T-ALL cell lines are highly dependent on de novo pyrimidine synthesis. (A) Volcano plot depicting the 22Q2 CRISPR (DepMap Public+Score, Chronos) genome-wide differential dependency (effect size) vs -log10(false discovery rate [FDR] q value) for T-ALL vs non–T-ALL cell lines. Dots correspond to genes. The DHODH gene is highlighted red. Differential dependency is estimated based on the 2-class comparison method implemented in DepMap (limma empirical Bayes; significance cutoff FDR q ≤ 0.10). (B) Scatter dot plot demonstrating the dependency on DHODH of the T-ALL, B-ALL, and AML cell lines in the CRISPR (DepMap 22Q2 Public+Score, Chronos) screen for 1086 cell lines. Dots represent cell lines. The 5 T-ALL cell lines in the screen are highlighted red, the 12 B-ALL cell lines are highlighted black, and the 26 AML cell lines are highlighted blue. The x-axis shows the DHODH Chronos dependency score (gene effect) per cell line. The y-axis shows the probability of dependency on DHODH per cell line. Dependency on DHODH is estimated by low negative (≤−0.5) Chronos gene effect scores. (C) Schematic outline of the de novo pyrimidine synthesis pathway and the role of DHODH, which is located in the inner mitochondrial membrane. Also shown is the location where uridine feeds into the pathway. (D) CRISPR KO of DHODH was performed in the Jurkat cell line. Parental and KO cells were seeded at equivalent densities, and supplemental uridine was withdrawn, at the time when DHODH-KO cells rapidly died while the parental cells continued to grow rapidly. dCTP, deoxycytosine triphosphate; dTMP, deoxythymidine 5′-monophosphate; dTTP, deoxythymidine 5′-triphosphate; dUDP, deoxyuridine diphosphate; dUMP, deoxyuridine monophosphate; dUTP, deoxyuridine triphosphate; GlcNAc, N-acetylglucosamine; OMP, orotidylate monophosphate.