Terpene Hydroperoxides in Allergic Contact Dermatitis: Synthesis, Lipid Peroxidation, and Immunological Implications

Abstract

Allergic contact dermatitis (ACD) is a type IV hypersensitivity reaction mediated by T cells. Contact allergens are typically too small to elicit an immune response on their own and must bind to larger macromolecules, classically proteins, to exert immunogenic effects. However, the potential role of lipids in facilitating these effects has remained largely unexplored. This work reports the successful synthesis and NMR characterisation of non-commercially available contact allergen hydroperoxides: limonene-2-hydroperoxide (Lim-2-OOH), linalool-6/7-hydroperoxides (Lin-6/7-OOH), and geraniol-6/7-hydroperoxides (Ger-6/7-OOH). Lipidomic analysis of HaCaT keratinocytes treated with Lim-2-OOH and Lin-6/7-OOH revealed remodeling of the lipidome, with upregulation of plasmanyl/plasmenyl phospholipids (O-/P-PC and O-/P-PE), sphingolipids (HexnCer), and triacylglycerols, alongside a decrease in polyunsaturated phospholipids (PC and PE). Incubation of phosphatidylcholine 16:0/18:1 (PLPC) liposomes with the synthesised hydroperoxides under biomimetic conditions induced oxidation predominantly yielding long-chain products, with lower levels of electrophilic short-chain products. The degree of lipid peroxidation was equal to or greater than that induced by hydrogen peroxide (H�O�), with Lim-2-OOH causing significantly more oxidation than all other hydroperoxides tested. Immunological studies demonstrated that Ger-6/7-OOH differentially modulated CD1 protein responses. CD1a tetramers loaded with Ger-6/7-OOH showed reduced tetramer-positive events, whereas CD1d tetramers loaded with Ger-6/7-OOH showed a marked increase in tetramer-positive T cells, comparable to the positive control �-GalCer. Functional assays further revealed that terpenes and their hydroperoxides exhibited diverse modes of interaction with CD1 proteins: some acted as agonists, eliciting T cell activation and cytokine responses; others acted as antagonists or displaced endogenous ligands, reducing CD1-TCR binding and functional outputs. Overall, this work demonstrates that terpene hydroperoxide contact allergens can modify lipids under biomimetic conditions by promoting lipid peroxidation, and can also directly engage CD1-mediated pathways to initiate immunogenic effects independently of classical haptenation mechanisms. These findings highlight a potential role for unconventional T cells in ACD and expand our understanding of lipid-driven immunogenicity.