ObjectivesGenetic variants in the transcription factor STAT4 are associated with increased susceptibility to systemic lupus erythematosus (SLE) and a more severe disease phenotype. This study aimed to clarify how the SLE-associated intronic STAT4 risk allele rs7574865[T] affects the function of immune cells in SLE.MethodsPeripheral blood mononuclear cells (PBMCs) were isolated from 52 genotyped patients with SLE. Phosphorylation of STAT4 (pSTAT4) and STAT1 (pSTAT1) in response to interferon (IFN)-α, IFN-γ or interleukin (IL)-12, total levels of STAT4, STAT1 and T-bet, and frequency of IFN-γ+ cells on IL-12 stimulation were determined by flow cytometry in subsets of immune cells before and after preactivation of cells with phytohaemagglutinin (PHA) and IL-2. Cellular responses and phenotypes were correlated to STAT4 risk allele carriership. Janus kinase inhibitors (JAKi) selective for TYK2 (TYK2i) or JAK2 (JAK2i) were evaluated for inhibition of IL-12 or IFN-γ-induced activation of SLE PBMCs.ResultsIn resting PBMCs, the STAT4 risk allele was neither associated with total levels of STAT4 or STAT1, nor cytokine-induced pSTAT4 or pSTAT1. Following PHA/IL-2 activation, CD8+ T cells from STAT4 risk allele carriers displayed increased levels of STAT4 resulting in increased pSTAT4 in response to IL-12 and IFN-α, and an augmented IL-12-induced IFN-γ production in CD8+ and CD4+ T cells. The TYK2i and the JAK2i efficiently blocked IL-12 and IFN-γ-induced activation of PBMCs from STAT4 risk patients, respectively.ConclusionsT cells from patients with SLE carrying the STAT4 risk allele rs7574865[T] display an augmented response to IL-12 and IFN-α. This subset of patients may benefit from JAKi treatment.

BackgroundGenetic variants in the transcription factor STAT4 are associated with increased susceptibility to systemic lupus erythematosus (SLE) and a more severe disease phenotype.ObjectivesThis study aimed to clarify how the SLE-associated intronic STAT4 risk variant rs7574865[T] affects the function of immune cells in SLE.MethodsPeripheral blood mononuclear cells (PBMCs) were isolated from 52 SLE patients in remission (SLEDAI-2K≤4). STAT4 and STAT1 protein levels and phosphorylation status in response to interferon (IFN)-α, IFN-γ, or interleukin (IL)−12 were determined, before and after pre-activation of cells with phytohaemagglutinin (PHA) and IL-2, in CD56dim NK cells, CD56bright NK cells, B cells, CD4+ T cells, CD8+ T cells and monocytes by flow cytometry. The frequency of IFN-γ+ cells upon IL-12 or PMA (phorbol 12-myristate-13-acetate) stimulation and the frequency of T-bet+ cells was determined in PHA/IL-2 pre-activated cells. Cellular responses and phenotypes were correlated with STAT4 risk allele carriership (rs7574865[T], 21 homozygous protective, 22 heterozygous and 9 homozygous risk) using an additive linear regression model. Janus kinase inhibitors (JAKi) selective for TYK2 (TYK2i, Compound 35) or JAK2 (JAK2i, BMS-911543) were evaluated for inhibition of IL-12 or IFN-γ-induced activation of SLE PBMCs.ResultsIn resting PBMCs, the STAT4 risk allele was neither associated with protein levels of STAT4 or STAT1, nor cytokine-induced phosphorylation of STAT4 (pSTAT4) or STAT1 (pSTAT1). However, following PHA/IL-2-activation, CD8+ T cells from STAT4 risk allele carriers displayed increased levels of STAT4 (p=0.04), resulting in increased pSTAT4 in response to IL-12 (p=0.003) and IFN-α (p=0.04). Analysis of T cell subsets revealed that the effect was seen in CD45RA+CD57– naïve and CD45RA–CD57– memory CD8+ T cells, but not in CD45RA–CD57+ memory or CD45RA+CD57+ effector CD8+ T cells. A slight increase in STAT4 protein levels and IL-12-induced pSTAT4 was also observed in CD4+ T cells from STAT4 risk allele carriers (p=0.08 and p=0.09, respectively). STAT4 risk allele carriers displayed an augmented IL-12-induced IFN-γ production in CD8+ and CD4+ T cells (p=0.03 for both), whereas PMA-induced IL-12 production was normal (p=0.31 and p=0.10, respectively). T-bet expression was not correlated to the STAT4 genotype. The TYK2i and the JAK2i efficiently blocked IL-12 and IFN-γ-induced activation of PBMCs from STAT4 risk patients, respectively.ConclusionsT cells from SLE patients carrying the STAT4 risk allele rs7574865[T] display an augmented response to IL-12 and IFN-α. This subset of patients may benefit from JAKi treatment.Disclosure of InterestN. Hagberg: None declared, M. Joelsson: None declared, D. Leonard: None declared, S. Reid: None declared, M.-L. Eloranta: None declared, J. Mo Employee of: AstraZeneca, M. Nilsson Employee of: AstraZeneca, A.-C. Syvänen: None declared, Y. Bryceson: None declared, L. Rönnblom: None declared