Fig. 2: Soquelitinib inhibits TCR-induced downstream signaling and activation.

a Diagram illustrates pathways that activate ITK and the downstream effects of that activation. Drawing created using BioRender. b Western blot analysis of whole-cell lysates from H9 cells stimulated with anti-CD3 for 30 s in the presence of increasing concentration of soquelitinib. Blots were probed with specific antibodies to total and phosphorylated PLCγ1 and ZAP-70. c Soquelitinib inhibits ERK and S6 phosphorylation. ERK and S6 phosphorylation was measured by flow cytometry following anti-CD3/28-crosslinking in human T cells. Each data point displays mean of fold change from three donors. Error bars represent SEM. d Soquelitinib suppresses IL-2 secretion in Jurkat cells in response to TCR stimulation. A representative curve is shown from one experiment with the mean of technical duplicates plotted. e Soquelitinib (≥1 μM) reduced the total number of cells while maintaining cell viability after six days in culture. Cell viability and cell expansion rate of human primary T cells measured by the Vi-Cell XL Cell Viability Analyzer (Beckman Coulter) after TCR stimulation. Data are means ± SEM of technical triplicates within an experiment and are representative of 15 human donors. Fig. 2c, d, e were created using Graphpad Prism. PI3K phosphatidyl inositol-3 kinase, AKT protein kinase B, NFkB nuclear factor kappa B, NFAT nuclear factor of activated T cells, ERK extracellular signal regulated kinase, mTOR molecular target of rapamycin, GATA3 GATA binding protein 3, IL interleukin, RpS6 ribosomal protein 6. *p < 0.05, ***p < 0.001.