Disrupting tRNA modifications to target mitochondrial vulnerabilities in drug-resistant leukemia cells Open Access

• TRMT5-mediated m1G modifications in tRNAs drive mitochondrial function and drug tolerance in AML via OXPHOS up-regulation.

• TRMT5 inhibition prevents OXPHOS up-regulation and synergises with cytarabine and venetoclax to overcome resistance.

Dysregulated RNA modifications contribute to cancer progression and therapy resistance, yet the underlying mechanism often remains unknown. Here, we perform CRISPR-based synthetic lethality screens to systematically explore the role of RNA modifications in mediating resistance to anti-leukaemic drugs. We identify the TRMT5-mediated formation of N1-methylguanosine (m1G) in the tRNA anticodon loop as essential for mediating drug tolerance to cytarabine and venetoclax in acute myeloid leukemia (AML). TRMT5 methylates nearly all mitochondrial and nuclear tRNAs with a guanosine at position 37, but its role in promoting drug tolerance specifically depends on its mitochondrial function. TRMT5 is essential for the dynamic upregulation of mitochondrial mRNA translation and oxidative phosphorylation (OXPHOS), which are critical for sustaining drug tolerance in leukemia cells. This mitochondrial dependency correlates with therapy outcomes in leukemia patients: lower expression of electron transport chain genes is linked to poorer outcomes in a cohort of nearly 100 AML patients undergoing first induction therapy. Finally, we demonstrate that targeted depletion of TRMT5 protein using a conditional degron, in conjunction with cytarabine and venetoclax treatment, synergistically induces cell death in drug-tolerant AML cells. Thus, our study reveals TRMT5 as promising drug target for therapy-resistant leukemia.