Functional heterogeneity in ETP-ALL reveals deranged developmental hierarchy with coexisting stem-like states and ineffectual lineage commitment. (A) Heatmap demonstrates expression of HSC, MPP, CMP, GMP, CLP, ETP, DN1, DN2, DN2-3, DN3A, DN3B, DN3-4, DN4, and DP signatures as defined by xcell²³  in individual leukemic cells. (B) t-SNE plot of all malignant cells using genes involved in HSC, CLP, and CMP progenitor programs. Clusters are derived using the Louvain algorithm. (C) RNA velocities projected on the t-SNE plot containing leukemic cells with root and endpoint cells highlighted (circles). (D) Heatmap depicting the expression of marker genes in the identified root and endpoint cells. (E) Violin plots depicting expression of key marker genes in root cells (top) and endpoint cells (bottom). Gene set enrichment analysis plots depicting the enrichment of HSC signature in root cells (left), differentiating T-lymphocyte signature (middle), and interferon-γ response (right) in endpoint cells. (F) t-SNE plot of leukemic cells colored based on the predicted cell-cycle phase (left), and relative percentage of cell-cycle phase in stacked bar plot for roots and endpoint states (right). (G) Heatmap depicting clustered transcriptional regulons (predicted transcription factor activity based on target gene expression) in root and endpoint cells (“Materials and methods”). (H-I) Heatmaps demonstrate expression of HSC, MPP, CMP, GMP, CLP, ETP, DN1, DN2, DN2-3, DN3A, DN3B, DN3-4, DN4, and DP signatures in ETP-ALL and T-ALL PDX models. *NOTCH1 mutated. #PTEN deleted.