Phototoxicity Test Research Articles

Cellular damage photosensitized by dasatinib, radical-mediated mechanisms and photoprotection in reconstructed epidermis

Dasatinib (DAS) is an anticancer drug employed in the treatment of certain hematological malignancies. Although DAS has been mainly developed for oral administration, it has recently garnered attention for its possible topical application. The use of topical drugs can cause photosensitivity, which is not listed as an adverse reaction for DAS. Since DAS absorbs UVA, it could potentially induce photosensitivity reactions and lead to oxidative damage to cellular targets. This study aims to investigate whether DAS exhibits phototoxic reactions on primary cellular targets in both solution and artificial skin, mimicking topical drug administration. It also examines the potential generation of highly reactive intermediates like organic radicals and ROS, which could trigger photosensitivity reactions. Upon DAS irradiation in the UVA region, the first transient species detected was the diradicaloid triplet excited state (3DAS∗) with an absorption maximum of around 490 nm, which was quenched by oxygen to produce singlet oxygen. Quenching experiments with linoleic acid and 3-methylindole indicated that radical-mediated (Type I) photosensitized damage to lipids and proteins is possible. However, the lack of triplet quenching with guanosine suggests that the Type II mechanism also plays a role in the photooxidation of biomolecules. Accordingly, the neutral red uptake phototoxicity test (photoirritation factor of 5) and the comet assay, revealed that this drug is photo(geno)toxic to cells. Moreover, investigations on lipid photoperoxidation, and protein and DNA photooxidation strongly support that different cellular compartments are potential targets for DAS-induced phototoxicity.Regarding its potential application in topical dermatological formulations, an O/W emulsion of DAS was prepared and tested in reconstructed human epidermis, and a significant phototoxicity was also demonstrated. Fortunately, this undesired side effect disappeared upon formulation of DAS along with a sunscreen. Thus, for topical treatments, the photosensitivity reactions induced by DAS can be prevented by using formulations including appropriate UVA filters.

Performance and mechanisms of constructed wetland integrated microbial fuel cell for remediation and detoxification of leather tannery wastewater

Several previous studies concerned of microbial fuel cells integrated into constructed wetlands, nevertheless, their application as a convenient treatment for wastewater is still developing. In this experimental investigation, five CW-MFC systems were similarly designed, setup, and operated in a batch mode for two subsequent cycles. Each cycle lasted for 10 days to evaluate the performance of CW-MFC system for the remediation of real leather tannery wastewater (LTW). Four CW-MFCs were planted, each with different type of vegetation including Conocarpus, Arundo donax, Canna lily, and Cyperus papyrus in CW1-MFC, CW2-MFC, CW3-MFC, and CW4-MFC, respectively. The fifth CW5-MFC was maintained unplanted and considered as the control system. The performance of each CW-MFCs systems was evaluated mainly based on the removal of organic content (COD), total dissolved solid (TDS) elimination, and power generation. The results demonstrated that the four types of plants maintained healthy and no sign of wilting was observed during the 20 days of monitoring. For the first cycle of batch operation, maximum removal efficiencies of COD were 99.8%, 99.5%, 99.7%, 99.6% and 99.5% with power outputs of 10,502.8, 10,254.6, 9956.4, 10,029.6, and 9888.0 mW/m3, while, maximum TDS elimination were 46.7%, 39.7%, 60.8%, 55.5%, and 13.8% observed in CW1-MFC, CW2-MFC, CW3-MFC, CW4-MFC, and CW5-MFC, respectively. Very comparable results were observed in the second operation cycle. Results of phototoxicity test indicated that the germination of Hordeum vulgare and Triticum aestivum were 100% watered with treated effluent compared to 90% accomplished with tap water as the control solution for both types of seeds.

Adaption of in vitro and in chemico phototoxicity tests for tattoo pigments and the effect of adsorption of the phototoxic contaminant benzo[a]pyrene to carbon black

Phototoxicity plays an important role in tattoo toxicology due to the skin's daily exposure to sunlight and the permanent deposition of pigments in skin. Here, we investigated in chemico reactive oxygen species generation using 2′,7′-dichlorodihydrofluorescein diacetate and in vitro phototoxicity towards skin fibroblasts based on the OECD Test Guideline 432.Proficiency substances, titanium dioxide (TiO2, anatase 30 nm, anatase 100 nm, rutile 30 nm), pigment red 22 (PR 22) with and without purification and carbon black (CB) were investigated. Phototoxic properties of benzo[a]pyrene (B[a]P) were investigated with and without adsorption to CB. Pigments were characterised by dynamic light scattering.All proficiency substances except for norfloxacin were predicted successfully and only TiO2 anatase exhibited phototoxicity in the cell-based approach. B[a]P showed the expected phototoxicity in contrast to B[a]P bound to carbon black. Upon UVA irradiation, only TiO2 pigment anatase and PR 22 generated ROS in the cell-free assay. DLS measurements showed a high degree of polydispersity/agglomeration.OECD Test Guideline 432 should be applied with an incubation time of 24 h when testing highly insoluble pigments. The in chemico ROS assay introduced below might serve as a quick screening test but may be further adapted to predict the in vitro outcome.

In Silico Phototoxicity Prediction of Drugs and Chemicals by using Derek Nexus and QSAR Toolbox.

Phototoxicity testing is crucial for evaluating the potential harmful effects of pharmaceuticals and chemicals on human skin when exposed to sunlight. Traditional in vivo models involving mice, rats, guinea pigs, as well as in vitro assays such as the 3T3 Neutral Red Uptake phototoxicity assay and methods based on the use of reconstructed human epidermis, have been established for phototoxicity testing. While these approaches are extremely valuable, they are costly in terms of both time and resources. Consequently, in silico approaches based on the use of predictive software tools can offer more rapid and cost-effective phototoxicity screening solutions. With this goal in mind, the current study evaluated two in silico tools - Derek Nexus 6.1.0/Derek Knowledge Base 2020 1.0 (Lhasa Limited, UK) and the QSAR Toolbox (v 4.5) developed by the Organisation for Economic Co-operation and Development (OECD) - for their capacity to predict the phototoxicity of several substances from diverse classes. Derek Nexus and the QSAR Toolbox were both found to be very useful for predicting the phototoxicity of drugs and other chemicals. Derek Nexus predicted phototoxicity of the compounds, with a sensitivity of 63%, specificity of 93%, Positive Predictive Values of 90% and Negative Predictive Value of 69%, overall accuracy of 77% and balanced accuracy of 78%. The QSAR Toolbox achieved sensitivity of 73%, specificity of 85%, Positive Predictive Value of 85% and Negative Predictive Value of 74%, overall accuracy of 79% and balanced accuracy of 79%. The results show that Derek Nexus and the QSAR Toolbox can be usefully incorporated in the workflow of phototoxicity testing for pharmaceuticals and chemicals.

Comparison of two biological systems used for phototoxicity testing: Cellular and tissue

The OECD has approved two similar methods for testing the phototoxic potency of chemicals. The first method, OECD 432, is based on the cytotoxicity properties of materials to the mouse 3T3 (clone A31) cell line (fibroblasts) after exposure to light. The second method, OECD 498, is based on the same properties but using reconstructed human epidermis - EpiDerm (stratified keratinocytes). The aim of this study was to compare these two methods using statistical tests (specificity, sensitivity, negative predictive value, positive predictive value and accuracy) and non-statistical characteristics (e.g. price and experimental duration, amount of material, level of complications, cell type, irradiation dose). Both tests were performed according to the relevant guidelines using the same 11 control substances. Higher performance values were observed for OECD 432 in both phototoxic and non-phototoxic classifications. The accuracy of OECD 432 was 90.9%, while that of OECD 498 was 72.7%. OECD 432 was also shorter and less expensive. On the other hand, OECD 498 was less complicated, and used human cells with stratum corneum, which better reflects real skin. This method can also be used with oily substances that are poorly soluble in water. However, both methods are important for testing the phototoxic properties of materials, and can be used alone or in a tiered strategy.

KoCVAM-led development of phototoxicity alternative test method using reconstructed human epidermis model (KeraSkin™)

Phototoxicity is an acute toxic reaction induced by topical skin exposure to photoreactive chemicals followed by exposure to environmental light and thus chemicals that absorb UV are recommended to be evaluated for phototoxic potential. There are currently three internationally harmonized alternative test methods for phototoxicity. One of them is the in vitro Phototoxicity: RhE Phototoxicity test method (OECD TG498).Korean center for the Validation of Alternative Methods (KoCVAM) developed an in vitro phototoxicity test method using a KeraSkin™ reconstructed human epidermis model (KeraSkin™ Phototoxicity Assay) as a ‘me-too’ test method of OECD TG498.For the development and optimization of KeraSkin™ Phototoxicity Assay, the following test chemicals were used: 6 proficiency chemicals in OECD TG498 (3 phototoxic and 3 non-phototoxic), 6 reference chemicals in OECD Performance Standard No. 356 (excluding the proficiency test chemicals, 3 phototoxic and 3 non-phototoxic) and 13 additional chemicals (7 phototoxic and 6 non-phototoxic). Based on the test results generated from the test chemicals above, the overall predictive capacity of KeraSkin™ Phototoxicity Assay was calculated. In particular, the assay exhibited 100 % accuracy, 100 % sensitivity, and 100 % specificity. Therefore, it fulfills the requirements to be included as a ‘me-too’ test method in OECD TG498.

Surfactants for stabilization of dermal emulsions and their skin compatibility under UVA irradiation: Diacyl phospholipids and polysorbate 80 result in high viability rates of primary human skin cells

Phospholipids are versatile formulation compounds with high biocompatibility. However, no data on their effect on skin in combination with UVA radiation exist. Thus, it was the aim of this work to (i) develop o/w nanoemulsions (NEs) differing in surfactant type and to investigate their physicochemical stability at different storage temperatures, (ii) establish a standardized protocol for in vitro phototoxicity testing using primary human skin cells and (iii) investigate the phototoxicity of amphoteric phospholipids (S45, S75, E80, S100, LPC80), sodium lauryl ether sulfate (SLES) and polysorbate 80 (PS80). Satisfying systems were developed with all surfactants except S100 due to low zeta potential (-21.4 mV ± 4.69). SLES and PS80-type NEs showed the highest stability after eight weeks; temperature-dependent variations in storage stability were most noticeable for phospholipid surfactants. For phospholipid-based NEs, higher phosphatidylcholine content led to unstable formulations. Phototoxicity assays with primary skin fibroblasts confirmed the lack of UVA-related phototoxicity but revealed cytotoxic effects of LPC80 and SLES, resulting in cell viability as low as 2.7 % ±0.78 and 1.9 % ±1.57 compared to the control. Our findings suggest that surfactants S45, S75 and PS80 are the most promising candidates for skin-friendly emulsifiers in sensitive applications involving exposure to UV light.

MLtox, online phototoxicity prediction webpage

Phototoxicity, sometimes in the literature referred to as photo-irritation, is a chemically induced reaction requiring light. While it is generally accepted that phototoxicity testing can be performed in the majority of cases in vitro (i.e. without the use of experimental animals), these tests may sometimes provide contradictory predictions. Understanding the mechanisms of initiating events based on the molecule's structure and its ability to reach the excited state and consequently generate ROS enables the creation of predictive QSAR for this adverse outcome. The ability to predict the phototoxicity potential via a QSAR model is beneficial in reducing the number of mechanical in vitro/in chemico tests needed to demonstrate absence of phototoxicity and it is very helpful in the overall safety assessment process.The QSAR prediction model presented here focused on developing a robust platform freely available on the web via the link http://mltox.online to provide interpretable predictions of the phototoxicity of tested molecules. Great attention was devoted to interpretability and explainability of the prediction results. The web application allows the user to input a chemical by CAS number, SMILES code or trivial name. The user can choose between simple prediction or advanced tools options. These extended tools include the artificial intelligence explainability of model prediction using XSMILES (interactive visualization technique to support the interpretation of SMILES) and SHAP values (impact each element on the prediction). The comprehensive tools in question allow the user to explore the properties of phototoxic substances and to understand the prediction outcomes better.

Chlorin E6-Curcumin-Mediated Photodynamic Therapy Promotes an Anti-Photoaging Effect in UVB-Irradiated Fibroblasts.

Skin photoaging due to ultraviolet B (UVB) exposure generates reactive oxygen species (ROS) that increase matrix metalloproteinase (MMP). Chlorin e6-photodynamic therapy (Ce6-PDT), in addition to being the first-line treatment for malignancies, has been shown to lessen skin photoaging, while curcumin is well known for reducing the deleterious effects of ROS. In the current study, PDT with three novel Ce6-curcumin derivatives, a combination of Ce6 and curcumin with various linkers, including propane-1,3-diamine for Ce6-propane-curcumin; hexane-1,6-diamine for Ce6-hexane-curcumin; and 3,3'-((oxybis(ethane-2,1-diyl))bis(oxy))bis(propan-1-amine) for Ce6-dipolyethylene glycol (diPEG)-curcumin, were studied for regulation of UVB-induced photoaging on human skin fibroblast (Hs68) and mouse embryonic fibroblast (BALB/c 3T3) cells. We assessed the antiphotoaging effects of Ce6-curcumin derivatives on cell viability, antioxidant activity, the mechanism of matrix metalloproteinase-1 and 2 (MMP-2) expression, and collagen synthesis in UVB-irradiated in vitro models. All three Ce6-curcumin derivatives were found to be non-phototoxic in the neutral red uptake phototoxicity test. We found that Ce6-hexane-curcumin-PDT and Ce6-propane-curcumin-associated PDT exhibited less cytotoxicity in Hs68 and BALB/c 3T3 fibroblast cell lines compared to Ce6-diPEG-curcumin-PDT. Ce6-diPEG-curcumin and Ce6-propane-curcumin-associated PDT showed superior antioxidant activity in Hs68 cell lines. Further, in UVB-irradiated in vitro models, the Ce6-diPEG-curcumin-PDT greatly attenuated the expression levels of MMP-1 and MMP-2 by blocking mitogen-activated protein kinases (MAPKs), activator protein 1 (AP-1), and tumor necrosis factor-α (NF-κB) signaling. Moreover, Ce6-diPEG-curcumin effectively inhibited inflammatory molecules, such as cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expression, while accelerating collagen synthesis. These results demonstrate that Ce6-diPEG-curcumin may be a potential therapy for treating skin photoaging.

Photodynamic therapy potential of cobalt phthalocyanine in triple-negative breast cancer

Triple-negative breast cancer (TNBC) has the worst prognosis among breast cancers. The deficiency of hormone receptors (ER-/PR-/HER-2-) limits treatment options. Photodynamic therapy (PDT) is a safe treatment method for target-specific applications and has no toxic side effects. In this study, the efficacy of PDT on TNBC cells after activation of phthalocyanine derivative CoPc-COOH with RED light was investigated. The cytotoxicity of CoPc-COOH in MDA-MB-231 cells was evaluated at 24, 48, and 72 h. To detect phototoxic activity, cells were exposed simultaneously to a dose range of CoPc-COOH at 48th hours and 20, 30, 40, and 50 J/cm2 RED light in a laser source with an output fluorescence of 12 mW/cm2. The apoptotic effect of single CoPc-COOH, RED light, and their combinations were examined by using the Annexin V method and the ROS generation potential using the carboxy-H2-DCFH-DA test in flow cytometry. DCF fluorescence intensity of control and dose groups were taken under the fluorescence microscope. The IC50 values of CoPc-COOH in MDA-MB-231 cells at the 24th, 48th, and 72nd hours were calculated as 131.04 µM, 66.75 µM, and 59.74 µM, respectively. In the phototoxicity test, 50 J/cm2 application increased the IC50 dose of CoPc-COOH to 73.73 µM; 20, 30, 40 J/cm2 application reduced the doses to 2.31 µM, 0.20 µM, and 28.14 µM, respectively. Compared to the untreated group, it was determined that ROS formation and apoptotic stimulation were triggered significantly in treatment groups. A combination of CoPc-COOH and 40 J/cm2 showed the highest synergistic effect.

Cellular photo(geno)toxicity of gefitinib after biotransformation.

Gefitinib (GFT) is a selective epidermal growth factor receptor (EGFR) inhibitor clinically used for the treatment of patients with non-small cell lung cancer. Bioactivation by mainly Phase I hepatic metabolism leads to chemically reactive metabolites such as O-Demethyl gefitinib (DMT-GFT), 4-Defluoro-4-hydroxy gefitinib (DF-GFT), and O-Demorpholinopropyl gefitinib (DMOR-GFT), which display an enhanced UV-light absorption. In this context, the aim of the present study is to investigate the capability of gefitinib metabolites to induce photosensitivity disorders and to elucidate the involved mechanisms. According to the neutral red uptake (NRU) phototoxicity test, only DF-GFT metabolite can be considered non-phototoxic to cells with a photoirritation factor (PIF) close to 1. Moreover, DMOR-GFT is markedly more phototoxic than the parent drug (PIF = 48), whereas DMT-GFT is much less phototoxic (PIF = 7). Using the thiobarbituric acid reactive substances (TBARS) method as an indicator of lipid photoperoxidation, only DMOR-GFT has demonstrated the ability to photosensitize this process, resulting in a significant amount of TBARS (similar to ketoprofen, which was used as the positive control). Protein photooxidation monitored by 2,4-dinitrophenylhydrazine (DNPH) derivatization method is mainly mediated by GFT and, to a lesser extent, by DMOR-GFT; in contrast, protein oxidation associated with DMT-GFT is nearly negligible. Interestingly, the damage to cellular DNA as revealed by the comet assay, indicates that DMT-GFT has the highest photogenotoxic potential; moreover, the DNA damage induced by this metabolite is hardly repaired by the cells after a time recovery of 18h. This could ultimately result in mutagenic and carcinogenic effects. These results could aid oncologists when prescribing TKIs to cancer patients and, thus, establish the conditions of use and recommend photoprotection guidelines.

56 Monitoring Diesel Exhaust Particles by Lock-in Thermography

Abstract Diesel exhaust particles (DEPs) can deposit onto the respiratory epithelial surface upon inhalation. In vitro exposures of various lung models to DEPs have been performed under the air-liquid interface (ALI) conditions in previous studies, however, methods to quantify DEPs onto/into cells during and after exposure are still challenging and not well established. In our study, a new particle detection system, i.e. lock-in thermography (LIT), was explored. LIT is a heat- sensitive imaging method that applies the light illumination to induce the heat of carbon-based particles for detection. By testing a standard DEPs sample (SRM2975), a positive linear relationship (R2 = 0.98) was established between the thermal emission signals and DEPs levels ranging from 0 to 30 µg/mL, with limit of detection (LOD) at 1.00 µg/mL. Moreover, phototoxicity tests showed that no cytotoxic effects were observed on human lung epithelial (A549) cells upon excitation light wavelength of 525 and 660 nm for 1 – 2 mins. Taken together, it suggests that the LIT can be used for detecting DEPs exposed with A549 cells at the relatively low concentration. Further improvement and optimization of the LIT system are ongoing to evaluate DEPs exposed lung cells.

Development of a GCN-based model to predict <i>in vitro</i> phototoxicity from the chemical structure and HOMO-LUMO gap

The interaction between sunlight and drugs can lead to phototoxicity in patients who have received such drugs. Phototoxicity assessment is a regulatory requirement globally and one of the main toxicity screening steps in the early stages of drug discovery. An in silico-in vitro approach has been utilized mainly for toxicology assessments at these stages. Although several quantitative structure-activity relationship (QSAR) models for phototoxicity have been developed, in silico technology to evaluate phototoxicity has not been well established. In this study, we attempted to develop an artificial intelligence (AI) model to predict the in vitro Neutral Red Uptake Phototoxicity Test results from a chemical structure and its derived information. To accomplish this, we utilized an open-source software library, kMoL. kMoL employs a graph convolutional neural networks (GCN) approach, which allows it to learn the data for the specified chemical structure. kMoL also utilizes the integrated gradient (IG) method, enabling it to visually display the substructures contributing to any positive results. To construct this AI model, we used only the chemical structure as a basis, then added the descriptors and the HOMO-LUMO gap, which was obtained from quantum chemical calculations. As a result, the assortment of chemical structures and the HOMO-LUMO gap produced an AI model with high discrimination performance, and an F1 score of 0.857. Additionally, our AI model could visualize the substructures involved in phototoxicity using the IG method. Our AI model can be applied as a toxicity screening method and could enhance productivity in drug development.

Flow cytometry as an alternative method to evaluate genotoxicity of natural cosmetic actives.

To suggest the expansion of the use of in vitro safety techniques recommended by the OECD guidelines and to propose the use of the automation of the in vitro mammalian micronucleus test method by flow cytometry to assess the genotoxic potential of Centella asiatica, Horse Chestnut, Witch Hazel, Blend, Ecoblend, and Caffeine extracts due to their widespread use in commercial products. Flow cytometer analysis was performed using the Accuri™ C6 equipment and analyzed using the FlowJo software. Cytotoxicity tests followed OECD 129 guidelines and Phototoxicity followed OECD/GD 432 guidelines. The results showed that the cytotoxicity assay presented a decrease in cell viability when cells were exposed to Centella asiatica from a concentration of 5.0%, horse chestnut 2.5%, Witch hazel 2.5%, Blend 3.13%, and Caffeine 3%, while Ecoblend at the tested concentrations did not show cytotoxicity. In the phototoxicity test, the samples at the tested concentrations showed a PIF <2 being considered potentially non-phototoxic. Finally, in the genotoxicity automated assay, samples were considered potentially non-genotoxic. In vitro methods are of paramount importance for the development of pre-clinical tests and the use of test automation helps to reduce the time for analysis and dissemination of results, being a determining factor for the prospect of new compounds.

Absence of phototoxicity/photoirritation potential of bergamottin determined In Vitro using OECD TG 432

The phototoxic potential of a number of furocoumarins is well established. On the other hand, studies have shown that bergamottin, a furocoumarin containing a bulky, hydrophobic side chain, has significantly less or is even absent of phototoxicity potential. The OECD Test Guideline 432 3T3/Neutral Red Uptake (NRU) in vitro phototoxicity test has shown to be a highly predictive test for identifying compounds that exhibit no phototoxicological potential. In this study using OECD 432, the established phototoxic furocoumarin 5-methoxypsoralen (5-MOP), 8-methoxypsoralen (8-MOP) and psoralen were phototoxic, whereas bergamottin showed no phototoxic potential. When compared to 5-MOP, 8-MOP and psoralen, bergamottin was clearly negative at molar-adjusted concentrations that were more than 9 times higher than those that produced phototoxicity in 8-MOP; nearly 16 times than those for psoralen and more than 36 times higher than those for 5-MOP. These data using in vitro 3T3 NRU Phototoxicity Test (OECD 432) are supportive of earlier studies showing bergamottin does not exhibit phototoxicological properties. The detection and quantification of bergamottin should therefore not contribute to the potential marker furocoumarins for risk management interventions intended to reduce the phototoxicity of natural furocoumarin containing preparations.

A proposed NAM-based tiered phototoxicity testing and human risk assessment framework for agrochemicals

Phototoxicity testing is required by European regulations for agrochemicals with UV/visible molar extinction/absorption coefficient (MEC) higher than 10 L x mol−1 x cm−1 in the 290–700 nm wavelength range. Furthermore, regulations identify a need of considering human exposure in case of positive results. While in vitro OECD test guidelines are available for hazard characterisation, there is no guidance on how to utilise positive results in human exposure risk assessments. Our goal was to take a first step towards developing a NAM based tiered testing approach and a framework for non-dietary acute human dermal risk assessment for phototoxicity to agrochemicals. The proposed framework can be divided into a few steps: 1) use the OECD updated MEC values of 1000 L x mol−1 x cm−1 as trigger for phototoxicity testing; 2) establish a reference concentration (RfC) from in vitro phototoxicity studies using BMC approach, 3) estimate potential exposure to skin, target organ for phototoxicity, using EFSA exposure models, product specific labels and skin penetration values, and 4) phototoxicity risk assessment; 5) refinement to RfC and/or exposure estimates can be considered. Finally, case studies of a nematicide and an herbicide active substance are provided to illustrate the proposed framework.

Validation of a Light Source for Phototoxicity in in vitro Conditions

Phototoxicity is a chemically induced skin irritation in the presence of sunlight. Phototoxic substances after irradiation with sunlight absorb and convert light energy into chemical energy, leading to the formation of highly reactive oxygen species with toxic effects. Recently LED technologies made possible their application for novel effective solar simulators, to be used in biology and medicine. We study the possibilities of application of LED sunlight simulator (Helios-iO, model LE-9ND55-H – 5500K) for in vitro testing of synthetic and natural products for phototoxicity. Cytotoxicity/phototoxicity was assessed by validated BALB/3T3 clone A31 Neutral Red Uptake Assay. The morphological alterations in BALB/3T3 cells induced by the Radachlorin® and irradiated with dose 2.4 J, were analyzed by fluorescent microscopy. The physical characteristics of the lamp were determined and the intensity distribution of the LED light over a model of a 96-well plate at a distance of 25 cm from the lamp diode matrix center was estimated. The obtained results show that the light emitted by the solar simulator used is similar to the natural sun light. The biological testing results reveal the high efficiency of the solar simulator in an in vitro experimental system for phototoxicity testing.

Evaluation of phototoxicity induced by the anticancer drug rucaparib

Rucaparib (RCP) is a potent selective inhibitor of both PARP-1 and PARP-2 enzymes that induces synthetic lethality in cancer cells. It is used for the treatment of breast and ovarian tumors harboring deleterious germline or somatic cancer susceptibility genes mutations. Although RCP has an indole chromophore in its structure, it displays a bathochromic shift of the absorption band towards the UVA region of sunlight, thus extending the active fraction of solar light able to produce photosensitivity reactions. In this context, it is highly interesting to study the photo(geno)toxicity disorders associated with this drug, bearing in mind that, for dermatologists it is crucial to understand the toxicity mechanism to improve clinical management. In the present work, RCP has shown to be potentially phototoxic, as observed in the neutral red uptake phototoxicity test. Moreover, this significant phototoxicity is attributed to both proteins and genomic DNA, as revealed in the protein photooxidation and comet assays. The results obtained are highly relevant concerning RCP photosafety and become clinically important in the context of identification of the cutaneous adverse events that can be associated with the targeted therapies. Interestingly, this is the first example of a PARP inhibitor able to induce photosensitized damage to biomolecules.

Determination of the Phototoxicity Potential of Commercially Available Tattoo Inks Using the 3T3-neutral Red Uptake Phototoxicity Test

Tattooing is an ancient practice and its popularity has been increasing in the recent years. After tattooing, complications may occur related to compose tattoo inks. In this study, the phototoxicity potential of the blue, red and black colors of the most commonly used three different commercially-available tattoo ink brands have been examined by performing in vitro 3T3-neutral red uptake (NRU) phototoxicity test. In the study, the phototoxicity of serial diluted concentrations of tattoo inks were evaluated with in vitro 3T3-NRU phototoxicity test method according to OECD guide 432. The data obtained from the NRU test result were uploaded to Phototox software (version 2.0) and the phototoxicity potentials of tattoo inks were determined via the calculation of the mean photo effect (MPE) and photo irritation factor (PIF) values. The red, black and blue colors of three different commercially available tattoo inks did not cause a cytotoxic activity on BALB/c 3T3 cells with 3T3-NRU test. The IC50 values could not be determined +ultraviolet (UV) and -UV conditions. PIF values could not be calculated and MPE values were <0.1, which predicts the absence of phototoxic effect for all of the tested tattoo inks. All tested inks were evaluated as non-phototoxic according to the results of MPE values calculated using Phototox software. However, test results should be verified by other phototoxicity test methods to obtain a comprehensive evaluation of phototoxic complications of different tattoo inks.

Safety and efficacy of combined essential oils for the skin barrier properties: In vitro, ex vivo and clinical studies.

To evaluate the safety and the synergistic effects of tea tree, lavender, eucalyptus and tangerine essential oils in combination on the skin using in vitro, ex vivo and clinical studies. The phototoxicity was predicted using 3T3 neutral red uptake phototoxicity test (OECD TG 432). Skin penetration was evaluated by confocal Raman microspectroscopy using direct application of essential oils to pig ears. For the clinical studies, 40 participants were enrolled and randomized in three groups: (1) lavender, eucalyptus and tangerine, (2) the same essential oils plus melaleuca and (3) placebo group. The skin was evaluated by noninvasive techniques before and after a 90-day period of topical use. The essential oils were non-phototoxic, but the tangerine oil showed dose-dependent cytotoxicity (IC50: 33.1µg/ml), presenting 35% of penetration in the viable epidermis. On the contrary, 17.7µg/ml in combination was applied per day in the clinical study and the penetration rate for the combinations (10%, 1.77µg/ml achieving the viable epidermis) guaranteed the safety, since in the clinical study, the application of the four essential oils improved skin barrier and morphologic skin characteristics, as well as increased skin hydration and decreased sebum levels, with no unwanted effects reported. All essential oils studied were considered non-cytotoxic or non-phototoxic separately except tangerine, which present a dose-dependent cytotoxicity. Finally, the essential oils in combination in an appropriate amount were safe and effective in the improvement of the hydrolipidic balance and morphological properties of the skin.