Regulatory T cells contribute to the immunosuppressive phenotype of neutrophils in a mouse model of chronic lymphocytic leukemia.

BACKGROUND: Impaired neutrophil activity is an important issue in chronic lymphocytic leukemia (CLL), as it contributes to a dysfunctional immune response leading to life-threatening infections in patients. Some features typical of CLL neutrophils, e.g., the B-cell-supportive secretion profile, have already been described. However, most of these studies were performed on cells isolated from peripheral blood. It is still unclear which molecular factors and cell types are involved in shaping neutrophil function and phenotype in the CLL microenvironment. Since regulatory T cells (Treg) play an important role in CLL progression and influence the activity of neutrophils, we investigated the crosstalk between Treg and neutrophils in the spleen using a murine model of CLL. METHODS: In this work, we used an Eµ-TCL1 mouse model of human CLL. For our in vivo and ex vivo experiments, we inoculated wild-type mice with TCL1 leukemic cells isolated from Eµ-TCL1 transgenic mice and then monitored disease progression by detecting leukemic cells in peripheral blood. We analyzed both the phenotype and activity of neutrophils isolated from the spleens of TCL1 leukemia-bearing mice. To investigate the interrelation between Treg and neutrophils in the leukemia microenvironment, we performed experiments using TCL1-injected DEREG mice with Treg depletion or RAG2KO mice with adoptively transferred TCL1 cells alone or together with Treg. RESULTS: The obtained results underline the plasticity of the neutrophil phenotype, observed under the influence of leukemic cells alone and depending on the presence of Treg. In particular, Treg affect the expression of CD62L and IL-4 receptor in neutrophils, both of which are crucial for the function of these cells. Additionally, we show that Treg depletion and IL-10 neutralization induce changes in the leukemia microenvironment, partially restoring the "healthy" phenotype of neutrophils. CONCLUSIONS: Altogether, the results indicate that the crosstalk between Treg and neutrophils in CLL may play an important role in CLL progression by interfering with the immune response.

In Situ Programming of CAR-T Cells: A Pressing Need in Modern Immunotherapy.

Chimeric antigen receptor-T (CAR-T) cell-based therapy has become a successful option for treatment of numerous hematological malignancies, but also raises hope in a range of non-malignant diseases. However, in a traditional approach, generation of CAR-T cells is associated with the separation of patient's lymphocytes, their in vitro modification, and expansion and infusion back into patient's bloodstream. This classical protocol is complex, time-consuming, and expensive. Those problems could be solved by successful protocols to produce CAR-T cells, but also CAR-natural killer cells or CAR macrophages, in situ, using viral platforms or non-viral delivery systems. Moreover, it was demonstrated that in situ CAR-T induction may be associated with reduced risk of the most common toxicities associated with CAR-T therapy, such as cytokine release syndrome, immune effector cell-associated neurotoxicity syndrome, and "on-target, off-tumor" toxicity. This review aims to summarize the current state-of-the-art and future perspectives for the in situ-produced CAR-T cells. Indeed, preclinical work in this area, including animal studies, raises hope for prospective translational development and validation in practical medicine of strategies for in situ generation of CAR-bearing immune effector cells.

A Specific CD44lo CD25lo Subpopulation of Regulatory T Cells Inhibits Anti-Leukemic Immune Response and Promotes the Progression in a Mouse Model of Chronic Lymphocytic Leukemia.

Regulatory T cells (Tregs) are capable of inhibiting the proliferation, activation and function of T cells and play an important role in impeding the immune response to cancer. In chronic lymphocytic leukemia (CLL) a dysfunctional immune response and elevated percentage of effector-like phenotype Tregs have been described. In this study, using the Eµ-TCL1 mouse model of CLL, we evaluated the changes in the Tregs phenotype and their expansion at different stages of leukemia progression. Importantly, we show that Tregs depletion in DEREG mice triggered the expansion of new anti-leukemic cytotoxic T cell clones leading to leukemia eradication. In TCL1 leukemia-bearing mice we identified and characterized a specific Tregs subpopulation, the phenotype of which suggests its role in the formation of an immunosuppressive microenvironment, supportive for leukemia survival and proliferation. This observation was also confirmed by the gene expression profile analysis of these TCL1-specific Tregs. The obtained data on Tregs are consistent with those described so far, however, above all show that the changes in the Tregs phenotype described in CLL result from the formation of a specific, described in this study Tregs subpopulation. In addition, functional tests revealed the ability of Tregs to inhibit T cells that recognize model antigens expressed by leukemic cells. Moreover, inhibition of Tregs with a MALT1 inhibitor provided a therapeutic benefit, both as monotherapy and also when combined with an immune checkpoint inhibitor. Altogether, activation of Tregs appears to be crucial for CLL progression.