SIRT3 overexpression and epigenetic silencing of catalase regulate ROS accumulation in CLL cells activating AXL signaling axis

Mitochondrial metabolism is the key source for abundant ROS in chronic lymphocytic leukemia (CLL) cells. Here, we detected significantly lower superoxide anion (O₂⁻) levels with increased accumulation of hydrogen peroxide (H₂O₂) in CLL cells vs. normal B-cells. Further analysis indicated that mitochondrial superoxide dismutase (SOD)2, which converts O₂⁻ into H₂O₂ remained deacetylated in CLL cells due to SIRT3 overexpression resulting its constitutive activation. In addition, catalase expression was also reduced in CLL cells suggesting impairment of H₂O₂-conversion into water and O₂ which may cause H₂O₂-accumulation. Importantly, we identified two CpG-islands in the catalase promoter and discovered that while the distal CpG-island (−3619 to −3765) remained methylated in both normal B-cells and CLL cells, variable degrees of methylation were discernible in the proximal CpG-island (−174 to −332) only in CLL cells. Finally, treatment of CLL cells with a demethylating agent increased catalase mRNA levels. Functionally, ROS accumulation in CLL cells activated the AXL survival axis while upregulated SIRT3, suggesting that CLL cells rapidly remove highly reactive O₂⁻ to avoid its cytotoxic effect but maintain increased H₂O₂-level to promote cell survival. Therefore, abrogation of aberrantly activated cell survival pathways using antioxidants can be an effective intervention in CLL therapy in combination with conventional agents.