Whole-genome CRISPR/Cas9 screen identifies cell cycle regulation as a vulnerability in HMGA2-expressing cells. (A) Western blot validation of HMGA2 protein expression in Cas9 OCI-AML5 15 days after infection with HMGA2-YFP and YFP vectors. (B) After infection with HMGA2-YFP– and YFP-expressing vectors, the percentage of YFP-expressing cells was monitored by flow cytometry during the course of CRISPR/Cas9 whole-genome screen. (C) Schematic overview of the CRISPR/Cas9 whole-genome screen performed in Cas9 OCI-AML5 cells containing the Extended Knockout sgRNA library. (D) Volcano plot representing results of whole-genome CRISPR/Cas9 screen performed in OCI-AML5 cells overexpressing HMGA2 compared with YFP control vector. Most significant genes belonging to the Gene Ontology (GO) terms 1901987 (regulation of cell cycle phase transition, P value = 5.0E-16, false discovery rate [FDR] q-value = 1.1E-13) and 0006281 (DNA repair, P value = 1.2E-13, FDR q-value = 1.9E-11) that enriched in synthetic lethal interaction are depicted. ATR, CHEK1, and WEE1 are also depicted. Asterisks indicate genes belonging to both GO terms. (E) Nonexhaustive list of significant GO terms associated with synthetic lethality in cells overexpressing HMGA2 compared with YFP control cells. (F) Gene Set Enrichment Analysis plot comparing HMGA2 high (RPKM >2; n = 39) vs HMGA2 null (RPKM = 0; n = 83) transcriptomic signatures in primary AML specimens. Results obtained for G2/M checkpoint hallmark gene set. DOX, doxycycline; ER, endoplasmic reticulum; NES, normalized enrichment score.