Expression analysis of CD8+ T cells following high-avidity or low-avidity T cell receptor (TCR) stimulation in the presence or absence of a DOT1L inhibitor

Adoptive T cell therapy (ACT) is a promising therapeutic approach for cancer patients. The use of allogeneic T cell grafts will improve its applicability and versatility provided that inherent allogeneic responses are controlled. T cell activation is finely regulated by multiple signaling molecules that are transcriptionally controlled by epigenetic mechanisms. Through extensive chemical probe screening, we found that inhibiting DOT1L, a histone H3-lysine 79 methyltransferase, alleviated allogeneic T cell responses. DOT1L inhibition reduced miR-181a expression, which increased the ERK phosphatase DUSP6 expression. The inhibition of DOT1L or ectopic expression of DUSP6 in T cells prevented the development of graft-versus-host disease while retaining potent antitumor activity in multiple ACT models. These results suggest that DOT1L inhibition may enable the safe and effective use of allogeneic antitumor T cells by suppressing unwanted immunological reactions in ACT. To further explore how DOT1L inhibition differentially affects high- and low-avidity stimulation-mediated T cell responses, gene expression profiles of DMF5- (high affinity) or cl.413- (low affinity) TCR-transduced T cells with or without SGC0946 treatment were analyzed following TCR stimulation. Human CD8+ T cells from three different healthy donors were transduced with HLA-A2/MART127-35-specific T cell receptor DMF5 (high affinity) or cl.413 (low affinity) and were cultured in the presence of a DOT1L inhibitor SGC0946 or control DMSO. T cells were then stimulated with artificial antigen presenting cells expressing HLA-A2 loaded with MART127-35 peptide. RNA was collected 24 hours after stimulation and gene expression profiles were analyzed by Affymetrix Human Gene 2.0 ST Array (gene-level analysis).