T- and B-Cell Subset Reconstitution As Predictor of Relapse in Pediatric and Young Adult Allo-Hematopoietic Cell Transplantation Patients

Background:

Immune reconstitution (IR) following allogeneic hematopoietic cell transplantation (allo-HCT), particularly involving CD4+ T cells and B cells, is associated with improved outcomes, including decreased non-relapse mortality, reduced incidence of graft-versus-host disease, and better overall and event-free survival. Despite these positive associations, a reliable cellular predictor for relapse following HCT has not yet been identified. Identifying such predictors is critical for improving post-transplant monitoring and intervention strategies. This analysis investigates the relationship between the reconstitution of lymphocyte subsets and relapse in pediatric and young adult patients with hematological malignancies, aiming to identify potential predictors of relapse and enhance the understanding of post-transplant immune dynamics.

Methods:

Standard flow cytometric immunophenotyping was performed over two years on patients who received allo-HCT for malignant indications (n=74) between 2020 and 2023. The study examined the reconstitution of immune cells, specifically B (CD19+), T (CD3+), NK (CD16+CD56+) and NKT (CD3+CD56+) cells. Additionally, within the CD4+ and CD8+ T cell compartments, various T cell subsets were assessed, including regulatory (Treg; CD3+CD4+CD25+CD127low), naïve (TN; CD45RO-CD197+), central memory (TCM; CD45RO+CD197+), effector memory (TEM; CD45RO+CD197-), terminal effector (TTE; CD45RO-CD197-), and activated (HLA-DR+) T cells. Pre-relapse cell populations were compared to those of the non-relapsing patients across standardized post-HCT timepoints. Any subset exhibiting a divergent trend between the two groups at any timepoint was evaluated for statistical significance using Student's t-test with a false discovery rate (FDR) threshold of p<0.05 after adjustment via the Benjamini-Hochburg method.

Results:

Relapse occurred in 18% of patients (n=13), with a median time to relapse of 270 days post-HCT. Of the 25 patients who received bone marrow (BM, 34%), 4 relapsed (16%); among the 20 patients who received cord blood (CB, 27%), 6 relapsed (30%); none of the 3 patients who received peripheral blood stem cells (PBSC, 4%) relapsed; and of the 26 patients who received T cell-depleted grafts (TCD, 35%), 3 relapsed (12%). Importantly, all cord blood recipients with B cell levels exceeding 50% of total lymphocytes at day 100 subsequently relapsed (n=4). 12 patients had previously undergone HCT (16%), 1 of which relapsed (8%). There were no significant differences in total CD4+ or CD8+ T cell populations between the relapse and non-relapse groups at any timepoint. However, after day 100, CD4+ and CD8+ TEM cell levels were reduced in patients who relapsed (p= 0.035; 0.03). Additionally, NKT cell counts were diminished post day 100 in patients who later relapsed (p=0.00004). After day 200 post-HCT, CD4+ TN, CD8+ TN, and CD4+ TCM central memory cell levels were decreased in patients pre-relapse (p=0.034; 0.0008; and 0.0128, respectively). Although Treg counts were not significantly different between groups, they consistently trended lower in the relapse group.

Conclusions:

The findings indicate that specific lymphocyte subsets, including elevated B cell levels between days 50 and 100 in cord blood recipients, and reduced TEM, TN, and TCM T cell levels, as well as diminished NKT cell counts post day 100, are associated with relapse in pediatric and young adult patients post-allo-HCT. These findings are consistent with previous studies that identified CD4+ and B-cell reconstitution as significant predictors of outcomes, supporting the role of immune cell subsets in relapse risk. This suggests a potential utility of monitoring these lymphocyte subsets as biomarkers for relapse risk. Due to the limited occurrence of relapse in this cohort, future work should focus on validating these findings in larger cohorts, allowing for more thorough analysis of known covariates of relapse, including disease risk index, CR status, and minimal residual disease status. Such studies must continue exploring the underlying immunologic mechanisms of relapse, aiming to refine prognostic tools and therapeutic strategies for relapse prevention in allo-HCT recipients.