Background
Acute myeloid leukemia (AML) is a lethal adult hematologic malignancy with a 5-year survival rate of 30%. The standard of care is cytotoxic chemotherapy which often results in remission, but most patients relapse. Recent findings support a rationale that immunotherapy may hold promise as a treatment for this malignancy. Our lab has shown that in a subset of patients, cytotoxic T cells lose their proliferation and cytokine production ability. T cell dysfunction called exhaustion is observed in chronic viral infections and cancers including AML. Epigenetic remodeling is a key feature of exhausted T cells. Changes in DNA methylation have been shown to be central to establishment of exhaustion. Blocking DNA methylation can restore T cell expansion in a chronic antigen setting. Hypomethylating agents (HMAs) such as Azacytidine (Aza) are approved for clinical use to treat AML. Studies have shown that Aza can modulate the immune system and may impact exhaustion.
Methods
AML mice were treated for 3 weeks with either Aza or control (PBS). Blood was collected at several time points and different tissues harvested at the end to evaluate T cell, myeloid and stem cell markers by flow cytometry. Bisulphite sequencing and proliferation assays were performed using spleens. Plasma was evaluated for cytokine expression using a multiplex ELISA. GraphPad PRISM 10 was used to conduct statistical analysis.
Results
Using a spontaneous mouse model of AML, we find that Aza treatment led to a decrease in tumor burden and spleen weight. It also caused a shift towards a stem like progenitor state and decrease in Tregs percent, suggesting Aza can modulate T cell function. Additionally, Aza caused a shift of different hematopoietic progenitor cell populations in the bone marrow. Pro-inflammatory cytokines associated with exhaustion were decreased after Aza treatment. Bisulphite sequencing in CD8 and CD4 T cells of AML mice treated with Aza revealed significant differentially methylated regions after treatment. Aza also led to hypomethylation of promoters for several genes, cytokines and transcription factors significant for T cell function, including TCF-7 and IL-15, indicating these markers may be critical for Aza resistance.
Conclusions
We showed potential ways by which Aza can impact effector T cell function and has the potential to restore T cell exhaustion- which is a key barrier to effective cancer immunotherapy. Ongoing studies focus on combining checkpoint blockade and other drugs that are used clinically in AML and evaluating their effect on exhaustion in patient samples.