Porphyrin-Induced Protein Oxidation and Aggregation as a Mechanism of Porphyria-Associated Cell Injury.

Genetic porphyrias comprise eight diseases caused by defects in the heme biosynthetic pathway that lead to accumulation of heme precursors. Consequences of porphyria include photosensitivity, liver damage and increased risk of hepatocellular carcinoma, and neurovisceral involvement, including seizures. Fluorescent porphyrins that include protoporphyrin-IX, uroporphyrin and coproporphyrin, are photo-reactive; they absorb light energy and are excited to high-energy singlet and triplet states. Decay of the porphyrin excited to ground state releases energy and generates singlet oxygen. Porphyrin-induced oxidative stress is thought to be the major mechanism of porphyrin-mediated tissue damage. Although this explains the acute photosensitivity in most porphyrias, light-induced porphyrin-mediated oxidative stress does not account for the effect of porphyrins on internal organs. Recent findings demonstrate the unique role of fluorescent porphyrins in causing subcellular compartment-selective protein aggregation. Porphyrin-mediated protein aggregation associates with nuclear deformation, cytoplasmic vacuole formation and endoplasmic reticulum dilation. Porphyrin-triggered proteotoxicity is compounded by inhibition of the proteasome due to aggregation of some of its subunits. The ensuing disruption in proteostasis also manifests in cell cycle arrest coupled with aggregation of cell proliferation-related proteins, including PCNA, cdk4 and cyclin B1. Porphyrins bind to native proteins and, in presence of light and oxygen, oxidize several amino acids, particularly methionine. Noncovalent interaction of oxidized proteins with porphyrins leads to formation of protein aggregates. In internal organs, particularly the liver, light-independent porphyrin-mediated protein aggregation occurs after secondary triggers of oxidative stress. Thus, porphyrin-induced protein aggregation provides a novel mechanism for external and internal tissue damage in porphyrias that involve fluorescent porphyrin accumulation.

Phototoxicity of B-RAF inhibitors: Exclusively due to UVA radiation and rapidly regressive.

BACKGROUND: New targeted melanoma therapies such as B-RAF inhibitors have shown high and promising clinical benefit but have cutaneous side-effects, including photosensitivity, which is triggered in the UVA radiation spectrum. However, visible spectrum implication has not yet been investigated. We conducted a study to determine whether visible light also contributes to the phototoxicity action spectrum of vemurafenib. The secondary end points were to determine the time to complete regression of the phototoxicity post-vemurafenib discontinuation and whether there was a significant difference between the UVA radiation immediate reactivity cut-offs, in patients treated with vemurafenib vs. those treated with dabrafenib. METHOD: This prospective, observational study included patients with B-RAF mutant metastatic melanoma: 34 patients treated with vemurafenib and 9 with dabrafenib. RESULTS: The visible-light phototest results in patients treated with vemurafenib were all negative before and after 2 months of treatment. The UVA radiation phototests conducted 1 or 2 weeks post-vemurafenib discontinuation in 4 patients showed a normalised UVA-radiation reactivity cut-off. UVA radiation phototests after 2 months of treatment were conducted for all patients. The UVA radiation reactivity cut-off had been lowered for 30 patients (88%) on vemurafenib and 3 patients (33%) on dabrafenib. The median UVA radiation reactivity cut-off was 12 J/cm(2) for the patients on vemurafenib and 20 J/cm(2) for the patients on dabrafenib. CONCLUSION: B-RAF inhibitor phototoxicity is exclusively triggered by UVA radiation and resolves rapidly post-treatment discontinuation. A significant difference between the UVA immediate reactivity cut-offs, vemurafenib vs. dabrafenib, explains the difference in the clinical photosensitivity rates reported in the clinical trials.

Exposure to phototoxic NSAIDs and quinolones is associated with an increased risk of melanoma.

PURPOSE: Ultraviolet radiation exposure is the most important exogenous risk factor for cutaneous malignancies. It is possible that phototoxic drugs promote the development of cutaneous melanoma (CM) by intensifying the effect of ultraviolet light on the skin. We investigated the association between the use of common systemic phototoxic drugs and development of CM. METHODS: This study was a case-control study in a Dutch population-based cohort. The drug dispensing data was obtained from PHARMO, a Dutch drug dispensing and hospital admissions registry, and linked to PALGA, the nationwide pathology network of the Netherlands. The cases were patients diagnosed with pathologically confirmed primary CM between 1991 and 2004. Controls were sampled from the PHARMO population. Exposure to systemic phototoxic drugs was measured and included antimicrobial agents, diuretics, antipsychotic drugs, antidiabetic drugs, cardiac drugs, antimalarials and nonsteroidal anti-inflammatory drugs (NSAIDs). A multivariate conditional logistic regression analysis was performed to study the association between exposure to phototoxic drugs and CM. RESULTS: The study population included 1,318 cases and 6,786 controls. Any phototoxic drug during the study period was dispensed for 46 % of the cases and 43 % of the controls (p = 0.012). The use of quinolones [odds ratio (OR) 1.33, 95 % confidence interval (CI) 1.01-1.76] and propionic acid derivative NSAIDs (OR 1.33, 95 % CI 1.14-1.54) had a positive association with CM. CONCLUSIONS: Our study shows that the use of phototoxic drugs is associated with an increased risk of developing CM. Even a short-term use of phototoxic quinolones and propionic acid derivative NSAIDs may increase the risk for CM. Patient education to promote sun-protective behaviour is essential to avoid immediate adverse effects and possible long-term effects of phototoxic drugs.

Photosensitizer adhered to cell culture microplates induces phototoxicity in carcinoma cells.

In vitro experiments in plastic receptacles are the basis of characterization of new photosensitizers (PSs) for the photodynamic therapy. We recently reported that lipophilic PSs adhere to cell culture microplates in a kinetic-like manner (Engelhardt et al., 2011). In the current study, we examined the interaction and phototoxic effects of the microplate-adhered PS in cancer cells. Therefore, we preloaded microplates with hypericin, Foscan, PVP-hypericin, or aluminum (III) phthalocyanine tetrasulfonate chloride (AlPCS4) for 24 hours and measured the PS distribution after addition of A431 human carcinoma cells: following another 24 hours up to 68% of hypericin were detected in the cell fraction. The hydrophilic PVP-hypericin and AlPCS4 also diffused into the cells, but the quantities of PS adherence were considerably lower. Microplate-adhered Foscan appeared not to be redistributed. In contrast to the hydrophilic PSs, the cellular phototoxicity of microplate-adhered lipophilic PS was high, independent of whether the PS (i) was pre-loaded onto microplates or (ii) added simultaneously with the cells or (iii) one day after cell seeding. Based on these results, we suggest testing lipophilic PS dyes for their adherence to microplates. Furthermore, the ability of plastic materials to (reversibly) store PSs might represent a new approach for the PS delivery or the development of antimicrobial coatings.

Phototoxicity in an ultraviolet ink manufacturing plant.

Ultraviolet (UV) dyes are used as inks in garment printing. Hypersensitivity reactions to these compounds have been reported in the literature. The authors report a case of reaction to UV ink in a patient already on corticosteroid therapy. The patient's clinical course was reviewed along with images of wounds that subsequently developed. The affected areas were debrided and covered with Vaseline gauze and silver impregnated dressings. Epithelium was salvaged in many areas, and regrowth occurred over several weeks in regions of deeper injury. The concurrent use of steroids and the rapidity of the onset of symptoms were not characteristic of hypersensitivity dermatitis, which has previously been reported. The cause of the wounds was likely phototoxicity from radical subtypes in the ink that catalyze the reaction when exposed to UV light.

Phototoxicity is not associated with photochemical tissue bonding of skin.

BACKGROUND AND OBJECTIVE: We have developed a light-activated method called photochemical tissue bonding (PTB) for closing wounds using green light and a photosensitizing dye (Rose Bengal-RB) to initiate photochemical crosslinking of wound surface proteins. These studies were designed to determine whether RB causes phototoxicity during closure of skin incisions with PTB. STUDY DESIGN/MATERIALS AND METHODS: RB phototoxicity was evaluated after sealing incisions in porcine skin ex vivo and rabbit skin in vivo using PTB (1 mM RB, 100 J/cm(2), 532 nm, 0.3 or 0.5 W/cm(2).) Dead cells were identified by pyknotic nuclei and eosinophilic cytoplasm on H&E-stained sections. The influence on RB phototoxicity of penetration of RB into the wound wall (by confocal microscopy), RB concentration in the tissue (by extraction), and fluence of 532 nm reaching depths in skin (calculated from skin optical properties) were investigated. RESULTS: No significant differences were found in the percent dead cells in PTB-treated and control incisions in porcine skin at 24 hours or in rabbit skin at 2 hours and 3 and 7 days after surgery. RB was retained in a approximately 100 microm wide band next to the wound wall. The mean RB concentration within this band was 0.42+/-0.03 mM. Monte Carlo modeling of light distribution indicated that the fluence rate decreased from the subsurface peak to 0.5 W/cm(2) in the mid-dermis (approximately 350 microm.) In vitro RB phototoxicity to dermal fibroblasts yielded an LD(50) of 0.50+/-0.09 J/cm(2) when the cells contained 0.46 mM RB. CONCLUSIONS: PTB does not cause phototoxicity when used to repair skin wounds even though the RB concentration and 532 nm fluence in the mid-dermis during PTB are much greater than the LD(50) for RB phototoxicity in vitro. These results indicate that phototoxicity is not a concern when using PTB for tissue repair.

Theoretical study of ibuprofen phototoxicity.

The photochemical properties and degradation of the common nonsteroid anti-inflammatory drug ibuprofen is studied by means of hybrid density functional theory. Computed energies and properties of various species show that the deprotonated form dominates at physiological pH, and that the species will not be able to decarboxylate from a singlet excited state. Instead, decarboxylation will occur, with very high efficiency, provided the deprotonated compound can undergo intersystem crossing from an excited singlet to its excited triplet state. In the triplet state, the C-C bond connecting the carboxyl group is elongated, and the CO2 moiety detaches with a free energy barrier of less than 0.5 kcal/mol. Depending on the local environment, the decarboxylated product can then either be quenched through intersystem crossing (involving the possible formation of singlet oxygen) and protonation, or serve as an efficient source for superoxide anions and the formation of a peroxyl radical that will initiate lipid peroxidation.

[Eczematous skin diseases]. / Ekzemás megbetegedések.

The skin, as one of the most important barriers of the human body, protects the inner homeostasis from the harmful environmental influences as well as physical, chemical and biological factors. When the impact of these factors exceeds the tolerance and reproducing capacity of the skin, pathological alterations will develop. If follows from this that dermatology can surely be considered to be a part of environmental medicine. Eczematous diseases are mostly pathological pictures of varied mechanisms developing as a result of environmental influences (irritants, contact allergens, microbes). Since their clinical appearance is similar, it is a serious professional challenge to diagnose them. In this article we present the clinical features, provoking factors of these skin diseases as well as therapeutical possibilities.

Analytical studies on the prediction of photosensitive/phototoxic potential of pharmaceutical substances.

PURPOSE: Phototoxic responses after administration of photosensitive pharmaceutics have been recognized as undesirable side effects, and predicting potential hazardous side effects is gaining importance as new drugs are introduced to the market. In this work, we characterize the photochemical/photobiological properties of model compounds to develop an effective screening method for the prediction of phototoxic/photosensitive potential. METHODS: Twenty-one known photosensitive/phototoxic compounds and five weak/nonphototoxic compounds were subjected to ultraviolet (UV) spectral analyses and photochemical evaluation including the determination of produced reactive oxygen species (ROS) and photostability study. The photooxidation of linoleic acid was also monitored in the presence of tested compounds, guided on the formation of thiobarbituric acid reactive substances. RESULTS: Most photosensitive/phototoxic drugs tested, even weak UV absorbers, at a concentration of 200 microM showed significant production of ROS under 18 h light exposure (30,000 lx). On the other hand, ROS generated from weak/nonphototoxic compounds, including strong UV absorber benzocaine, were low or negligible. Although exposure of quinine to light resulted in significant degradation (half-life, t1/2=6.4 h), it was dramatically attenuated by the addition of ROS scavengers, especially sodium azide (t1/2=122.6 h). Furthermore, concomitant exposure of photosensitive/phototoxic compounds (200 microM) and linoleic acid (1 mM) for 18 h led to the marked formation of lipoperoxide. CONCLUSION: Results indicated that known photosensitive/phototoxic compounds tested have the ability to generate ROS under light exposure, and this photochemical reaction could be associated with their photoinstability and/or phototoxic responses. Based on these findings, determination of ROS, generated from photoirradiated compounds, may be an effective predictive model in recognizing their photosensitive/phototoxic potential.

Protection of photoreceptor cells from phototoxicity by transplanted retinal pigment epithelial cells expressing different neurotrophic factors.

Transplantation of cells or tissues and the intravitreal injection of neurotrophic factors are two methods that have been used to treat retinal diseases. The purpose of this study was to examine the effects of combining both methods: the transplantation of retinal pigment epithelial (RPE) cells expressing different neurotrophic factors. The neutrophic factors were Axokine, brain derived-neurotrophic factor (BDNF), and basic fibroblast growth factor (bFGF). The enhanced green fluorescence protein (eGFP) gene was used as a reporter gene. These genes were transduced into RPE cells by lipofection, selected by antibiotics, and transplanted into the subretinal space of 108 rats. The rats were examined at 1 week and 3 months after the transplantation to determine whether the transduced cells were present, were expressing the protein, and were able to protect photoreceptors against phototoxicity. The survival of the transplanted cells was monitored by the presence of eGFP. The degree of protection was determined by the thickness of the outer nuclear layer. Our results showed that the degree of photoreceptor protection was different for the different types of neurotrophic factors at 1 week. After 3 months, the number of surviving transplanted cell was markedly reduced, and protection was observed only with the BDNF-transduced RPE cells. A significant degree of rescue was also observed by BDNF-transduced RPE cells in the nontransplanted area of the retina at both the early and late times. Lymphocytic infiltration was not detected in the vitreous, retina, and choroid at any time. We conclude that the transplantation of BDNF-transduced RPE cells can reduce the photoreceptor damage induced by phototoxicity in the transplanted area and weakly in the nontransplanted area.

Photosensitivity to exogenous agents.

OBJECTIVE: To better understand cutaneous photosensitivity reactions, a review of its etiologic factors, clinical characteristics, pathogenesis, and treatment modalities was undertaken. METHODS: Articles discussing the above aspects of phototoxic and photoallergic reactions were used to demonstrate what is currently known about photoinduced reactions and how to treat them. RESULTS: Upon interaction of solar UV radiation with the chemical that is present in significant levels on the skin, one of two known reactions may occur in susceptible patients: a phototoxicity and/or photoallergy. Phototoxic and photoallergic reactions can be diagnosed separately on the basis of pathogenesis, clinical characteristics, and histology. Examples of drugs capable of inducing a phototoxic reaction include amiodarone, retinoids, nonsteroidal antiinflammatory agents, diuretics, and antibiotics. Substances known to cause a photoallergic response are fragrances, sunscreens, topical antimicrobials, NSAID, and psychiatric medications, such as chlorpromezine. CONCLUSION: Photoinduced reactions produced by exogenous chemicals are common skin disorders. Definitive therapy requires identifying and removing the offending agent, either the photosensitizing chemical or light. The use of fully protective clothing and a sunscreen of high SPF are important measures when light exposure is inevitable.

In vitro phototoxic properties of new 6-desfluoro and 6-fluoro-8-methylquinolones.

A representative set of potent antibacterial 6-desfluoro-8-methylquinolones, in which the C-6 fluorine atom is replaced by -NH(2) or -H, and their 6-fluoro counterparts, were investigated to evaluate their phototoxic potential and to explore the mechanism behind their phototoxicity. The capacity to photosensitize biological substrates (lipids, proteins, DNA) has been analyzed, as well as their photocytotoxicity on red blood cells and 3T3 murine fibroblasts. The results obtained show that the quinolones studied are able to photosensitize red blood cell lysis in an oxygen-dependent way and induce a high decrease in cell viability after UVA irradiation. A major correlation with phototoxicity lies in the structure of the individual antibacterials and their hydrophobicity; in particular, 6-amino derivatives are less phototoxic than corresponding unsubstituted and fluorinated compounds. Cellular phototoxicity was inhibited by the addition of free radical and hydroxyl radical scavengers (BHA, GSH and DMTU), suggesting the involvement of a radical mechanism in their cytotoxicity. A good correlation was observed between lipid peroxidation and phototoxicity, indicating that the test compounds exert their toxic effects mainly in the cellular membrane. Preliminary experiments on pBR322 DNA show that these derivatives do not photocleave DNA, differently from the two photogenotoxic fluoroquinolones, ciprofloxacin and lomefloxacin, used as reference compounds.

Cutaneous consequences of photodynamic therapy.

Photodynamic therapy (PDT) has several cutaneous complications: photosensitivity is well known, but the other complications are rarely reported, Since late 1997, we have studied the dermatologic complications of using porfimer sodium PDT to treat either Barrett esophagus with high-grade dysplasia or gastroesophageal cancer in 72 consecutive patients. Cutaneous complications of PDT included serious phototoxicity requiring oral corticosteroid treatment (22 patients; 31%), herpes zoster (HZ) requiring hospitalization and intravenous antiviral treatment (1 patient; 1%), and erythema multiforme drug reaction related to porfimer sodium (1 patient; 1%). PDT-associated dermatologic complications were common and were not related to cutaneous photosensitivity.