Assessing the preclinical potential of the selective p38δ silencing in human skin organoids (HSO) and ex vivo human skin explant model of atopic dermatitis.

Summary: Abstract Body: p38δ genetic deletion caused an inflammatory skin phenotype in an irritant contact dermatitis mouse model. However, the p38δ role in atopic dermatitis (AD) pathophysiology remains unexplored. To circumvent the lack of selective p38δ inhibitors, we employed a rationally designed hydrophobic conjugate docosanoic acid (DCA)-linked fully chemically stabilized small interfering RNA (siRNA) approach to silence p38δ in HSO and ex vivo human skin explant model of AD. In vitro screening identified two lead siRNAs, si-p38δ-1 and si-p38δ-2, that yielded the highest p38δ knockdown (KD) efficacy and potency versus the non-targeting control. DCA-si-p38δ-1- and -2-mediated p38δ KD in HSO disrupted epidermal morphogenesis and caused marked epidermal hyperplasia during the early stages of the organotypic epidermal development and hyperkeratosis at later stages, as well as gross tissue disturbances and aberrant filaggrin and desmoglein 1 expression patterns. Next, we efficiently silenced p38δ in ex vivo human skin explants using intradermal injections of DCA-si-p38δ-1 and -2 in the absence and presence of TH2 (IL4+IL13) cytokine stimulation. Remarkably, p38δ expression was significantly upregulated by the TH2 cytokine treatment. Multiplex chemokine/ cytokine array and bulk RNA sequencing data showed dysregulation of expression of genes related to epidermal inflammation, differentiation, lipid storage and transport, iron homeostasis and transport, and glycoprotein biosynthesis and metabolism upon p38δ KD in unstimulated and TH2 cytokine-stimulated ex vivo skin. These data support the p38δ role in epidermal morphogenesis and differentiation and in regulating AD-like inflammatory responses in human skin. Furthermore, the siRNA drug approach outlined in this work provides novel opportunities to modulate other disease targets in the skin. Katy Trampel¹, Katherine Gross², Daniel O'Reilly², K Russell³, Anastasia Khvorova², Bethany E. Perez White³, Tatiana Efimova¹ 1. Biomedical Engineering, Northwestern University, Chicago, IL, United States. 2. University of Massachusetts Chan Medical School, Worcester, MA, United States. 3. Northwestern University Feinberg School of Medicine, Chicago, IL, United States. Translational Studies: Cell and Molecular Biology