Paul Vigny

Photosensitized reactions of nucleic acids

The main effects of near-ultraviolet and visible light on cellular DNA are reviewed with emphasis on base lesions, oligonucleotide single-strand breaks and DNA-protein cross-links. Model system photosensitization reactions of DNA are also discussed. This includes photodynamic effects, menadione-mediated photooxidation, photoionization of antibiotics, the photochemistry of 5-halogenopyrimidines and urocanic acid.

Photochemotherapy using pyridopsoralens

Aiming to decrease the acute side effects and genotoxic hazards of PUVA, pyrido (3,4-C) psoralen (PP) and 7-methyl pyrido (3,4-C) psoralen (MPP) were synthesized and studied. Their UVA maximum absorption lies at 325 and 330 nm, respectively. Their photostability is comparable to that of 8-MOP. They complex to DNA in the dark, and, in the presence of UVA, produce only monoadditions to DNA, as shown by fluorescence and DNA denaturation-renaturation studies. In diploid eukaryotic yeast they are more effective than 8-MOP for the induction of lethal effects and mitochondrial damage. Their mutagenic activity per unit dose of UVA is in the same range as that of 8-MOP. However, per viable cell they are clearly less mutagenic than 8-MOP. This difference is also observed for recombinogenic activity. No oxygen effect is observed. In mammalian cells the following ranges of effectiveness are found: inhibition of DNA synthesis in human fibroblasts: MPP greater than PP greater than 8-MOP; mutagenic activity in V79 Chinese hamster cells: MPP greater than PP greater than 8-MOP; cell transforming ability in C3H embryonic mouse cells: MPP greater than 8-MOP greater than PP as a function of UVA dose, and: 8-MOP greater than MPP greater than PP as a function of survival; induction of sister chromatic exchanges (SCE) per unit dose: MPP greater than PP greater than 8-MOP in the linear part of the induction curve, and : 8-MOP greater than PP greater than MPP at the maximum level of SCE obtained.(ABSTRACT TRUNCATED AT 250 WORDS)

Chemical structure of psoralen-nucleic acid photoadducts

Resume Un certain nombre de structures chimiques de produits de photoaddition de psoralenes avec le DNA ou ses constituants a pu etre obtenu recemment dans quelques laboratoires. Le but de ce bref article est de presenter letat actuel des connaissances dans ce domaine.

CHARACTERIZATION OF PHOTOCYCLOADDITION PRODUCTS FROM REACTION BETWEEN THYMIDINE and THE MONOFUNCTIONAL 3‐CARBETHOXYPSORALEN

The photochemical reaction of 3‐carbethoxypsoralen. a monofunctional furocoumarin. with thymidine was investigated as a model system for its photoaddition to DNA. Near UV irradiation (320 nm > λ > 400 nm) of a mixture of thymidine and 3‐carbethoxypsoralen as a dry film gave rise to two main nucleoside diastereoisomers which were isolated by reverse phase high performance liquid chromatography. The structure of these products was assigned on the basis of UV absorption, fluorescence.‘H‐NMR and plasma desorption mass spectra analysis. The results are consistent with 1:1 C, cycloadducts involving the 5,6 double bond of thymine and the 4′, 5′double bond of 3‐carbethoxypsoralen. These two cycloadducts of cis‐syn stereoconfiguration show opposite circular dichroism suggesting a diastereoisomeric relationship.

SPECTROSCOPIC PROPERTIES OF PSORALEN DERIVATIVES SUBSTITUTED BY CARBETHOXY GROUPS AT THE 3,4 AND/OR 4,5' REACTION SITE

Abstract— The newly synthesized linear psoralen derivative 3‐carbethoxypsoralen has been shown recently to behave as a monofunctional derivative and has attracted some interest in the psoriasis treatment. In a first attempt to understand, by the fluorescence technique, the molecular mechanism by which it interacts with DNA, a spectroscopic study of the molecule was undertaken. The fluorescence of 3‐carbethoxypsoralen at room temperature resembles that of 8‐methoxypsoralen with a ten times higher quantum yield. 365 nm irradiation of dilute solutions of 3‐carbethoxypsoralen rapidly leads to the formation of two types of highly fluorescent photoproducts. Type 1 photoproducts (λemmax= 425 nm, λexcmax= 360 nm) have been identified as the result of the addition of a solvent molecule to the 4,5 reaction site of the molecule. Their fluorescence intensity is a hundred times higher for 3‐carbethoxypsoralen than for 8‐methoxypsoralen. They become negligible when the 4,5 reaction site carries also a carbethoxy group. Type 2 photoproducts exhibit a somewhat different emission (λemmax = 443 nm, λexcmax= 413 nm). They are probably the result of an opening of the furocoumarin molecule. The implications of the peculiar photochemical properties of 3‐carbethoxypsoralen are discussed in view of its biological activity. In addition, the use of fluorescence in monitoring the photobinding of psoralens to DNA is also discussed

Saponins from Steganotaenia araliacea.

Six saponins have been isolated and identified from the leaves of Steganotaenia araliacea. They were identified as 3-O-[beta-D-galactopyranosyl(1----2)-(beta-D-galactopyranosyl (1----3))-beta-D-glucuronopyranosyl]-21-O-tigloyl and -21-O-angeloyl-R1-barrigenol, 3-O-[beta-D-glucopyranosyl(1----2)-(beta-D-xylopyranosyl (1----3))-beta-D-glucuronopyranosyl]-21-O-tigloyl and -21-O-angeloyl-R1-barrigenol, 3-O-[beta-D-glucopyranosyl(1----2)-(beta-D-glucopyranosyl-(1----3))-(alp ha-L- rhamnopyranosyl(1----4))-beta-D-glucopyranosyl] steganogenin and 3-O-[(beta-D-galactopyranosyl(1----2)-beta-D-glucuronopyranosyl]-2 8-O- beta-D-glucopyranosyl olean-12-ene-28-oic acid. Steganogenin is a new 17,22-seco-oleanolic acid derivative. The structures of the saponins were established by analysis of their 1H and 13C NMR spectra with the help of 2D-experiments and by Californium Plasma Desorption Mass Spectrometry.

Chemical structure of 3-carbethoxypsoralen-DNA photoadducts

The enzymatic digest from salmon sperm DNA photochemically modified by the monofunctional 3-carbethoxypsoralen was analyzed by high-performance liquid chromatography. The modified nucleosides extracted from DNA were compared with model compounds obtained from irradiation in the dry state of mixtures of 3-carbethoxypsoralen with 2-deoxyribonucleosides whose chemical structures had previously been characterized. The main photoadducts formed in DNA are two cis-syn diastereoisomers formed via a C4-cycloaddition reaction involving the 4, 5 double bond of 3-carbethoxypsoralen and the 5,6 double bond of 2-deoxythymidine. Among them, the most polar one accounts for 72%. Under the same conditions, photoadducts formed between 3-carbethoxypsoralen and 2deoxycytidine account for less than 1%.

Photobiological activities of 1,6-dioxapyrene in pro- and eukaryotic cells

The photobiological effect of a new pyrene derivative, 1,6-dioxapyrene (1,6-DP), was studied in Salmonella typhimurium (strain TA100) and in the diploid strain D7 of the yeast Saccharomyces cerevisiae. In Salmonella, 1,6-DP shows little mutagenicity in the dark in comparison to benzo[a]pyrene (B[a]P). This mutagenic activity decreases in the presence of liver S9 homogenates from Aroclor induced XVIInc/Z mice. However, in combination with 365 nm (UVA) radiation and in the absence of S9 mix, 1,6-DP behaves as an effective photodynamic compound inducing lethal and mutagenic effects in both organisms. In yeast, its activity, like that of B[a]P, is highly dependent on the presence of oxygen. For the same incident dose of UVA, 1,6-DP is, however, at least 6 times more effective than B[a]P in inducing cytotoxic and mutagenic effects. At equitoxic doses, 1,6-DP is as photomutagenic as B[a]P, suggesting that in both cases mutagenicity is due to similar mechanisms. Spectrophotometric measurements indicate physical interaction of 1,6-DP with DNA in the dark. Laser flash photolysis experiments show that 1,6-DP generates singlet oxygen with a quantum yield of 0.17. In vitro 1,6-DP produces oxidative damage to guanine bases specific for singlet oxygen mediated reactions. Alkaline step elution analysis of 1,6-DP plus UVA treated yeast cells indicates a decrease in average molecular weights in DNA and an induction of single strand breaks (ssb) originating from alkali labile sites. This effect is enhanced by D2O and is thus likely to be due to the production of singlet oxygen. The strand breaks appear to differ from those induced by gamma-rays because little, if any, repair of these ssb occurs during 30 min of post-treatment incubation in complete growth medium. These results suggest that the photobiological effects of 1,6-DP are due to oxidative damage in DNA mostly induced by singlet oxygen.

TRIPLET EXCITED STATES OF 4',5'‐PHOTOMONOADDUCTS OF 3‐ CARBETHOXYPSORALEN and 8‐METHOXYPSORALEN WITH DNA NUCLEOSIDES

Abstract— Triplet absorption spectra, triplet extinction coefficient and intersystem crossing for 4,5‐monocycloadducts of 3‐carbethoxypsoralen (3‐CPs) with thymidine (dThd) and uridine (dUrd) in ethanol have been investigated in order to elucidate whether their triplet state properties could be the limitating step for a further photoreaction of 3‐CPs monoadducts with DNA nucleosides. The comparison between the triplet characteristics of 4,5‐monoadducts of 3‐CPs and those of 8‐methoxypsoralen (8‐MOP) shows that the quantum yield is much higher in the case of 3‐CPs than for 8‐MOP. The monofunctionality of 3‐CPs cannot therefore be ascribed to the triplet excited states properties of its monoadducts. It is likely that steric hindrance introduced by the bulky carbethoxy group remains a reasonable explanation.

PHOTOREACTION OF MONOFUNCTIONAL 3-CARBETHOXYPSORALEN WITH DNA: IDENTIFICATION AND CONFORMATIONAL STUDY OF THE PREDOMINANT cis-syn FURAN SIDE MONOADDUCT TO THYMINE

The main DNA‐3‐CPs furan side adduct has been isolated following acidic hydrolysis of photomodified DNA and subsequent reversed‐phase high performance liquid chromatography purification. This photoadduct has been identified as a thymine‐3‐CPs C4‐cycloadduct Thy<5 4′6 5′, >3‐CPs based on its optical spectroscopic features and of its plasma desorption mass spectrometric characteristics. Moreover, its reversed‐phase high performance liquid chromatography retention time is in agreement with a cis‐syn stereochemistry as compared to the chromatographic properties of the cis‐syn model adduct Thy< 5 4′6 5′ >3‐CPs previously identified. Further support for the cis‐syn stereoconfiguration assignment was provided by fluorescence quenching experiments using iodide and silver ions as external and internal quenchers respectively. These data strongly indicate that the Thy< 5 4′6 5′ >3‐CPs photoadduct is located inside the DNA helix in agreement with its cis‐syn stereochemistry.