T-cell PD1 expression and chimerism in the marrow after transplant. (A) Day 100 PD1 expression on marrow and blood CD4 T cells, stratified by disease relapse status and compared using the Mann-Whitney test. PD1 analysis for conventional CD4 T cells and regulatory CD4 T cells included in supplemental Figure 3A. The analysis included only samples obtained before the occurrence of relapse diagnosis. ∗P < .05. (B) UMAP visualization of single-cell RNA sequencing of day 100 posttransplant bone marrow aspirates from patient 9 (n = 20 943 cells) and patient 26 (n = 16 131 cells), showing principal CD45⁺ cell-type clusters. (C) Single-cell chimerism analysis of marrow cells from patient 26. Violin plots showing gene expression (log-normalized counts) of XIST (female donor cells) and RPS4Y1 (male host cells). Owing to dropout, further characterization was performed using Souporcell¹⁵ to cluster genotypes by single-nucleotide variants, with proportions of host contributions to the major lymphocyte and myeloid compartments shown in the bar plot in panel D. CD4 T cells are separated into conventional CD4 T cells and Tregs. pDCs, plasmacytoid dendritic cells; UMAP, uniform manifold approximation and projection.