Fig. 3

The Interferon gene signature (IGS) based on the expression of six type I IFN-related genes (IFI27, IFI44L, IFIT1, ISG15, RSAD2, SIGLEC1) on peripheral blood throughout the disease course. Before the initiation of tofacitinib therapy, a positive type I IGS was observed. Three months after tofacitinib initiation, the IGS turned negative. Eighteen months after starting tofacitinib, the IGS was stably negative, but a mild increase in the expression of IFI44L, IFIT1, and RSAD2 was observed. The IGS was assessed by calculating the expression of six type I interferon-stimulated genes (ISGs) (IFI27, IFI44L, IFIT1, ISG15, RSAD2, and SIGLEC1). Real-Time PCR was performed using gene-specific custom-designed FAM probes (TibMolBiol, Roche, Germany) in duplicate. Gene copy number were calculated as follow: (1) standard curves were prepared for each gene with 10-fold serial dilutions of the synthetic control; (2) curves were generated through linear regression analysis. Values were normalised with the geometric means of two internal controls (TBP and HPRT1) and expressed as gene score (GS). IFN Score (IS) was calculated as the geometric mean of the GS of the six ISGs. The IS cut-off value was determined based on 26 healthy donors (HD) using the mean + 2 standard deviations of their GS to classify as positive/negative the IS analysis (normal IS values < 0.7). HD normal reference values and standard deviation for each gene are reported in the graph (in grey). The bar blot illustrates gene scores on the y-axis, with individual genes listed on the x-axis. Coloured bars represent the patient (red for baseline, green and blue for three and eighteen months after tofacitinib initiation, respectively), while grey bars represent HD