AKR1B1 is a member of aldo–keto-reductase (AKR) superfamily which catalyze the reduction of carbonyl groups to hydroxyl groups in NADPH-dependent ways. Previous studies have shown that AKR1B1 promotes cancer progression, but its exact role in acute leukemia was unclear. Cell counting and Luminescent Cell Viability Assay were performed to measure the cell proliferation and viability. Soft-Agar Colony Formation (CFU) assay was conducted to measure the capacity of single cells to form colonies in vitro. Cell apoptosis, cell cycle, and cell differentiation were assessed by flow cytometry. Western blotting and RT-qPCR were utilized to examine AKR1B1 expression in acute leukemia cells. In vivo leukemia growth and mouse survival were evaluated using a model of xenotransplantation mice. We explored the AKR1B1 effect and mechanism in acute leukemia cells using RNA-sequencing technology and transcriptomic analysis. AKR1B1 is highly expressed in acute leukemia cells. Knockdown of AKR1B1 inhibited acute leukemia cell proliferation, colony-forming capability, and cell cycle and promoted apoptosis. Additionally, xenograft experiments proved that knockdown of AKR1B1 delayed the progression of acute leukemia cell in vivo. RNA-sequencing data analysis demonstrated that AKR1B1 was involved in the epigenetic silencing of H3K27me3-targeted genes. EZH2 inhibitor UNC1999 combined with knockdown of AKR1B1 showed synergistic inhibitory effect on acute leukemia cells. AKR1B1 is essential for the leukemogenesis and may serve as a potential therapeutic target to treat acute leukemia patients.