Cleavage of abasic sites in DNA by intercalator-amines

Reactive oxygen species lead to oxidative damage of the nucleobase and sugar components of nucleotides in double-stranded DNA. The 2-deoxyribonolactone (or oxidized abasic site) lesion results from oxidation of the C-1' position of DNA nucleotides and has been implicated in DNA strand scission, mutagenesis, and covalent cross-linking to DNA binding proteins. We previously described a strategy for the synthesis of DNA-containing deoxyribonolactone lesions. We now report an improved method for the site specific photochemical generation of deoxyribonolactone sites within DNA oligonucleotides and utilize these synthetic oligonucleotides to characterize the products and rates of DNA strand scission at the lactone lesion under simulated physiological conditions. A C-1' nitroveratryl cyanohydrin phosphoramidite analogue was synthesized and used for the preparation of DNA containing a photochemically "caged" lactone precursor. Irradiation at 350 nm quantitatively converted the caged analogue into the deoxyribonolactone lesion. The methodology was validated by RP-HPLC and MALDI-TOF mass spectrometry. Incubation of deoxyribonolactone-containing DNA under simulated physiological conditions gave rise to DNA fragmentation by two consecutive elimination reactions. The DNA-containing products resulting from DNA cleavage at the deoxyribonolactone site were isolated by PAGE and unambiguously characterized by MALDI-TOF MS and chemical fingerprinting assays. The rate of DNA strand scission at the deoxyribonolactone site was measured in single- and double-stranded DNA under simulated physiological conditions: DNA cleavage occurred with a half-life of approximately 20 h in single-stranded DNA and 32-54 h in duplex DNA, dependent on the identity of the deoxynucleotide paired opposite the lesion site. The initial alpha,beta-elimination reaction was shown to be the rate-determining step for the formation of methylene furanone and phosphorylated DNA products. These investigations demonstrated that the deoxyribonolactone site represents a labile lesion under simulated physiological conditions and forms the basis for further studies of the biological effects of this oxidative DNA damage lesion.

show abstract

“…Thus, authentic 5 was produced by strand cleavage of a DNA oligonucleotide (14, see Experimental Section) containing an AP site (39,40).…”

Section: Isolation and Maldi-tof Ms Characterization Of The Products mentioning

confidence: 99%

Half-Life and DNA Strand Scission Products of 2-Deoxyribonolactone Oxidative DNA Damage Lesions

Zheng

Sheppard

2004

Chem. Res. Toxicol.

View full text Add to dashboard Cite

show abstract

“…However, in parallel to inhibiting APE1 activity, polyazacyclophane ligands also induce an enzyme‐independent cleavage of AP sites through a β‐elimination mechanism, due to the presence of secondary amino groups in the ligand structure (Scheme , bottom) . The latter process, reminiscent of the action of AP lyases (class I endonucleases), was also observed for oligopeptides KWK and KWKK, as well as several other small‐molecule ligands endowed with primary or secondary amino groups, eponymously termed “artificial AP lyases” . It has been proposed that such molecules may interfere with the normal BER process because of the accumulation of “dirty ends” (products of β‐ and β,δ‐elimination) that cannot be utilized as substrates by DNA polymerases, and therefore, also increase the cytotoxic effect of DNA‐alkylating drugs …”

Section: Introductionmentioning

confidence: 99%

Interaction of Functionalized Naphthalenophanes with Abasic Sites in DNA: DNA Cleavage, DNA Cleavage Inhibition, and Formation of Ligand–DNA Adducts

Caron

Duong

Guillot

et al. 2019

Chemistry A European J

View full text Add to dashboard Cite

Ligands interacting with abasic (AP) sites in DNA may generate roadblocks in base‐excision DNA repair (BER) due to indirect inhibition of DNA repair enzymes (e.g., APE1) and/or formation of toxic byproducts, resulting from ligand‐induced strand cleavage or covalent cross‐links. Herein, a series of 12 putative AP‐site ligands, sharing the common naphthalenophane scaffold, but endowed with a variety of substituents, have been prepared and systematically studied. The results demonstrate that most naphthalenophanes bind to AP sites in DNA and inhibit the APE1‐induced hydrolysis of the latter in vitro. Remarkably, their APE1 inhibitory activity, as characterized by IC50 and KI values, can be directly related to their affinity and selectivity to AP sites, as assessed by means of fluorescence melting experiments. On the other hand, the molecular design of naphthalenophanes has a crucial influence on their intrinsic AP‐site cleavage activity (i.e., ligand‐catalyzed β‐ and β,δ‐elimination reactions at the AP site), as illustrated by the compounds either having an exceptionally high AP‐site cleavage activity (e.g., 2,7‐BisNP‐S, 125‐fold more efficacious than spermine) or being totally devoid of this activity (four compounds). Finally, the unprecedented formation of a stable covalent DNA adduct upon reaction of one ligand (2,7‐BisNP‐NH) with its own product of the AP‐site cleavage is revealed.

show abstract

“…It has been demonstrated that many photoactive DNA dyes are able to intercalate between nucleobases and nick plasmid DNA in the presence of molecular oxygen and visible light (62–65). The search for molecules with desirable photophysical properties, which can function as efficient artificial photonucleases and biomolecular probes, has resulted in the discovery of a large number of DNA intercalators that can cleave ds‐DNA via photochemical pathways (66–72). Even though DAP has proven to possess interesting photosensitizing characteristics upon irradiation with visible light, it is important to determine whether it has any functional role in biological systems.…”

Section: Resultsmentioning

confidence: 99%

“…The excited‐state intermediates responsible for the photocytotoxicity of DAP remain unknown; however, preliminary experiments show that sodium azide, a known scavenger of singlet oxygen, results in the complete inhibition of the DNA photocleavage by DAP (supplementary material). It is well known that intercalator aromatic amines usually cleave DNA through the generation of reactive oxygen species (67,70–73). The potential presence of DAP as a contaminant in consumer products and with subsequent topical human exposure on sun‐exposed skin requires further investigation of its harmful effects and photodamage to human cells.…”

Section: Discussionmentioning

confidence: 99%

Photoinduced DNA Cleavage and Cellular Damage in Human Dermal Fibroblasts by 2,3‐Diaminophenazine^¶

Abuzakhm

Turró

2005

Photochem & Photobiology

View full text Add to dashboard Cite

Aromatic amines, such as o‐phenylenediamine (OPD), have been used extensively in commercial hair dyes and in the synthesis of agricultural pesticides. Air oxidation of OPD results in the formation of 2,3‐diaminophenazine (DAP). Although the mutagenic toxicity of DAP has been shown in both prokaryotic and eukaryotic systems, its phototoxicity remains largely unexplored. This study focuses on the pH‐dependent photophysical properties of DAP and demonstrates its ability to photoinduce DNA damage to pUC19 plasmid in vitro. The photocytotoxicity of DAP toward human skin fibroblasts was also measured. DAP exhibits weak intercalative binding to double‐stranded DNA with a binding constant Kb= 3.5 × 103M−1. Furthermore, upon irradiation with visible light, DAP is able to nick plasmid DNA in the presence of oxygen. The concentration of DAP that resulted in 50% cell death was 172 ± 9 μM in the dark and 13 ± 1 μM after irradiation of the DAP‐treated cell cultures with visible light (400–700 nm, 30 min, 5 J/cm2). The 13‐fold increase in toxicity upon exposure to visible light shows the need for further study of the photocytotoxicity of contaminants such as DAP.

show abstract

Cleavage of abasic sites in DNA by intercalator-amines

Cited by 14 publications

References 49 publications

Half-Life and DNA Strand Scission Products of 2-Deoxyribonolactone Oxidative DNA Damage Lesions

Half-Life and DNA Strand Scission Products of 2-Deoxyribonolactone Oxidative DNA Damage Lesions

Interaction of Functionalized Naphthalenophanes with Abasic Sites in DNA: DNA Cleavage, DNA Cleavage Inhibition, and Formation of Ligand–DNA Adducts

Photoinduced DNA Cleavage and Cellular Damage in Human Dermal Fibroblasts by 2,3‐Diaminophenazine^¶

Contact Info

Product

Resources

About

Cleavage of abasic sites in DNA by intercalator-amines

Cited by 14 publications

References 49 publications

Half-Life and DNA Strand Scission Products of 2-Deoxyribonolactone Oxidative DNA Damage Lesions

Half-Life and DNA Strand Scission Products of 2-Deoxyribonolactone Oxidative DNA Damage Lesions

Interaction of Functionalized Naphthalenophanes with Abasic Sites in DNA: DNA Cleavage, DNA Cleavage Inhibition, and Formation of Ligand–DNA Adducts

Photoinduced DNA Cleavage and Cellular Damage in Human Dermal Fibroblasts by 2,3‐Diaminophenazine¶

Contact Info

Product

Resources

About

Photoinduced DNA Cleavage and Cellular Damage in Human Dermal Fibroblasts by 2,3‐Diaminophenazine^¶