Fig. 6: BLUE compounds reduce ISG expression in PBMCs.
From: Molecular glues that inhibit deubiquitylase activity and inflammatory signaling
a,b, Type I IFN signaling gene expression analysis (67 genes normalized for housekeeping genes: ACTB, GAPDH, HPRT1 and RPLP0) of healthy control PBMCs stimulated with IFNα2. Volcano plot of genes increased upon addition of IFNα2 with negative control AP-5-145 compared to DMSO (no IFN) condition (a). Effect of JMS-175-2 + IFN stimulation compared to AP-5-145 + IFN (b). The blue line indicates a P value of 0.05. Data points are the means from three independent experiments. c, CXCL10 protein levels in supernatant from IFNα2-stimulated healthy PBMCs (n = 3) quantified by ELISA and shown as a percentage of a control treated with IFN + DMSO (100%). The bar graph shows the average of three independent experiments. d, PBMCs were isolated from participants with SSc and treated with DMSO, AP-5-145, FX-171-C or JMS-175-2 for 16 h without IFN stimulation. Secreted CXCL10 in supernatant is shown as a percentage of the DMSO control. The bar graph shows the average of data from seven SSc donors. e, Type I IFN signaling gene expression analysis of unstimulated SSc PBMCs, treated with 2 µM AP-5-145, JMS-175-2 or FX-171-C. Heat map showing log2 mean fold change in ISG expression by treatment, compared to AP-5-145, according to qPCR SuperArray. ΔCt was calculated against the geometric mean of four housekeeping genes, followed by the ΔΔCt (fold change) relative to AP-5-145 and log2 transformation. Heat maps represent the mean fold change from nine SSc donors. P values were calculated using a Student’s t-test (two-tailed distribution and equal variances between the two samples). In a–d, paired, two-tailed Student’s t-tests were used to compare between treatment conditions for statistical significance. Error bars represent ±s.e.m.
