Single-Cell Analyses Unveil a Potential Mechanism Promoting Graft-Versus-Leukemia Response in Haploidentical Transplantation Using Peripheral Stem Cells Plus Cord Blood Grafts through Early Immune Reconstruction of CD8+ Infg+ T Cells

Abstract

Background

Haploidentical donor stem cell transplantation (haplo-SCT) by using G-CSF-primed peripheral stem cells combined with bone marrow grafts (PBSC+BM) represents a widely employed strategy for acute myeloid leukemia (AML). While leukemic relapse is one of the main reasons for the failure of SCT. Surprisingly, our clinical data found that haploidentical peripheral stem cells combined with cord blood (haplo-PBSC+cord) transplantation had significantly lower incidence of relapse than haplo-PBSC+BM transplantation. Nonetheless, the underlying mechanism needs to be explored.

Method

From 2021 to 2022, bone marrow samples at 1 and 3 months post-transplantation from 4 AML patients who underwent haplo-PBSC+cord transplantation and 3 AML patients who received haplo-PBSC+BM transplantation were obtained. We used single-cell RNA-sequence (scRNA-seq) technology for analysis of hematopoietic reconstitution disparity between the PBSC+cord and PBSC+BM groups.

Results

Significant disparities manifest in the initial stages of hematopoietic reconstitution between the PBSC+cord and the PBSC+BM groups. Notably, at 1 month and 3 months post-transplantation, the PBSC+cord group demonstrated conspicuous reconstitution of lymphoid and erythroid cells, with CD8+ T cells exhibiting the most pronounced effect. Differential gene analysis revealed that T cells in the PBSC+cord group display elevated expression of graft-versus-leukemia (GVL)-related cytokines, including IFN-γ, CCL3, and IL1B, when compared to the PBSC+BM group. Furthermore, noteworthy differences in T cell subsets were observed between the two groups. Within 1 month and 3 months post-transplantation, the PBSC+cord group experienced up-regulation of CD8+ effector memory T cells (TEM), characterized by heightened expression of the IFNG gene. Gene Set Variation Analysis (GSVA) unveiled significant enrichment of CD8+ IFNG+ TEM in T cell activation, cytotoxicity, cytokine secretion, and other pathways, suggesting its potential GVL effect. On the contrary, the PBSC+BM group demonstrated a higher proportion of CD4+ T cells, γδT cells, and MAIT cells at 1 month and 3 months post-transplantation. Among CD8+ T cells, the PBSC+BM group showcased a more robust reconstruction of CD8+ TIGIT+ T cell subsets, which highly express exhaustion-related genes like TIGIT, LAG3, PDCD1 (PD-1), HAVCR2 (TIM-3), and exhibited increased apoptosis-related pathway activity.

Conclusion

Our investigation identified a potential mechanism promoting GVL response in the immune reconstitution of haplo-PBSC+cord transplantation.