The effect of flanking bases on direct and triplet sensitized cyclobutane pyrimidine dimer formation in DNA depends on the dipyrimidine, wavelength and the photosensitizer

Abstract

Cyclobutane pyrimidine dimers (CPDs) are the major products of DNA produced by direct absorption of UV light, and result in C to T mutations linked to human skin cancers. Most recently a new pathway to CPDs in melanocytes has been discovered that has been proposed to arise from a chemisensitized pathway involving a triplet sensitizer that increases mutagenesis by increasing the percentage of C-containing CPDs. To investigate how triplet sensitization may differ from direct UV irradiation, CPD formation was quantified in a 129-mer DNA designed to contain all 64 possible NYYN sequences. CPD formation with UVB light varied about 2-fold between dipyrimidines and 12-fold with flanking sequence and was most frequent at YYYR and least frequent for GYYN sites in accord with a charge transfer quenching mechanism. In contrast, photosensitized CPD formation greatly favored TT over C-containing sites, more so for norfloxacin (NFX) than acetone, in accord with their differing triplet energies. While the sequence dependence for photosensitized TT CPD formation was similar to UVB light, there were significant differences, especially between NFX and acetone that could be largely explained by the ability of NFX to intercalate into DNA.

The major mechanism of melanoma mutations is based on deamination of cytosine in pyrimidine dimers as determined by circle damage sequencing

“…S3C) were dominated by a T in the second position and by having another pyrimidine as the first base. This pattern is similar to one obtained after in vitro irradiation of DNA with UVB ( 25 ).…”

Ability of the Putative Decomposition Products of 2,3‐dioxetanes of Indoles to Photosensitize Cyclobutane Pyrimidine Dimer (CPD) Formation and its Implications for the “Dark” (Chemisensitized) Pathway to CPDs in Melanocytes†

“…To further confirm that NFX, TOP and IOP were indeed photosensitizing CPD formation, we carried out high-resolution sequencing gel experiments with a 5 0 -[ 32 P]-end-labeled 42-mer sequence containing the eight possible GPyPyG and APyPyA sites (Figure S4). We found that the major bands induced by UVA + sensitizer + T4-pdg occurred at ATTA and to a much lesser extent at GTTG, and undetectable at C-containing CPD sites in accord with our previous work on flanking sequence effects on photosensitized CPD formation with NFX (34). At the ATTA site, CPDs formed with decreasing efficiency in the order NFX>TOP>IOP with a ratio of 1 > 0.24 > 0.08, respectively.…”

“…The assay is based on the ability of T4 pyrimidine dimer glycosylase (T4-pdg) to cleave the phosphodiester backbone of DNA at CPDs by a lyase mechanism following cleavage of the glycosidic bond of the 5 0pyrimidine (30,31). T4-pdg can also efficiently cleave abasic sites (32) and formamido-dA (FAPYdA) products ( 33), but a recent study of photosensitization by NFX and acetone using T4pdg only showed cleavage at the 5 0 -pyrimidine of dipyrimidines indicating that photosensitization does not produce significant amounts of these other products (34). To quantify the number of CPDs produced, use was made of the fact that introduction of just one single-strand break in supercoiled circular duplex DNA (Form I) converts it to relaxed open circular DNA (Form II) which has a lower electrophoretic mobility in an agarose gel and can be quantified by imaging of an ethidium bromide-stained gel.…”

Lignin Nanoparticles Deliver Novel Thymine Biomimetic Photo-Adducts with Antimelanoma Activity

“…Spectroscopical data of novel compounds were consistent with the expected structures, as well as full comparable with data previously reported for photo-adducts deriving from N-1 methyl thymine and thymidine derivatives [27][28][29][30][31][32][33][34][35][36][37]. Again, CPD-TS isomers prevailed with respect to the CS counterpart (Table 2, entries 4-6) [41][42][43]. As a general trend, the total yield of photo-adducts increased by increasing the carbon length of the spacer.…”