Abstract Acute myeloid leukemia (AML) is an aggressive blood cancer with high rates of relapse and limited second-line treatment options, resulting in a five-year survival of 20-30%. A defining feature of AML cells is their prominent nucleoli, which demarcate sites of active ribosomal RNA (rRNA) synthesis. rRNAs are specialized non-coding RNAs that form the catalytic cores of ribosomes, and their transcription by RNA polymerase 1 (Pol1) is the rate-determining step in ribosome biogenesis. We hypothesized that AML cells require increased rRNA synthesis and protein translational capacities to support their growth and survival. To test this premise, we developed a “FISH-Flow” assay to measure the relative levels of nascent and mature rRNAs in different cell-types across the human hematopoietic tree. Profiling over twenty bone marrow aspirates from healthy donors and AML patients, we found that rRNA synthesis varies widely throughout normal hematopoiesis – peaking in the stem/progenitor cell populations (CMPs, GMPs, MEPs) – and is elevated by 1.5 to 2-fold in AML cells. Our analysis of publicly available single-cell RNA-seq data further showed that the Pol1 gene signature is overexpressed in AML cells. To assess whether rRNA “hyper-synthesis” is essential for AML cell fitness, we generated a HoxA9 transgene-driven mouse AML cell-line with POLR1A (the catalytic subunit of Pol1) fused to an FKBP degron domain, which enables targeted degradation of Pol1 by a small molecule dTAG. Pol1 depletion led to a rapid decrease in rRNA synthesis, G1 cell cycle arrest, and monocytic differentiation, as evidenced by downregulation of the stem cell marker c-Kit and upregulation of the mature monocyte marker Ly6C. Additionally, dTAG treatment synergized strongly with Bcl-2 inhibitors to induce apoptosis in these cells. This synergy did not extend to other AML chemotherapy agents, such as daunorubicin or cytarabine, indicating a unique, synthetically lethal interaction between the rRNA synthesis and Bcl-2 signaling pathways in AML cells. In summary, we demonstrate that rRNA synthesis and Pol1 expression are amplified in AML cells to sustain an oncogenic, stem-like state that renders them more resistant to apoptotic stimuli. Notably, our findings identify rRNA synthesis as a therapeutic vulnerability in AML cells and uncover a novel synthetic lethality with Bcl-2 inhibition. Citation Format: Putzer J. Hung, Jill A. Henrich, Charles Antony, Vikram R. Paralkar, . Regulation of ribosomal RNA synthesis in acute myeloid leukemia abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 1365.
Hung et al. (Fri,) studied this question.