A Mouse Model for Immunotherapeutic Reversal of Leukemia-Induced T Cell Dysfunction
Authors: Ramsay Alan G. , Gorgun Gullu , Holderried Tobias A.W. , Zahrieh David , Liu Fenglong , Quackenbush John , Croce Carlo M. , Gribben John G. 
Published: April 2020

Abstract
Understanding the elusive mechanisms of tumor-driven immune evasion will aid the refinement of existing cancer immunotherapy strategies and identify novel treatments. To date, pre-clinical animal models that closely model human cancer, including the immune suppressive mechanisms utilized by cancer cells, have been under-characterized. The identification and use of such models should allow better predictions of successful human responses to immunotherapy. As a model for changes induced in non-malignant cells by cancer, we examined T cell function in Eμ-TCL1 transgenic mice as they developed leukemia from 12-months of age. Transgenic expression of TCL1 in B cells had no demonstrable effect on T cells, however, mice with leukemia had decreased in vivo antigen specific T cell activation, suppressed T cell mitogenic proliferation and impaired induction of idiotype specific CD8 T cells capable of killing CLL cells compared to control WT mice (age-matched throughout study) or Eμ-TCL1 transgenic mice without CLL. Leukemic mice also had dysfunctional T cell lymphokine production (Th2-preponderant). To understand the molecular basis for the observed functional defects and to compare changes seen in mice and patients with CLL we performed gene expression profiling. Analysis of highly purified CD4 and CD8 T cells in CLL mice demonstrated altered gene expression profiles compared to WT mice or to young Eμ-TCL1 mice without disease. Of note, infusion of CLL cells into young Eμ-TCL1 mice induced gene expression changes comparable to those seen in mice with developed leukemia, demonstrating a causal relationship between leukemia and the T cell defects. Analysis of gene expression changes in T cells in CLL mice compared with those in patients was performed using RESOURCERER, a database for annotating and linking microarray resources within and across species and identified 50 overlapping genes in CD4 T cells and 45 overlapping genes in CD8 T cells. The majority of differentially expressed genes in CD4 T cells from both mice and patients with CLL were involved in cell proliferation and activation pathways with increase in Lck. Multiple defects within the actin cytoskeletal formation pathways were identified in both CD4 and CD8 T cells including Cdc42. Integrity of the T cell cytoskeleton is essential to regulate the dynamic signaling required for T cell activation and effector function in response to immunological recognition of antigen-presenting cells (APCs). T cell conjugates from mice with leukemia had suppressed antigen-dependent F-actin accumulation and early T cell signaling at the immune synapse with CLL cells (APCs) compared to WT mice conjugates. Moreover, we have demonstrated that infusion of CLL cells into young mice induces this T cell defect, demonstrating an in vivo immunomodulating mechanism utilized by tumor cells. Treatment of both CLL cells and autologous T cells from leukemic mice with lenalidomide (0.5 μM for 24 h) enhanced the formation of the F-actin immune synapse and recruitment of tyrosine-phosphorylated proteins irrespective of the presence of exogenous antigen. Of note, the capacity to repair immunological recognition with this agent was associated with increased recruitment of the cytoskeletal signaling molecules Lck and Cdc42 to the immunological synapse, regardless of whether the gene was increased or decreased on gene expression profiling. These results demonstrate that leukemia cells induce changes in multiple T cell pathways regulating antigen recognition and effector function. The similarities with human CLL including reversible immunological synapse dysfunction with an immunomodulating drug validates the use of Eμ-TCL1 mice as a model for further analyses of ways to prevent and reverse cancer-induced immune dysfunction. The use of this model to understand and reverse the molecular changes in T cells induced by leukemia will likely have broad applications to maximize immune responses in patients.