CBFA2T3 expression is rapidly induced during relapse of low-CBFA2T3 non-CBF AML patients. (A) Comparison of CBFA2T3 expression levels in matched primary and relapsed samples from 11 cytogenetically normal non-CBF AML patients (GSE66525). (B, left) Scatter plot showing inverse correlation between CBFA2T3 expression changes and its expression levels in primary samples. (B, right) Principal component analysis identified 2 sample clusters, one of which (“Pre-LSC”) comprised all low-CBFA2T3 patients at diagnosis. Green dots denote samples from diagnosis, and red dots denote relapsed samples, with dashed lines defining the 2 samples from a single individual. (C) To quantify the total “change” between each patient’s diagnosis and relapse sample, we calculated the patient-specific Euclidean distance between diagnosis (Dx) and relapsed (Rel) samples using all genes. The Wilcoxon rank sum test showed a significant difference between LSC and pre-LSC groups. The points corresponding to each patient are colored by their CBFA2T3 expression at diagnosis. (D) Expression levels of genes indicated at the top in primary and relapsed samples within the low-CBFA2T3 pre-LSC and high-CBFA2T3 LSC sample clusters. (E) GSEA results showing enriched expression of HSC/LSC and cell cycle genes and depleted expression of IFN-γ and innate immune genes in LSC cluster vs pre-LSC cluster. NES, normalized enrichment score. (F) Comparison of expression levels of NM_005187 and NM_175931 CBFA2T3 transcripts in a single-cell RNA-Seq (scRNA-Seq) dataset profiling human HSPCs expressing different cell surface markers. (G) Comparison of expression changes of NM_005187 and NM_175931 CBFA2T3 transcripts in primary and relapsed AML samples in the GSE83533 dataset.