OMIP-090: A 20-parameter flow cytometry panel for rapid analysis of cell diversity and homing capacity in human conventional and regulatory T cells.

The panel was developed and optimized for monitoring changes in homing capacity and functional diversity of human CD4+ conventional and regulatory T cell subsets. The analysis was based on expression of only surface markers in freshly isolated peripheral blood mononuclear cells (PBMCs) to reduce at minimum any alteration due to permeabilization or freezing/thawing procedures. We included markers to assess the distribution of naïve and memory populations based on the expression of CD45RA, CCR7, CD25, CD28 and CD95 in both conventional and regulatory T cells. The identification of major functional subsets was performed using CCR4, CCR6, CCR10, CXCR3 and CXCR5. Homing capacity of these subsets to skin, airway tract, gut and inflammatory lesions could finally be assessed with the markers CLA, CCR3, CCR5 and integrin ß7. The panel was tested on freshly isolated PBMCs from healthy donors and patients with allergic rhinitis or autoimmune disorders.

Enhanced NF-κB signaling in type-2 dendritic cells at baseline predicts non-response to adalimumab in psoriasis.

Biologic therapies have transformed the management of psoriasis, but clinical outcome is variable leaving an unmet clinical need for predictive biomarkers of response. Here we perform in-depth immunomonitoring of blood immune cells of 67 patients with psoriasis, before and during therapy with the anti-TNF drug adalimumab, to identify immune mediators of clinical response and evaluate their predictive value. Enhanced NF-κBp65 phosphorylation, induced by TNF and LPS in type-2 dendritic cells (DC) before therapy, significantly correlates with lack of clinical response after 12 weeks of treatment. The heightened NF-κB activation is linked to increased DC maturation in vitro and frequency of IL-17+ T cells in the blood of non-responders before therapy. Moreover, lesional skin of non-responders contains higher numbers of dermal DC expressing the maturation marker CD83 and producing IL-23, and increased numbers of IL-17+ T cells. Finally, we identify and clinically validate LPS-induced NF-κBp65 phosphorylation before therapy as a predictive biomarker of non-response to adalimumab, with 100% sensitivity and 90.1% specificity in an independent cohort. Our study uncovers important molecular and cellular mediators underpinning adalimumab mechanisms of action in psoriasis and we propose a blood biomarker for predicting clinical outcome.