Clonidine: irritant and allergic contact dermatitis assays

Clonidine in petrolatum, clonidine transdermal device, and its placebo device were assayed for relative irritancy potential (21-day cumulative irritancy assay) and allergic contact dermatitis potential (Draize repeat insult patch test assay). Both clonidine in petrolatum and the clonidine transdermal device appear to be of minimal irritancy potential. Only the clonidine device demonstrated, in the Draize assay (and in clinical trials), allergic contact dermatitis potential. Its relatively delayed onset of demonstration of allergic contact sensitization in clinical trials requires investigation as to mechanism.     

Identification of contact sensitizers by animal assay

Guinea pig testing constitutes the first step in evaluating the allergenicity of new chemicals and products. Some of the most commonly used animal predictive tests are reviewed. The guinea pig maximization test, which is the recommended test method in Sweden, is described in detail and the interpretation of results obtained with this test is discussed. In the guinea pig maximization test the sensitization capacity of a substance is examined by the use of maximized conditions for i he exposure, i.e. the potential ability of the material to induce a contact allergy is determined. The extent to which an allergen causes contact dermatitis in exposed persons depends on the mode of use and various environmental factors.     

Route of contact sensitization and in vitro lymphocyte transformation

A guinea pig skin extract conjugate with dinitrofluorobenzene elicited significant in vitro transformation of cultured lymphnode lymphocytes from 19 of 27 guinea pigs sensitized by footpad injection of dinitrochlorobenzene in Freund's complete adjuvant, as compared to only 1 of 26 guinea pigs topically sensitized to dinitrochlorobenzene. Topically sensitized guinea pigs appear to be more appropriate models for contact allergy in man than guinea pigs sensitized by other methods. Other sensitization procedures are likely to produce more heterogeneous forms of sensitization, with features of contact allergy, tuberculin-type allergy, antibody-mediated hypersensitivity and cutaneous basophile hypersensitivity.     

A new animal model for contact dermatitis: the hairless guinea pig.

Allergic and irritant contact reactions were evaluated in the recently identified hairless guinea pig, Crl:IAF(HA)BR, a mutant from the Hartley strain. The cutaneous changes were observed macro- and microscopically. The irritant contact dermatitis was induced by croton oil, 2,4-dinitrochlorobenzene (DNCB), or anthralin. Both hairless and hairy guinea pigs developed similar reactions to these chemicals. The density of the epidermal Langerhans cells (LC) of hairless guinea pigs was significantly higher than that in the hairy strain. Allergic contact sensitization was easily induced with DNCB. Photoallergic contact sensitization was also induced with tetrachlorosalicylanilide (TCSA) but not with tribromosalicylanilide (TBS). However, by administration of cyclophosphamide before sensitization, positive photocontact responses were seen with TBS. These results indicate that hairless guinea pigs can be used as animal models for investigation of immunologic and nonimmunologic contact reactions.     

Contact and respiratory sensitization by chemical allergens: Uneasy relationships

A variety of chemicals are able to cause allergic disease in susceptible individuals. Chemical-induced allergy may take several forms, chief among them being skin sensitization or allergic contact dermatitis and respiratory hypersensitivity. It has been found that contact and respiratory chemical allergens induce in mice qualitatively divergent immune responses consistent with the preferential activation of discrete Thelper (Th) cell subpopulations, Th1 and Th2 cells, respectively. Despite the selective stimulation of Th2type responses by chemical respiratory allergens, such materials are nevertheless able to induce contact sensitization in mice, which is a form of delayed-type hypersensitivity usually considered to be effected by Th1 cells. Indeed, the available evidence indicates that chemicals known to cause respiratory allergy in humans test positive in both mouse and guinea pig assays used for the prospective identification of skinsensitizing chemicals. The activity of chemical respiratory sensitizers in such tests has important-implications for the toxicological evaluation and classification of potential chemical allergens. Paradoxically, however, although chemical respiratory allergens show a potential for contact sensitization in predictive tests, this hazard rarely translates into an important risk of allergic contact dermatitis in humans.     

A review of contact dermatitis associated with transdermal therapeutic systems

The literature is reviewed for contact dermatitis associated with transdermal therapeutic systems. Clonidine, nitroglycerin, scopolamine, estradiol and testosterone are utilized in such applications, and fentanyl is under investigation. Most cutaneous reactions are limited to localized dermatitis; however, generalized systemic effects may occur. Investigators are reporting skin-related side-effects in up to 50% of patients using transdermal clonidine; however, with the other agents, this is demonstrated much less frequently. Reactivation of the dermatitis via oral medication, following sensitization to the patch, is noted in rare instances.     

Oral substitution in patients sensitized by transdermal clonidine treatment

The safety of oral clonidine substitution in 29 patients with confirmed localized allergic contact dermatitis due to transdermal clonidine treatment was evaluated. Only 1 patient had a skin reaction to oral clonidine. The dose of topical clonidine-inducing sensitization, and the oral challenge dose administered later, appear to be insufficient to elicit a systemic reaction in most patients. An examination of current patch test technology seems indicated in view of the fact that a number of patients who gave a negative reaction to 48-h applications of 9% clonidine in petrolatum responded positively to 7-day application of transdermal clonidine.     

Sensitization of mice to topically applied drugs: albuterol chlorpheniramine, clonidine and nadolol

Allergic contact dermatitis from drugs is a significant obstacle to the development of transdermal drug delivers systems. Protocols for the sensitization of mice to drugs are needed to test methods for the prevention of allergic contact dermatitis. CBA/J female mice were sensitized to the drugs albuterol, chlorpheniramine, clonidine and nadolol by topical application. Sensitization was achieved by application of drug at 5% (w/v) 10 shaven dorsal skin for 5 days in a hydroxyethylcellulose vehicle. Contact serialization was determined by measuring the car swelling response to application of l%. drug in vehicle. Control mice treated by application of vehicle alone did not exhibit an ear swelling response to drug. Supplementation of the mice with vitamin A boosted the ear swelling response, as did application of drug to dorsal versus abdominal skin. Although plasma amounts of retinol were higher in vitamin A supplemented versus control mice, the rate of drug (albuterol and nadolol) permeation was not significantly different between vitamin A supplemented and control mice. Permeability of dorsal skin for nadolol was twice that of ventral skin, which may explain the differences in sensitization at these sites. This sensitization protocol should be useful in the development of hypoallergenic transdermal drug delivery systems.     

Immunosuppressive effects of clonidine on the induction of contact sensitization in the balb/c mouse

The clonidine transdermal therapeutic system (clonidine-TTS) has been associated with a significant incidence of allergic contact sensitization. This incidence was not predicted by premarket skin sensitization testing in animals or humans. One possible explanation lies in recent findings in guinea pigs that clonidine exposure could inhibit the elicitation of skin reactions to unrelated strong contact sensitizers. However, these studies also showed that clonidine pretreatment did not appear to affect the induction of contact sensitization. On this basis, we sought to specifically evaluate the induction phase of sensitization to clonidine as an alternative means of assessing its sensitization properties. The method selected was the assay of in situ lymphocyte proliferation in lymph nodes draining the sites of clonidine exposure, a method recently promoted as an alternative means to assess contact allergenic potential. Utilizing various induction application techniques and regimens, we were consistently unable to demonstrate clonidine's allergenic potential through such an assessment of lymphocyte proliferation. We were also unable to demonstrate sensitization by in vivo ear swelling or in vitro lymphocyte blastogenesis assay techniques. However, a subsequent assessment of the effect of clonidine exposure on the induction of sensitization to unrelated strong contact allergens demonstrated a consistent 40-70% inhibition of the proliferative response to the contact allergens oxazolone and trinitrochlorobenzene. This was similar to the degree of suppression produced by the corticosteroids fluocinonide and hydrocortisone when they were tested at 80 and 10 times lower concentrations. In addition, we observed a comparable inhibition of the ear swelling response to oxazolone. These data extend our knowledge of the immunomodulatory effects of clonidine and offer additional mechanistic insights into the failure of short-term predictive patch-test methods to detect this chemical's potential to induce allergic contact sensitization.     

Adverse skin reactions to nicotine in a transdermal therapeutic system

Nicotine in a transdermal therapeutic system (TTS) has been introduced recently to help people to stop smoking. 14 volunteers (10 male, 4 female) with a history of former adverse skin reactions to this device were investigated. Skin tests for contact urticaria and patch tests for contact allergy were done with the individual components of the TTS. Contact sensitization to nicotine was identified in 5 and to the TTS itself in 1 individual. Irritant reactions due to occlusion were present in 9 subjects. The optimal test agent and concentration for elucidating the adverse skin reaction was an aqueous solution of 10% nicotine base. Atopy, as diagnosed by history and skin prick tests to common inhalant allergens, and contact sensitization to standard patch test allergens, were each identified in 6 subjects. Nicotine should be added to the expanding list of transdermally delivered drugs which may elicit contact dermatitis.     

A single accidental exposure may result in a chemical burn, primary sensitization and allergic contact dermatitis

It is known from experimental studies that antigenic potency and the concentration of antigen determine whether exposure to an antigen will result in sensitization. A single accidental exposure to concentrated antigen may therefore induce primary sensitization. The purpose of this report was to collect clinical cases in which a single exposure had resulted in contact dermatitis suspected to be allergic. Only patients without previous relevant skin symptoms were included. Patch testing was used to demonstrate sensitization. 6 patients developed occupational allergic contact dermatitis from accidental exposure. Patch testing revealed allergy to diglycidylether of bisphenol A epoxy resin, poly functional aziridine hardener, methyl acrylate, phenol-formaldehyde resin, and methylchloroisothiazolinone/methylisothiazolinone (Kathon LX), respectively. Furthermore, 2 patients developed allergic contact dermatitis from their first exposure to tear gas chemicals, namely omega-chloroacetophenone and ortho-chlorobenzylidene malonitrile. A single exposure can therefore induce both sensitization and subsequent allergic contact dermatitis without further exposure. The allergens described must be considered strong allergens. The skin should immediately be cleaned if an accidental splash with such an allergen has taken place.     

Skin lesions due to okra (Hibiscus esculentus L): proteolytic activity and allergenicity of okra

The present report describes experimental studies on the proteolytic activity of the secretion on the surface of okra pods and the allergenicity of okra components, to clarify the etiology of skin lesions due to okra. Proteolytic activity was detected on the surface of immature okra pods and seemed to be sufficient to cause the skin lesions. Further, in vivo, intradermal injection of the enzyme solution prepared from immature okra pods led to increased capillary permeability in guinea pigs, in contrast to heated preparations. The fraction purified by preparative paper chromatography from an ethyl acetate extract of okra pods showed moderate allergenicity in the guinea pig maximization test. The present experimental evidence supports our suggestions from previous surveys that the proteolytic enzyme of okra may be responsible for development of skin lesions, and that allergic contact dermatitis may also play a part in addition to irritant contact dermatitis.     

Use of the local lymph node assay in the evaluation of the sensitizing potential of pharmaceutical process intermediates

The murine local lymph node assay (LLNA) has recently been developed to determine the contact sensitization potential of chemicals. Since its original development, the LLNA results have been the subject of extensive comparisons with guinea pig and human data. The investigations described here were designed to explore the ability of the LLNA to identify accurately, pharmaceutical process intermediates (PIs) known to cause contact allergy in humans. To that end, 16 PIs previously tested in the guinea-pig maximization test (GPMT) were tested in the LLNA. Another PI known to be a contact sensitizer in humans was tested only in the LLNA. Cases of contact sensitization in humans were reported only for PIs that were extreme sensitizers in the GPMT and had low EC3 values (concentration of the test substance required to generate a threefold increase in lymph node cell proliferation) in the LLNA. These data provide additional evidence that the LLNA is able to discriminate skin sensitizers from chemicals that do not possess a significant skin sensitization potential and is thus a useful method for hazard identification. In addition, this method also offers important animal welfare benefits and may also be useful for risk assessment purposes.     

Activity of human contact allergens in the murine local lymph node assay

The murine local lymph node assay (LLNA) is a predictive test for the identification of chemicals that have the potential to cause skin sensitization. Since its original development, the assay has been the subject of national and international evaluation studies and extensive comparisons with guinea pig tests and human data. On the basis of these investigations, the LLNA has recently been endorsed by ICCVAM (Interagency Coordinating Committee on the Validation of Alternative Methods) as a stand-alone method for skin sensitization hazard identification. At the same time, ICCVAM confirmed that, although the LLNA is not an in vitro method, it does represent a refinement in the way animals are used and can provide a means for reducing the number of animals used in sensitization hazard assessment. The investigations described here were designed to explore further the ability of the LLNA to identify accurately those chemicals that cause allergic contact dermatitis in humans. To that end we have measured, amongst 3 independent laboratories, LLNA responses induced by a total of 18 test chemicals, 11 of which are known to cause skin sensitization and 7 of which are believed not to be associated with any significant evidence of allergic contact dermatitis in humans. The LLNA correctly classified 16 of the 18 materials. The 11 chemicals tested which are associated with allergic contact dermatitis in humans were found to be positive in the LLNA. Of the 7 materials believed to be non-sensitizers, 5 were negative in the LLNA and 2 produced positive results. Collectively, these data provide additional evidence that the LLNA is able to discriminate skin sensitizers from those chemicals which do not possess a significant skin sensitization potential and thus provides a method for hazard identification that offers important animal welfare benefits.     

Experimental nickel sensitization in the guinea pig: comparison of different protocols

3 different sensitization protocols were compared for inducing delayed-type nickel contact hypersensitivity in guinea pigs. Open epicutaneous sensitization (OE) induced nickel allergy in 11/22 (50%) guinea pigs. When intradermal injections of Freund's complete adjuvant into the nickel-painted skin was added to the same protocol. 4/13 (31 %) became sensitized. The guinea pig maximization protocol induced nickel allergy in only 7/31 (23%) of the animals. Compared with the 2 other methods, animals sensitized with open epicutaneous applications reacted more rapidly (maximum at 6 h) and strongly (2+ reaction in 12/22 of animals) in previous patch lest sites upon systemic (i p.) nickel challenge. Open epicutaneous sensitization of guinea pigs should he a useful model for studying cellular and immunological mechanisms in nickel contact sensitivity.     

Sensitizing potential in mice, guinea pig and man of the preservative Euxyl K 400 and its ingredient methyldibromo glutaronitrile

The allergenicity of the preservative Euxyl K 400 and its principal allergen methyldibromo glutaronitrile (MDBGN) (1,2-dibromo-2,4-dicyanobutane) was investigated using 3 animal models; in mice, the local lymph node assay (LLNA) and in guinea pigs, the guinea pig maximization test (GPMT) and the cumulative contact enhancement test (CCET) with a dose-response protocol included. Previous attempts to define the sensitization capacity of these chemicals have given conflicting results. For comparison, the frequency and causes of positive patch test reactions to Euxyl K 400 and MDBGN were studied in patients referred to an occupational dermatology clinic. This investigation showed that Euxyl K 400 and MDBGN can give rise to contact allergy in man and that the relevant cases found mainly had similar exposure as non-occupational cases. A contact allergenic potential could be detected for MDBGN in 2 animal models, i.e., the CCET and the LLNA, and also for Euxyl K 400 in the LLNA. However, statistical analysis of the results from the GPMT with MDBGN failed to detect the sensitizing potential of this particular allergen. The results indicate that to be able to detect the allergenic potential of Euxyl K 400 and MDBGN, a predictive test method with multiple topical applications at induction is required. It is therefore important that an investigator is aware of the possibility of using various predictive test models for investigation of potential contact allergens.     

Testing of controlled-release transdermal dosage forms. Product development and clinical trials

In vitro techniques have been developed that are predictive of drug permeation through skin. Methods also have been developed to determine the skin irritation potential of a drug substance during the preclinical stages of transdermal dosage form development. Clinical studies are performed to test for skin irritation, contact sensitization, plasma levels of drug, and efficacy of the dosage form. In theory, these studies should be predictive of a dosage form's performance in routine clinical use. This may not be the case, however; contact sensitization cannot be predicted with absolute certainty beforehand in animals. This article discusses various aspects of testing transdermal dosage forms and how the results of these tests correlate with what happens when a transdermal dosage form is put into routine clinical use.     

Sensitization patterns in Compositae‐allergic patients with current or past atopic dermatitis

Background. An association between Compositae sensitization and atopic dermatitis has been suggested on the basis of case reports and clinical studies. Objectives. To describe the characteristics of sensitization in Compositae‐allergic patients with current and/or past atopic dermatitis. Patients/materials/methods. Consecutive Compositae‐sensitive patients were selected for analysis if they had a history of (i) present and/or past atopic dermatitis or (ii) childhood flexural eczema or (iii) childhood eczema of any kind and a positive prick test result. Results. Fifty‐one persons (35 females and 16 males) were included. The mean age was lower and the percentage of males was slightly higher than in non‐atopics. Testing with sesquiterpene lactone mix, parthenolide and Compositae mix 6% or 5% detected 96% of the patients. Occupational sensitization occurred in 22%. The sensitizing pattern did not differ much from that of non‐atopics, except that dandelion was an important allergen in children. Cobalt allergy was the most frequent other contact allergy, occurring in 37%. Conclusions. Persons with current or past atopic dermatitis may become sensitized to Compositae at any age, both occupationally and non‐occupationally. They should be screened for Compositae allergy on equal terms with non‐atopics, except that dandelion extract should always be tested in children. Co‐sensitization to cobalt was frequent, but probably not related to the plant allergy.     

Genetic factors in nickel allergy evaluated in a population-based female twin sample

Environmental exposures are important for development of allergic contact dermatitis, but genetic factors have been proposed to be of additional importance for contact sensitization. Recently genetic factors were shown to be of significance for hand eczema. In this study, a sample of twins recruited on the basis of hand eczema has been evaluated with respect to influence of genetic factors on development of nickel sensitization. A total of 1076 individual twins were patch tested and underwent clinical examination, and in the final genetic statistical analysis 630 females were available, of which 146 had a positive patch test to nickel. The aggregation of nickel allergy among twin pairs was measured by the casewise concordance and the twin odds ratio. The twin odds ratio were adjusted for effects of risk factors known to be associated with nickel allergy, namely, wet work, atopic dermatitis, and self-reported hand eczema. There was a small tendency for larger odds ratio among monozygotic twins than among dizygotic twins, which was not statistically significant. As a result of the statistical analysis, it is concluded that allergic nickel contact dermatitis is mainly caused by environmental and only to a lesser degree genetic factors. The selection of twins on the basis of hand eczema may theoretically influence the prevalence of nickel allergy and concordance estimates, which should be considered before extrapolating the data to a random population-based twin sample.     

Induction of tolerance to poison ivy urushiol in the guinea pig by epicutaneous application of the structural analog 5-methyl-3-n-pentadecylcatechol

Previous studies have established that epicutaneous application of 5-methyl-3-n-pentadecylcatechol (5-Me-PDC), a synthetic analog of a poison ivy urushiol component, leads to immune tolerance to 3-n-pentadecylcatechol (PDC) in mice. The induction of tolerance by 5-Me-PDC may be mediated by a protein conjugate formed via selective reaction of thiol nucleophiles present on the carrier macromolecule with the corresponding o-quinone derived from the parent catechol. In order to examine further the tolerogenic properties of 5-Me-PDC, we have extended our studies to the guinea pig, the generally accepted experimental species for the study of contact allergy. The results have established that specific immune tolerance to poison ivy urushiol is induced following 2 epicutaneous applications of the PDC analog. Furthermore, we were able to show that the treated animals remained tolerant for at least 6 weeks, a period of time comparable to that observed following the intravenous administration of the O,O-bis-acetyl derivative of PDC. The data point to the possibility of developing a therapeutically effective topical tolerogen for poison ivy contact dermatitis.