Monitoring PD-1 Phosphorylation to Evaluate PD-1 Signaling during Antitumor Immune Responses
Xia Bu, Vikram R. Juneja, Carol Reynolds, Kathleen M. Mahoney, Melissa T. Bu, Kathleen A. McGuire, Seth Maleri, Ping Hua, Baogong Zhu, Sarah R. Klein, Edward Greenfield, Philippe Armand, Jerome Ritz, Arlene H. Sharpe, Gordon J. Freeman
Abstract
Abstract PD-1 expression marks activated T cells susceptible to PD-1–mediated inhibition but not whether a PD-1–mediated signal is being delivered. Molecular predictors of response to PD-1 immune checkpoint blockade (ICB) are needed. We describe a monoclonal antibody (mAb) that detects PD-1 signaling through the detection of phosphorylation of the immunotyrosine switch motif (ITSM) in the intracellular tail of mouse and human PD-1 (phospho–PD-1). We showed PD-1+ tumor-infiltrating lymphocytes (TILs) in MC38 murine tumors had high phosphorylated PD-1, particularly in PD-1+TIM-3+ TILs. Upon PD-1 blockade, PD-1 phosphorylation was decreased in CD8+ TILs. Phospho–PD-1 increased in T cells from healthy human donors after PD-1 engagement and decreased in patients with Hodgkin lymphoma following ICB. These data demonstrate that phosphorylation of the ITSM motif of PD-1 marks dysfunctional T cells that may be rescued with PD-1 blockade. Detection of phospho–PD-1 in TILs is a potential biomarker for PD-1 immunotherapy responses.