898 The proteasome inhibitor PTTC restores the lipid barrier in a harlequin ichthyosis in vitro skin model

Harlequin ichthyosis is a life threatening rare skin condition characterized by aberrant differentiation of the epidermis leading to a severe barrier defect. It is caused by mutations in ABCA12 a lipid transporter involved in transport of glucosylceramides from the lamellar body to the lipid lamellae. The current therapeutic approach consists of topical emollients and oral administration of retinoids causing side effects, so there is an unmet need for new treatments. The ubiquitin-proteasome pathway is responsible for maintaining protein homeostasis. 3,5-Ditert-butyl-4-hyroxyhydrocinnamate (PTTC) is a novel proteasome inhibitor which has been trialed in humans for rosacea andin vitro for psoriasis. We hypothesized that PTTC could restore the epidermal barrier in a Harlequin ichthyosis 3D organotypic (OT) models established using CRispR-Cas9 deletion of ABCA12. After assessing concentrations/toxicity of PTTC in an MTT assay, 3D OTs generated from ABCA12 KO and WT keratinocytes were treated with PTTC at two concentrations (10 and 20 μM) and compared to negative controls treated with vehicle only (n=4). The OTs were stained with H&E, Nile Red and differentiation markers such as K10, K14, TGM1 and involucrin. Epidermal thickness significantly decreased in KO OTs comparing to WT for all the PTTC concentrations. However, at 10 μM an increase of neutral and polar lipid translocation toward the stratum corneum was seen, as well as normalisation of expression of K10, K14 and involucrin compared to the controls and KO OTs treated with 20 μM PTTC. This suggests that 10 μM PTTC has a positive effect in restoring epidermal function and differentiation in ABCA12 KO OTs. These results are promising and we are considering further in vivo studies.