The Intracellular TCR Signaling Pathway As a Novel Therapeutic Target in AITL Patients

Angioimmunoblastic T cell lymphoma (AITL) is a distinct subtype of peripheral T cell lymphoma (PTCL) with 5-year overall survival rates of less than 30%. Although frequent somatic mutations of the epigenetic related genes have been identified in AITL, the molecular pathogenesis mechanisms are still unknown to this disease. T-cell receptor (TCR) signaling is pivotal in T-cell development and function. The T-cell receptor presents on the surface of T lymphocytes, which plays a pivotal in T-cell development and function. Engagement of the TCR by antigen presented on major histocompatibility complex (MHC) molecules regulated cellular proliferation, differentiation and cytokine production. In our study, twelve paired AITL samples were subjected to whole exome sequencing. The somatic TCR signaling related mutations were frequently observed in these patients, such as RHOA (50%), PLCg1 (41.7%), ZAP70 (41.7%), CD28 (25%) and Itk (16.7%). Furthermore, phosphorylated ZAP70, Itk and PLCg1 immunohistochemistry staining analysis were performed in 33 AITL paraffin sections. More than half of the patients exhibited constitutively activated intracellular TCR signaling pathway, which might gain significant attention for its possible pathogenesis role in AITL patients. To further illustrated this hypothesis, the knockdown effect of TCR signaling key regulatory kinase ZAP70 and Itk on cellular functions was by shRNA transfection in T cell lymphoma cell lines. The suppression of these target gene expressions significantly compromises proliferation, adhesion, invasion, and migration of malignant T cells. Meanwhile, the pro-apoptotic effect and G2/M phase cell cycle arrest were also induced by ITK and ZAP70 shRNA transfection. Western Blotting analysis demonstrated that the activation of intracellular TCR and related downstream signal pathways was also strongly reduced post shRNA genetic inhibition. More importantly, the similar results were also confirmed using specific Itk inhibitor BMS-509744 and PLS-6-63. Additionally, the anti-tumor and pro-apoptotic activities of Itk inhibitor also were demonstrated using xenograft mice models. Taken together, the constitutively active TCR signaling pathway suggested a novel candidate therapeutic target for the treatment of AITL.