The molecular mechanism underlying the autoimmune-associated PTPN22 R620W variation and the quest for therapeutics

PTPs involved in modulation of signal transduction through the T cell receptor (TCR) are promising targets for human autoimmunity. Here we will focus on the lymphoid tyrosine phosphatase LYP, a critical negative modulator of TCR signaling encoded by the PTPN22 gene. A missense C1858T single nucleotide polymorphism in the PTPN22 gene recently emerged as a major risk factor for multiple human autoimmune diseases. In T cells, LYP forms a complex with the negative regulatory kinase Csk and is a critical negative regulator of signaling through the T cell receptor. The C1858T SNP results in the LYP-R620W variation within the LYP-Csk interaction motif. LYP-W620 exhibits reduced interaction with Csk and is a gain-of-function inhibitor of TCR signaling. While strong genetic and functional evidence has suggested that LYP is a promising candidate drug target for treatment of human autoimmunity, the molecular mechanism of the autoimmune-associated R620W variation remains unknown.;We hypothesize that an inhibitor of LYP could potentially revert the gain-of-function effect and restore normal TCR signaling levels in carriers of LYP-W620. While a small molecule inhibitor would be useful for both understanding the role of LYP in the immune system and for validating LYP as a drug target, the development of specific, cell-permeable PTP inhibitors is currently hampered by the lack of methods for high-throughput screening (HTS) to identify cell-permeable leads.;Here we present the mechanism by which the pathogenic R620W polymorphism of LYP causes a gain-of-function form of the phosphatase. We also propose two approaches to identify candidate cell-permeable inhibitors of PTPs. We show that through a novel screen of a library of drug-like small-molecule compounds, we were able to identify a non-phospho-mimetic cell-permeable inhibitor of LYP which increases TCR signaling and T cell activation. We also propose the first cell-based assay to directly monitor PTP activity at the single-cell level. Our assay can be adapted to screening of inhibitor libraries for any PTP of interest to identify cell-permeable inhibitors.