| 1. |
- Andresen Bergström, Moa, 1978, et al.
(författare)
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Conjugated dienes as prohaptens in contact allergy: in vivo and in vitro studies of structure-activity relationships, sensitizing capacity, and metabolic activation.
- 2006
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Ingår i: Chemical research in toxicology. - : American Chemical Society (ACS). - 0893-228X .- 1520-5010. ; 19:6, s. 760-9
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Tidskriftsartikel (refereegranskat)abstract
- There is a great interest in developing in vitro/in silico methods for the prediction of contact allergenic activity. However, many proposed methods do not take the activation of prohaptens to sensitizers by skin metabolism into account. As a consequence, consumer products containing potent sensitizers could be marketed. To identify prohaptens, studies regarding their structure-activity relationships and the mechanisms of their activation must be conducted. In the present investigation, we have studied the structure-activity relationships for alkene prohaptens. A series of seven alkenes (1-7), all of the same basic structure but with variation in the number and position(s) of the double bond(s), were designed and screened for sensitizing capacity using the murine local lymph node assay. Compounds 1-7 were also incubated with liver microsomes in the presence of glutathione to trap and identify reactive metabolites. The metabolic conversion of three alkenes (9-11) to epoxides (12-15) was also studied along with comparison of their sensitizing capacity. Our results show that conjugated dienes in or in conjunction with a six-membered ring are prohaptens that can be metabolically activated to epoxides and conjugated with GSH. Related alkenes containing isolated double bonds and an acyclic conjugated diene were shown to be weak or nonsensitizers. For the first time, the naturally occurring monoterpenes alpha-phellandrene, beta-phellandrene, and alpha-terpinene were demonstrated to be prohaptens able to induce contact allergy. The difference in sensitizing capacity of conjugated dienes as compared to alkenes with isolated double bonds was found to be due to the high reactivity and sensitizing capacity of the allylic epoxides metabolically formed from conjugated dienes. We recommend that these structure-activity relationship rules are incorporated into in silico predictive databases and propose that the prediction of contact allergenic activity of suspected prohaptens is based on assessment of susceptibility to metabolic activation and chemical reactivity of potential metabolites.
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| 2. |
- Andresen Bergström, Moa, 1978, et al.
(författare)
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Metabolic epoxidation of an alpha,beta-unsaturated oxime generates sensitizers of extreme potency. Are nitroso intermediates responsible?
- 2007
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Ingår i: Chemical Research in Toxicology. - : American Chemical Society (ACS). - 0893-228X .- 1520-5010. ; 20:6, s. 927-936
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Tidskriftsartikel (refereegranskat)abstract
- Metabolic activation of inherently nonprotein-reactive compounds (prohaptens) in the skin can lead to development of contact allergy, a chronic skin disease. The prohapten hypothesis has existed for more than 20 years; yet, detailed knowledge regarding the mechanisms of activation as well as what structural moieties can be transformed to protein-reactive sensitizers is still limited. Today, the consideration of cutaneous bioactivation is important when developing nonanimal-based assays for prediction of contact allergenic activity, as only methods that include skin metabolism are able to detect prohaptens as sensitizers. We have studied the mechanism of activation of the prohapten carvoxime (1), a strongly sensitizing but in itself poorly protein-reactive alpha,beta-unsaturated oxime. alpha,beta-Unsaturated oximes represent a novel class of prohaptens, which previously have never been investigated for potential metabolic activation. To identify reactive metabolites formed from (1), liver microsomal incubations in the presence of glutathione were carried out. Putative reactive metabolites were synthesized, and their allergenic activity, chemical reactivity toward nucleophiles, and ability to elicit a contact allergenic response in animals induced with 1 were assessed. We found that 1 is metabolically activated by epoxidation of the allylic carbon-carbon double bond. The alpha,beta-epoxy oxime metabolites were found to be sensitizers of extreme potency in the local lymph node assay and highly reactive toward nucleophilic amino acids and a model peptide. One of the two diastereomeric epoxy metabolites also elicited an allergic reaction in mice sensitized to 1, in the mouse ear swelling test. Furthermore, this study presents strong indications that the basis of the high reactivity and sensitizing capacity observed for the alpha,beta-unsaturated oximes is related to their ability to form highly reactive nitroso intermediates by tautomerization. To our knowledge, the formation of nitrosoalkenes by oxidative metabolism of alpha,beta-unsaturated oximes has not been shown so far.
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| 3. |
- Bauer, Brigitte, 1978, et al.
(författare)
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Modification and expulsion of keratins by human epidermal keratinocytes upon hapten exposure in vitro.
- 2011
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Ingår i: Chemical research in toxicology. - : American Chemical Society (ACS). - 1520-5010 .- 0893-228X. ; 24:5, s. 737-43
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Tidskriftsartikel (refereegranskat)abstract
- Allergic contact dermatitis is the most prevalent form of human immunotoxicity. It is caused by reactive low molecular weight chemicals, that is, haptens, coming in contact with the skin where hapten-peptide complexes are formed, activating the immune system. By using sensitizing fluorescent thiol-reactive haptens, that is, bromobimanes, we show how keratinocytes respond to hapten exposure in vitro and reveal, for the first time in a living system, an exact site of haptenation. Rapid internalization and reaction of haptens with keratin filaments were visualized. Subsequently, keratinocytes respond in vitro to hapten exposure by release of membrane blebs, which contain haptenated keratins 5 and 14. Particularly, cysteine 54 of K5 was found to be a specific target. A mechanism is proposed where neoepitopes, otherwise hidden from the immune system, are released after hapten exposure via keratinocyte blebbing. The observed expulsion of modified keratins by keratinocytes in vitro might play a role during hapten sensitization in vivo and should be subject to further investigations.
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| 4. |
- Bell, Catherine C., et al.
(författare)
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T-cells from HLA-B*57 : 01+ human subjects are activated with abacavir through two independent pathways and induce cell death by multiple mechanisms
- 2013
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Ingår i: Chemical Research in Toxicology. - : American Chemical Society (ACS). - 0893-228X .- 1520-5010. ; 26:5, s. 759-766
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Tidskriftsartikel (refereegranskat)abstract
- Susceptibility to abacavir hypersensitivity has been attributed to possession of the specific human leukocyte antigen allele HLA-B*57:01. HLA-B*57:01-restricted activation of CD8+ T-cells provides a link between the genetic association and the iatrogenic disease. The objectives of this study were to characterize the functionality of drug-responsive CD8+ T-cell clones generated from HLA-B*57:01+ drug-naive subjects and to explore the relationship between abacavir accumulation in antigen presenting cells and the T-cell response. Seventy-four CD8+ clones expressing different Vβ receptors were shown to proliferate and kill target cells via different mechanisms when exposed to abacavir. Certain clones were activated with abacavir in the absence of antigen presenting cells. Analysis of the remaining clones revealed two pathways of drug-dependent T-cell activation. Overnight incubation of antigen presenting cells with abacavir, followed by repeated washing to remove soluble drug, activated approximately 50% of the clones, and the response was blocked by glutaraldehyde fixation. In contrast, a 1 h antigen presenting cell pulse did not activate any of the clones. Accumulation of abacavir in antigen presenting cells was rapid (less than 1 h), and the intracellular concentrations were maintained for 16 h. However, intracellular abacavir was not detectable by mass spectrometry after pulsing. These data suggest that T-cells can be activated by abacavir through a direct interaction with surface and intracellular major histocompatibility complex (MHC) molecules. With the former, abacavir seemingly participates in the MHC T-cell receptor binding interaction. In contrast, the latter pathway likely involves MHC binding peptides displayed as a consequence of abacavir exposure, but not abacavir itself.
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| 5. |
- Busquets, Rosa, et al.
(författare)
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Biomonitoring of Dietary Heterocyclic Amines and Metabolites in Urine by Liquid Phase Microextraction: 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), a Possible Biomarker of Exposure to Dietary PhIP
- 2013
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Ingår i: Chemical Research in Toxicology. - : American Chemical Society (ACS). - 1520-5010 .- 0893-228X. ; 26:2, s. 233-240
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Tidskriftsartikel (refereegranskat)abstract
- Heterocyclic amines (HCAs) are mutagenic/carcinogenic compounds formed at ng/g levels during frying meat or fish. The effect of the normal intake of dietary HCAs in humans and their involvement in the etiology of cancer are currently unknown. In this work, a new extraction method, liquid phase microextraction (LPME) with hollow fibers, and LC-MS/MS have been used for the first time to determine HCAs and metabolites in nonspiked human urine following a single meal of chicken cooked at 180 degrees C for 6 min. The total intake of HCAs was estimated to be 6 mu g, of which 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) accounted for about 1 mu g. The concentrations of PhIP in nonhydrolyzed urine samples ranged from 11.7 to 59.4 pg/g. The total amount of PhIP in urine ranged between 9.3 and 21.1 ng, which corresponds to 0.91-2.1% of the ingested PhIP. In addition, the urine levels of 4'-OH-PhIP (2-amino-1-methyl-6-(4'-hydroxy)phenylimidazo[4,5-b]pyridine) and 5-OH-PhIP (2-amino-1-methyl-6-(5-hydroxy)phenylimidazo[4,5-b]pyridine) also showed a narrow variation between the samples. The analysis of urine samples after acid hydrolysis did not give additional information but showed a notable increase in norharman in some cases. The obtained results suggest PhIP in urine as a possible biomarker of exposure to HCAs and the LPME and LC-MS/MS method as an appropriate strategy to biomonitor HCAs in urine.
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| 6. |
- Carlsson, Henrik, et al.
(författare)
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Adductomic Screening of Hemoglobin Adducts and Monitoring of Micronuclei in School-Age Children
- 2017
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Ingår i: Chemical Research in Toxicology. - : American Chemical Society (ACS). - 0893-228X .- 1520-5010. ; 30:5, s. 1157-1167
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Tidskriftsartikel (refereegranskat)abstract
- Electrophilic compounds/metabolites present in humans, originating from endogenous processes or exogenous exposure, pose a risk to health effects through their reactions with nucleophilic sites in proteins and DNA, forming adducts. Adductomic approaches are developed to screen for adducts to biomacromolecules in vivo by mass spectrometry (MS), with the aim to detect adducts corresponding to unknown exposures from electrophiles. In the present study, adductomic screening was performed using blood samples from healthy children about 12 years old (n = 51). The frequencies of micronuclei (MN) in erythrocytes in peripheral blood were monitored as a measure of genotoxic effect/genotoxic exposure. The applied adductomic approach has been reported earlier by us and is based on analysis of N-terminal valine adducts in hemoglobin (Hb) by liquid chromatography tandem mass spectrometry (LC-MS/MS). High resolution MS was introduced for refined screening of previously unknown N-terminal Hb adducts. Measured adduct levels were compared with MN frequencies using multivariate data analysis. In the 51 individuals, a total of 24 adducts (whereof 12 were previously identified) were observed and their levels quantified. Relatively large interindividual variations in adduct levels were observed. The data analysis (with partial least-squares regression) showed that as much as 60% of the MN variation could be explained by the adduct levels. This study, for the first time, applies the combination of these sensitive methods to measure the internal dose of potentially genotoxic chemicals and genotoxic effects, respectively. The results indicate that this is a valuable approach for the characterization of exposure to chemical risk factors for the genotoxic effects present in individuals of the general population.
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| 7. |
- Carlsson, Henrik, et al.
(författare)
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Characterization of a Hemoglobin Adduct from Ethyl Vinyl Ketone Detected in Human Blood Samples
- 2015
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Ingår i: Chemical Research in Toxicology. - : American Chemical Society (ACS). - 0893-228X .- 1520-5010. ; 28:11, s. 2120-2129
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Tidskriftsartikel (refereegranskat)abstract
- Electrophiles have the ability to form adducts to nudeophilic sites in proteins and DNA. Internal exposure to such compounds thus constitutes a risk for toxic effects. Screening of adducts using mass spectrometric methods by adductomic approaches offers possibilities to detect unknown electrophiles present in tissues. Previously, we employed untargeted adductomics to detect 19 unknown adducts to N-terminal valine in hemoglobin (Hb) in human blood. This article describes the characterization of one of these adducts, which was identified as the adduct from ethyl vinyl ketone (EVK). The mean adduct level was 40 +/- 12 pmol/g Hb in 12 human blood samples; adduct levels from acrylamide (AA) and methyl vinyl ketone (MVK) were quantified for comparison. Using L-valine p-nitroanilide (Val-pNA), introduced as a model of the N-terminal valine, the rate of formation of the EVK adduct was studied, and the rate constant determined to 200 M(-1)h(-1) at 37 degrees C. In blood, the reaction rate was too fast to be feasibly measured, EVK showing a half-life <1 min. Parallel experiments with AA and MVK showed that the two vinyl ketones react approximately 2 x 10(3) times faster than AA. The EVK-Hb adduct was found to be unstable, with a half-life of 7.6 h. From the mean adduct level measured in human blood, a daily dose (area under the concentration-time-curve, AUC) of 7 nMh EVK was estimated. The AUC of AA from intake via food is about 20 times higher. EVK is naturally present in a wide range of foods and is also used as a food additive. Most probably, naturally formed EVK is a major source to observed adducts. Evaluation of available toxicological data and information on occurrence of EVK indicate that further studies of EVK are motivated. This study illustrates a quantitative strategy in the initial evaluation of the significance of an adduct detected through adduct screening.
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| 8. |
- Carlsson, Henrik, et al.
(författare)
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LC–MS/MS Screening Strategy for Unknown Adducts to N-Terminal Valine in Hemoglobin Applied to Smokers and Nonsmokers
- 2014
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Ingår i: Chemical Research in Toxicology. - : American Chemical Society (ACS). - 0893-228X .- 1520-5010. ; 27:12, s. 2062-2070
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Tidskriftsartikel (refereegranskat)abstract
- Electrophilically reactive compounds have the ability to form adducts with nucleophilic sites in DNA and proteins, constituting a risk for toxic effects. Mass spectrometric detection of adducts to N-terminal valine in hemoglobin (Hb) after detachment by modified Edman degradation procedures is one approach for in vivo monitoring of exposure to electrophilic compounds/metabolites. So far, applications have been limited to one or a few selected reactive species, such as acrylamide and its metabolite glycidamide. This article presents a novel screening strategy for unknown Hb adducts to be used as a basis for an adductomic approach. The method is based on a modified Edman procedure, FIRE, specifically developed for LC-MS/MS analysis of N-terminal valine adducts in Hb detached as fluorescein thiohydantoin (FTH) derivatives. The aim is to detect and identify a priori unknown Hb adducts in human blood samples. Screening of valine adducts was performed by stepwise scanning of precursor ions in small mass increments, monitoring four fragments common for the FTH derivative of valine with different N-substitutions in the multiple-reaction mode, covering a mass range of 135 Da (m/z 503-638). Samples from six smokers and six nonsmokers were analyzed. Control experiments were performed to compare these results with known adducts and to check for artifactual formation of adducts. In all samples of smokers and nonsmokers, seven adducts were identified, of which six have previously been studied. Nineteen unknown adducts were observed, and 14 of those exhibited fragmentation patterns similar to earlier studied FTH derivatives of adducts to valine. Identification of the unknown adducts will be the focus of future work. The presented methodology is a promising screening tool using Hb adducts to indicate exposure to potentially toxic electrophilic compounds and metabolites.
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| 9. |
- Chavan, Swapnil, et al.
(författare)
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Predicting Chemical-Induced Liver Toxicity Using High-Content Imaging Phenotypes and Chemical Descriptors : A Random Forest Approach
- 2020
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Ingår i: Chemical Research in Toxicology. - : American Chemical Society (ACS). - 0893-228X .- 1520-5010. ; 33:9, s. 2261-2275
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Tidskriftsartikel (refereegranskat)abstract
- Hepatotoxicity is a major reason for the withdrawal or discontinuation of drugs from clinical trials. Thus, better tools are needed to filter potential hepatotoxic drugs early in drug discovery. Our study demonstrates utilization of HCI phenotypes, chemical descriptors, and both combined (hybrid) descriptors to construct random forest classifiers (RFCs) for the prediction of hepatotoxicity. HCI data published by Broad Institute provided HCI phenotypes for about 30 000 samples in multiple replicates. Phenotypes belonging to 346 chemicals, which were tested in up to eight replicates, were chosen as a basis for our analysis. We then constructed individual RFC models for HCI phenotypes, chemical descriptors, and hybrid (chemical and HCI) descriptors. The model that was constructed using selective hybrid descriptors showed high predictive performance with 5-fold cross validation (CV) balanced accuracy (BA) at 0.71, whereas within the given applicability domain (AD), independent test set and external test set prediction BAs were equal to 0.61 and 0.60, respectively. The model constructed using chemical descriptors showed a similar predictive performance with a 5-fold CV BA equal to 0.66, a test set prediction BA within the AD equal to 0.56, and an external test set prediction BA within the AD equal to 0.50. In conclusion, the hybrid and chemical descriptor-based models presented here should be considered as a new tool for filtering hepatotoxic molecules during compound prioritization in drug discovery.
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| 10. |
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