Acute myeloid leukemia (AML) is an aggressive and lethal blood cancer. AML cells are particularly vulnerable to disturbed telomere integrity. Imetelstat is a covalently lipidated 13-mer thiophosphoramidate oligonucleotide that competitively inhibits telomerase activity. Imetelstat has shown clinical efficacy in essential thrombocythemia, myelofibrosis, and myelodysplastic syndromes. Here, we show that imetelstat effectively diminishes AML burden and prolongs survival in a comprehensive, randomized Phase II-like preclinical trial in AML patient-derived xenografts (PDX). AML subgroups containing mutations in NRAS and gene expression signatures associated with oxidative stress demonstrate sustained responses to imetelstat. Genome-wide CRISPR/Cas9 editing identifies two key mediators of imetelstat efficacy, late endosomal/lysosomal adaptor, MAPK and MTOR activator 2 (LAMTOR2), and fatty acid desaturase 2 (FADS2). At the molecular level, imetelstat interferes with endogenous G-quadruplex structures, and induces alterations in lipid metabolism leading to lipid peroxidation and increased levels of reactive oxygen species. Pharmacological inhibition of ferroptosis prevents imetelstat-induced cell death. Standard induction chemotherapy induces oxidative stress that sensitizes AML patient samples to imetelstat resulting in significant delay or prevention of AML relapse.