Analysis of Fluoroquinolone-mediated Photosensitization of 2′-Deoxyguanosine, Calf Thymus and Cellular DNA: Determination of Type-I, Type-II and Triplet-Triplet Energy Transfer Mechanism Contribution¶

Abstract

Fluoroquinolone (FQ) antibacterials are known to exhibit photosensitization properties leading to the formation of oxidative damage to DNA. In addition, photoexcited lomefloxacin (Lome) was recently shown to induce the formation of cyclobutane pyrimidine dimers via triplet-triplet energy transfer. The present study is aimed at gaining further insights into the photosensitization mechanisms of several FQ including enoxacin (Enox), Lome, norfloxacin (Norflo) and ofloxacin (Oflo). This was achieved by monitoring the formation of DNA base degradation products upon UVA-mediated photosensitization of 2deoxyguanosine, isolated and cellular DNA. Oflo and Norflo act mainly via a Type-II mechanism whereas Lome and, to a lesser extent, Enox behave more like Type-I photosensitizers. However, the extent of oxidative damage was found to be relatively low. In contrast, it was found that cyclobutane thymine dimers represent the major class of damage induced by Enox, Lome and Norflo within isolated and cellular DNA upon UVA irradiation. This striking observation confirms that FQ are able to promote efficient triplet energy transfer to DNA. The levels of photosensitized formation of strand breaks, alkali-labile sites and oxidative damage to cellular DNA, as measured by the comet assay, were confirmed to be rather low. Therefore, we propose that the phototoxic effects of FQ are mostly accounted for energy transfer mechanism rather than by Type-I or -II photosensitization processes.

Photoinduced Damage to Cellular DNA: Direct and Photosensitized Reactions†

“…There is an abundant literature on the TTET-mediated formation of CPDs in ds DNA by several photosensitizers, including acetone (142), acetophenone (54,142), benzophenone (54,143), carprofen (144), fenofibric acid (143), fluoroquinolones (92,93,145), ketoprofen (143) and three pyridopsoralen derivatives (146,147). The predominant CPD photoproduct of the latter photosensitized reactions was found to be cis-syn T<>T, in agreement with the fact that thymine exhibits the lowest triplet energy (E T ) among nucleobases (148).…”

“…As will be discussed, later triplet-triplet energy transfer (TTET) process is documented for instance for several phototoxic drugs. Interestingly, chromatographic measurements showed that, in calf thymus DNA triplet transfer photosensitization using ketone compounds leads to the predominant formation of T< >T (90%) together with lower amounts of T< >C and C< >T as observed in UVA-irradiated cells (54,93). An alternative explanation is a direct excitation mechanism that would result from the weak absorption of DNA in the UVA range.…”

“…The decrease by about 43 kJ mol )1 of the thymine E T value with respect to that of the isolated base was suggested to result from the occurrence of p-stacking and base pairing in DNA (92). Several examples of UVA-sensitized formation of CPDs by fluoroquinolones in the DNA of isolated cells (93,149,150) and mouse skin (151) are available. Thus, UVA-excited lomefloxacin has been found to generate in human keratinocytes CPDs that were revealed as T4 endonuclease V-sensitive sites (149).…”

“…Evidence has been also provided using an immunohistochemical detection approach for the lomefloxacin-sensitized formation of CPDs in the skin of mice XPA + ⁄ + and XPA ) ⁄ ) upon UVA irradiation (151). Further detailed information was gained on the photosensitizing properties of several fluoroquinolones on the DNA of THP-1 human monocytes (93). It was found that in most cases TTET was the predominant mechanism of photosensitized damage to DNA over the formation of 8-oxodGuo and DNA strand breaks.…”

Combined Use of In Vitro Phototoxic Assessments and Cassette Dosing Pharmacokinetic Study for Phototoxicity Characterization of Fluoroquinolones

“…Thus, these previous findings were partly consistent with the results from the IBP assay on FQs. Some FQs, including enoxacin, NFLX, CPFX, LFLX, and SPFX, induce DNA damage after exposure to UV, the major mechanisms of which may involve generation of singlet oxygen, radical chain reaction, and formation of thymine cyclobutane dimers as described in previous reports (27,28). Considering the present findings and previous reports, irradiated FQ-induced DNA damage may mainly occur through type II photochemical reaction.…”