencing technology in this review.

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ROYAL SOC CHEMISTRY  

The oxidative damage of DNA is associated with aging and the development of various diseases. Although nucleoside-derived radicals play an important role in DNA oxidation, their analysis methods are limited. Herein, we propose a fluorometric detection and structural analysis of radicals on the surface of oxidatively damaged DNA using a profluorescent nitroxide probe combined with liquid chromatography-fluorometry and high-resolution tandem mass spectrometry.

DOI： 10.1039/d1cc04998d

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：MDPI  

MTH1 is an enzyme that hydrolyzes 8-oxo-dGTP, which is an oxidatively damaged nucleobase, into 8-oxo-dGMP in nucleotide pools to prevent its mis-incorporation into genomic DNA. Selective and potent MTH1-binding molecules have potential as biological tools and drug candidates. We recently developed 8-halogenated 7-deaza-dGTP as an 8-oxo-dGTP mimic and found that it was not hydrolyzed, but inhibited enzyme activity. To further increase MTH1 binding, we herein designed and synthesized 7,8-dihalogenated 7-deaza-dG derivatives. We successfully synthesized multiple derivatives, including substituted nucleosides and nucleotides, using 7-deaza-dG as a starting material. Evaluations of the inhibition of MTH1 activity revealed the strong inhibitory effects on enzyme activity of the 7,8-dihalogenated 7-deaza-dG derivatives, particularly 7,8-dibromo 7-daza-dGTP. Based on the results obtained on kinetic parameters and from computational docking simulating studies, these nucleotide analogs interacted with the active site of MTH1 and competitively inhibited the substrate 8-oxodGTP. Therefore, novel properties of repair enzymes in cells may be elucidated using new compounds.

DOI： 10.3390/ijms22031274

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Royal Society of Chemistry (RSC)  

<p>We herein demonstrated for the first time the direct recognition of duplex DNA bearing the 5-methyl-2′-deoxycytosine and 2′-deoxyguanosine base pair by triplex DNA formation.</p>

DOI： 10.1039/d1ra02831f

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

γ-Amido-modified 2'-deoxynucleoside triphosphates (dNTPs) and nucleoside triphosphates (NTPs) are becoming increasingly important as biological tools. We herein describe the simple and easy synthesis of γ-amido-dNTPs and -NTPs from commercially available corresponding dNTPs and NTPs in a one-pot reaction using water-soluble carbodiimide and ammonia solution. We examined the effects of synthesized γ-amido-dNTPs on the DNA polymerase reaction. The results obtained showed the incorporation of these derivatives into the DNA primer while maintaining nucleobase selectivity; however, their incorporation efficiency by DNA polymerase was lower than that of dNTP. This is the first study to demonstrate the successful synthesis of four sets of γ-amido-dNTPs and clarify their properties.

DOI： 10.1248/cpb.c21-00497

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

The triplex DNA forming method is an attractive tool as a gene-targeting agent. Using artificial nucleoside analogues based on C-nucleoside, stable and selective triplex DNA can be formed in a specific region of duplex DNA, and its biotechnology applications will greatly expand. In this study, we designed and synthesized novel C-nucleoside analogues based on the pyrimidine skeleton, 3MeAP-d(Y-Cl) and 3MeAP-d(Y-H), capable of recognizing a CG mismatch site that is not recognized by natural nucleosides. After incorporating them into the oligonucleotides, their triplex forming abilities were evaluated by gel-shift assay. Although it was only one sequence, the 3'-GZG-5' sequence, the stability of the CG mismatch site recognition was greatly improved compared with previous nucleoside analogues.

DOI： 10.1016/j.bmc.2020.115782

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ROYAL SOC CHEMISTRY  

We recently found the translocation of double-stranded DNA into the nucleus. We herein describe the concept of novel booster oligodeoxynucleotides including 2 '-deoxy uridine, which release antigene oligonucleotides in the nucleus by enzymatic digestion. This system exhibited stronger hTERT mRNA expression inhibitory activity than single-stranded antigene oligonucleotides.

DOI： 10.1039/d0cc04240d

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：EXCERPTA MEDICA INC-ELSEVIER SCIENCE INC  

The immune response after transcatheter aortic valve implantation (TAVI) in comparison to that after surgical aortic valve replacement (SAVR) remains to be fully elucidated. In a 2-part study, we assessed laboratory data obtained before, immediately after, and 24 and 48 hours after SAVR (128 patients; age >= 80 [mean 82] years) or transfemoral TAVI (102 patients; age >= 80 [mean 86] years) performed for aortic stenosis. In-hospital mortalities were similar (3% vs 0%), but leukocyte counts and aspartate aminotransferase and creatine kinas concentrations were decreased immediately and 24 hours after surgery (all, p <0.001). We performed cytokine profiling in a SAVR group (11 patients; mean age, 77 years) and transfemoral TAVI group (12 patients; mean age, 84 years). By measuring normalized concentrations of 71 cytokines at 3 time points, we found a significant difference (defined as fold change >1.7 and p <0.05 [by Mann-Whitney U-test]) in 23 cytokines. The differentially expressed cytokines fell into 3 hierarchical clusters: cluster A (high increase after SAVR and suppressed increase after TAVI only immediately after surgery [CCL2, CCL4, and 2 others]), cluster B (high increase after SAVR and suppressed increase after TAVI at 2 time points [IL-1Ra, IL-6, IL-8, IL-10, and 5 others]), and cluster C (various patterns [TRAIL, CCL11, and 8 others]). Gene enrichment analysis identified multiple pathways associated with the inflammatory responses in SAVR and altered responses in TAVI, including cellular responses to tumor necrosis factor (p = 0.0035) and interleukin-1 (p = 0.0062). In conclusion, a robust inflammatory response follows SAVR, and a comparatively attenuated response follows TAVI. (C) 2020 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.amjcard.2020.04.037

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ROYAL SOC CHEMISTRY  

Expansion of the triplex DNA forming sequence is required in the genomic targeting fields. Basically, triplex DNA is formed by the interaction between the triplex-forming oligonucleotides and homo-purine region with the target duplex DNA. The presence of the base pair conversion sites hampers stable triplex formation. To overcome this limitation, it is necessary to develop an artificial nucleic acid to recognize the base conversion sites, and the CG and TA base pairs. We describe the synthesis of C-nucleoside analogues and an evaluation of the ability of triplex formation. Consequently, the combined use of the novel C-nucleoside analogues, AY - AY-d(Y-NH2), AY-d(Y-Cl) and (I)AP-d(Y-Cl), is capable of recognizing duplex DNA including the TA or dUA base pair.

DOI： 10.1039/d0ob00420k

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ELSEVIER  

Tumorigenesis induced by oxidative stress is thought to be initiated by mutagenesis, but via an indirect mechanism. The dose-response curves for agents that act by this route usually show a threshold, for unknown reasons. To gain insight into these phenomena, we have analyzed the dose response for mutagenesis induced by the oral administration of potassium bromate, a typical oxidative-stress-generating agent, to gpt delta mice. The agent was given orally for 90 d to either Nrf2+ or Nrf2-knockout (KO) mice and mutants induced in the small intestine were analyzed. In Nrf2+mice, the mutant frequency was significantly greater than in the vehicle controls at a dose of 0.6 g/L but not at 0.2 g/L, indicating that a practical threshold for mutagenesis lies between these doses. At 0.6 g/L, the frequencies of G-to-T transversions (landmark mutations for oxidative stress) and G-to-A transitions were significantly elevated. In Nrf2-KO mice, too, the total mutant frequency was increased only at 0.6 g/L. G-to-T transversions are likely to have driven tumorigenesis in the small intestine. A site-specific G-to-T transversion at guanine (nucleotide 406) in a 5'-TGAA-3' sequence in gpt, and our primer extension reaction showed that formation of the oxidative DNA base modification 8-oxo-deoxyguanosine (8-oxo-dG) at nucleotide 406 was significantly increased at doses of 0.6 and 2 g/L in the gpt delta mice. In the Apc oncogene, guanine residues in the same or similar sequences (TGAA or AGAA) are highly substituted by thymine (G-to-T transversions) in potassium bromate-induced tumors. We propose that formation of 8-oxo-dG in the T(A)GAA sequence is an initiating event in tumor formation in the small intestine in response to oxidative stress.

DOI： 10.1016/j.mrgentox.2020.503136

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PHARMACEUTICAL SOC JAPAN  

N-Acetyl-7-nitroindoline has a characteristic reaction in that its acetyl group is photo-activated to acetylate amines to form amides. In this study, the N-acetyl-7-nitroindoline part was connected to the 2'-deoxyribose part at the 3- or 5-position or to a glycerol unit at the 3-position through an ethylene linker (1, 2, and 3, respectively). They were incorporated into the oligodeoxynucleotides, and their photo-reactivities toward the complementary RNA were evaluated. The acetyl group of 1 was photo-activated to form the deacelylated nitroso derivative without affecting the RNA strand. The photoreaction with 2 suggested acetylation of the RNA strand. In contrast, compound 3 formed the photo-cross-linked adduct with the RNA. These results have shown the potential application of N-acetyl-7-nitroindoline unit in aqueous solutions.

DOI： 10.1248/cpb.c20-00594

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

The efficient synthesis of the highly-deuterated thymidine-d(9) by the glycosylation reaction between a deuterated nucleobase and deuterated sugar is described. It is also incorporated into the oligonucleotides using a DNA synthesizer. Using this approach, it is possible to make highly-deuterated oligonucleotides for biological studies and structural analyses. (C) 2019 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.tetlet.2019.151037

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：TAYLOR & FRANCIS INC  

8-OxodGTP is generated by the reaction between dGTP and reactive oxygen species and a considered mutagenic nucleotide. It can be incorporated into the duplex DNA during replication processes by the DNA polymerase, and thus the repair enzyme removes oxodGTP from the nucleotide pools in living cells. On the other hand, the gamma-modified triphosphates show interesting properties for use as biological tools. Therefore, the gamma-N-pyrenylalkyl-oxodGTP derivatives were synthesized and their effect on the enzymatic reactions were evaluated. The gamma-N-pyrenylmethyl-oxodGTP was found to be accepted by the DNA polymerase just like oxodGTP, but showed a competitive inhibition property for the human oxodGTPase.

DOI： 10.1080/15257770.2019.1586919

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Blackwell Publishing Inc.  

This unit describes the detailed synthetic protocol for the preparation of the phosphoramidite units of the 2′-deoxy-4-aminopyridinylpseudocytidine derivatives. These C-nucleoside derivatives are useful units for the incorporation into triplex forming oligonucleotides (TFOs) to form the stable triplex DNA containing the CG interrupting sites. Commercially available 1-methyl-2′-deoxypseudouridine is prepared from thymidine and 5-iodo-uracil by a simple method, that is, coupling of glycal and 5-iodo-1-methyluracil by the Heck reaction, followed by desilylation and diastereoselective reduction. The carbonyl group at the 4 position of the pseudouridine derivative is activated by 3-nitorotriazole and treated with the corresponding aromatic amine compounds to produce the 2′-deoxy-4-aminopyridinylpseudocytidine derivatives. These derivatives are then successfully converted to the phosphoramidite units and incorporated into the oligodeoxynucleotides. © 2019 by John Wiley &amp
Sons, Inc.

DOI： 10.1002/cpnc.80

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PHARMACEUTICAL SOC JAPAN  

The adenosine triphosphate derivatives of 2-oxo-1,3-diazaphenoxazine (dAdapTP) showed a significant discrimination ability for the template strand including that between 8-oxo-2'-deoxyguanosine (8-oxodG) and 2'-deoxyguanosine (dG) by the single nucleotide primer extension reaction using the Klenow Fragment. In this study, we synthesized new dAdapTP derivatives, i.e., 2-amino-dAdapTP, 2-chloro-dAdapTP and 2-iodo-dAdapTP, to investigate the effect on the selectivity and efficiency of incorporation for the primer extension reaction using a variety of DNA polymerases. In contrast to the previously tested dAdapTP, the selectivity and efficiency of the 2-halo-dAdapTP incorporation were dramatically decreased using the Klenow Fragment. Moreover, the efficiency of the 2-amino-dAdapTP incorporation into the T-containing template was almost the same with that of dAdapTP. In the case of the Bsu DNA polymerase, the efficiency of all the dAdapTP derivatives decreased compared to that using the Klenow Fragment. However, the incorporation selectivity of dAdapTP had improved against the oxodG-containing template for all the template sequences including the T-containing template. Moreover, 2-amino-dAdapTP showed a better efficiency than dAdapTP using the Bsu DNA polymerase. The 2-amino group of the adenosine unit may interact with syn-oxodG at the active site of the Bsu DNA polymerase during the single primer extension reaction.

DOI： 10.1248/cpb.c19-00453

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PHARMACEUTICAL SOC JAPAN  

The 4-vinylpyrimidin-2-one nucleoside (T-vinyl) forms a cross-link with the RNA containing uracil at the complementary site at a high reaction rate. To obtain the stable T-vinyl derivative so that its reactivity is protected until it access to the target site, several derivatives were investigated, and the 2-thiopyridinyl- and 2-thiopyrimidinyl T-vinyl derivatives were determined to be good candidates. The 2-thiopyrimidinyl T-vinyl derivative was found to more efficiently cross-link with mRNA albeit having a better stability than the 2-thiopyridinyl T-vinyl derivative. The investigation using the luciferase (Luc) mRNA, the synthetic mRNA and non-cellular translation system revealed that the translation is terminated at the end of the cross-linked duplex between the mRNA and the oligoribonucleotide (ORN). Thus, the 2-thiopyrimidinyl T-vinyl derivative has successfully demonstrated both a good stability and high efficiency for the cross-linking reaction, and expanded its applicability in biological applications.

DOI： 10.1248/cpb.c19-00337

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記述言語：英語   掲載種別：論文集(書籍)内論文   出版者・発行元：Springer Singapore  

The sequence-specific triplex formation against duplex DNA offers a potential basis for genome targeting technology, such as diagnostics, regulation of gene expression and sequencing technologies. In an antiparallel triplex DNA, a purine-rich triplex forming oligonucleotide (TFO) consisting of a dG, dA or T forms two reverse Hoogsteen hydrogen bonds with a GC, AT or AT base pair of the duplex DNA, respectively, with a high selectivity in a sequence specific manner. However, there is no natural nucleoside which can recognize the inverted CG and TA base pair of the duplex DNA. Therefore, the development of recognition molecules for the CG and TA inversion sites with a high stability and selectivity has been demanded for the triplex forming technology. In this chapter, we describe the design and synthesis of W-shaped nucleoside analogues (WNA-βT) and pseudo-dC derivatives (MeAP-ΨdC) for selective recognition of the TA and CG base pair, respectively, to expand the triplex-forming sequence.

DOI： 10.1007/978-981-13-1912-9_15

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：OXFORD UNIV PRESS  

The antigene strategy based on site-specific recognition of duplex DNA by triplex DNA formation has been exploited in a wide range of biological activities. However, specific triplex formation is mostly restricted to homo-purine strands within the target duplex DNA, due to the destabilizing effect of CG and TA inversion sites where there is an absence of natural nucleotides that can recognize the CG and TA base pairs. Hence, the design of artificial nucleosides, which can selectively recognize these inversion sites with high affinity, should be of great significance. Recently, we determined that 2-amino-3-methylpyridinyl pseudo-dC ((3Me)AP-Psi dC) possessed significant affinity and selectivity toward a CG inversion site and showed effective inhibition of gene expression. We now describe the design and synthesis of new modified aminopyridine derivatives by focusing on small chemical modification of the aminopyridine unit to tune and enhance the selectivity and affinity toward CG inversion sites. Remarkably, we have newly found that 2-amino-4-methoxypyridinyl pseudo-dC ((4OMe)AP-Psi dC) could selectively recognize the CG base pair in all four adjacent base pairs and form a stable triplex structure against the promoter sequence of the human gene including multiple CG inversion sites.

DOI： 10.1093/nar/gky704

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PHARMACEUTICAL SOC JAPAN  

The antiparallel triplex DNA is formed by the interaction between purine-rich triplex forming oligonucleotides (TFOs) and the homo-purine region within a duplex DNA. The formation of such a structure with the genome DNA promises to control the gene expression in a living cell. In this study, in an attempt to enhance the stability of the triplex DNAs, we have designed the N-2-arylated deoxyguanosine derivatives. Among these analogues, we found that the TFOs containing N-2-phenyl-2'-deoxyguanosine (PhdG) showed a stable and selective triplex DNA formation with the GC base pair as compared to the natural dG/GC triplet. However, the multiple incorporation of PhdG into the TFOs hampered the stable triplex DNA, instead, showed a tendency to form a higher order structure. Therefore, we concluded that the stable and selective triplex DNA formation is expected by the replacement of dG by PhdG in the purine-rich TFO sequence.

DOI： 10.1248/cpb.c18-00043

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記述言語：日本語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SOC SYNTHETIC ORGANIC CHEM JPN  

Triplex DNA is a powerful tool for the genome researches and genome targeting technologies. However, there is an intrinsic limitation to form the stable triplex DNA at any duplex DNA sequence. Recently, we have developed the artificial nucleoside analogue to recognize the CG base pair that cannot be stabilized by natural nucleic acids. Interestingly, the artificial nucleic acid was designed on the basis of the natural T-CG triplex formation. Consequently, we succeeded in expanding the triplex DNA recognition code and achieved the inhibition of the gene expression by the novel antigene triplex forming oligonucleotides.

DOI： 10.5059/yukigoseikyokaishi.76.482

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

Triplex formation against a target duplex DNA has the potential to become a tool for the genome research. However, there is an intrinsic restriction on the duplex DNA sequences capable of forming the triplex DNA. Recently, we demonstrated the selective formation of the stable antiparallel triplexes containing the CG inversion sites using the 2'-deoxy-1-methylpseudocytidine derivative (Psi dC), whose amino group was conjugated with the 2-aminopyridine at its 5-position as an additional hydrogen bonding unit (AP-Psi dC). The 1-N of 2-aminopyridine was supposed to be protonated to form the hydrogen bond with the guanine of the CG inversion site. In this study, to test the effect of the 3-substitution of the 2-aminopyridine unit of AP-Psi dC on the triplex stability, we synthesized the 3-halogenated 2-aminopyridine derivatives of AP-Psi dC. The pKa values 1-N of the 2-aminopyridine unit of AP-Psi dC as the monomer nucleoside were determined to be 6.3 for 3-CH3 ((Me)AP-Psi dC), 6.1 for 3-H (AP-Psi dC), 4.3 for 3-Cl ((Cl)Ap-Psi dC), 4.4 for 3-Br ((Br)AP-Psi dC), and 4.7 for 3-I ((I)AP-Psi dC), suggesting that all the halogenated AP-Psi dCs are not protonated under neutral conditions. Interestingly, although the recognition selectivity depends on the sequence context, the TFO having the sequence of the 3'-G-((I)AP-Psi dC)-A-5' context showed the selective triplex formation with the CG inversion site. These results suggest that the protonation at the 1-N position plays an important role in the stable and selective triplex formation of AP-Psi dC derivatives in any sequences. (C) 2017 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmc.2017.05.035

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記述言語：中国語   出版者・発行元：PHARMACEUTICAL SOC JAPAN  

8-Oxo-2'-deoxyguanosine (8-oxo-dG) is a representative of nucleoside damage, which is generated by the reaction of the 8 position of dG with reactive oxygen species. Abundant 8-oxo-dG in DNA exhibits genotoxicity and has been linked to aging and disease, such as cancer. As the metabolism of cancer cells is much faster than that of normal cells, the oxidized product of the oligonucleotides and the nucleotide pool produces 8-oxo-dG and 8-oxo-2'-deoxyguanosine triphosphate (8-oxo-dGTP), respectively. Human oxoguanine glycosylase (hOGG1) shows base excision activity for 8oxo-dG in duplex DNA. On the other hand, human mutT homologue protein (hMTH1, also known as NUDT1) is important for oxidized nucleotide removal including 8-oxo-dGTP, and it is reported that the presence of hMTH1 is not essential for normal cells but is required for the survival of cancer cells. Therefore, we designed and synthesized 8 halogenated 7-deaza-2'-deoxyguanosine triphosphate (8-halo-7-deaza-dGTP) derivatives as mimics of 8-oxo-dGTP in order to interact with hMTH1. The 8-halo-7-deaza-dGTP derivatives were poor substrates for but strong binders to hMTH1. Interestingly, they exhibited strong competitive inhibition of hMTH1 in the hydrolysis of 8-oxo-dGTP. This inhibitory effect is caused by the slower rate of hydrolysis due to possible small enzyme structural changes. Although the detailed inhibition mechanism of the hydrolysis activity of hMTH1 is unknown, this result is the first to demonstrate the potential of nucleoside triphosphate derivatives as antitumor agents.

DOI： 10.1248/yakushi.16-00231-2

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-V C H VERLAG GMBH  

The 2,13-dimethoxy[5] helicene-spermine ligand 8b possesses an axial chirality. The racemic 8b was bound to B-DNA by the accompanying induction of its (P)-chirality together with the B-to-Z helicity change of the duplex DNA, [(dC-dG)(3)](2). The (P)-chirality of the bound 8b, in turn, transitioned to the (M)-chirality according to the Zhelicity of the DNA. These results illustrate the chirality synchronization between the DNA and the ligand.

DOI： 10.1002/chem.201605276

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1248/cpb.c16-00310

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-V C H VERLAG GMBH  

The sequence-specific formation of triplex DNA offers a potential basis for genome-targeting technologies. In an antiparallel triplex DNA, the sequence-specificity is established by the formation of specific base triplets (G-GC, A-AT, and T-AT) between a triplex-forming oligonucleotide (TFO) and a duplex DNA. However, there are no natural nucleosides that can selectively recognize the inverted CG and TA base pairs. Therefore, the recognition of the CG and TA inversion sites to form a stable triplex DNA has been a long-standing goal for the triplex-forming technology. We now describe the design and synthesis of pseudo-deoxycytidine (Psi dC) derivatives for selective recognition of the CG base pair to expand the triplex-forming sequence. The aminopyridine-bearing Psi dC derivatives showed high selectivity and affinity toward the CG base pair in all neighboring base contexts. Remarkably, 3-methyl-2-aminopyridinyl-Psi dC ((Me)AP-Psi dC) formed a stable triplex with the promoter sequence of the hTERT gene containing four CG inversion sites, and effectively inhibited its transcription in human cancer cells. Thus, (Me)AP-Psi dC is expected to serve as a new starting point of triplex-forming oligonucleotides for a wide variety of genome-targeting applications.

DOI： 10.1002/anie.201606136

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

8-OxodG (8-oxo-2'-deoxyguanosine) is representative of nucleoside damage and shows a genotoxicity. To significantly reveal the contributions of 7-NH and C8-oxygen to the mutagenic effect of 8-oxodG by DNA polymerases, we evaluated the effects of the 8-halo-7-deaza-dG (8-halogenated 7-deaza-2'-deoxyguanosine) derivatives by DNA polymerases. 8-Halo-7-deaza-dGTPs were poorly incorporated by both KF (exo) and human DNA polymerase beta opposite dC or dA into the template DNA. Furthermore, it was found that KF(exo) was very sensitive to the introduction of the C8-halogen, while polymerase beta can accommodate the C8-halogen resulting in an efficient dCTP insertion opposite the 8-halo-7-deaza-dG in the template DNA. These results indicate that strong hydrogen bonding between 7-NH in the 8-oxo-G nucleobase and 1-N in the adenine at the active site of the DNA polymerase is required for the mutagenic effects. Whereas, I-deaza-dGTP shows an antiproliferative effect for the HeLa cells, suggesting that it could become a candidate as a new antitumor agent. (C) 2016 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmc.2016.06.030

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-V C H VERLAG GMBH  

hMTH1 (8-oxo-2-deoxyguanine triphosphatase) hydrolyzes oxidized nucleoside triphosphates; its presence is non-essential for survival of normal cells but is required for survival of cancer cells. In this study, 8-halogenated-7-deaza-2-deoxyguanosine triphosphate (8-halogenated-7-deazadGTP) derivatives were synthesized. Interestingly, these triphosphates were poor substrates for hMTH1, but exhibited strong competitive inhibition against hMTH1 at nanomolar levels. This inhibitory effect is attributed to slower rate of hydrolysis, possibly arising from enzyme structural changes, specifically different stacking interactions with 8-halogenated-7-deazadGTP. This is the first example of using nucleotide derivatives to inhibit hMTH1, thus demonstrating their potential as antitumor agents.

DOI： 10.1002/cbic.201500589

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

Recent studies indicate that oxidative damage to RNA results in dysfunction of translation and eventual pathogenesis. A representative oxidized base in RNA is 8-hydroxyguanosine (8-oxo-rG), however, unlike its DNA counterpart (8-oxo-dG), its role in pathogenesis has not attracted much attention until recently. The 2'-deoxyadenosine derivative with a diazaphenoxazine skeleton at the 6 -amino group (Adap) was shown to be selective for 8-oxo-dG in DNA. In this study, the 2'-0-methoxy derivative of Adap (2'-0MeAdap) was designed as a selective molecule for 8-oxo-rG in RNA. 8-Oxo-rG in the homopurine RNA was selectively recognized by the ODN probe incorporating Adap. In contrast, although it was not possible by the Adap-containing ODN prove due to the instability of the corresponding duplex, 8-oxorG in homopyrimidine RNA was selectively detected by the 2'-0MeRNA probe incorporating 2'-0MeAdap. (C) 2016 Published by Elsevier Ltd.

DOI： 10.1016/j.bmc.2016.02.001

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PHARMACEUTICAL SOC JAPAN  

This paper reports the preparation of 14-mer triplex-forming oligonucleotides (TFOs) containing a 2-O-methyl-1-beta-phenyl-alpha-propargyl-ribose unit, which was conjugated with azide-modified molecules via a click reaction. Modification of these TFOs with pyrene assisted triplex formation, improving the stability of the triplex DNA and the anti-proliferative effects against A549 cells.

DOI： 10.1248/cpb.c15-00570

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.bmcl.2015.05.064

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-V C H VERLAG GMBH  

Cellular DNA continuously suffers various types of damage, and unrepaired damage increases disease progression risk. 8-Oxo-2-deoxyguanine (8-oxo-dG) is excised by repair enzymes, and their analogues are of interest as inhibitors and as bioprobes for study of these enzymes. We have developed 8-halogenated-7-deaza-2-deoxyguanosine derivatives that resemble 8-oxo-dG in that they adopt the syn conformation. In this study, we investigated their effects on Fpg (formamidopyrimidine DNA glycosylase) and hOGG1 (human 8-oxoguanine DNA N-glycosylase 1). Relative to 8-oxo-dG, Cl- and Br-deaza-dG were good substrates for Fpg, whereas they were less efficient substrates for hOGG1. Kinetics and binding experiments indicated that, although hOGG1 effectively binds Cl- and Br-deaza-dG analogues with low K-m values, their lower k(cat) values result in low glycosylase activities. The benefits of the high binding affinities and low reactivities of 8-oxo-dG analogues with hOGG1 have been successfully applied to the competitive inhibition of the excision of 8-oxoguanine from duplex DNA by hOGG1.

DOI： 10.1002/cbic.201402690

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-V C H VERLAG GMBH  

Non-natural RNA modifications have been widely used to study the function and structure of RNA. Expanding the study of RNA further requires versatile and efficient tools for site-specific RNA modification. We recently established a new strategy for the site-specific modification of RNA based on a functionality-transfer reaction between an oligodeoxynucleotide (ODN) probe and an RNA substrate. 2-Deoxy-6-thioguanosine was used to anchor the transfer group, and the 4-amino group of cytosine or the 2-amino group of guanine was specifically modified. In this study, 2-deoxy-4-thiothymidine was adopted as a new platform to target the 6-amino group of adenosine. The (E)-pyridinyl vinyl keto transfer group was attached to the 4-thioT in the ODN probe, and it was efficiently and specifically transferred to the 6-amino group of the opposing adenosine in RNA in the presence of CuCl2. This method expands the available RNA target sites for specific modification.

DOI： 10.1002/cbic.201500084

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-V C H VERLAG GMBH  

The adenosine derivative of 2-oxo-1,3-diazaphenoxazine (Adap) exhibits a superb ability to recognize and form base pairs with 8-oxo-2'-deoxyguanosine (8-oxo-dG) in duplex DNA. In this study, the triphosphate of Adap (dAdapTP) was synthesized and tested for single nucleotide incorporation into primer strands using the Klenow Fragment. The efficiency of dAdapTP incorporation into 8-oxo-dG-containing templates was more than 36-fold higher than with dG-containing templates, and provides better discrimination than does the incorporation of natural 2'-deoxyadenosine triphosphate (dATP). The selective incorporation of dAdapTP into 8-oxo-dG templates was therefore applied to the detection of 8-oxo-dG in human telomeric DNA sequences extracted from H2O2-treated HeLa cells. The enzymatic incorporation of dAdapTP into 8-oxo-dG-containing templates may provide a novel basis for sequencing oxidative DNA damage in the genome.

DOI： 10.1002/anie.201412086

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-V C H VERLAG GMBH  

Sequence-specific recognition of duplex DNA mediated by triple helix formation offers a potential basis for oligonucleotide therapy and biotechnology. However, triplex formation is limited mostly to homopurine strands, due to poor stabilization at CG or TA base pairs in the target duplex DNA sequences. Several non-natural nucleosides have been designed for the recognition of CG or TA base pairs within an antiparallel triplex DNA. Nevertheless, problems including low selectivity and high dependence on the neighboring bases remain unsolved. We thus synthesized N-2-arylmethyl isodC derivatives and incorporated them into triplex-forming oligonucleotides (TFOs) for the selective recognition of the CG base pair within antiparallel triplex DNA. It was shown that an isodC derivative bearing a 2-amino-6-methylpyridine moiety (AP-isodC) recognizes the CG base pair with high selectivity in antiparallel triplex DNA irrespective of the flanking base pairs.

DOI： 10.1002/cbic.201402328

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ELSEVIER SCI LTD  

Pancreatic beta cells secrete insulin in response to glucose levels and thus are involved in controlling blood glucose levels. A line of pancreatic beta cells "MIN6" has been used in studies related to the function of beta cells and diabetes therapy. Regulating gene expression in MIN6 cells could accelerate these studies, but an efficient method for the transfection of nucleic acids targeted to MINE cells is required. We report here on a liposome-based carrier targeted to pancreatic beta cells (Multifunctional envelope-type nano device for pancreatic beta cells, beta-MEND). We identified a lipid composition for use in preparing the beta-MEND, which permits the particles to be efficiently internalized into MIN6, as evidenced by flow cytometry analyses. Intracellular observation by confocal laser scanning microscopy showed that the beta-MEND efficiently delivered the oligo nucleic acids to the cytosol of MINE cells. Moreover, using a beta-MEND encapsulating a 2'-O-Methyl RNA complementary to a microRNA that suppresses insulin secretion, the knockdown of the targeted microRNA and an up-regulation of insulin secretion were observed in MINE. Thus, the beta-MEND holds promise as an efficient system for delivering nucleic acids targeted to MIN6 and can contribute to research and therapy aimed at diabetes. (C) 2014 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.biomaterials.2014.04.017

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記述言語：英語   出版者・発行元：AMER CHEMICAL SOC  

DOI： 10.1021/bc500145s

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

8-Oxo-2'-deoxyguanosine (8-oxo-dG) is a nucleoside resulting from oxidative damage and is known to be mutagenic. 8-Oxo-dG has been related to aging and diseases, including neurological disorders and cancer. Recently, we reported that a fluorescent nucleoside derivative, adenosine-1,3-diazaphenoxazine (Adap), forms a stable base pair with 8-oxo-dG in DNA with accompanying efficient quenching. In this study, a new Adap derivative having an additional 2-amino group on the adenosine moiety (2-amino-Adap) was designed with the anticipation of additional hydrogen bonding with the 8-oxo group of 8-oxo-dG. The properties of the ODN containing 2-amino-Adap were evaluated by measuring thermal stability and fluorescence quenching. In contrast to the previously designed Adap, the base-pairing and fluorescence quenching properties of 2-amino-Adap varied depending on the ODN sequence, and there was no clear indication of an additional hydrogen bond with 8-oxo-dG. Instead, the base pairing of 2-amino-Adap with dG was significantly destabilized compared with that of Adap with dG, resulting in improved selectivity for 8-oxo-dG in the human telomere DNA sequence. Thus, the telomere-targeting ODN probe containing 2-amino-Adap displayed selective, sensitive and quantitative detection of 8-oxo-dG in the human telomere DNA sequence in a light-up detection system using SYBR Green. (C) 2014 Published by Elsevier Ltd.

DOI： 10.1016/j.bmc.2014.01.024

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

8-Halogenated-7-deaza-2'-deoxyguanosines (8-halo-7-deaza-dG) were designed to structurally mimic 8-oxo-2'-deoxyguanosine (8-oxo-dG), which is representative of an oxidized nucleoside. It has been shown by NMR that the conformation around the N-glycosidic bond of (8-halo-7-deaza-dG) is preferably syn, similar to 8-oxo-dG. The base pairing properties of 8-halo-7-deaza-dG were studied by measuring the thermal denaturation temperature of the duplexes, showing that their base pair with dC is destabilized compared with natural dG. These results also support their preference for syn conformation. Unlike 8-oxo-dG, 8-halo-7-deaza-dG did not form a stable base pair with dA, most likely due to the lack of N7-H hydrogen bonding with dA. In conclusion, the newly-designed 8-halo-7-deaza-dG analogs resemble 8-oxo-dG in its shape and preference for syn conformation, but they do not form Hoogsteen base pair with the opposing dA. (C) 2014 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.tet.2014.01.047

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：OXFORD UNIV PRESS  

Modified nucleosides in natural RNA molecules are essential for their functions. Non-natural nucleoside analogues have been introduced into RNA to manipulate its structure and function. We have recently developed a new strategy for the in situ modification of RNA based on the functionality transfer reaction between an oligodeoxynucleotide probe and an RNA substrate. 2'-Deoxy-6-thioguanosine (6-thio-dG) was used as the platform to anchor the transfer group. In this study, a pyridinyl vinyl ketone moiety was newly designed as the transfer group with the expectation that a metal cation would form a chelate complex with the pyridinyl-2-keto group. It was demonstrated that the (E)-pyridinyl vinyl keto group was efficiently and specifically transferred to the 4-amino group of the opposing cytosine in RNA in the presence of NiCl2 with more than 200-fold accelerated rate compared with the previous system with the use of the diketo transfer group. Detailed mechanistic studies suggested that NiCl2 forms a bridging complex between the pyridinyl keto moiety and the N7 of the purine residue neighboring the cytosine residue of the RNA substrate to bring the groups in close proximity.

DOI： 10.1093/nar/gku538

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：OXFORD UNIV PRESS  

The formation of interstrand cross-links in nucleic acids can have a strong impact on biological function of nucleic acids; therefore, many cross-linking agents have been developed for biological applications. Despite numerous studies, there remains a need for cross-linking agents that exhibit both efficiency and selectivity. In this study, a 4-vinyl-substituted analog of thymidine (T-vinyl derivative) was designed as a new cross-linking agent, in which the vinyl group is oriented towards the Watson-Crick face to react with the amino group of an adenine base. The interstrand cross-link formed rapidly and selectively with a uridine on the RNA substrate at the site opposite to the T-vinyl derivative. A detailed analysis of cross-link formation while varying the flanking bases of the RNA substrates indicated that interstrand cross-link formation is preferential for the adenine base on the 5'-side of the opposing uridine. In the absence of a 5'-adenine, a uridine at the opposite position underwent cross-linking. The oligodeoxynucleotides probe incorporating the T-vinyl derivative efficiently formed interstrand cross-links in parallel-type triplex DNA with high selectivity for dA in the homopurine strand. The efficiency and selectivity of the T-vinyl derivative illustrate its potential use as a unique tool in biological and materials research.

DOI： 10.1093/nar/gkt197

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ELSEVIER  

Clinical use of arsenic trioxide (As2O3), which can induce the remission of relapsed or refractory acute promyelocytic leukemia, is often limited because of its cardiotoxicity. Symptoms of cardiotoxicity include acute cardiac conduction disturbances, such as QT prolongation. The present study was undertaken to evaluate the effects of alpha-lipoic acid (LA) on acute As2O3-induced ECG abnormalities (QTc interval prolongation) in anesthetized guinea pigs. Intravenous injection of As2O3 in guinea pigs caused QTc interval prolongation, which was significantly attenuated by co-treatment with LA (0.35, 3.5 and 35 mg/kg) in a dose-dependent manner. In isolated guinea pig cardiomyocytes, the decrease in I-Ks current induced by As2O3 (1 mu M) was rapidly restored to the basal level by the addition of LA (10 mu M). Consistent with this finding, the As2O3-induced QTc interval prolongation was also improved rapidly by post-treatment with LA in guinea pigs. Electrospray ionization time-of-flight mass spectrometry analysis detected an expected peak of arsenic-LA complex in vitro, indicating that LA and As2O3 form a new compound in vivo. In addition, pre-treatment with a chelating agent, British anti-Lewisite (BAL, 3.5 or 35 mg/kg), also attenuated the As2O3-induced QTc interval prolongation. In this study, co-and post-treatments with LA and pre-treatment with BAL ameliorated As2O3-induced acute QT prolongation in anesthetized guinea pigs. Because LA and probably BAL may bind to As2O3, these agents may exert protective effects through their chelating activity. Further studies are needed to determine whether LA is beneficial as a prophylactic or rescue agent for acute promyelocytic leukemia patients treated with As2O3. (C) 2013 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.ejphar.2013.02.027

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

We have previously developed the innovative bicyclic nucleoside analogues (WNA) for the formation of the triplex DNA. The WNA analogue consists of an aromatic ring and a recognition base on the bicyclic skeleton, and the recognition of the CG or TA interrupting sites has been achieved by the WNA analogues. However, the stabilization ability of the WNA analogue is dependent on its neighboring nucleobases within the TFO. We hypothesized that the sequence dependency might arise from the fixed conformation of the bicyclic ring of the WNA. Thus, it was expected that an open-linker between the sugar part and the nucleobase might produce the flexibility and improve the stabilizing effect of the nucleobase analogues. We now report the design and synthesis of a new nucleoside analogue as an open-form of WNA-beta T, the 1'-phenyl-2'-OMe-ribose derivative, connecting the thymine base to the ribose part through a methylene linker (1) or an ethylene linker (2). TFO containing the 3'-dA-1-dG context recognized the CG interrupting site, and that with the 3'-dG-1-dG context recognized the GC site. In contrast, 2 displayed a stabilizing effect on all four base pairs with some preferences for the TFO containing 3'-dA-2-dG and 3'-dG-2-dG. These results suggested that a flexible linker between the nucleobase and the ribose part may improve the sequence dependency for the triplex formation. (c) 2012 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.tet.2012.11.016

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：TAYLOR & FRANCIS INC  

6-O-7-N-Bis(diphenylcarbamoyl)-2-N-phenoxyacetyl-5'-O-dimethoxytrityl-2'-O-{[(triisopropyl-silyl)oxy]methyl}-8-oxoguanosine-3'-yl-beta-cyanoethyl-N,N-diisopropylphosphoramidite (5) was synthesized as a new phosphoramidite precursor unit for the synthesis of RNA. Compound 5 was successfully incorporated into the middle of the RNA sequences, and the synthesized RNAs were identified by MALDI-TOF mass measurements. Their properties were evaluated for formation of the RNA duplex and RNA/DNA heteroduplex. ORNs 1 and 4 containing 8-oxo-G can form base pairs with rC or dC in an anti conformation, while it can also interact with rA or dA in a syn conformation in the RNA duplex or RNA/DNA heteroduplex. The described synthetic method is therefore a useful procedure for the synthesis of ORN containing 8-oxo-G and contributes to the study of 8-oxo-G in RNA.

DOI： 10.1080/15257770.2013.767461

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ROYAL SOC CHEMISTRY  

The development of novel nucleoside analogues for the formation of triplex DNA containing pyrimidine-purine inversion sites has been a challenging field. In this paper, we describe the design and synthesis of non-natural nucleoside analogues, N-substituted-2'-deoxy-5-methylisocytidine derivatives, and their evaluation for triplex formation. It has been shown that N-(guanidinoethyl)-2'-deoxy-5-methylisocytidine exhibits selective recognition of a CG interrupting site and potentiates the formation of antiparallel triplexes.

DOI： 10.1039/c3ob40472b

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：TAYLOR & FRANCIS INC  

(1S,3S,4R)-1-Phenyl-1-thymidyl-3-hydroxy-4-hydroxymethylcyclopentane (10) and their analogs were synthesized, incorporated into the oligodeoxynucleotides, and their properties were evaluated for the formation of duplex and triplex DNA. The known chiral cyclopentanone derivative was converted into the corresponding ketimine sulfonamide derivative, which was subjected to a stereoselective PhLi addition. The formed sulfonamide was hydrolyzed to afford the primary amino group, on which the thymine moiety was built. The benzyl protecting groups were removed to form the nucleoside analog having a phenyl group and the thymine unit at the 1' position of a carbocyclic skeleton (10). In the estimation of the oligodeoxynucleotides incorporating 10 for duplex and triplex formation, the carbocyclic nucleoside analog 10 did not show the stabilizing effect for duplex formation; on the other hand, it stabilized the triplex. Therefore, the skeleton of the phenyl-substituted carbocyclic nucleoside analog 10 may be a platform for the formation of stable triplex DNA.

DOI： 10.1080/15257770.2012.737970

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-V C H VERLAG GMBH  

Molecules that can target duplex DNA with sequence selectivity have the potential to be useful tools in genomic research and also as therapeutic agents. Homopurinehomopyrimidine stretches in duplex DNA can be recognized by homopurine or homopyrimidine TFOs (triplex-forming oligonucleotides) through the formation of triplex DNA. We have previously developed bicyclic nucleoside analogues (WNAs) for the formation of stable triplexes in the formation of stable antiparallel triplexes containing a TA or a CG interrupting site. In this study, we investigated the effects on triplex DNA formation of ortho-, meta-, and para-methyl substituent groups on the aromatic ring of the WNA analogue. It was found that the homopurine TFO containing meta- and para-methyl-substituted WNA-beta T (mMe-WNA-beta T, pMe-WNA-beta T) stabilized triplexes containing a TA interrupting site or a GC site, respectively. Interestingly, the ortho-methyl-substituted WNA-beta T (oMe-WNA-beta T) efficiently promoted DNA strand displacement to form the TFO/pyrimidine duplex. A detailed investigation showed that the duplex was in the antiparallel orientation and that its formation took place prior to triplex formation with the need for a magnesium cation. NOESY measurements indicated a significant difference in the rotation flexibilities of the phenyl rings of WNA-beta Ts: that is, the conformation of the ortho-methylated phenyl ring was stable in a temperature-independent manner. It was speculated that the initial formation of a ternary complex was followed by strand displacement and then the formation of the TFO/pyrimidine duplex together with the TFO2/pyrimidine triplex formation during the early stage, and that the equilibrium shifted to the triplex during the later stage. Although the detailed role is still uncertain, the fixed phenyl ring of oMe-WNA-beta T might play a role in the displacement reaction.

DOI： 10.1002/cbic.201200066

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER CHEMICAL SOC  

Repair glycosylases locate and excise damaged bases from DNA, playing central roles in preservation of the genome and prevention of disease. Two key glycosylases, Fpg and hOGG1, function to remove the mutagenic oxidized base 8-oxoG (OG) from DNA. To investigate the relative contributions of conformational preferences, leaving group ability, enzyme-base hydrogen bonding, and nucleobase shape on damage recognition by these glycosylases, a series of four substituted indole nucleosides, based on the parent OG nonpolar isostere 2Cl-4F-indole, were tested as possible direct substrates of these enzymes in the context of 30 base pair duplexes paired with C. Surprisingly, single-turnover experiments revealed that Fpg-catalyzed base removal activity of two of the nonpolar analogs was superior to the native OG substrate. The hOGG1 glycosylase was also found to catalyze removal of three of the nonpolar analogs, albeit considerably less efficiently than removal of OG. Of note, the analog that was completely resistant to hOGG1-catalyzed excision has a chloro-substituent at the position of NH7 of OG, implicating the importance of recognition of this position in catalysis. Both hOGG1 and Fpg retained high affinity for the duplexes containing the nonpolar isosteres. These Studies show that hydrogen bonds between base and enzyme are not needed for efficient damage recognition and repair by Fpg and underscore the importance of facile extrusion from the helix in its damaged base selection. In contrast, damage removal by hOGG1 is sensitive to both hydrogen bonding groups and nucleobase shape. The relative rates of excision of the analogs with the two glycosylases highlight key differences in their mechanisms of damaged base

DOI： 10.1021/ja208510m

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

We have recently reported that Adap (adenosine-1,3-diazaphenoxazine) is an artificial nucleoside analogue for the specific recognition by multiple hydrogen bonding and that its fluorescence is selectively quenched with 8-oxo-2'-deoxyguanosine (8-oxo-dG) in DNA. We now report the development of a new OFF-to-ON type FRET probe, in which one strand contains Adap and another contains natural nucleotides for the formation of a less stable double strand. Each strand was labeled with Cy3 or BHQ2 at the 5'-end or 3'-end, respectively. It was expected in this system that fluorescence of the duplex probe is first quenched by FRET, but the target DNA strand containing 8-oxo-dG at the complementary site of Adap would enhance the displacement reaction of the less stable duplex probe that results in the fluorescence recovery. The results showed that the duplex probe containing the Adap-T base pair exhibited a complete discrimination between 8-oxo-dG and dG in DNA by fluorescence enhancement. (C) 2011 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmcl.2011.10.093

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Blackwell Publishing Inc.  

Chemically modified oligonucleotides play a significant role for genomic research. Modified nucleosides, such as with a fluorescent dye, can be obtained by chemical synthesis. Site-specifically modified long nucleic acids are obtained by ligation of chemically modified short oligonucleotides with enzyme, photochemistry, or catalytic DNA. The functionality-transfer ODN (FT-ODN), which contains 2'-deoxy-6-thioguanosine (6-thio-dG) functionalized with the 2-methyliden-1,3-diketone group, is hybridized with the target RNA to trigger the selective functionalization of the 4-amino group of the cytosine base at pH 7 or the 2-amino group of the guanine base at pH 9.4 or at pH 7.4 in the presence of NiCl2. In particular, the functionality-transfer reaction (FTR) under the alkaline conditions or neutral conditions in the presence of NiCl2 proceeds rapidly and selectively to lead to the modification of the target guanine. The transfer reaction of the acetylene-containing diketone group produces the acetylene-modified RNA, which can be subjected to the Cu(I)-catalyzed "click chemistry" with a variety of azide compounds for highly specific, internal modification of RNA. © 2012 by John Wiley &amp
Sons, Inc.

DOI： 10.1002/0471142700.nc0449s48

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ROYAL SOC CHEMISTRY  

Triplex forming oligonucleotides (TFOs) are some of the most promising tools in the antigene strategy for the development of gene targeting therapeutics. However, the stable triplex formation is restricted to the homopurine sequences consisting of purine nucleosides, dG and dA. Therefore, the T or dC nucleoside in the homopurine strand inhibits the stable triplex formation. We have developed W-shaped nucleoside analogues (WNAs) for the formation of the unnatural type triplex DNA, with sequences containing the interrupting site in an antiparallel triplex formation. In the present study, we tested the antigene effect of TFOs having WNA-beta T, which increased the stability of the triplex formation with a target sequence including the TA interrupting site. We designed the GU TFO (WNA) and GU TFO (natural) for targeting sequences of the Bcl-2 or survivin oncogene. The gel shift assay showed that the TFO (WNA) formed more stable triplexes than the natural TFO. Remarkably, the Bcl-2- or survivin-targeted TFO (WNA) inhibited the cell proliferation and induced a caspase-dependent apoptosis. It was confirmed that the survivin-targeted TFO (WNA) more effectively decreased the number of survivin products in the A549 cell than the natural TFOs.

DOI： 10.1039/c2ob26431e

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ROYAL SOC CHEMISTRY  

O-6-Methyl-20-deoxyguanosine (O-6-Me-dG) is a mutagenic nucleotide in DNA. O-6-Me-dG in DNA was rapidly and selectively modified by a functionality transfer reaction using the ODN incorporating 6-S-functionalized thioguanosine. Subsequent labelling of O-6-Me-dG with the fluorescent FAM or biotin group via click chemistry has permitted the sensitive and selective detection of O-6-Me-dG in DNA.

DOI： 10.1039/c2cc17621a

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記述言語：英語   出版者・発行元：MARY ANN LIEBERT INC  

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記述言語：英語   出版者・発行元：AMER CHEMICAL SOC  

DOI： 10.1021/ja204536k

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER CHEMICAL SOC  

The selective detection of 8-oxo-2'-deoxyguanosine (8-oxo-dG) in DNA without chemical or enzymatic treatment is an attractive tool for genomic research. We designed and synthesized the non-natural nucleoside analogue, the adenosine-1,3-diazaphenoxazine (Adap) derivative, for selective recognition of 8-oxo-dG in DNA. This study clearly showed that Adap has a highly selective stabilizing effect on the duplex containing the Adap 8-oxo-dG base pair. Furthermore, the fluorescent property of Adap was shown to be useful for the selective detection of 8-oxo-dG in the duplex DNA. To the best of our knowledge, this is the first successful demonstration of a non-natural nucleoside with a high selectivity for 8-oxo-dG in DNA.

DOI： 10.1021/ja200327u

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記述言語：英語   出版者・発行元：ROYAL SOC CHEMISTRY  

As the knowledge of the biological functions of RNA expands, the demand for research tools to investigate intracellular RNA is increasing. Oligonucleotides can be rationally designed for the target RNA sequence, and therefore, have become a reliable platform for the development of specific molecules for RNA. The chemical modification of RNA has a strong impact on RNA research; the fluorescent labeling of RNA is useful to monitor RNA production, processing, relocation in the cell, interaction with other intracellular components and degradation, etc. Chemical modification may affect the RNA function through a variety of pathways, and therefore, would be potentially useful for biological research, therapeutic approach and artificial manipulation of the RNA function. This tutorial review starts with an introduction of the biological relevance of modified RNA, and focuses on the recent progress of the oligodeoxynucleotide probes for the covalent modifications of RNA. The prospects of this new technology are also discussed.

DOI： 10.1039/c1cs15066a

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ROYAL SOC CHEMISTRY  

The internal modification of RNA has been successfully achieved by the functionality transfer reaction (FTR) and following click chemistry with diverse azide compounds. The benefits of the FTR have been demonstrated by its specificity, rapidity, broad applicability, and procedure simplicity.

DOI： 10.1039/c1cc10582e

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

Potent peroxidase-like activity of the beta-ketoenamine (1)-dicopper (II) complex (2) for the chemiluminescence (CL) of luminol either in the presence or absence of H2O2 has been previously demonstrated by our group. In this study, the beta-ketoenamine (1) as the ligand unit for copper(II) was incorporated into the oligonucleotide (ODN) probes. It has been shown that the catalytic activity of the ODN probes conjugating the ligand-Cu(II) complex is activated by hybridization with the target DNA with the complementary sequence. Thus, this study has successfully demonstrated the basic concept for the sensitive detection of nucleic acids by CL based on the template-inductive activation of the catalytic unit for CL. (C) 2010 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmc.2010.10.007

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

We previously reported that oligodeoxynucleotides containing 2-amino-6-vinylpurine (2-AVP: 1) exhibit efficient selective cross-linking to cytosine. In this study, the 2'-OMe nucleoside analogue (2) of 2-AVP was designed in order to increase its affinity to RNA and enhance metabolic stability. It has been demonstrated that 2'-OMe oligonucleotides bearing 2 achieve highly selective cross-linking to the thymine base in DNA and show higher antisense effect on luciferase production in cell lysate. (C) 2010 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmcl.2010.08.027

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記述言語：英語   出版者・発行元：AMER CHEMICAL SOC  

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1021/bc100131j

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Elsevier  

DOI： 10.1016/j.bmc.2010.04.025

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

We have previously reported that the 2-amino-6-vinylpurine (AVP) nucleoside exhibits a highly efficient and selective crosslinking reaction toward cytosine and displayed an improved antisense inhibition in cultured cells. In this study, we further investigated the alkyl-connected AVP nucleoside analogs for more efficient crosslinking to the cytosine base (rC) of the target RNA. We synthesized three AVP analogs which connect the 2-amino-6-vinylpurine unit to the 2'-deoxyribose through a methylene, an ethylene, or a butylene linker. The ODN incorporating the AVP analog with the methylene or the butylene linker showed a slightly higher crosslinking to the target rC of RNA than the original AVP with no linker. In contrast, the AVP with the ethylene linker formed a selective and efficient crosslink to the rC of the target RNA. (C) 2010 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmc.2010.03.008

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：OXFORD UNIV PRESS  

Site-specific modification of RNA is of great significance to investigate RNA structure, function and dynamics. Recently, we reported a new method for sequence- and cytosine-selective chemical modification of RNA based on the functional group transfer reaction of the 1-phenyl-2-methylydene-1,3-diketone unit of the 6-thioguanosine base incorporated in the oligodeoxynucleotide probe. In this study, we describe that the functionality transfer rate is greatly enhanced and the selectivity is shifted to the guanine base when the reaction is performed under alkaline conditions. Detailed investigation indicated that the 2-amino group of the enolate form of rG is the reactant of the functionality transfer reaction. As a potential application of this efficient functionality transfer reaction, a pyrene group as a relatively large fluorescent group was successfully transferred to the target guanine base of RNA with a high guanine and site selectivity. This functionality transfer reaction with high efficiency and high site-selectivity would provide a new opportunity as a unique tool for the study of RNA.

DOI： 10.1093/nar/gkp930

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：WILEY-V C H VERLAG GMBH  

DNA containing alternating purine and pyrimidine repeats has the potential to adopt the Z-DNA structure, one of the well-studied structures besides A- and B-DNA. Despite a number of molecular models that have been proposed to explain the mechanism for B -> Z transition, there is continued discussion on the mechanism and physiological role of this transition. In this study, we have found that the bis(2-naphthyl)-maleimide-spermine conjugate (3c) exhibits a remarkable ability to cause the B -> Z, transition of d(CGCGCG)(2) at low salt concentrations. Using isothermal titration calorimetry (ITC) we show that the B -> Z transition induced by 3c is both enthalpically and entropically favorable. The ligand might effect the dehydration of B-DNA, which leads to the B -> Z transition. Interestingly, an intermediate CD between the B and Z forms was observed in the pH-dependent transition in the presence of the ligand. The unique structure and characteristics of the ligand designed in this investigation will be useful for the study of Z-DNA.

DOI： 10.1002/chem.201000947

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

We have previously developed W-shaped nucleoside analogs (WNA) for recognition of TA and CG interrupting sites, which are the intrinsic limitation for the formation of a stable triplex DNA by the natural triplex-forming oligonucleotide (TFO). However, the stabilization effect of WNA is dependent on the neighboring nucleobases at both sides of the WNA analogs within the TFO. Considering that the base is located at the hindered site constructed of three bases of the target duplex and the TFO, it was expected that replacement of the pyrimidine base of the WNA analog with a smaller pyrazole ring might avoid steric repulsion to produce a greater stability for the triplex. In this study, the new WNA analogs bearing the pyrazole ring, 3-aminopyrazole (AP), and 4-methyl-3-pyrazole-5-on (MP) were synthesized, incorporated into the TFOs, then their stabilizing effects on the triplexes were evaluated. A remarkable success was illustrated by the fact that the TFO containing WNA-beta AP in the 3'G-WNA-G-5' sequence formed a stable triplex with selectivity to the CG interrupting site where the previous WNA-beta C did not induce the triplex formation. (C) 2009 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmc.2009.08.040

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER CHEMICAL SOC  

Efficient methods for the covalent modification of large RNA molecules should find significance utility as innovative biological tools as well as therapeutic methods. In this study, the development of a general method for site-specific RNA modification guided by the functional ODN template has been investigated. The ODN probe containing 6-thioguanosine was modified by the methylenediketone derivative to form the S-functionalized ODN. Site-specific and cytosine-selective RNA modifications were achieved by the functionality-transfer reaction from the sulfur atom of the functionalized probe to the amino group of the cytosine base of the target strand. It was shown that the base and site selectivity were due to the close proximity of the reactants in the DNA-RNA duplexes.

DOI： 10.1021/bc900009p

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

The 8-oxoG-clamp, a specific fluorescent probe for 8-oxo-deoxyguanosine (8-oxo-dG), was incorporated into the oligodeoxynucleotide (ODN) within or at the 3 '-end of the purine and the pyrimidine sequences. Based on the UV-melting temperature, the 8-oxoG-clamp showed slightly lower stabilizing effects on the duplexes containing 8-oxo-dG at the complementary site than that with dG. On the other hand, 8-oxo-dG in DNA was selectively detected by fluorescence quenching of the 8-oxoG-clamp. (C) 2008 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmcl.2008.12.036

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

Efficient methods for the modification of RNA molecules have been expected as innovative biological tools and therapeutic methods. In this study, the development of a general method for site-specific RNA modification by the functionality-transfer ODN probes has been investigated. Site-specific and cytosine-selective RNA modifications were achieved by the functionality-transfer reaction. It was shown that the base and site-selectivity were due to the close proximity of the reactants in the DNA-RNA duplexes.

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

We have previously reported that the 2-amino-6-vinylpurine (AVP) in oligonucleotide (ODN) showed the highly selective and effective cross-linking reaction toward cytosine within target complementary sequences. Recently, we revealed that the PEGylated ODN containing AVP analogues exhibited the effective antisense effect in the cultured cell. However, subsequent studies showed that the cross-linking ability of AVP for cytosine was affected by the target RNA sequences. We therefore developed new intelligent nucleoside analogues connected AVP to sugar through the ethylene linker (et-AVP), that exhibited highly effective cross-linking ability to rC in the target RNA. In this paper, we described the synthesis of the PEGylated ODN containing cross-linking nucleoside analogues and properties of it about RNase H activity and antisense effect in cell lysate. As a result, the PEGylated ODN containing et-AVP showed the effective antisense effect in the non-cell system for targeting the luciferase mRNA by non-RNase-H mechanism.

DOI： 10.1093/nass/nrp084

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：TAYLOR & FRANCIS INC  

6-S-{2-[(2-ethylhexyl)oxycarbonyl]ethyl)}-3',5'-O-bis(tert-butyldimethylsilyl)-2'-deoxy-6-thiogua nosine (2) was synthesized in high yield from the corresponding 6-O-mesitylenesulfonyl derivative by the reaction with 2-ethylhexyl 3-mercapto-propionate. The phosphoramidite precursor derived from 2 was successfully applied to an automated DNA synthesizer to produce 2'-deoxy-6-thioguanosine containing ODN. The results showed that 2-ethylhexyl 3-mercaptopropionate is useful as an odor less reagent and also as an S-protecting group of 2'-deoxy-6-thioguanosine.

DOI： 10.1080/15257770903155576

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

We have previously developed W-shaped nucleoside analogs (WNAs) having a nucleobase and an aromatic ring for the formation of the unnatural triplex DNA. Modification of an aromatic ring of WNA is highly effective regarding the stability of the triplex DNA. In this study, we designed new WNA analogs having the p-aminobenzene as an aromatic ring, which were synthesized via the Curtius-Yamada rearrangement. Based on the evaluation of the triplex formation with p-amino-WNA-TFO, it has been shown that the amino group may produce a non-selective interaction with the phosphate backbone of the target duplexes. These results indicate that the amino modification is useful to overcome the sequence-dependence of the TFO containing WNA analogs. (c) 2008 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.tet.2008.05.096

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記述言語：英語   掲載種別：研究論文（国際会議プロシーディングス）   出版者・発行元：ACAD SCI CZECH REPUBLIC, INST ORGANIC CHEM & BIOCHEMISTRY  

Chemical modifications of DNA or RNA cause significance impact on gene expression. Our group has been investigating artificial strategies to perform an efficient reaction to a target nucleobase within the hybridized complex as an innovative biological tool to manipulate the gene at a single nucleobase level. This study was aimed at developing a useful tool for a specific modification of DNA or RNA by the use of the ODN incorporating S-vinyl thioguanosine analog. It has been demonstrated that the diketone unit is transferred from S-vinyl thioguanosine analog to the amino group of cytidine at the target site of the complementary ODN. The new functional-group transfer reaction may be applicable to any sequences and have potential applications as an innovative method for efficient and selective modification of the target cytidine in DNA and RNA.

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掲載種別：研究論文（学術雑誌）  

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

1'-Phenyl substituted ribonucleoside analogs with all four nucleobases have been synthesized by the conventional N-glycosidation method.

DOI： 10.1016/S0385-5414(07)81201-9

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER CHEMICAL SOC  

A substantial fraction of mutations that arise in the cell comes from oxidative damage to DNA bases. Oxidation of purine bases at the 8-position, yielding 8-oxo-G and 8-oxo-A, results in conformational changes (from anti to syn) that cause miscoding during DNA replication. Here we describe the synthesis and biophysical and biochemical properties of low-polarity shape mimics of 8-oxopurines, and we report that these new analogues exhibit remarkable mimicry of the mutagenic properties of the natural damaged bases. A 2-chloro-4-fluoroindole nucleoside (1) was designed as an isosteric analogue of 8-oxo-dG, and a 2-chloro-4-methylbenzimidazole nucleoside (2) as a mimic of 8-oxo-dA. The nucleosides were prepared by reaction of the parent heterocycles with Hoffer's chlorodeoxyribose derivative. Structural studies of the free nucleosides 1 and 2 revealed that both bases are oriented syn, thus mimicking the conformation of the oxopurine nucleosides. Suitably protected phosphoramidite derivatives were prepared for incorporation into synthetic DNAs, to be used as probes of DNA damage responses, and 5'-triphosphate derivatives (3 and 4) were synthesized as analogues of damaged nucleotides in the cellular nucleotide pool. Base pairing studies in 12-mer duplexes showed that 1and 2 have low affinity for polar pairing partners, consistent with previous nonpolar DNA base analogues. However, both compounds pair with small but significant selectivity for purine partners, consistent with the idea that the syn purine geometry leads to pyrimidine-like shapes. Steady-state kinetics studies of 1 and 2 were carried out with the Klenow fragment of Escherichia coli DNA Pol I (exo(-)) in single-nucleotide insertions. In the DNA template, the analogues successfully mimicked the mutagenic behavior of oxopurines, with 1 being paired selectively with adenine and 2 pairing selectively with guanine. The compounds showed similar mutagenic behavior as nucleoside triphosphate analogues, being preferentially inserted opposite mutagenic purine partners. The results suggest that much of the mutagenicity of oxopurines arises from their shapes in the syn conformation rather than from electrostatic and hydrogen-bonding effects. The new analogues are expected to be generally useful as mechanistic probes of cellular responses to DNA damage.

DOI： 10.1021/ja071970q

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

Efficient and specific targeting of DNA sequences by synthetic ligands is a major goal in chemical biology. Triplex-forming oligonucleotides (TFOs) are sequence-specific DNA-binding agents, but are limited to homopurine/homopyrimidine target sequences. We have previously reported two useful analogues (WNA: W-shaped nucleoside analogues), WNA-bT and WNAbC, which recognize a TA and a CG interrupting site forming triplexes with high stability and selectivity, respectively. However, their ability to form triplexes depended on their neighbouring bases in the TFO. Subsequent studies have shown that the sequence-dependency of the WNA analogues, for the formation of triplexes, has been partially solved by use of a WNA analogue bearing a substituted aromatic ring. Investigations into the effects of the substituted aromatic ring of WNA derivatives on the stability of triplexes led to the discovery of strand invasion by the TFO incorporating the new WNA analogue to form a highly stable duplex.

DOI： 10.1093/nass/nrm128

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掲載種別：研究論文（学術雑誌）  

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

Triplex-forming oligonucleotides (TFOs) are sequence-specific DNA-binding agents, but their recognizable duplexes are limited to homopurine/homopyrimidine sequences because of interruption of pyrimidine base in the purine strand. This problem has not been fully solved despite numerous studies. We have previously reported that the novel nucleoside analogues (WNA: W-shaped nucleoside analogues), WNA-betaT and WNA-betaC, can recognize a TA and a CG interrupting sites to form triplexes with high stability and selectively, respectively. However, further investigations have shown that the triplex formation using the WNA derivatives is dependent on its neighbouring bases of the TFOs. In this study, the new WNA analogues having a variety aromatic ring were synthesized to evaluate effects on sequence dependency. And it was found that changes in the substituent of aromatic ring of WNA derivatives altered the binding selectivity and stability in the triplex formation.

DOI： 10.1093/nass/nrl092

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

Triplex-forming oligonucleotides (TFOs) are powerful tools for genomic research. The most stable triplex is formed by the interaction between TFOs and homopurine/homopyrimidine sequences in the target duplex, but the triplex DNA is hampered by one pyrimidine base in the homopurine tract. Previously, we developed novel nucleoside analogues (WNA: W-shaped nucleoside analogues) to recognize pyrimidine/purine inversion sites (TA or CG interrupting sites) and determined two useful WNA analogues, WNA-betaT and WNA-betaC. However, subsequent study showed that the triplex formation using the WNA analogues was dependent on its neighbouring bases of the TFOs. In this study, the new WNA analogues having a different aromatic ring were synthesized to evaluate effects on sequence dependency. It has been found that o-bromo, m-bromo-, and p-cyano-substituted WNA-betaT derivatives are selective to a TA interrupting site to form triplexes with high stability in the sequences where original WNA-betaT could not recognize.

DOI： 10.1093/nass/49.1.173

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：TAYLOR & FRANCIS INC  

Recently, we have developed new base analogs (WNA) and demonstrated that WNA-beta T with thymine and WNA-beta; C with cytosine stabilize non-natural antiparallel triplexes with a TA or a CG interrupting site, respectively. However, limitations in recognizable sequences with the WNA-containing TFO were also found. The objective of this study is to search better WNA analogs for expansion of triplex recognition codes to general duplex sequences. In this study, we designed new WNA analogs by systematic modification of the aromatic part and the recognition part. The new WNA analogs with the benzene ring substituted with bromide or cyanide have determined for selective stabilization of triplexes at a TA interrupting site, and general formation of triplexes having a TA interrupting site has been achieved.

DOI： 10.1081/NCN-200060309

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER CHEMICAL SOC  

Triplex-forming oligonucleotides (TFOs) are potential DNA-targeting molecules and would become powerful tools for genomic research. As the stabilization of the TFO is partially provided by hydrogen bonds to purine bases, the most stable triplexes form with homopurine/homopyrimidine sequences, and a pyrimidine base in the purine strand of the duplex interrupts triplex formation. If a TFO can recognize sequences including such an interrupting site, the target regions in the genome would be expanded to a greater extent. However, this problem has not been generally solved despite extensive studies. We have previously reported a new base analogue (WNA) constructed of three parts, a benzene ring, a heterocyclic ring, and a bicyclic skeleton to hold these two parts. In this study, we have further investigated modification of WNA systematically and determined two useful WNA analogues, WNA-betaT and WNA-betaC, for selective stabilization of triplexes at a TA and a CG interrupting site, respectively. The triplexes with WNA analogues have exhibited an interesting property in that they are more stable than natural-type triplexes even at low Mg2+ concentration. From comparison of the results with H-WNA-betaT lacking benzene and those with WNA-H without thymine, it has been suggested that benzene is a major contributor for triplex stability and thymine provides selectivity. Thus, it has been successfully demonstrated that WNA-betaT/TA and WNA-betaC/CG combinations may expand triplex recognition codes in addition to the natural A/AT and G/GC base triplet codes. The results of this study will provide useful information for the design of new WNA analogues to overcome inherent problems for further expansion of triplex recognition codes.

DOI： 10.1021/ja037211z

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

Triplex-forming oligonucleotides (TFOs) are potential DNA-targeting molecules, but their recognizable duplexes are limited to homopurine:homopyrimidine sequences by interruption of pyrimidine bases in the purine strand. Despite numerous studies, this problem has not been generally solved. We have recently demonstrated that the new nucleoside analogues, WNA-betaT and WNA-betaC exhibit selective stabilization of the triplexes at a TA and a CG interrupting site, respectively. However, subsequent investigations have shown that there are limitations in recognizable sequences by these analogs. In this study, we performed further systematic investigation on WNA analogs by synthesizing a variety of WNA analogs having 5-substituted cytosine and uracil, and found that WNA-betaFU exhibit high CG-selectivity.

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

Novel nucleoside analogs have been desired for selective formation of non-natural type triplexes containing TA or CG interrupting sites. We have previously reported that the W-shaped nucleic acid (WNA) would be a useful skeleton to develop new base analog for the formation of non-natural triplexes of antiparallel motif. In this study, we have found that triplex forming oligonucleotide (TFO) incorporating the new WNA analog, WNA-beta T, formed more stable triplex than natural triplex with high selectivity to the TA site. It is also noted that the TFO containing WNA-beta T did not form aggregates at the physiological condition of K+.

DOI： 10.1093/nass/2.1.35

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PERGAMON-ELSEVIER SCIENCE LTD  

Novel nucleoside analogs have been designed for selective formation of anti-parallel triplexes including a TA or a CG interrupting site. The new compounds are constructed of a W-shape bicyclic nucleic acid (WNA) bearing an aromatic ring as a stacking motif and a guanine for the formation of Hoogesteen hydrogen bonds, and are expected to effect triplex stabilization by both stacking and complementary hydrogen bonds. Purine-rich triplex-forming oligodeoxynucleotide (TFO) incorporating the new analog, WNA-7 betaG, formed a stable triplex with high selectivity to the TA site. (C) 2001 Elsevier Science Ltd. All rights reserved.

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

Novel nucleoside analogs have been designed for selective formation of antiparallel triplexes including a TA or a CG interrupting site. The new compounds are constructed of a W-shape bicyclic nucleic acid (WNA) bearing an aromatic ring as a stacking motif and a guanine for the formation of Hoogesteen hydrogen bonds, and are expected to effect triplex stabilization by both stacking and complementary hydrogen bonds. Purine-rich triplex-forming oligodeoxynucleotide (TFO) incorporating the new analog, WNA-7 beta G, formed a stable triplex with high selectivity to the AT site.

DOI： 10.1093/nass/1.1.23

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