Abstract 3539: IL-12 overcomes T-cell immune suppression by the B-ALL microenvironment
Authors: Hunter Rae , Imbach Kathleen , Dougan Jodi , Do Priscilla , Townsel Ashley , Dreaden Erik , Gibson Gregory , Henry Curtis , Porter Christopher 
Published: June 2022

Abstract
Leukemia remains one of the leading causes of disease-related deaths in children. Recent clinical advancements in both cell-directed and antibody therapies for B-cell leukemias demonstrate the importance of the immune system in eliciting anti-leukemia activity. Yet, there are still patients who suffer relapse with these treatments, which underscores the need to better understand immune evasion by leukemia cells and the need to develop more effective therapies.
Although IL-12 has been well-studied in the context of pathogenic infections and immune cells, we previously published that IL-12 potently promotes B-ALL clearance in immune competent mice via increased T-cell activation. To further characterize the role of IL-12 in leukemia control, we examined the bone marrow of immune competent mice with B-ALL using an extensive flow cytometry panel. From these studies, we observed both an increase in T-cell and dendritic cell (DC) numbers and augmented DC activation in rIL-12p70 treated WT mice. Furthermore, rIL-12p70 treatment of WT mice with B-ALL exhibited increased GM-CSF, IL-2, IL-1β, CXCL2, CCL12, and IFN-gamma levels in the bone marrow microenvironment. In these experiments, we also analyzed gene expression profiles in T-cells. The expression of exhaustion-associated genes, including Lag3, Pcdc1, and Tigit, was higher in T-cells recovered from untreated mice with B-ALL relative to those from mice without leukemia. These data are consistent with observations made from single cell RNA-sequencing studies of children with B-ALL, where it was revealed that immune cell exhaustion was present at higher levels in those with measurable disease after induction therapy. Treatment of mice with IL-12 abrogated the leukemia induced increases in exhaustion-associated gene expression.
Mechanistically, we observed that when T-cells are stimulated in vitro in B-ALL supernatant there is a reduction in the surface expression of the T-cell activation marker, CD44, and marker of degranulation, CD107b. Jurkat T-cells expressing NFAT or NF-kB reporter plasmids were used to assess activation of T-cell signaling pathways. In addition to suppressing CD44 and CD107b, we also found that B-ALL-secreted factors also suppress the activation of signaling pathways downstream of the TCR.
Considering clinical implications of this discovery, we further explored IL-12 ex vivo in combination with the clinically approved bi-specific T-cell engager, blinatumomab. Notably, B-ALL-secreted factors reduced blinatumomab-mediated T-cell lysis of target cells, which was restored with recombinant IL-12 treatment.
Overall, this data shows the potential for IL-12 to be used as an immunotherapeutic in conjunction with other current therapies to overcome T-cell suppression by B-ALL.
Citation Format: Rae Hunter, Kathleen Imbach, Jodi Dougan, Priscilla Do, Ashley Townsel, Erik Dreaden, Gregory Gibson, Curtis Henry, Christopher Porter. IL-12 overcomes T-cell immune suppression by the B-ALL microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3539.