Fig. 1: CD19 loss is associated with loss of B cell identity.

A CART19 cohort of PDX samples analyzed by CyTOF. Created in BioRender⁸¹. B Median CD19 expression in CR (n = 6), Negative (n = 11), Positive (n = 4), and Refractory (n = 4) pre-CART19 samples. C UMAP based on developmental classifier protein expressions in Lin-/ B+ fraction of healthy BM (left) and projection of tumor cells from pre-CART19 patients that achieved durable CR (n = 6), suffered CD19^neg relapse (n = 11), and post-CD19 loss (n = 8), respectively. IBI Immature-BI, IBII Immature-BII, IBIII Immature-BIII, MBI Mature-BI, MBII Mature-BII, ENBI Early-non-BI, ENBII Early-non-BII, NBIII Non-BIII.. D Developmental classification of samples in (C). The pro-BII population is significantly more abundant in the CR group (CR vs. Negative pre-CART19 p-value: 0.0005; CR vs. Negative post-CART19 p-value: 0.0017), while early-non-BI is significantly more abundant between the pre- and post-CD19^neg relapse groups (p-value = 0.0202). E Median protein expression of CD19, IKAROS, and PAX5 in pro-BII-like tumor cells in CR (n = 6), Negative pre-CART19 (n = 11), and Negative post-CART19 (n = 7) samples. F Cohort of pre-and post-CART19 B-ALL samples analyzed by single-cell CITE-seq. G UMAP based on most variables genes in Lin-/ B+ fraction of healthy BM. Space occupied by healthy clusters is depicted. H Projection of B-ALL cells from 9 samples onto healthy BM-defined UMAP space. I B-ALL cells from CR (n = 3), Negative pre-CART19 (n = 3), and Negative post-CART19 (n = 3) samples were associated with their closest healthy cluster based on k-nearest neighbor assignment. Cluster 6 is significantly less abundant in the Negative pre-CART19 group (Negative pre-CART19 vs. CR p-value < 0.0001; Negative pre-CART19 vs. Negative post-CART19 < 0.0001), while cluster 9 is significantly more abundant in the Negative pre-CART19 group (Negative pre-CART19 vs. CR p-value < 0.0001; Negative pre-CART19 vs. Negative post-CART19 < 0.0001). J–K IKZF1 gene expression (J) and single-cell enrichment score for HSC/MPP gene signature (K) in pro-B-like B-ALL cells (cluster 6). N represents individual patient samples. Patient samples analyzed by mass cytometry or single-cell CITE-seq were not performed in replicates. Boxplots in (B) and (E) extend from the 25th to the 75th percentiles, with a line in the middle representing the median and whiskers extending from the minimum to the maximum values. Curves in (D) and (I) show mean ± SEM. Violin plots in (J) and (K) show the median (solid line) and 25^th and 75^th quantile (dash lines). The statistical tests used were one-way ANOVA followed by Tukey’s multiple comparisons tests (B); two-way ANOVA followed by Tukey’s multiple comparisons tests (D), (E), and (I); and two-sided Wilcoxon rank sum test followed by Bonferroni’s multiple comparisons test in (J) and (K).