Supplementary Figure 11: Characterization of leukemogenic evolution in the patient cohort.

(a) Representative sorting strategy for HSCs (green), LSCs (yellow), and blasts (orange) from a single patient with AML (SU353). HSCs are characterized immunophenotypically as CD34⁺CD38⁻CD99⁻TIM3⁻. LSCs are characterized immunophenotypically as CD34⁺CD38⁻CD99⁺TIM3⁺. Blasts are characterized as CD99⁺TIM3⁺CD45^midSSC^high. Depending on the sample, blasts can be CD34⁺ or CD34⁻. (b) Preleukemic HSC purification strategies for all patients with AML analyzed by ATAC-seq and RNA-seq. Healthy bone marrow after CD34 magnetic bead enrichment is included as a normal control. (c) Mutation frequencies observed in the full patient cohort (Fig. 5a; n = 39). This cohort is biased toward samples harboring internal tandem duplications in FLT3. This lesion leads to a higher white blood cell count and, therefore, more cells to work with, making these samples more attractive. (d) Variant allele frequencies in HSCs isolated from patients with AML (n = 39) as determined by targeted amplicon sequencing to depth >500 reads. *P < 0.05, **P < 0.01, χ² test. (e,f) Principal-component analysis of RNA-seq data (e) and ATAC-seq data (f) showing the spread of AML cell types across the myelopoietic continuum. The position of each normal hematopoietic cell type is shown using cartoon cells. These normal hematopoietic cell type positions represent the average for all biological and technical replicates.