Unravelling the role of JAK3/TEC kinase signalling with ritlecitinib in alopecia areata lesional skin and alopecia areata-induced human hair follicles ex vivo

Summary: Abstract Body: JAK3/TEC signalling is clinically relevant in alopecia areata (AA). Ritlecitinib is an FDA-approved inhibitor of JAK3/TEC signalling. JAK3/ TEC family kinases regulate T-cell activity by acting downstream of cytokines such as IL-2 and T-cell receptors (TCR), respectively. Here we aim to further dissect their role in AA pathogenesis. TCR activation in healthy hair follicles (HFs) with αCD3/αCD28+IL-2 significantly up-regulated intra- and peri-follicular T-cell numbers, and proliferation, and enhanced secretion of cytokines, including the key AA pathogenic factor IFNγ, and the cytolytic molecules perforin, granzyme A and B, granulysin, and FasL. Furthermore, it significantly increased bulbar MHC class I and ectopic MHC class II expression in the outer root sheath. All these effects were prevented by ritlecitinib 2µM treatment. Qualitative analysis of chronic lesional AA compared to healthy scalp skin showed increased intra- and peri-bulbar pSTAT5+CD3+, pSTAT3+, and pSTAT6+ cell numbers (JAK3 signalling) along with more CD8+GranzymeB+ T-cells. IRF4 expression, a downstream target of TCR-TEC, was also increased in the HF epithelium and in infiltrates around AA HFs. In lesional AA skin, under re-stimulation with αCD3/αCD28+IL-2, ritlecitinib decreased follicular and perifollicular pSTAT3+ cell numbers and IRF4+CD3+ cell numbers as well as percentages amongst CD3 cells. Thus, JAK3/TEC signalling is active in lesional AA skin and can be inhibited by ritlecitinib treatment. This, together with the ability of ritlecitinib to interfere with αCD3/αCD28+IL-2-induced peri-follicular T-cell expansion and HF immune privilege collapse in HFs ex vivo, highlights the clinical mechanism by which JAK3/TEC inhibition reduces symptoms of AA. Alexandre Lejeune¹, Joana Viola-Söhnlein², Thomas Rouillé², Elizabeth Kieras³, David Martin³, Jean-Baptiste Telliez³, Leslie J Berg⁴, Ilaria Piccini², Karin Pappelbaum², Janin Edelkamp², Marta Bertolini² 1. Pfizer Inc, Paris, France. 2. QIMA Life Sciences, Münster, Germany. 3. Pfizer Inc, Cambridge, MA, United States. 4. University of Colorado, Aurora, CO, United States. Translational Studies: Cell and Molecular Biology