Conjugated Dienes as Prohaptens in Contact Allergy:  In Vivo and in Vitro Studies of Structure−Activity Relationships, Sensitizing Capacity, and Metabolic Activation

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.

Fragrance Compound Geraniol Forms Contact Allergens on Air Exposure. Identification and Quantification of Oxidation Products and Effect on Skin Sensitization

“…In previous autoxidation studies, limonene oxide and linalylacetate epoxide (unpublished results) were found to be non-sensitizers in the LLNA. These epoxides are sterically hindered and, therefore, less reactive in contrast to less substituted allylic epoxides, which we recently showed to be highly reactive and sensitizing (34). Formate 5 was a very weak sensitizer, and gave an EC3 value of 4.4 M. This result is consistent with previous results from the study of autoxidation of ethoxylated surfactants, where a non-sensitizing ethoxylated formate was identified in the oxidation mixture of air-exposed poly(oxyethylene) alcohols (35).…”

α-Terpinene, an Antioxidant in Tea Tree Oil, Autoxidizes Rapidly to Skin Allergens on Air Exposure

“…Thus, α-terpinene (1) can be considered as a prehapten as well as a prohapten since we have previously shown a metabolic activation of this compound. 18 Quantification of α-Terpinene and Oxidation Products after Air Exposure. The concentration of α-terpinene (1) decreased rapidly when exposed to air at room temperature.…”

“…18 As they formed glutathione-trapped epoxides in microsomal incubations, it was postulated that these compounds act as prohaptens. 18 Because of their similarity in structure, α-and β-phellandrene will probably autoxidize via pathways similar to those of α-terpinene (1), i.e., forming reactive compounds with allergenic effects.…”

“…16,17 We have previously shown that α-terpinene (1) is a prohapten and a moderate sensitizer in the murine local lymph node assay (LLNA). 18 α-Terpinene (1) has structural features in common with other monoterpene prehaptens (e.g., limonene). Accordingly, it is expected to autoxidize on air exposure to form allergenic compounds.…”

Study on the cytochrome P450-mediated oxidative metabolism of the terpene alcohol linalool: Indication of biological epoxidation

“…Oxidation by CYP sometimes results in more reactive products such as epoxides (Guengerich, 2003). Epoxides are able to react with nucleophilic sites of skin proteins to form haptens, which may result in allergic reactions (Bergstrom et al 2006). We reported recently, that the enzymatic product Á 3 -carene-epoxide was the result of such a P450-mediated bio-activation reaction of the monoterpene Á 3 -carene (Duisken et al 2005a).…”