Selectively targeting the malignant clone through the cellular redox state

Antioxidant compounds that can mimic the activity of endogenous enzymes via redox cycling offer a novel method of targeted cancer treatment. We investigated the therapeutic effect of a mitochondrial- targeted, MnSOD mimetic, Mn(III) mesotetrakis (N-n-butoxyethylpyridinium-2yl) porphyrin, (MnTnBuOE) within myeloid lineage hematopoietic malignancies and utilize the hematopoietic disorder myelodysplastic syndrome (MDS) for the presentation of mechanistic insights to MnTnBuOE efficacy. Consistent with our previous report, MnTnBuOE treatment offers protection of normal HSCs demonstrated by an increase in both CD34+CD38- and CD34+CD38+ stem/progenitor cell viability. However, similar treatment shows significant cytotoxicity mediated by MnTnBuOE induced hydrogen peroxide within acute myeloid leukemia (AML) and MDS models. Transcriptional array analysis indicates that MnTnBuOE treatment initiates activator protein 1 (AP-1) activity. DNA mediated pull down and immunoprecipitation identifies JunB as a transcriptional mediator of MnTnBuOE induced cytotoxicity. Stable knock down of JunB provides protection against MnTnBuOE induced cytotoxicity confirming JunB’s critical role in MnTnBuOE induced hematopoietic malignant cell death. Precipitation with biotin-1,3-cyclopentadione combined with LC-MS/MS analysis of JunB identified Cys285 as the critical cysteine within the DNA binding region of JunB that is susceptible to oxidation by hydrogen peroxide. This, in concert with co-immunoprecipitation of JunB placed Ref-1 as a protein specifically involved in the redox-modification of JunB. Lentiviral mediated knock down of Ref-1 indicates that the interaction between Ref-1 and JunB is critical for MnTnBuOE induced cytotoxicity. The AP-1 ability to bind consensus promoter sequence regardless of methylation status, indicates MnTnBuOE mediated activation of JunB as a potential alternative therapy for aggressive or chemoresistant malignancies in a p53 independent manner. The finding that MnTnBuOE can exert selective cytotoxicity in malignant cells via redox-dependent differential activation of molecular pathways controlling cell fate will significantly aid further development of safe, effective redox-active therapeutics to prevent normal tissue injury and enhance chemotherapeutic efficacy.