Single-cell DNA sequencing of samples with co-occurring WT1 and NRAS mutations. (A) Box plot of the NRAS VAF in WT1 and NRAS comutated patients with FLT3-ITD, FLT3-TKD, and patients with FLT3 wild-type. (B-H) Inferred clone relationships derived from tapestri single-cell DNA sequencing. Each oval represents a mutational event. Each shaded circle represents a subclone with accumulated mutational events along its path from the top of the tree. The size of each shaded circle is proportional to the size of the clone population. The first number within each colored circle is the percentage of each clone among the total number of tumor cells included in the single-cell data analysis for that sample and the second number is the absolute number of cells included in that subclone. (B) FLT3-ITD and NRAS mutations arose in mutually exclusive subclones of a WT1 mutated founder clone. The 3 detected FLT3-ITDs may represent a single subclone because of difficulties in mapping the FLT3-ITD reads. A TET2 mutation observed by bulk sequencing was not detected by single-cell sequencing. (C-F) FLT3-TKD and RAS pathway mutations arise in mutually exclusive subclones from a WT1 mutated founder clone. Sample C-05-1893: FLT3-ITD mutation was observed by bulk sequencing but was not detected by single-cell sequencing; sample C-08-2671: FLT3-TKD and PTPN11 mutations were not detected by bulk sequencing. (G-H) Patients with non-FLT3-mutation. In 1 sample, the prominent subclone harbors an NRAS mutation; in the other sample, NRAS was in the prominent founder clone, followed by the accumulation of WT1 mutations in the subclones. Sample C-03-1696: NPM1 was detected with low coverage and DNMT3A mutations were not covered by bulk sequencing.