Fig 1: LZTR1-mediated drug resistance can be overcome by modulating RAS-GTP abundance. A, Scatter plots of RNA expression levels [shown as normalized reads per kilobase of transcript per million reads mapped (RPKM)] plotted against drug-sensitivity measured ex vivo as the area under the curve (AUC) from AML patient samples harboring FLT3-ITD mutation. n = 111. *, P < 0.05. Pearson correlation coefficients are shown as (r) values. Statistical significance is indicated. B, Schema of the MOLM-13 cell line xenograft experiment. Luciferase-expressing MOLM-13 cells treated with anti-LZTR1 or control sgRNAs were systemically engrafted with 1 × 105 cells/animal via a tail-vein injection after sublethal (225 cGy) irradiation. Seven days later, animals were treated with either vehicle or 30 mg/kg/day of gilteritinib via oral gavage. Animals underwent bioluminescence imaging weekly. C, Kaplan–Meier curve of experiment in B. n = 7–10. **, P < 0.01; ***, P < 0.001. D, Schema of protein-based degrader of RAS proteins. A chimeric protein consisting of a high-affinity target-binding domain for RAS proteins (DARPin K27) is fused to an engineered E3 ligase adapter (SPOP) to confer ubiquitin-mediated degradation of RAS proteins. E, Western blot demonstrating levels of LZTR1 as well as total and GTP-bound KRAS/NRAS/MRAS and RIT1 in MOLM-13 cells ± LZTR1 KO upon doxycycline-mediated induction of the RAS biodegrader (performed in biological triplicate). EV, empty vector. F, Seventy-two-hour cell proliferation assay on parental and LZTR1 KO MOLM-13 cells ± induction of RAS degradation. Cell viability was measured in triplicate using CellTiter-Glo. G, 2D synergy plots using the Zero Interaction Potency (ZIP) model of control sgRNA (“WT”) or anti-LZTR1 sgRNA (“LZTR1 KO”) MOLM-13 cells treated for 72 hours with BI-3406 and/or gilteritinib at various concentrations. Western blot demonstrating p-MEK, p-ERK, and total MEK, ERK, KRAS/NRAS/MRAS, and RIT1 levels (H) as well as RAS-GTP levels (I) in MOLM-13 cells ± LZTR1 KO treated with increasing concentration of DMSO or BI-3406 alone 4 hours after drug treatment.