Epigenetic and phenotypic signatures of T-cell response to blinatumomab in pediatric relapsed and refractory B-ALL.

Abstract

2528 Background: By forming an immunological synapse between T cells and tumor antigen, bispecific T cell engagers (BiTEs) like blinatumomab have shown great promise in treating B-cell acute lymphoblastic leukemia (B-ALL). However, many relapsed and refractory (R/R) patients fail to achieve long-term survival, with 40% not surviving past 24 months. Prolonged T cell activation with blinatumomab therapy may lead to changes in differentiation that leave the T cell population unable to elicit a sustained anti-tumor response. A deeper understanding of the dynamics of the T cell compartment in R/R B-ALL patients will lead to improved treatment strategies and optimized patient selection for blinatumomab therapy. Methods: To characterize T cell persistence and response in this context, we assessed memory and exhaustion phenotypes in blinatumomab-treated T cells isolated from 10 R/R pediatric B-ALL patients treated with blinatumomab. CD8+ T cells were isolated from peripheral blood and bone marrow samples and analyzed for memory and exhaustion phenotypes via flow cytometry. Absolute lymphocyte counts were measured and linked to the sample flow cytometry data to assess expansion and contraction of T cell memory subsets throughout the course of therapy. Whole genome enzymatic methyl sequencing was performed on post-treatment PD-1 High and PD-1 Low CD8 T cells to determine the multipotency of the patient T cell compartment after blinatumomab treatment. Results: After 7 days of continued blinatumomab infusion, patient T cells demonstrated a significant expansion of terminally differentiated and effector memory T cells. Notably, we observed that non-responders had a high tumor burden at the start of the therapy and possessed a large population of naïve CD8 T cells that failed to expand. These CD8 T cells exhibited a significant increase in expression of TIM-3 and PD-1 compared to responders after the 7-day infusion. Methylation analysis of post-treatment CD8 T cells showed decreased methylation of exhaustion regulators IKZF1 and CD300a in non-responders compared to the responders. Additionally, in vitro treatment of T cells with blinatumomab induced T-cell exhaustion in a target-dependent manner. Conclusions: Blinatumomab therapy in pediatric B-ALL patients induced variable epigenetic and phenotypic changes to the T cell compartment indicative of exhaustion, corresponding to differences in T cell expansion and persistence between patients. Our study is the first to link epigenetic changes in exhaustion regulators with response variability in blinatumomab-treated patients. Furthermore, our findings highlight a potential role of baseline T cell composition and tumor burden in determining therapeutic outcomes. These insights provide a novel framework for improving patient stratification and treatment strategies to mitigate T cell exhaustion in blinatumomab therapy.