Assessment of Pre- and Pro-haptens Using Nonanimal Test Methods for Skin Sensitization

Abstract

Because of ethical and regulatory reasons, several nonanimal test methods to assess the skin sensitization potential of chemicals have been developed and validated. In contrast to in vivo methods, they lack or provide limited metabolic capacity. For this reason, identification of pro-haptens but also pre-haptens, which require molecular transformations to gain peptide reactivity, is a challenge for these methods. In this study, 27 pre- and pro-haptens were tested using nonanimal test methods. Of these, 18 provided true positive results in the direct peptide reactivity assay (DPRA; sensitivity of 67%), although lacking structural alerts for direct peptide reactivity. The reaction mechanisms leading to peptide depletion in the DPRA were therefore elucidated using mass spectrometry. Hapten-peptide adducts were identified for 13 of the 18 chemicals indicating that these pre-haptens were activated and that peptide binding occurred. Positive results for five of the 18 chemicals can be explained by dipeptide formations or the oxidation of the sulfhydryl group of the peptide. Nine of the 27 chemicals were tested negative in the DPRA. Of these, four yielded true positive results in the keratinocyte and dendritic cell based assays. Likewise, 16 of the 18 chemicals tested positive in the DPRA were also positive in either one or both of the cell-based assays. A combination of DPRA, KeratinoSens, and h-CLAT used in a 2 out of 3 weight of evidence (WoE) approach identified 22 of the 27 pre- and pro-haptens correctly (sensitivity of 81%), exhibiting a similar sensitivity as for directly acting haptens. This analysis shows that the combination of in chemico and in vitro test methods is suitable to identify pre-haptens and the majority of pro-haptens.

Molecular docking predictions of fragrance binding to human leukocyte antigen molecules

“…Similar chemical effects (oxidation of weakly reactive chemicals) can be induced by cytochrome P450, although the activity of these enzymes in human skin is not very high . Of the fragrance compounds studied here, citral, d ‐limonene and linalool were found to be prehaptens, eugenol and isoeugenol are prohaptens, and cinnamyl alcohol and geraniol are both prehaptens and prohaptens . In agreement with these observations, recent experiments showed that benzyl benzoate, benzyl salicylate and benzyl cinnamate remained chemically stable when exposed to forced degradation by oxidation, and remained non‐reactive to cysteine .…”

Application of IATA – A case study in evaluating the global and local performance of a Bayesian network model for skin sensitization

“…Chemicals identified as Michael acceptors or S N 2 electrophiles have an excellent probability of having their sensitization potential predicted correctly. This fits with what is currently known about the ability of the non-animal assays leading into the network; mainly that the DPRACys and KeratinoSens™ assays tend to work best with soft electrophiles [38]. While LLNA class prediction was poor for those with an S N 2 domain, it was excellent for Michael acceptors at 76 86 66 %, i.e.…”

Keratinocytes improve prediction of sensitization potential and potency of chemicals with THP-1 cells_suppl

“…using the available methods show limitations in the demonstration of concentration-dependency and have limited capacities to detect certain prohaptens and for potency prediction (Piroird et al, 2015;Natsch et al, 2015;Nukada et al, 2012;Teunis et al, 2014;Jaworska et al, 2015;Urbisch et al, 2016;Adler et al, 2011).…”